Amino Acids: Origin of the canonical twenty  amino acids required for life

Amino acid synthesis requires solutions to four key biochemical problems

1. Nitrogen fixation
Nitrogen is an essential component of amino acids. Earth has an abundant supply of nitrogen, but it is primarily in the form of atmospheric nitrogen gas (N2), a remarkably inert molecule. Thus, a fundamental problem for biological systems is to obtain nitrogen in a more usable form. This problem is solved by certain microorganisms capable of reducing the inert N = N triple-bond molecule of nitrogen gas to two molecules of ammonia in one of the most amazing reactions in biochemistry. Nitrogen in the form of ammonia is the source of nitrogen for all the amino acids. The carbon backbones come from the glycolytic pathway, the pentose phosphate pathway, or the citric acid cycle.


2. Selection of the 20 canonical bioactive amino acids
Why are 20 amino acids used to make proteins ( in some rare cases, 22) ?  Why not more or less ? And why especially the ones that are used amongst hundreds available? In a progression of the first papers published in 2006, which gave a rather shy or vague explanation, in 2017, the new findings are nothing short than astounding.  In January 2017, the paper : Frozen, but no accident – why the 20 standard amino acids were selected, reported:

" Amino acids were selected to enable the formation of soluble structures with close-packed cores, allowing the presence of ordered binding pockets. Factors to take into account when assessing why a particular amino acid might be used include its component atoms, functional groups, biosynthetic cost, use in a protein core or on the surface, solubility and stability. Applying these criteria to the 20 standard amino acids, and considering some other simple alternatives that are not used, we find that there are excellent reasons for the selection of every amino acid. Rather than being a frozen accident, the set of amino acids selected appears to be near ideal. Why the particular 20 amino acids were selected to be encoded by the Genetic Code remains a puzzle."

3. Homochirality
In amino acid production, we encounter an important problem in biosynthesis—namely, stereochemical control. Because all amino acids except glycine are chiral, biosynthetic pathways must generate the correct isomer with high fidelity. In each of the 19 pathways for the generation of chiral amino acids, the stereochemistry at the a -carbon atom is established by a transamination reaction that includes pyridoxal phosphate (PLP) by transaminase enzymes, which however were not extant on a prebiotic earth, which creates an unpenetrable origin of life problem. One of the greatest challenges of modern science is to understand the origin of the homochirality of life: why are most essential biological building blocks present in only one handedness, such as L-amino acids and D-sugars ?


4. Amino acid synthesis regulation
Biosynthetic pathways are often highly regulated such that building blocks are synthesized only when supplies are low. Very often, a high concentration of the final product of a pathway inhibits the activity of allosteric enzymes ( enzymes that use cofactors ) that function early in the pathway to control the committed step. These enzymes are similar in functional properties to aspartate transcarbamoylase and its regulators. Feedback and allosteric mechanisms ensure that all 20 amino acids are maintained in sufficient amounts for protein synthesis and other processes.

Glycine: 
Phosphoglycerate dehydrogenase
http://en.wikipedia.org/wiki/Phosphoglycerate_dehydrogenase
Phosphoserine transaminase
http://en.wikipedia.org/wiki/Phosphoserine_transaminase
Phosphoserine phosphatase
http://en.wikipedia.org/wiki/Phosphoserine_phosphatase
Serine hydroxymethyltransferase
http://en.wikipedia.org/wiki/Serine_hydroxymethyltransferase

Valine
Acetolactate synthase enzyme
http://www.ebi.ac.uk/interpro/entry/IPR004789
Acetohydroxy acid isomeroreductase
http://www.ebi.ac.uk/interpro/entry/IPR013023
Dihydroxy-acid dehydratase
http://en.wikipedia.org/wiki/Dihydroxy-acid_dehydratase

Leucine and Isoleucine
Acetolactate synthase
https://en.wikipedia.org/wiki/Acetolactate_synthase
2-isopropylmalate synthase
https://en.wikipedia.org/wiki/2-isopropylmalate_synthase
3-Isopropylmalate dehydratase
https://en.wikipedia.org/wiki/3-Isopropylmalate_dehydratase
3-Isopropylmalate dehydrogenase
https://en.wikipedia.org/wiki/3-Isopropylmalate_dehydrogenase
Transaminase
https://en.wikipedia.org/wiki/Transaminase

Tryptophan and the riddle of making amino acids by hypercomplex enzymatic reactions

https://reasonandscience.catsboard.com/t1740p25-amino-acids-origin-of-the-canonical-twenty-amino-acids-required-for-life#6700

If you can't make a brick, you can't make a house. Naturalistic scenarios are all based on ad-hoc anecdotal pseudo-scientific claims. Without having the right twenty left-handed amino acids,  life essential proteins cannot be made, nor protein complexes, nor biological cells. The pathway of tryptophan synthesis is perhaps the most thoroughly studied of any biosynthetic sequence, particularly in terms of its genetic organization and expression.

Never, in any simulated Origin of Life experiment, the amino acid Tryptophan was synthesized. Why? The biosynthesis pathway to make tryptophan is the most biochemically expensive and most complicated process of all life essential amino acid pathways, and tightly regulated. Glucose feeds the Glycolysis pathway, which utilizes nine enzymatic steps and enzymes to produce a subproduct, which enters the Shikimake pathway, which uses another seven enzymes, to make chorismate, which enters the Tryptophan biosynthesis pathway, and after another five steps and enzymes, finally produces Tryptophan. So, in total, 21 enzymes. 

Some enzymes in the pathway are highly sophisticated, veritable multienzyme nanomachines, like a paper called the bacterial tryptophan synthase, which channels the substrates through a long interconnecting tunnel with a clear logic: the substrate is not lost from the enzyme complex and diluted in the surrounding milieu. This phenomenon of direct transfer of enzyme-bound metabolic intermediates, or tunnelling, increases the efficiency of the overall pathway by preventing loss and dilution of the intermediate. Smart, hah ??!!

You can have a closer look at the entire pathway to make tryptophan here:
https://reasonandscience.catsboard.com/t1740-origin-of-the-canonical-twenty-amino-acids-required-for-life#5939

Now let me tell you why this is so relevant. Tryptophan is the substrate, used to make one of the most important electron carriers used in life: NAD:

Nicotinamide adenine dinucleotide (NAD) in origin of life scenarios
https://reasonandscience.catsboard.com/t2708-nicotinamide-adenine-dinucleotide-nad-in-origin-of-life-scenarios#6060

NAD is pivotal for cell life, first as a reusable redox coenzyme for energy production, second as a consumable substrate in enzymatic reactions regulating crucial biological processes, including gene expression, DNA repair, cell death and lifespan, calcium signaling, glucose homeostasis, and circadian rhythms.  (NADPH) is an essential electron donor in all organisms. It provides the reducing power that drives numerous anabolic reactions, including those responsible for the biosynthesis of all major cell components. It is also the driving force of most biosynthetic enzymatic reactions, including those responsible for the biosynthesis of all major cell components, such as DNA and lipids. The consumption of NADP+ is connected to the consumption of NAD+ and to the regulation of various major biological activities such as DNA repair, gene expression, apoptosis, nitrogen fixation, and calcium homeostasis.  (NADPH) is an essential electron donor in all organisms. It provides the reducing power that drives numerous anabolic reactions, including those responsible for the biosynthesis of all major cell components.

Free-living bacteria synthesize tryptophan (Trp), which is an essential amino acid for mammals, some plants, and lower eukaryotes. The Trp synthesis pathway appears to be highly conserved, and the enzymes needed to synthesize tryptophan are widely distributed across the three domains of life. This pathway is one of three that compose aromatic amino acids from chorismate.   As another point of distinction, the Trp pathway is the most biochemically expensive of the amino acid pathways, and for this reason, it is expected to be tightly regulated.   

Tryptophan (Trp) biosynthesis is a biologically expensive, complicated process. In fact, the products of four other pathways are essential contributors to carbon or nitrogen during tryptophan formation. Thus, the principal pathway precursor, chorismate, is also the precursor of the other aromatic amino acids, phenylalanine and tyrosine, as well as serving as the precursor of aminobenzoic acid and several other metabolites. 

Chorismate biosynthesis occurs via the shikimate pathway: 

1. DAHP synthase
2. 3-dehydroquinate synthase
3. 5-Dehydroquinate Synthetase
4. Shikimate dehydrogenase
5. Shikimate kinase
6. 3-enolpyruvylshikimate-5-phosphate synthase
7. Chorismate synthase

The Aromatic Amino Acids Are Synthesized from Chorismate
Indeed, chorismate is common to the synthesis of cellular compounds having benzene rings, including these amino acids, the fat-soluble vitamins E and K, folic acid, and coenzyme Q and plastoquinone (the two quinones necessary to electron transport during respiration and photosynthesis, respectively). Lignin, a polymer of nine-carbon aromatic units, is also a derivative of chorismate. Lignin and related compounds can account for as much as 35% of the dry weight of higher plants; clearly, enormous amounts of carbon pass through the chorismate biosynthetic pathway.

Enzymes and reactions in the Tryptophan biosynthesis pathway:
Tryptophan is synthesized in five steps from chorismate. Each step requires a specific enzyme activity. The four intermediates between chorismate and tryptophan serve no function other than as precursors of tryptophan.  

That means, these enzymes have no other purpose, and could not have been co-opted from other synthesis pathways. Their exclusive and necessity in intermediate stages make them irreducible, and the pathway as a whole is irreducibly complex. 

6. Anthranilate synthase
7. Anthranilate phosphoribosyltransferase
8. Phosphoribosylanthranilate isomerase
9. Indole-3-glycerol-phosphate synthase
10 + 11. Tryptophan synthase

Origin of Tryptophan biosynthesis and abiotic scenarios
Aromatic amino acid biosynthesis proceeds via a long series of reactions, most of them concerned with the formation of the aromatic ring before branching into the specific routes to phenylalanine, tyrosine, and tryptophan. Chorismate, the common intermediate of the three aromatic amino acids, is derived in eight steps from erythrose-4-phosphate and phosphoenolpyruvate.

How Structure Arose in the Primordial Soup
Tryptophan, in comparison, has a complex structure and is comparatively rare in the protein code, like a Y or Z, leading scientists to theorize that it was one of the latest additions to the code. 6

How Structure Arose in the Primordial Soup
May 19, 2015
In research published online in March in the Journal of Molecular Evolution, Fournier showed that the last chemical letter added to the code was a molecule called tryptophan 2

Ancestral Reconstruction of a Pre-LUCA Aminoacyl-tRNA Synthetase Ancestor Supports the Late Addition of Trp to the Genetic Code.1 
2015 Mar 20
The genetic code was likely complete in its current form by the time of the last universal common ancestor (LUCA). Several scenarios have been proposed for explaining the code’s pre-LUCA emergence and expansion, and the relative order of the appearance of amino acids used in translation. One co-evolutionary model of genetic code expansion proposes that at least some amino acids were added to the code by the ancient divergence of aminoacyl-tRNA synthetase (aaRS) families. 

It is important to remark that secular science puts the origin and appearance of amino acids prior to LUCA. That said, it's clear that any allusion to evolution is prohibited by the fact that there was no evolution prior to DNA replication.

1. https://www.ncbi.nlm.nih.gov/pubmed/25791872
2. https://www.scientificamerican.com/article/how-structure-arose-in-the-primordial-soup/
3. http://2009.igem.org/wiki/images/a/a7/The_tryptophan_biosynthetic_pathway.pdf
4. http://sci-hub.tw/https://www.ncbi.nlm.nih.gov/pubmed/18486479
5. Origins of Life on the Earth and in the Cosmos SECOND EDITION, page 117
6. https://www.scientificamerican.com/article/how-structure-arose-in-the-primordial-soup/
7. https://www.nature.com/scitable/topicpage/an-evolutionary-perspective-on-amino-acids-14568445
8. https://en.wikipedia.org/wiki/Tryptophan

Tryptophan

Enzymes and reactions in the Tryptophan biosynthesis pathway:

Tryptophan is synthesized in five steps from chorismate. Each step requires a specific enzyme activity. The four intermediates between chorismate and tryptophan serve no function other than as precursors of tryptophan. these enzymes have no other purpose, and could not have been co-opted from other synthesis pathways. Their exclusive and necessity in intermediate stages make them irreducible, and the pathway as a whole is irreducibly complex.Enzymes and reactions in the Tryptophan biosynthesis pathway:

1. DAHP synthase
2. 3-dehydroquinate synthase
3. 5-Dehydroquinate Synthetase
4. Shikimate dehydrogenase
5. Shikimate kinase
6. 3-enolpyruvylshikimate-5-phosphate synthase
7. Chorismate synthase8. Anthranilate synthase
9. Anthranilate phosphoribosyltransferase
10. Phosphoribosylanthranilate isomerase
11. Indole-3-glycerol-phosphate synthase
12 + 11. Tryptophan synthase

The bacterial tryptophan synthases are multienzyme nanomachines that catalyze the last two steps in L-tryptophan biosynthesis. 4

Aromatic amino acid biosynthesis proceeds via a long series of reactions, most of them concerned with the formation of the aromatic ring before branching into the specific routes to phenylalanine, tyrosine, and tryptophan. Chorismate, the common intermediate of the three aromatic amino acids, is derived in eight steps from erythrose-4-phosphate and phosphoenolpyruvate.

It is important to remark that secular science puts the origin and appearance of amino acids prior to LUCA. That said, it's clear that any allusion to evolution is prohibited by the fact that there was no evolution prior to DNA replication.

1. https://www.ncbi.nlm.nih.gov/pubmed/25791872
2. https://www.scientificamerican.com/article/how-structure-arose-in-the-primordial-soup/
3. http://2009.igem.org/wiki/images/a/a7/The_tryptophan_biosynthetic_pathway.pdf
4. http://sci-hub.tw/https://www.ncbi.nlm.nih.gov/pubmed/18486479
5. Origins of Life on the Earth and in the Cosmos SECOND EDITION, page 117
6. https://www.scientificamerican.com/article/how-structure-arose-in-the-primordial-soup/
7. https://www.nature.com/scitable/topicpage/an-evolutionary-perspective-on-amino-acids-14568445
8. https://en.wikipedia.org/wiki/Tryptophan

Scientists find biology's optimal 'molecular alphabet' may be preordained

https://phys.org/news/2019-09-scientists-biology-optimal-molecular-alphabet.html?fbclid=IwAR18uckj_nrJ1p6B8Gi7n3QRW-WoAnEi3RH57SrOXxoj6yvjx5CkofkWqAY

There are millions of possible types of amino acids that could be found on Earth or elsewhere in the universe, each with its own distinctive chemical properties. Indeed, scientists have found these unique chemical properties are what give biological proteins, the large molecules that do much of life's catalysis, their own unique capabilities. The team had previously measured how the CAA set compares to random sets of amino acids and found that only about 1 in a billion random sets had chemical properties as unusually distributed as those of the CAAs.

Dimas A. M. Zaia A Few Experimental Suggestions Using Minerals to Obtain Peptides with a High Concentration of L-Amino Acids and Protein Amino Acids 10 December 2020

The peptides/proteins of all living beings on our planet are mostly made up of 19 L-amino acids and glycine, an achiral amino acid. Arising from endogenous and exogenous sources, the seas of the prebiotic Earth could have contained a huge diversity of biomolecules (including amino acids), and precursors of biomolecules. 

My comment: Agreed. There is NO REASON or function whatsoever that molecules could have benefitted from, by performing a selection of amino acids used in life.

Thus, how were these amino acids selected from the huge number of available amino acids and other molecules? What were the peptides of prebiotic Earth made up of? How were these peptides synthesized? Minerals have been considered for this task, since they can preconcentrate amino acids from dilute solutions, catalyze their polymerization, and even make the chiral selection of them.

My comment: How could mineral select the chiral form of amino acids? I have never seen a science paper solving that issue. 

However, until now, this problem has only been studied in compartmentalized experiments. There are separate experiments showing that minerals preconcentrate amino acids by adsorption or catalyze their polymerization

My comment: The polymerization would/could have occurred by binding one amino acid to another at any of the possible binding sites, like on the side chain.

or separate L-amino acids from D-amino acids. Based on the [GADV]-protein world hypothesis, as well as the relative abundance of amino acids on prebiotic Earth obtained by Zaia, several experiments are suggested. The main goal of these experiments is to show that using minerals it is possible, at least, to obtain peptides whose composition includes a high quantity of L-amino acids and protein amino acids (PAAs). These experiments should be performed using hydrothermal environments and wet/dry cycles. In addition, for hydrothermal environment experiments, it is very important to use one of the suggested artificial seawaters, and for wet/dry environments, it is important to perform the experiments in distilled water and diluted salt solutions.

My comment: The only problem is that distilled water was not the medium in the prebiotic earth....

Finally, from these experiments, we suggest that, without an RNA world or even a pre genetic world, a small peptide set could emerge that better resembles modern proteins.

https://www.mdpi.com/2073-8994/12/12/2046

Why are 20 amino acids used to make proteins? Why not more or less ? And why especially the ones that are used amongst hundreds available?
Science is absolutely clueless about how the optimal set of amino acids to make proteins emerged. Francis Crick attributed  the formation of the genetic code to an example of a frozen accident.  According to this hypothesis, the genetic code was formed through a random, highly improbable combination of its components formed by an abiotic route. Besides the problem to explain the origin of the genetic code, about which Eugene Koonin wrote: " we cannot think of a more fundamental problem in biology ", there is another, not less enigmatic situation : Why are 20 amino acids used to make proteins ( in some rare cases, 22) ?  Why not more or less ? And why especially the ones that are used amongst hundreds available? 

The study's experiment points to chemical evolution having prefabricated some amino acid chains useful in living systems before life had evolved a way to make proteins. The preference for the incorporation of the biological amino acids over non-biological counterparts also adds to possible explanations for why life selected for just 20 amino acids when 500 occurred naturally on the Hadean Earth. 14

In a progression of the first papers published in 2006, which gave a rather shy or vague explanation, in 2017, the new findings are nothing short than astounding.  In January 2017, the paper : Frozen, but no accident – why the 20 standard amino acids were selected, reported:

" Amino acids were selected to enable the formation of soluble structures with close-packed cores, allowing the presence of ordered binding pockets. Factors to take into account when assessing why a particular amino acid might be used include its component atoms, functional groups, biosynthetic cost, use in a protein core or on the surface, solubility and stability. Applying these criteria to the 20 standard amino acids, and considering some other simple alternatives that are not used, we find that there are excellent reasons for the selection of every amino acid. Rather than being a frozen accident, the set of amino acids selected appears to be near ideal. Why the particular 20 amino acids were selected to be encoded by the Genetic Code remains a puzzle." 

It remains a puzzle as so many other things in biology which find no answer when answers are constraint to a set of possible explanations, where an intelligent causal agency is excluded a priori. Only intelligence actively selects. Attributes, that unguided, random events lacks and is too unspecific,  but an intelligent creator can employ to create life. The authors also write about natural selection and evolution, a mechanism that has no place to explain the origin of life.

The paper continues: 
" Here, I argue that there are excellent reasons for using (or not using) each possible amino acid and that the set used is near optimal.
Biosynthetic cost
Protein synthesis takes a major share of the energy resources of a cell. Table 1 shows the cost of biosynthesis of each amino acid, measured in terms of number of glucose and ATP molecules required. These data are often nonintuitive. For example, Leu costs only 1 ATP, but its isomer Ile costs 11. Why would life ever, therefore, use Ile instead of Leu, if they have the same properties? Larger is not necessarily more expensive; Asn and Asp cost more in ATP than their larger alternatives Gln and Glu, and large Tyr cost only two ATP, compared to 15 for small Cys. The high cost of sulfur-containing amino acids is notable. " 

This is indeed completely counterintuitive and does not conform with naturalistic predictions.

" Burial and surface
Proteins have close-packed cores with the same density as organic solids and side chains fixed into a single conformation. A solid core is essential to stabilize proteins and to form a rigid structure with well-defined binding sites. Nonpolar side chains have therefore been selected to stabilize close-packed hydrophobic cores. Conversely, proteins are dissolved in water, so other side chains are used on a protein surface to keep them soluble in an aqueous environment. " 

The problem here is that molecules and an arrangement of correctly selected variety of amino acids would bear no function until life began. Functional subunits of proteins, or even fully operating proteins by their own would only have function, after life began, and the cells intrinsic operations were on the go. It is as if molecules had the inherent drive to contribute to life to have the first go, which of course is absurd. The only rational alternative is that a powerful creative agent had foresight, and new which arrangement and selection of amino acids would fit and work to make life possible.

"Which amino acids came first?
It is plausible that the first proteins used a subset of the 20 and a simplified Genetic Code, with the first amino acids acquired from the environment." 

Why is plausible? It is not only not plausible, but plain and clearly impossible. The genetic code could not emerge gradually, and there is no known explanation how it emerged. The author also ignores that the whole process of protein synthesis requires all parts of the process fully operational right from the beginning. A gradual development by evolutionary selective forces is impossible.

" Energetics of protein folding
Folded proteins are stabilized by hydrogen bonding, removal of nonpolar groups from water (hydrophobic effect), van der Waals forces, salt bridges and disulfide bonds. Folding is opposed by loss of conformational entropy, where rotation around bonds is restricted, and introduction of strain. These forces are well balanced so that the overall free energy changes for all the steps in protein folding are close to zero." 

Foresight and superior knowledge would be required to know how to get a protein fold that bears function, and where the forces are outbalanced naturally to get an overall energy homeostatic state close to zero.

" Conclusion
There are excellent reasons for the choice of every one of the 20 amino acids and the nonuse of other apparently simple alternatives. If all else fails, one can resort to chance or a ‘frozen accident’, as an explanation. " 

Or to design ?!



A quantitative investigation of the chemical space surrounding amino acid alphabet formation 9
21 January 2008
To date, explanations for the origin and emergence of the alphabet of amino acids encoded by the standard genetic code have been largely qualitative and speculative. Here, with the help of computational chemistry, we present the first quantitative exploration of nature’s ‘‘choices’’ set against various models for plausible alternatives. Specifically, we consider the chemical space defined by three fundamental biophysical properties (size, charge, and hydrophobicity) to ask whether the amino acids that entered the genetic code exhibit a higher diversity than random samples of similar size drawn from several different definitions of amino acid possibility space. In summary, the question of whether early life selected a non-randomly diverse alphabet of amino acids remains far from clear in this initial inquiry into the chemical space of prebiotic amino acids.

Did Evolution Select a Nonrandom ‘‘Alphabet’’ of Amino Acids? 2
26 January 2011
The last universal common ancestor of contemporary biology (LUCA) used a precise set of 20 amino acids as a standard alphabet with which to build genetically encoded protein polymers. Considerable evidence indicates that some of these amino acids were present through nonbiological syntheses prior to the origin of life, while the rest evolved as inventions of early metabolism.

Invention indicates teleology, which there is no justification to bring it into the game. There could also not yet have been evolutionary forces at work since evolution depends on a full setup and extant proteome, which origin is what is being tried to be elucidated.  

One possibility is that natural selection favored a set of amino acids that exhibits clear, nonrandom properties—a set of especially useful building blocks.

So did lifeless matter have the goal to become useful? Useful for what ? for life? So did lifeless matter have the inherent drive to group and transform itself to form building blocks , later used to make molecular machines, that would drive life?

Calculating the expected characteristics for a random alphabet of amino acids
Building from these assumptions, we performed three specific tests: we compared (in terms of coverage) (i) the full set of 20 genetically encoded amino acids for size, charge, and hydrophobicity with equivalent values calculated for a sample of 1 million alternative sets (each also comprising 20 members) drawn randomly from the pool of 50 plausible prebiotic candidates

Conclusions:
we see a consistent, unambiguous pattern; random chance would be highly unlikely to represent the chemical space of possible amino acids with such breadth and evenness in charge, size, and hydrophobicity (properties that define what protein structures and functions can be built).

MAPPING AMINO ACIDS TO UNDERSTAND LIFE'S ORIGINS 7
Jan 13, 2014
Only 20 standard amino acids are used to build proteins, but why exactly nature "chose" these particular amino acids is still a mystery. One step towards solving this is to explore the “amino acid space”, the set of possible or hypothetical amino acids that might have been used instead.

Cartography of amino acids
The number of amino acid structures generated surpasses all previous estimates. Using the method with the single fuzzy formula produced 120,000 plausible structures and using ten fuzzy formulas narrows this down to a more biologically relevant set of nearly 4,000 amino acids. This shows that there were a staggering amount of options available that could have possibly been used for building the genetically encoded amino acid set – and yet there are only 20. They compared the output of both methods to databases of biological alpha amino acids beyond the 20 genetic ones, as well as to amino acids found in carbonaceous meteorites.

Beyond Terrestrial Biology: Charting the Chemical Universe of α‑Amino Acid Structures 8
October 23, 2013
Nonbiological processes can and often do produce far more than 20 amino acids, including α-amino acids beyond those found in the genetic code, as well as β-, γ-, and δ-amino acids and others


Generic structural types of amino acids, shown as their zwitterions with respect to their core α-amino acid motif, using standard notation that R is a side chain of variable structure and composition: 
(Ia) and (Ib) are the L- and D-stereoisomers of a simple α-amino acid; (Ic) is an α,α-dialkyl amino acid with two alkyl side chains bound to the α carbon; (IIa) and (IIb) show two of the many variations that become possible when an extra carbon atom is inserted between amino and carboxyl functional groups so as to form a β-amino acid; (III) is a γ-amino acid; (IVa), (IVb), and (IVc) illustrate secondary, tertiary, and quaternary amines, respectively. All genetically encoded amino acids are of type (Ia) with the exception of proline, which is of type (IVa).

75 to 100 different amino acids have been detected in the Murchison meteorite to date, and improvements in analytical sensitivity continue to reveal a far greater diversity of molecular structure than was previously suspected in both meteoritic samples and prebiotic simulations. Despite this molecular diversity, the products of abiotic chemistry can account for only around half of the 20 genetically encoded amino acids. The amino acids selenocysteine and pyrrolysine, which fit this description, are currently entering the genetic code as the 21st and 22nd coded amino acid within some lineages. In this context, it is noteworthy that diverse biological systems use far more types of amino acids than the 20 into which genes are decoded. These additional amino acids fall into various categories, including secondary metabolites, post-translational modifications, amino acids used in nonribosomal peptide synthesis, and intermediates of the metabolic pathways by which the standard 20 are synthesized and degraded. The total number of amino acids occurring in biological systems is unknown; however, estimates range into the hundreds or thousands, with the majority found in plants and microbes.

Since abiotic synthesis and metabolism can each produce many amino acids besides those found in the genetic code,  20 for use in genetic coding were selected from a much larger pool of possible chemical structures. While simple algorithms have been used to calculate the total number of possible alkyl amino acids, the incorporation of heteroatoms (i.e., atoms other than carbon or hydrogen) vastly increases the potential for molecular diversity and the corresponding challenges for exploration. An important challenge here is to understand whether physical or chemical principles predict which 20 α-amino acids would be selected  from the near-infinite number of structural possibilities. Are other possible combinations better in some obvious functional respects, such as in the coverage of physical properties which might be useful in protein folding or catalysis? If so, then the outcome  may represent some degree of ″frozen accident″, as has been advanced for other aspects of the genetic code.

Does Life Use a Non-Random Set of Amino Acids? 1
Biology could conceivably have used a different amino acid alphabet, and there appears to be a fairly wide range from which it could have chosen. But is there anything special — is there anything unique or unusual — about the set of 20 amino acids (some organisms use one or two additional amino acids) that life does use? And, if there is, how might this fundamentally non-random contingency be explained?

Jonathan M. of Evolutionnews comments: 
If chance and necessity are seemingly inadequate, either on their own or in co-operation, what about the causal powers of agent causality? Such delicately balanced and finely-tuned parameters are routinely associated with purposive agents. Agents are uniquely endowed with the capacity of foresight, and have the capacity to visualise and subsequently actualise a complex and finely-tuned information-rich system, otherwise unattainable by chance and law. If, in every other realm of experience, such features are routinely attributed to intelligent causes, and we have seen no reason to think that this intuition is mistaken, are we not justified in positing and inferring that these systems we are finding in biology also originated at the will of a purposive conscious agent?

Paper Reports that Amino Acids Used by Life Are Finely Tuned to Explore “Chemistry Space” 3
June 5, 2015
A recent paper in Nature‘s journal Scientific Reports, “Extraordinarily Adaptive Properties of the Genetically Encoded Amino Acids,” has found that the twenty amino acids used by life are finely tuned to explore “chemistry space” and allow for maximal chemical reactions. Considering that this is a technical paper, they give an uncommonly lucid and concise explanation of what they did:

Extraordinarily Adaptive Properties of the Genetically Encoded Amino Acids 4
24 March 2015
We drew 10^8 random sets of 20 amino acids from our library of 1913 structures and compared their coverage of three chemical properties: size, charge, and hydrophobicity, to the standard amino acid alphabet. We measured how often the random sets demonstrated better coverage of chemistry space in one or more, two or more, or all three properties. In doing so, we found that better sets were extremely rare. In fact, when examining all three properties simultaneously, we detected only six sets with better coverage out of the 10^8 possibilities tested.

Luskin of Evolutionnews continues: That’s quite striking: out of 100 million different sets of twenty amino acids that they measured, only six are better able to explore “chemistry space” than the twenty amino acids that life uses. That suggests that life’s set of amino acids is finely tuned to one part in 16 million.

Nature continues: This is consistent with the hypothesis that natural selection influenced the composition of the encoded amino acid alphabet, contributing one more clue to the much deeper and wider debate regarding the roles of chance versus predictability in the evolution of life.


The number of random sets (out of 10^8, or 100,000,000) with better coverage than the encoded amino acids in one, two, or three properties. 
Note that the circles are not drawn to scale; an appropriately scaled circle representing the number of random sets with better coverage in all three properties than the encoded set would only cover an area approximately 1/ 100th of that of the period at the end of this sentence.

Well, or maybe there was neither evolution, nor natural selection, and if chance is not a good explanatory candidate, we might consider another option, commonly ignored by secular science: Selection by an intelligent agency with foresight and higher intelligence.

Frozen, but no accident – why the 20 standard amino acids were selected 6
13 January 2017
The 20 standard amino acids encoded by the Genetic Code were adopted during the RNA World, around 4 billion years ago. This amino acid set could be regarded as a frozen accident, implying that other possible structures could equally well have been chosen to use in proteins. Amino acids were not primarily selected for their ability to support catalysis, as the RNA World already had highly effective cofactors to perform reactions, such as oxidation, reduction and transfer of small molecules. Rather, they were selected to enable the formation of soluble structures with close-packed cores, allowing the presence of ordered binding pockets. Factors to take into account when assessing why a particular amino acid might be used include its component atoms, functional groups, biosynthetic cost, use in a protein core or on the surface, solubility and stability. Applying these criteria to the 20 standard amino acids, and considering some other simple alternatives that are not used, we find that there are excellent reasons for the selection of every amino acid. Rather than being a frozen accident, the set of amino acids selected appears to be near ideal.

Setting the common a priori mainstream science assumptions aside, the last sentence is remarkable. " the set of amino acids selected appears to be near ideal. ". 

Andrew J.Doing ( author of the paper ): Why the particular 20 amino acids were selected to be encoded by the Genetic Code remains a puzzle.

It remains a puzzle as so many other things in biology which find no answer by the ones that build their inferences on a constraint set of possible explanations, where an intelligent causal agency is excluded a priori. Selection is an active process, that requires intelligence. Attributes, that chance alone lacks, but an intelligent creator can employ to create life. The authors also write about natural selection and evolution, a mechanism that has no place to explain the origin of life. 

Here, I argue that there are excellent reasons for using (or not using) each possible amino acid and that the set used is near optimal.

Biosynthetic cost
Protein synthesis takes a major share of the energy resources of a cell [12]. Table 1 shows the cost of biosynthesis of each amino acid, measured in terms of number of glucose and ATP molecules required. These data are often nonintuitive. For example, Leu costs only 1 ATP, but its isomer Ile costs 11. Why would life ever therefore use Ile instead of Leu, if they have the same properties? Larger is not necessarily more expensive; Asn and Asp cost more in ATP than their larger alternatives Gln and Glu, and large Tyr costs only two ATP, compared to 15 for small Cys. The high cost of sulfur-containing amino acids is notable.

This is indeed completely counterintuitive and does not conform with naturalistic predictions.

Burial and surface
Proteins have close-packed cores with the same density as organic solids and side chains fixed into a single conformation. A solid core is essential to stabilise proteins and to form a rigid structure with well-defined binding sites. Nonpolar side chains have therefore been selected to stabilise close-packed hydrophobic cores. Conversely, proteins are dissolved in water, so other side chains are used on a protein surface to keep them soluble in an aqueous environment.

The problem here is that molecules and an arrangement of correctly selected variety of amino acids would bear no function until life began. Functional subunits of proteins, or even fully operating proteins by their own would only have function, after life began, and the cells intrinsic operations were on the go. It is as if molecules had the inherent drive to contribute to life to have a first go, which of course is absurd. The only rational alternative is that a powerful creator had foresight, and new which arrangement and selection of amino acids would fit and work to make life possible.

Which amino acids came first?
It is plausible that the first proteins used a subset of the 20 and a simplified Genetic Code, with the first amino acids acquired from the environment.

Why is plausible? It is not only not plausible, but plain and clearly impossible. The genetic code could not emerge gradually, and there is no known explanation how it emerged. The author also ignores that the whole process of protein synthesis requires all parts in the process fully operational right from the beginning. A gradual development by evolutionary selective forces is impossible.

Energetics of protein folding
Folded proteins are stabilised by hydrogen bonding, removal of nonpolar groups from water (hydrophobic effect), van der Waals forces, salt bridges and disulfide bonds. Folding is opposed by loss of conformational entropy, where rotation around bonds is restricted, and introduction of strain. These forces are well balanced, so that the overall free energy changes for all the steps in protein folding are close to zero.

Foresight and superior knowledge would be required to know how to get a protein fold that bears function, and where the forces are outbalanced naturally to get an overall energy homeostatic state close to zero.

Conclusion
There are excellent reasons for the choice of every one of the 20 amino acids and the nonuse of other apparently simple alternatives. If all else fails, one can resort to chance or a ‘frozen accident’, as an explanation.

Or to design ?!

The 20 standard amino acids - as general reasons for choosing side chains, we can now consider the 20 individually.

Side chain negative
• Aspartic acid - Asp - D 
• Glutamic acid - Glu - E 

Aspartic, Glutamic acid

The carboxyl group is stable, has a negative charge for strong electrostatic bonds, binds metals, is an excellent hydrogen bond acceptor, increases protein solubility and can transfer protons. Its twofold symmetry means that the entropic cost of fixing the carboxyl group is low.As Asp and Glu (and Asn and Gln) have the same functional groups, why do we have two pairs of these amino acids? At some sites, Asp and Glu (or Asn and Gln) can readily substitute for each other. However, this is not always the case. The first residue preceding the α-helix is the N-cap; Asp and Asn are strongly favoured here as they can accept hydrogen bonds from free backbone NH groups. Glu and Gln cannot do this, as their extra CH2 group pushes their functional groups out beyond the helix terminus. Similarly, at the second (N2) position of an α-helix, Gln and Glu are frequently found because they can loop around and hydrogen bond to the backbone NH groups. Asn and Asp are too short to form these rings. Glu to Asp or Gln to Asn can sometimes thus be very nonconservative substitutions, so all four are used.

Lysine, Arginine

Lys and Arg are used to give a protein solubility and for catalysis. They are rarely buried, not only because they have positively charged groups, but also because they are on the ends of long, flexible, straight chains, with four rotatable bonds. Their functional groups are also valuable when a positive charge is needed. Arg, in particular, is a superb hydrogen bond donor, with five NH bonds, polarised due to its delocalised positive charge.
Nonpolar residues, such as Leu and Ile, are also often found on the protein surface. One might expect this to be a problem for the protein, as they will reduce protein solubility. If there are Lys or Arg side chains nearby, however, nonpolar groups can form hydrophobic interactions with the CH2 groups in the Lys and Arg side chains. Nonpolar side chains would thus be less tolerated on the surface if charged groups had fewer CH2 groups.

Side chain positive
• Arginine - Arg - R 
• Lysine - Lys - K 
• Histidine - His - H 

Histidine

While the range of chemical reactions available to the 20 side chains is dismally poor, they are good at proton transfer. Rates of proton transfer are maximised when the general acid has a pKa that is the same as the environment. His, with a pKa of around 6.5, is easily tweaked by varying its environment, is an excellent side chain for general acid and base catalysis and is thus abundantly found in active sites requiring proton transfer. It is also often found binding metals.

[size=13]Lysine, Arginine[/size]

Lys and Arg are used to give a protein solubility and for catalysis. They are rarely buried, not only because they have positively charged groups, but also because they are on the ends of long, flexible, straight chains, with four rotatable bonds. Their functional groups are also valuable when a positive charge is needed. Arg, in particular, is a superb hydrogen bond donor, with five NH bonds, polarised due to its delocalised positive charge.
Nonpolar residues, such as Leu and Ile, are also often found on the protein surface. One might expect this to be a problem for the protein, as they will reduce protein solubility. If there are Lys or Arg side chains nearby, however, nonpolar groups can form hydrophobic interactions with the CH2 groups in the Lys and Arg side chains. Nonpolar side chains would thus be less tolerated on the surface if charged groups had fewer CH2 groups.

Uncharged polar (usually participate in hydrogen bonds as proton donors or acceptors):
• Glutamine - Gln - Q 
• Asparagine - Asn - N 
• Serine - Ser - S 
• Threonine - Thr - T 
• Tyrosine - Tyr - Y 

Serine, Threonine, Asparagine, Glutamine, Tyrosine
These amino acids frequently hydrogen bond to other side chains, amide groups in the backbone, to substrates or to ligands. The intrinsic benefit of hydrogen bond formation is offset by the conformational entropic cost of fixing the side chain in position and strain. Functional groups selected for hydrogen bonding are thus on the ends of short chains, with just one or two CH2 groups, so that the cost of restricting them is low and their biosynthetic cost is minimised.The aromatic ring in Tyr lowers the pKa of its OH, making it easier to form an O− group and act as an acid. Tyr can also form functional radicals, such as in ribonucleotide reductase, photosystem II and prostaglandin H synthase and transport electrons in pili.Allothreonine, with the opposite chirality at its Cβ, decomposes twice as fast, so threonine may have been selected for this reason.

Hydrophobic (normally buried inside the protein core):

Side chain nonpolar
• Alanine - Ala - A 
• Isoleucine - Ile - I 
• Leucine - Leu - L 
• Methionine - Met - M 
• Phenylalanine - Phe - F 
• Valine - Val - V 
• Proline - Pro - P 
• Glycine - Gly - G
• Cysteine - Cys - C 
• Tryptophan - Trp - W

Alanine

Ala is small and cheap. Its CH3 side chain has little surface to form van der Waals bonds, but it has a profound effect on the backbone, restricting it to either the α- or β-conformations. It is therefore more energetically favourable to put Ala into secondary structure than Gly. The formation of an isolated α-helix in water restricts rotations in side chains in all amino acids, except Ala and Gly. Hence, Ala has the highest preference to be in isolated α-helices .

Valine, Leucine, Isoleucine, Phenylalaline

These hydrocarbon side chains are clearly present to drive protein folding by forming hydrophobic cores, yet why exactly these were selected deserves an explanation. Firstly, why are there so many hydrocarbon side chains, rather than just, say, Leu? Multiple hydrocarbon side chains may be used as they are required to fill a protein core with no clashes and no holes. A variety of pieces are required to fit all the gaps within a protein core. Each can adopt a number of rotamers with similar energies , thus giving a range of possible shapes for each side chain. Thus, Leu and Ile are both needed to increase the range of possible hydrocarbon 3D shapes, despite Ile being more costly to synthesise . Val, Leu, Ile and Phe are striking in that they all have branched carbons, rather than straight chains. Using a branch gives one fewer dihedral angle to be fixed compared to a straight chain. Side chains therefore enter a protein core not just because they are hydrophobic, but also because they do not lose too much conformational entropy when they fold. Similarly, rings are rigid, so Phe has a large hydrocarbon surface and only two bonds to be fixed. Hydrocarbon side chains larger than Phe, Ile or Leu may not be used, as they would be less soluble as amino acids. A cyclohexane ring is also less soluble than a benzene ring, so Phe is used, rather than cyclohexylalanine.
Ile and Thr differ from the other 18 in that they have centres of chirality in their side chains. Alloisoleucine has the opposite chirality at its Cβ. The selection of isoleucine over alloisoleucine seems to be chance.

Methione

The explanation above for why branched side chains are preferred over straight chains fails for Met. Met has three bonds to be fixed if it is used in a protein core, giving less stabilisation than one might expect for its size. The first use of Met in protein is likely not to have been for protein stability, however, but rather for initiation of protein synthesis, using the AUG codon and formylMet. Occasional use of AUG would then allow Met to be found at other sites in a protein, such as in forming sulfur–aromatic interactions. It may also be useful in forming a hydrophobic core with its unique shape. Met is prone to oxidation at its sulfur, potentially leading to loss of protein function, but this would not have occurred before the Great Oxidation Event around 2.3 billion years. Met is also the most expensive amino acid to biosynthesise. Life is now stuck with this nonideal amino acid.

Cysteine

A key function of proteins is to bind metals. Soft metals, such as copper, zinc and cadmium, bind more tightly to sulfur than to oxygen. Cys may therefore have been selected for its metal binding properties, despite its high biosynthetic cost. In particular, Cys is commonly used to bind iron-sulfur clusters. These cofactors are found in a wide variety of metalloproteins, are playing crucial roles in metabolism and are very ancient. The SH side chain is also an effective nucleophile; it has a lower pKa than OH, readily forming S−.
Cys is the only side chain able to perform redox reactions, by forming disulfide bonds to stabilise folded proteins. Cys was selected in an anaerobic environment, however, where disulfide bond formation would have been rare or nonexistent. It is therefore not plausible that Cys was selected for its ability to form disulfides, and its subsequent use for this purpose, starting very approximately 1.5 billion years later after the Great Oxidation Event, is a lucky accident.

Maybe neither the atmosphere was anaerobic, and neither was it a lucky accident that Cys was selected.  

Proline

The unique structure of Pro, with no backbone NH group and its ring restricting its backbone, means that it is incompatible with many sites in a protein. Nevertheless, it can be highly stabilising, if its ring locks it into a desired conformation, such as within turns. Indeed, it has the lowest conformational entropic cost of folding of any amino acid. Pro is the simplest stable ring structure linking the Cα and N in an amino acid.

Tryptophan

Trp with its double aromatic ring is the least abundant amino acid and the most expensive to synthesise. It has some distinct properties: Trp, with an absorption maximum at around 280 nm, is well suited to be a UV-B chromophore The UVR8 protein uses an excitonically coupled Trp pyramid for this purpose in plants, for example, detecting potentially damaging levels of UV radiation. The Trp indole chromophore may therefore have been selected to help protect organisms from the intense UV radiation that was present on early Earth in the absence of an ozone layer.

Transition from prebiotic amino acid synthesis, to biosynthesis pathways in modern cells to synthesize amino acids

Little information in peer-reviewed scientific papers can be found to answer this key question: How could a supposed transition from prebiotic synthesis of amino acids, to the extremely complex biosynthesis pathways in modern cells have occurred? The sobering answer is: there are no answers. There is a huge unbridgeable gap. Following science paper describes the dilemma as follows:

On the Development Towards the Modern World: A Plausible Role of Uncoded Peptides in the RNA World 5
2009
Arguably one of the most outstanding problems in understanding the progress of early life is the transition from the RNA world to the modern protein based world. One of the main requirements of this transition is the emergence of mechanism to produce functionally meaningful peptides and later, proteins. What could have served as the driving force for the production of peptides and what would have been their properties and purpose in the RNA world? The answer may seem immediately clear; proteins are better enzymes than ribozymes.

The unwarranted insertion of teleology is implicitly clear and demonstrates the easy insertion of pseudo-explanations which do no justice and are inadequate to explain the monstrous problem that origin of life researchers face: Let's say it loud and clear. Unanimated matter does not select goal-oriented, what better works. Better for what ? for cellular functions? that's certainly true, but: do molecules have foresight or goals?  

However, modern proteins are only useful in their folded stated, whereas peptides need to reach a critical size and specific amino acid sequence before they fold into a functional biomolecules. It is more than likely that emerging peptides were not immediately useful as operational enzymes.

Then, why would they emerge AT ALL? 

Herein we describe two plausible roles that emerging peptides could have played, firstly support of the pre-existing RNA machinery. 

Based on what evidence do the authors suppose there was a pre-existing RNA machinery in place ? that are simply unwarranted made up assertions and scenarios. 

and secondly as early chemical catalysts. During the era of the RNA world, protein (or peptide) evolution would have been strongly coupled to the extant RNA infrastructure development.

There was no evolution at this stage. 

Thus, the emergence of peptides as a mechanism for supporting pre-existing RNA machinery, is a sound reason for peptides to be retained in an RNA world.

Is the prebiotic origin of amino acids by natural means plausible?

https://reasonandscience.catsboard.com/t1740p25-amino-acids-origin-of-the-canonical-twenty-amino-acids-required-for-life#9379

Chemical evolution of amino acids and proteins ? Impossible !!
https://www.youtube.com/watch?v=1L1MfGrtk0A

Chemical evolution of amino acids and proteins ? Impossible !!
https://reasonandscience.catsboard.com/t2887-chemical-evolution-of-amino-acids-and-proteins-impossible

Amino acids are monomers. (A monomer is a molecule that can react together with other monomer molecules to form a larger polymer chain or three-dimensional network 49) They are one of the principal components that make up proteins. 22 amino acids encode proteins. 20 in the standard genetic code and another 2 (selenocysteine, which is an essential amino acid component in selenoproteins, which are involved in a variety of cellular and metabolic processes 43 and pyrrolysine are restricted to a very small number of organisms and proteins. 42 )

Scitable gives us a short description of AAs:
Chemically, an amino acid is a molecule that has a carboxylic acid group 38 (COOH) and an amino group 39 (NH2) that are each attached/bonded to a central carbon atom, also called the α carbon. Each of the 20 amino acids has a specific side chain, known as an R group, that is also attached to the α carbon. 40 and a hydrogen atom.  The R groups, 41 the variable group or the side-chain, have a variety of shapes, sizes, charges, and reactivities. This allows amino acids to be grouped according to the chemical properties of their side chains. For example, some amino acids have polar side chains that are soluble in water; examples include serine, threonine, and asparagine. Other amino acids avoid water and are called hydrophobic, such as isoleucine, phenylalanine, and valine. The amino acid cysteine has a chemically reactive side chain that can form bonds with another cysteine. 23



A single amino acid – the subunit monomer of polypeptides and proteins.
NH2 is the amine group and the blue -COOH is the carboxylic acid group. The green R is a side-chain that is different for each of the 20 or so amino acids found in proteins.Attribution: Marc T. Facciotti (own work)

Alpha-amino acids are monomer units that are bonded ( polymerized ) and linked to polymer strands via a head-to-tail linkage that can fold ( depending on the sequence ) into complex functional 3D shapes. Once folded, often as a joint venture with other polymer strands, they form secondary, tertiary, and quaternary structures and catalytic pockets where in many cases complex metal clusters perform the catalytic reaction where the manufacturing of a compound takes place.

Origin of the proteinogenic ( protein creating ) amino acids used in life
There are many hypotheses about how the amino acids used in life could have originated. They are divided into terrestrial, and extraterrestrial origin. Terrestrial proposals are Spark discharge, Irradiation (UV, X-ray, etc.), Shock heating, and hydrothermal vents. Extraterrestrial amino acids have been observed in various types of carbonaceous chondrites, comets, and micrometeorites. Norio Kitadai and colleagues gave a good overview in their scientific paper: Origins of building blocks of life: A review from 2017 54 As we will see, none of them withstand scrutiny. Wherever one looks, there are problems.

Extraterrestrial origins
Norio Kitadai et al. 2017: 

To date, over 80 kinds of amino acids have been identified in carbonaceous chondrites, including 12 protein-amino acids of Ala, Asp, Glu, Gly, Ile, Leu, Phe, Pro, Ser, Thr, Tyr, and Val. 

Amongst the problems is that these amino acids come always in mixtures with non-proteinogenic aa's, and they are all chirally mixed ( L and R-handed chiral form)

Panspermia
One hypothesis is that amino acids amongst other biofriendly molecules were made in space, and delivered to our planet by meteorites, comets,  interplanetary dust particles, etc. It is called panspermia ( seeds everywhere in the universe ). There are good reasons to reject the idea. Nir Goldman and colleagues published an article in Nature magazine on the subject in 2010. They wrote:

Delivery of prebiotic compounds to early Earth from an impacting comet is thought to be an unlikely mechanism for the origins of life because of unfavorable chemical conditions on the planet and the high heat from impact.  5

Hugh Ross pointed out what seems to be one of the main problems:
What happens to comets and their supply of these molecules when they pass through Earth’s atmosphere and when they strike the planetary surface presents a big problem. Calculations and measurements show that both events generate so much heat (atmosphere passage generates 500°+ Centigrade while the collision generates 1,000°+ Centigrade) that they break down the molecules into components useless for forming the building blocks of life molecules. In 1974, comet 81P Wild passed within 500,000 miles of Jupiter, which caused the comet to be perturbed into orbiting within the inner solar system. This new orbit enabled NASA to send the Stardust Spacecraft to the comet in 2004 to recover samples, which were returned to Earth and analyzed for organic molecules. The only amino acid indisputably detected in the sample was glycine at an abundance level of just 20 trillionths of a mol per cubic centimeter. 7

Amino acids found in meteorites are racemic, that is, they come in right-handed, and left-handed helicity. Life uses 100% left-handed AAs. [url=https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4919777/#:~:text=Analyses of pieces of the,alanine%2C suggesting minimal terrestrial biological]10[/url]

Recently a new discovery made the rounds. Yasuhiro Oba and colleagues reported in Nature Communications:
the detection of nucleobases in three carbonaceous meteorites using state-of-the-art analytical techniques optimized for small-scale quantification of nucleobases down to the range of parts per trillion (ppt). 8

Liz Kruesi reported in an article in science news about the finding:
Space rocks that fell to Earth within the last century contain the five bases that store information in DNA and RNA. These “nucleobases” — adenine, guanine, cytosine, thymine and uracil — combine with sugars and phosphates to make up the genetic code of all life on Earth. The discovery adds to evidence that suggests life’s precursors originally came from space, the researchers say. Scientists have detected bits of adenine, guanine and other organic compounds in meteorites since the 1960s Researchers have also seen hints of uracil, but cytosine and thymine remained elusive, until now. “We’ve completed the set of all the bases found in DNA and RNA and life on Earth, and they’re present in meteorites,” says astrochemist Daniel Glavin of NASA’s Goddard Space Flight Center in Greenbelt, Md. 9

but here comes the cold shower from the same article:

In the new analysis, the researchers measured more than a dozen other life-related compounds, including isomers of the nucleobases.

That means the anthropogenic nucleobases are mixed with other isomeric molecules, not used in living cells. That raises the question, of how those used in life could have been joined, concentrated, selected and sorted out from those not used in life.

And PIERAZZO and colleagues wrote in the paper: Amino acid survival in large cometary impacts: 
It is clear that there are substantial uncertainties in estimates for both exogenous and endogenous sources of organics, as well as the dominant sinks. All of the likely mechanisms described here lead to extremely low global concentrations of amino acids, emphasizing the need for substantial concentration mechanisms4r for altogether different approaches to the problem of prebiotic chemical synthesis. 6 

This is what Stanley Miller had to say in an interview that was conducted in October, 1996
The amount of useful compounds you are going to get from meteorites is very small. The dust and comets may provide a little more. Comets contain a lot of hydrogen cyanide, a compound central to prebiotic synthesis of amino acids as well as purines. Some HCN came into the atmosphere from comets. Whether it survived impact, and how much, are open to discussion. I'm skeptical that you are going to get more than a few percent of organic compounds from comets and dust. It ultimately doesn't make much difference where it comes from. I happen to think prebiotic synthesis happened on the Earth, but I admit I could be wrong. There is another part of the story. In 1969 a carbonaceous meteorite fell in Murchison Australia. It turned out the meteorite had high concentrations of amino acids, about 100 ppm, and they were the same kind of amino acids you get in prebiotic experiments like mine. This discovery made it plausible that similar processes could have happened on primitive Earth, on an asteroid, or for that matter, anywhere else the proper conditions exist. 20

What about the synthesis of amino acids in hydrothermal vents?
Hugh Ross and Fazale Rana explain:
Laboratory experiments simulating a hot, chemically harsh environment modeled after deepsea hydrothermal vents indicate that amino acids, peptides, and other biomoleculars can form under such conditions. However, a team led by Stanley Miller has found that at 660 °F (350 °C), a temperature that the vents can and do reach, the amino acid half-life in a water environment is only a few minutes. (In other words, half the amino acids break down in just a few minutes.) At 480 °F (250 °C) the half-life of sugars measures in seconds. For a nucleobase to function as a building block for DNA or RNA it must be joined to a sugar. For polypeptides (chains of amino acids linked together by peptide bonds but with much lower molecular weight than proteins) the half-life is anywhere from a few minutes to a few hours. 21

Punam Dalai and colleagues inform:
The thermal and chemical gradients at hydrothermal vents on the Earth’s surface may have played an important role in thermodynamically favorable reactions for organic synthesis. These reactions may have been catalyzed by transition metal–sulfide minerals such as pyrite. However, destructive free radicals are also generated photo catalytically at the surface of these sulfides and at the surfaces of the ultramafic minerals that constitute peridotite and komatiite. 58

The Miller-Urey experiment
Miller and Urey performed the legendary in vitro spark-discharge experiments in 1953, attempting to produce amino acids under primitive earth conditions. 13 With that experiment, only a few weeks distant from Watson and Crick discovering the DNA structure, the modern era in the study of the Origin of life began. The hope was that this experiment would pave the road to finding naturalistic answers to life's origins. It has widely been heralded as evidence for the origin of the set of amino acids used in life, and based on it, many claim even today that abiogenesis did become a plausible explanation for the origin of life. After 50 years, in 2003 Jeffrey L. Bada and Antonio Lazcano commemorated the Miller-Urey experiment in an article published in Science magazine. They wrote:

But is the “prebiotic soup” theory a reasonable explanation for the emergence of life? Contemporary geoscientists tend to doubt that the primitive atmosphere had the highly reducing composition used by Miller in 1953. 14

Miller himself pointed out that:
There is no agreement on the composition of the primitive atmosphere; opinions vary from strongly reducing (CH4 + N2, NH3 + H2O, or CO2 + H2 + N2) to neutral (CO2 + N2 + H2O) 53

We don't know what the conditions were, so every laboratory experiment is flawed from the get-go. Nobody knows the influencing conditions, like geological, temperature, electromagnetic radiation, etc. contributing to the  Physico-chemical complexity of the earth influencing biochemical processes. There are several other fatal flaws. There is always a team of researchers, that tweak and fine-tune the experiment towards the desired outcome. On prebiotic earth, there was no such thing. 

Consider what the researchers had to do to set up the experiment: Jeffrey L. Bada and colleagues explained:
Numerous steps in the protocol described here are critical for conducting Miller-Urey type experiments safely and correctly. First, all glassware and sample handling tools that will come in contact with the reaction flask or sample need to be sterilized. Sterilization is achieved by thoroughly rinsing the items in question with ultrapure water and then wrapping them in aluminum foil, prior to pyrolyzing at 500 °C in air for at least 3 hr. Once the equipment has been pyrolyzed and while preparing samples for analysis, care must be taken to avoid organic contamination. The risk of contamination can be minimized by wearing nitrile gloves, a laboratory coat, and protective eyewear. Be sure to work with samples away from one's body as common sources of contamination include fingerprints, skin, hair, and exhaled breath. Avoid contact with wet gloves and do not use any latex or Nylon materials. 19
This is just the first step. Bada continues:  There are many additional notes worth keeping in mind when carrying out various steps in the protocol outlined here.

You know where this goes. This has nothing to do with what happened on early earth. There were no test drives multiple times, trial and error to get to optimal conditions. Get a bit of atmospheric pressure here, change the gas composition a bit there. Change the electromagnetic radiation or the temperature variations. There are basically innumerable possibilities of different atmospheric conditions, and having the just-right atmosphere depends on many different factors. One can resort to the number of possible earth-like candidates in the universe and claim, that one by chance could have the right conditions. While nobody knows the odds, some scientific papers have calculated numbers that are far from what could be considered a reasonably probable chance 12  In 2008, after Miller's death,  Adam P. Johnson and colleagues reexamined the boxes containing the dried residues from the apparatus from the second ( the volcanic experiment ) in 1953. 15 Miller identified five different amino acids, plus several unknowns in the extracts from this apparatus. Johnson et al. however, identified 22 amino acids and five amines. Several were not identified previously in Miller’s experiments. In 2011, the same researchers extended their analysis of Miller’s old flacons to include those from a spark-discharge experiment made in 1958. They reported:

The samples contained a large assortment of amino acids and amines, including numerous sulfur amino acids. This mixture might have been prominent on a regional scale (for example, near volcanoes), where these gases may have played a vital role in the localized synthesis of some of the first terrestrial organic compounds. 16 

A survey from the U.S.Government informed that:
Ninety-nine percent of the gas molecules emitted during a volcanic eruption are water vapor (H2O), carbon dioxide (CO2), and sulfur dioxide (SO2). The remaining one percent is comprised of small amounts of hydrogen sulfide, carbon monoxide, hydrogen chloride, hydrogen fluoride, and other minor gas species. [url=https://www.usgs.gov/faqs/what-gases-are-emitted-kilauea-and-other-active-volcanoes#:~:text=Ninety%2Dnine percent of the,and other minor gas species.]17[/url]

This composition is not conducive to producing amino acids on early earth. Unless the emissions were different, supposedly 3,9 Gya ago. Were they?

The gases released into the atmosphere by high-temperature volcanic eruptions have been dominated by H2O, CO2, and SO2 since at least 3600 Ma, and probably since at least ∼3900 Ma. Mantle-derived volcanic gases that entered the atmosphere from high-temperature volcanism would have provided low, but not zero, yields of prebiotic molecules during that interval. 18

Carol Turse informed in a science paper from 2013 that:
Variations of Miller’s experiments, some completed by Miller himself, have been completed that include aspects of hydrothermal vents, neutral atmospheres, reducing H2S atmospheres, as well as volcanic conditions  In each of these variations amino acids or organic precursors of amino acids are produced at some level. 51

but the following proteinogenic amino acids were never produced in any of the experiments: Cysteine, Histidine, Lysine, Asparagine, Pyrrolysine, Proline, Glutamine, Arginine, Threonine, Selenocysteine, Tryptophan, Tyrosine 52

Homochirality
The DNA and RNA ribose backbone has to be in right-handed chiral form. (Chirality is from Greek and means handedness). Amino acids, left-handed. Phospholipids, right-handed. That is essential for life, and its origin in biological systems demands an explanation. In nature, carbon compounds come in mixed chiral forms. In the cell, the complex protein machinery fabrics the materials in the right enantiomeric form, as life requires. Change Laura Tan and Rob Stadler bring it succinctly to the point. They write in:  The Stairway To Life:

In all living systems, homochirality is produced and maintained by enzymes, which are themselves composed of homochiral amino acids that were specified through homochiral DNA and produced via homochiral messenger RNA, homochiral ribosomal RNA, and homochiral transfer RNA. No one has ever found a plausible abiotic explanation for how life could have become exclusively homochiral.

In order for proteins to fold into functional 3D structures, the building blocks that make them, amino acids, must be chiral. As Wikibooks explains:
A tetrahedral carbon atom with four distinct groups is called asymetric, or chiral. The ability of a molecule to rotate plane-polarized light to the left, L (levorotary) or right, D (dextrorotary) gives it its optical and stereochemical fingerprint. 50

Biologically synthesized amino acids, for instance, occur exclusively in their lefthanded (levorotatory L) form, while the sugar backbone of nucleic acids, ribose, are all right-handed (dextrorotatory D), and phospholipid glycerol headgroups of archaea and bacteria exclusively homochiral.  (Bacteria and eukarya have membranes comprised of phospholipids with backbones of left-handed configurational stereochemistry, whereas archaea contain backbones of right-handed stereochemistry). Amino acids are not the same as their mirror image, analogously a left shoe will not fit properly on one's right foot no matter how someone rotates it. 

Daniel P. Glavin and colleagues elucidated in a scientific paper from 2020:
The observed homochirality in all life on Earth, that is, the predominance of “left-handed” or l-amino acids and “right-handed” or d-sugars, is a unique property of life that is crucial for molecular recognition, enzymatic function, information storage, and structure and is thought to be a prerequisite for the origin or early evolution of life. 32

Racemic, mixed proteins are non-functional.  If a chemist cooks up a bunch of amino acids or their precursor molecules in a laboratory, the result will always be a racemic mixture of left and right. Proteins made by mixtures of left and right-handed amino acids do not form well-defined tertiary and quaternary protein structures. Ribose must have been in its right-handed form for the first RNA molecules to be useful for functional structures, which cannot occur with random mixtures of right and left-handed nucleotides. Chemical reactions starting from racemic mixtures result always in mixed, nonchiral systems. The chemistry explaining how exclusive left-handed and right-handed molecules could have been formed is one of the biggest open questions, a profound mystery, persisting for over 150 years since Lous Pasteur discovered the right and left-handed chiral form (levorotatory and the dextrorotatory form) in chemistry. Hypotheses on the origin of homochirality in the living world can be classified into two major types: biotic and abiotic. The abiotic appearance of chiral materials hence leads to deeper questions. How can symmetry be originated in a universe that is not governed by physical laws that convey this symmetry?  Scientists do not know how the right-handed amino acids and left-handed sugars and either exclusively right-handed or left-handed backbones of phospholipids could have been instantiated prebiotically by accident. Since there was no prebiotic natural selection, the only alternative to conscious choice is an unguided random non-designed coincidence. Homochirality had to arise for amino acids and sugars and phospholipids simultaneously. Life uses its complex molecular machinery to instantiate just the right or lefthanded molecules. 

Benjamin List and David MacMillan were awarded the Nobel Prize in Chemistry in 2021. A science news article quoted them:  

“Why in the world is biology single-handed? Why do we have this preference in nature? We don’t know,” List said. “This handedness is transferred in the catalytic reaction onto the substrates so that you get more of these handed molecules. It’s a great gift, I would say, that nature provides these molecules for us.”

“Chirality, for me, is the most interesting question in physics and chemistry and maybe even in biology,” Felser says, adding that today’s announcement could be “inspiring for the younger generations to look more for symmetry violations in nature”. 34

Homochirality, its origin a scientifically longstanding unresolved issue
Donna G. Blackmond explains in her paper published in 2010:
There is one general feature of the molecules constituting all known living systems on Earth, and in particular of biopolymers, which needs to be explained within the problem of origins: their homochirality. Most molecules of life are homochiral, that is, they possess the same handedness or chirality. Homochirality of biological molecules is a signature of life. The chirality or sense of handedness of the amino acid molecules is an important problem.
Figure above shows two versions, or enantiomers, of the amino acid alanine. Each contains exactly the same number of elements with the same types of chemical bonds, and yet they are the mirror image of each other. A molecule that is not superimposable on its mirror image is chiral. When a molecule with a definite sense of handedness reacts chemically with one that is symmetric (or otherwise does not have a particular handedness), the left- and right-handed amino acids have similar properties. Likewise, the chemical properties of an interaction between two left-handed molecules or two right-handed molecules are the same. However, neither of these interactions is the same as when left- and right-handed molecules are interacting with each other. Hence, the handedness of biological molecules such as amino acids or nucleotides plays a role in their functionality.

 “Symmetry breaking” is the term used to describe the occurrence of an imbalance between left and right enantiomeric molecules. This imbalance is traditionally measured in terms of the enantiomeric excess, or ee, where ee are concentrations of the right and left-hand molecules, respectively. Proposals for how an imbalance might have come about may be classified as either terrestrial or extraterrestrial, and then subdivided into either random or deterministic (sometimes called “de facto” and “de lege” respectively). A trivial example is that any collection of an odd number of enantiomeric molecules has, by definition, broken symmetry. Fluctuations in the physical and chemical environment could result in transient fluctuations in the relative numbers of left- and right-handed molecules. However, any small imbalance created in this way should average out as the racemic state unless some process intervenes to sustain and amplify it. Thus, whether or not the imbalance in enantiomers came about by chance, arising on earth or elsewhere, an amplification mechanism remains the key to increasing enantiomeric excess and ultimately to approaching the homochiral state. 22


The two mirror-image enantiomers of the amino acid alanine

A. G. CAIRNS-SMITH exposes the problem in his book: Seven clues to the origin of life, on page 40:
A particularly clear case is in the universal choice of only 'left-handed' amino acids for making proteins, when, as far as one can see, 'right-handed' ones would have been just as good. Let me clarify this.
Molecules that are at all complex are usually not superposable on their mirror images. There is nothing particularly strange about this: it is true of most objects. Your right hand, for example, is a left hand in the mirror. It is only rather symmetrical objects that do not have 'right-handed' and 'left-handed' versions. When two or more objects have to be fitted together in some way their 'handedness' begins to matter. If it is a left hand it must go with a left glove. If a nut has a right-hand screw, then so must its bolt. In the same sort of way the socket on an enzyme will generally be fussy about the 'handedness' of a molecule that is to fit it. If the socket is 'left-handed' then only the 'left-handed' molecule will do. So there has to be this kind of discrimination in biochemistry, as in human engineering, when 'right-handed' and 'left-handed' objects are being dealt with. And it is perhaps not surprising that the amino acids for proteins should have a uniform 'handedness'. There could be a good reason for that, as there is good reason to stick to only one 'handedness' for nuts and bolts. But whether, in such cases, to choose left or right, that is pure convention. It could be decided by the toss of a coin. 24

A. G. CAIRNS-SMITH genetic takeover, page53
It is commonly believed that proteins of a sort or nucleic acids of a sort (or both) would have been necessary for the making of those first systems that could evolve under natural selection and so take off from the launching platform provided by prevital chemical processes. We have already come to a major difficulty here: Much of the point of protein and the whole point of nucleic acid would seem to be lost unless these molecules have appropriate secondary/tertiary structures, and that is only possible with chirally defined units. As we saw, the ‘abiotic‘ way of circumventing this problem (by prevital resolution of enantiomers) seems hopelessly inadequate, and ‘biotic’ mechanisms depend on efficient machinery already in action. 25

Sean Henahan interviewed Dr. Stanley L. Miller  in 1998. To the question:  What about the even balance of L and D (left and right oriented) amino acids seen in your experiment, unlike the preponderance of L seen in nature? How have you dealt with that question?, Miller answered:

All of these pre-biotic experiments yield a racemic mixture, that is, equal amounts of D and L forms of the compounds. Indeed, if you're results are not racemic, you immediately suspect contamination. The question is how did one form get selected. In my opinion, the selection comes close to or slightly after the origin of life. There is no way in my opinion that you are going to sort out the D and L amino acids in separate pools. My opinion or working hypothesis is that the first replicated molecule had effectively no asymmetric carbon. 30

Some claim that the problem of the origin of chiral molecules has been solved (May 2022), but as far as the scientific literature illucidates, that is not the case. Following are a few quotes:
Homochirality is a common feature of amino acids and carbohydrates, and its origin is still unknown. (September 24, 2020) 26
The origin of homochirality in L-amino acid in proteins is one of the mysteries of the evolution of life. (30 November 2018) 27
How L-chiral proteins emerged from demi-chiral mixtures is unknown.The lack of understanding of the origins of the breaking of demi-chirality found in the molecules of life on Earth is a long-standing problem (December 26, 2019) 28
How homochirality concerning biopolymers (DNA/RNA/proteins) could have originally occurred (i.e., arisen from a non-life chemical world, which tended to be chirality-symmetric) is a long-standing scientific puzzle.
(January 8, 2020) 29

Why only left-handed, and not right-handed amino acids? 
Apparently, there is no deeper functional reason or justification that makes it necessary that they are left, rather than right-handed, which are no less stable and no more reactive 45. All they need is to be pure, that is, not mixed with left-handed ones. 

Viviane Richter wrote an article for cosmosmagazine in 2015:

It didn’t have to be that way. When life first emerged, why did it choose left and not right? Steve Benner believes biology picked left by chance. Malcolm Walter, an astrobiologist at the Australian Centre for Astrobiology at the University of New South Wales agrees. He also doubts we’ll ever come up with a definitive answer for why biology decided to be a lefty. “It’s going to remain speculative for a very long time – if not forever!” 44

From prebiotic to biotic chirality determination
The formation of the left-handedness of amino acids is performed in cells by a group of enzymes called aminotransferase through a transaminase reaction. The transamination reaction involves the transfer of an amino group for example by one of these enzymes, Aspartate Transaminase AST  37 from a donor, like an aspartate amino acid,  to the carbon atom of an alpha-keto acid 35, the acceptor, so that once the alpha-keto acid ring receives that amino group it will be converted into a glutamate amino acid ( the product). An example of an alpha-keto acid is an alpha-ketoglutarate (  Alpha-ketoglutarate (AKG) is a key molecule in the Krebs cycle [ or tricarboxylic acid TCA ] cycle determining the overall rate of the citric acid cycle of the organism. 36) By losing the amino group, the aspartate amino acid is transformed into oxaloacetate. And by receiving an amino group, alpha-ketoglutarate is being transformed into glutamate. In order to perform this reaction,  AST requires pyridoxal 5′ phosphate (P5P) as an essential cofactor for maximum enzyme activity. P5P is the active metabolite of vitamin B6, therefore it is a  reduced vitamin B6 ( it is used in hundreds of enzymes ) P5P serves as a molecular shuttle for ammonia and electrons between the amino donor and the amino acceptor. 18 different proteinogenic amino acids can be used as the starting point of the reaction.   The reaction can be an anabolic reaction to make amino acids or catabolic to make waste products, like nitrogenous waste urea, and released from the body as a toxic product. Aspartate aminotransferase (AST) has high specificity to operate with alpha-ketoglutarate. 

This is a complex process. The literature on ASTs spans approximately 60 years, and much fundamental mechanistic information on PLP-dependent reactions has been gained from its study. 47 but even in 2019 it was still not fully understood despite being "one of the most studied enzymes of this category" 46

Aspartate Aminotransferase
Since left-handedness is life-essential,  AST is a key metabolic enzyme, and its origin has to be ancient and be part of the minimal proteome and enzymatic setup of the first life forms. It is found in bacterial to eukaryotic species.  

The authors, Mei Han, and colleagues, reported in a scientific paper from 2021:
Aspartate Aminotransferase is present in all of the free-living organisms AST is a much-conserved enzyme found in both prokaryotes and eukaryotes and is closely linked to purine’s biosynthesis salvage pathway as well as the glycolytic and oxidative phosphorylation pathways. 48

1. Craig Venter: Life: What A Concept! 2008 
2. A. G. Cairns-Smith:  Genetic Takeover: And the Mineral Origins of Life 
3. Robert M. Hazen: Fundamentals of Geobiology 2012 
4. Fred Hoyle: The Intelligent Universe   1983
5. Nir Goldman: Synthesis of glycine-containing complexes in impacts of comets on early Earth 12 September 2010 
6. PIERAZZO  Amino acid survival in large cometary impacts 1999 
7. Hugh Ross: Could Impacts Jump-Start the Origin of Life? November 8, 2010 
8.Yasuhiro Oba: Identifying the wide diversity of extraterrestrial purine and pyrimidine nucleobases in carbonaceous meteorites 26 April 2022 
9. About Liz Kruesi: All of the bases in DNA and RNA have now been found in meteorites 
10. Jamie E. Elsila: Meteoritic Amino Acids: Diversity in Compositions Reflects Parent Body Histories 2016 Jun 22 
11. E A Martell: Radionuclide-induced evolution of DNA and the origin of life 
12. Brian C. Lacki: THE LOG LOG PRIOR FOR THE FREQUENCY OF EXTRATERRESTRIAL INTELLIGENCES September 21, 2016  
13. Stanley L. Miller A Production of Amino Acids Under Possible Primitive Earth Conditions May 15, 1953 
14. JEFFREY L. BADA: Prebiotic Soup--Revisiting the Miller Experiment 
15. ADAM P. JOHNSON: The Miller Volcanic Spark Discharge Experiment 17 Oct 2008 
16. Eric T. Parker: Primordial synthesis of amines and amino acids in a 1958 Miller H2S-rich spark discharge experiment March 21, 2011 
17. [url=https://www.usgs.gov/faqs/what-gases-are-emitted-kilauea-and-other-active-volcanoes#:~:text=Ninety%2Dnine percent of the,and other minor gas species.]What gases are emitted by Kīlauea and other active volcanoes? [/url]
18. J W Delano: Redox history of the Earth's interior since approximately 3900 Ma: implications for prebiotic molecules Aug-Oct 2001 
19. Eric T. Parker: Conducting Miller-Urey Experiments 2014 Jan 21 
20. Dr. Stanley L. Miller: From Primordial Soup to the Prebiotic Beach An interview with exobiology pioneer 
21. Hugh Ross, Fazale Rana,  Origins of Life, page 73 
22. Donna G. Blackmond: The Origin of Biological Homochirality 2010 May; 2 
23. https://www.nature.com/scitable/definition/amino-acid-115/
24. A. G. CAIRNS-SMITH Seven clues to the origin of life, page 58 
25. A. G. CAIRNS-SMITH genetic takeover 1988  
26. Shubin Liu: Homochirality Originates from the Handedness of Helices September 24, 2020 
27. Tadashi Ando: Principles of chemical geometry underlying chiral selectivity in RNA minihelix aminoacylation 30 November 2018 
28. Jeffrey Skolnick:  On the possible origin of protein homochirality, structure, and biochemical function December 26, 2019 
29. Yong Chen: The origin of biological homochirality along with the origin of life January 8, 2020 
30. From Primordial Soup to the Prebiotic Beach An interview with exobiology pioneer, Dr. Stanley L. Miller, 
31. Change Laura Tan, Rob Stadler: The Stairway To Life: An Origin-Of-Life Reality Check  March 13, 2020 
32. Daniel P. Glavin:  The Search for Chiral Asymmetry as a Potential Biosignature in our Solar System November 19, 2019 
33. 
34. Davide Castelvecchi: ‘Elegant’ catalysts that tell left from right scoop chemistry Nobel 06 October 2021
35. About: Keto acid: 
36. Nan Wu: Alpha-Ketoglutarate: Physiological Functions and Applications 2016 Jan 24 
37. Daniel Nelson: Amino Group: Definition And Examples  2, November 2019 
38. What is Carboxylic Acid? 
39. Introduction to Amines – Compounds Containing Nitrogen 
40. alpha carbon: 
41. [url=http://www.chem.ucla.edu/~harding/IGOC/R/r_group.html#:~:text=R group%3A An abbreviation for,halogens%2C oxygen%2C or nitrogen.]R-group: [/url]
42. Guillaume Borrel: Unique Characteristics of the Pyrrolysine System in the 7th Order of Methanogens: Implications for the Evolution of a Genetic Code Expansion Cassette 
43. Rare, but essential – the amino acid selenocysteine June 19, 2017 
44. Viviane Richter [url= Why]https://cosmosmagazine.com/science/biology/why-the-building-blocks-in-our-cells-turned-to-the-left/]Why building blocks in our cells turned left[/url] 10 August 2015
45. https://www.scripps.edu/newsandviews/e_20040920/onpress.html
46. Kumari Soniya: Transimination Reaction at the Active Site of Aspartate Aminotransferase: A Proton Hopping Mechanism through Pyridoxal 5′-Phosphate 
47. Michael D. Toney: [url= Aspartate]https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3946379/]Aspartate Aminotransferase: an old dog teaches new tricks [/url]2013 Oct 9.
48. Mei Han: l-Aspartate: An Essential Metabolite for Plant Growth and Stress Acclimation 2021 Apr; 26 
49. Amino acids :https://bio.libretexts.org/Courses/University_of_California_Davis/BIS_2A%3A_Introductory_Biology_(Easlon)/Readings/04.3%3A_Amino_Acids
50. https://en.wikibooks.org/wiki/Structural_Biochemistry/Volume_5#Modified_Amino_Acids
51. Carol Turse: Simulations of Prebiotic Chemistry under Post-Impact Conditions on Titan 2013 Dec 17 
52. Miller–Urey experiment https://en.wikipedia.org/wiki/Miller%E2%80%93Urey_experiment
53. Stanley L. Miller: [url= Prebiotic]https://global.oup.com/us/companion.websites/fdscontent/uscompanion/us/pdf/Rigoutsos/I-SampleChap.pdf]Prebiotic Chemistry on the Primitive Earth[/url] 2006
54. Norio Kitadai: Origins of building blocks of life: A review 29 July 2017
55. STANLEY L. MILLER AND HAROLD C. UREY: Organic Compound Synthesis on the Primitive Earth: Several questions about the origin of life have been answered, but much remains to be studied 31 Jul 1959
56. Jessica Wimmer and William Martin: Likely energy source behind first life on Earth found ‘hiding in plain sight’ January 19, 2022
57. Leslie E Orgel †: The Implausibility of Metabolic Cycles on the Prebiotic Earth  January 22, 2008 
58. Punam Dalai: [url=https://pubs.geoscienceworld.org/msa/eleme

Soo Yeon Jeong and colleagues described the enzyme in a scientific paper from 2019: L-aspartate aminotransferase (AST) is highly conserved across species and plays essential roles in varied metabolic pathways. It also regulates the cellular level of amino acids by catalyzing amino acid degradation and biosynthesis. AST generally forms a homodimer consisting of two active sites in the vicinity of subunit interfaces; these active sites bind to its cofactor PLP and substrate independently. Each subunit is composed of three parts: large domain, small domain, and N-terminal arm. The active site is situated in the cavity formed by two large domains and one small domain. 17

Proteopedia informs: It is a homodimer that is 413 amino acids long and serves a critical role in amino acid and carbohydrate metabolism 18

This is an enzyme that operates based on high specificity. Soo Yeon Jeong reported in the conclusion remarks even: "We observed the mode of intercommunication during catalytic reactions between two protomers of the dimer." There are several life-essential enzymes, that operate based on intrinsic signaling and communication ( ribosomes, aminoacyl tRNA synthetases) which indicates the high sophistication of these enzymes.  pyridoxal 5′ phosphate is furthermore an integral part of the enzymatic reaction, which indicates interdependence. It performs various key functions. Considering this all together, it is remarkable evidence of an intelligently designed setup.

How were the 20 proteinogenic amino acids selected on early earth?
Science is absolutely clueless about how and why specifically this collection of amino acids is incorporated into the genetic code to make proteins. Why 20, and not more or less? ( in some rare cases, 22) considering that many different ones could have been chosen? Stanly Miller wrote in the  science paper from 1981: Reasons for the Occurrence of the Twenty Coded Protein Amino Acids:

There are only twenty amino acids that are coded for in protein synthesis, along with about 120 that occur by post-translational modifications. Yet there are over 300 naturally-occurring amino acids known, and thousands of amino acids are possible. The question then is - why were these particular 20 amino acids selected during the process that led to the origin of the most primitive organism and during the early stages of Darwinian evolution. Why Are beta, gamma and theta Amino Acids absent? The selection of a-amino acids for protein synthesis and the exclusion of the beta, gamma, and theta amino acids raises two questions. First, why does protein synthesis use only one type of amino acid and not a mixture of various α, β, γ, δ… acids? Second, why were the a-amino acids selected? The present ribosomal peptidyl transferase has specificity for only a-amino acids. Compounds with a more remote amino group reportedly do not function in the peptidyl transferase reaction. The ribosomal peptidyl transferase has a specificity for L-a-amino acids, which may account for the use of a single optical isomer in protein amino acids. The chemical basis for the selection of a-amino acids can be understood by considering the deleterious properties that beta, theta, and gamma-amino acids give to peptides or have for protein synthesis. 1



The question is not only why not more or less were selected and are incorporated in the amino acid "alphabet", but also how they could/would have been selected from a prebiotic soup, ponds, puddles, or even the archaean ocean?
The ribosome core that performs the polymerization, or catenation of amino acids, joining one amino acid monomer to another,  the ribosomal peptidyl transferase center, only incorporates alpha-amino acids, as Joongoo Lee and colleagues explain in a scientific article from 2020:

Ribosome-mediated polymerization of backbone-extended monomers into polypeptides is challenging due to their poor compatibility with the translation apparatus, which evolved to use α-L-amino acids. Moreover, mechanisms to acylate (or charge) these monomers to transfer RNAs (tRNAs) to make aminoacyl-tRNA substrates is a bottleneck. The shape, physiochemical, and dynamic properties of the ribosome have been evolved to work with canonical α-amino acids 11

There are no physical requirements that dictate, that the ribosome should/could not be constructed capable to incorporate β, γ, δ… amino acids. Indeed, scientists work on polymer engineering, designing ribosomes that use an expanded amino acid alphabet. A 3D printer uses specifically designed polyester filaments to be fed with, that can process them, and print various objects based on the software information that dictates the product form. If someone tries to use raw materials that are inadequate, the printer will not be able to perform the job it was designed for. The ribosome is a molecular 3D nano printer, as Jan Mrazek and colleagues elucidate in a science paper published in 2014

Structural and functional evidence point to a model of vault assembly whereby the polyribosome acts like a 3D nanoprinter to direct the ordered translation and assembly of the multi-subunit vault homopolymer, a process which we refer to as polyribosome templating. 12 where the reaction center is also specifically adjusted to perform its reaction with the specific set of α-amino acids. 

The materials that the machine is fed with, and the machine itself have both to be designed from scratch, in order to function properly. One cannot operate with the adequacy of the other. There is a clear interdependence that indicates that the amino acid alphabet was selected to work with the ribosome as we know it.

From Georga Tech: The preference for the incorporation of the biological amino acids over non-biological counterparts also adds to possible explanations for why life selected for just 20 amino acids when 500 occurred naturally on the Hadean Earth. “Our idea is that life started with the many building blocks that were there and selected a subset of them, but we don’t know how much was selected on the basis of pure chemistry or how many biological processes did the selecting. Looking at this study, it appears today’s biology may reflect these early prebiotic chemical reactions more than we had thought,” said Loren Williams,  professor in Georgia Tech’s School of Chemistry and Biochemistry 4

The authors mention 500 supposedly extant on early earth. Maybe they got that number from a scientific article about nonribosomal peptides (NRPs) which coincides with that number of 500. Areski Flissi and colleagues write:

Secondary metabolites (nonribosomal peptides) are produced by bacteria and fungi. In fact, >500 different building blocks, called monomers, are observed in these peptides, such as derivatives of the proteinogenic amino acids, rare amino acids, fatty acids or carbohydrates. In addition, various types of bonds connect their monomers such as disulfide or phenolic bonds. Some monomers can connect with up to five other monomers, making cycles or branches in the structure of the NRPs. 5

Stuart A. Kauffman (2018) gives us an entirely different perspective. He wrote on page 22, in the section Discussion: Using the PubChem dataset and the Murchison meteorite mass spectroscopy data we could reconstruct the time evolution and managed to calculate the time of birth of amino acids, which is about 165 million years after the start of evolution. ( They mean after the Big Bang)  a mere blink of an eye in cosmological terms. All this puts the Miller-Urey experiment in a very different perspective. the results suggest that the main ingredients of life, such as amino acids, nucleotides and other key molecules came into existence very early, about 8-9 billion years before life. 6

Why should the number of possible amino acids on early earth be restricted to 500? In fact, as Allison Soult, a chemist from the University of Kentucky wrote: Any ( large ) number of amino acids can possibly be imagined.  7 

Steven Benner goes along with the same reasoning (2008): Conceptually, the number of compounds in gas clouds, meteorites, Titan, and laboratory simulations of early Earth is enormous, too many for any but a super-human imagination to start puzzling over. Each of those n compounds (where n is a large number) can react with any of the other compounds (for the mathematically inclined, this gives n 2 reactions). Of course, each of these n 2 products can react further. Thus, any useful scientific method must begin by constraining the enormity of possibilities that observations present to focus the minds of us mortal scientists. 19

This number is defacto limitless. The universe should theoretically be able to produce an infinite number of different amino acids. The AA R sidechains can have any isomer combination. They can come right-handed, or left-handed, with one or two functional groups, with cyclic (cyclobutane, cyclopentane, and cyclohexane) and/or branched structures, they can come amphoteric, aliphatic, aromatic, polar, uncharged, positively and negatively charged, and so on. Furthermore: A carbon atom bonded to a functional group, like carbonyl,  is known as the α carbon atom. The second is β (α, β, γ, δ…) and so on, according to the Greek alphabetical order. It is conceivable that the protein alphabet would be made of β peptides. There is nothing that physically constrains or limits amino acids to have different configurations. In fact, we do know bioactive peptides that use β-amino acids do form polymer sequences 3  Every synthetic chemist will confirm this. There is also no plausible reason why only hydrogen, carbon, nitrogen, oxygen, and sulfur should/could be used in a pool of 118 elements extant in the universe. If the number of possible AA combinations to form a set is limitless, then the chance of selecting randomly a specific set of AAs for specific functions is practically zero. It would have never happened by non-designed means. 

Science Daily reported in 2018, claiming that quantum chemistry supposedly solved the mystery of why there are these 20 amino acids in the genetic code. They wrote: "The newer amino acids had become systematically softer, i.e., more readily reactive or prone to undergo chemical changes. The transition from the dead chemistry out there in space to our own biochemistry here today was marked by an increase in softness and thus an enhanced reactivity of the building blocks." 14 The pertinent follow-up question then is:  Why the soft amino acids were added to the toolbox in the first place? What exactly were these readily reactive amino acids supposed to react with?
They answered: " At least some of the new amino acids, especially methionine, tryptophan, and selenocysteine, were added as a consequence of the increase in the levels of oxygen in the biosphere. This oxygen promoted the formation of toxic free radicals, which exposes modern organisms and cells to massive oxidative stress. The new amino acids underwent chemical reactions with the free radicals and thus scavenged them in an efficient manner. The oxidized new amino acids, in turn, were easily repairable after oxidation, but they protected other and more valuable biological structures, which are not repairable, from oxygen-induced damage. Hence, the new amino acids provided the remote ancestors of all living cells with a very real survival advantage that allowed them to be successful in the more oxidizing, "brave" new world on Earth. "With this in view, we could characterize oxygen as the author adding the very final touch to the genetic code" 

There are several problems with this hypothesis. 
1. if the prebiotic atmosphere were oxygenated, organic molecules like RNA and DNA would have been susceptible to thermal oxidation and photo-oxidation and would have readily been destroyed. 
2. Twelve of the proteinogenic amino acids were never produced in any lab experiment 15 
3. there was no selection process extant to sort out those amino acids best suited and used in life. ( those used are better than 2 million possible alternative amino acid "alphabets"  
4. There was no concentration process to collect the amino acids at one specific assembly site. 
5. There was no enantiomer selection process 
6. They would have disintegrated, rather than complexified 
7. There was no process to purify them. 

John Maynard Smith, a British biologist wrote in The Major Transitions in Evolution in 1997: Why does life use twenty amino acids and four nucleotide bases? It would be far simpler to employ, say, sixteen amino acids and package the four bases into doublets rather than triplets. Easier still would be to have just two bases and use a binary code, like a computer. If a simpler system had evolved, it is hard to see how the more complicated triplet code would ever take over. The answer could be a case of “It was a good idea at the time.” A good idea of whom?  If the code evolved at a very early stage in the history of life, perhaps even during its prebiotic phase, the numbers four and twenty may have been the best way to go for chemical reasons relevant at that stage. Life simply got stuck with these numbers thereafter, their original purpose lost. Or perhaps the use of four and twenty is the optimum way to do it. There is an advantage in life’s employing many varieties of amino acid, because they can be strung together in more ways to offer a wider selection of proteins. But there is also a price: with increasing numbers of amino acids, the risk of translation errors grows. With too many amino acids around, there would be a greater likelihood that the wrong one would be hooked onto the protein chain. So maybe twenty is a good compromise. Do random chemical reactions have knowledge to arrive at a optimal conclusion or a " good compromise"? 16

No, of course, chemical reactions have no knowledge, no know-how, no foresight, no goals. 

Optimality of the amino acid set that is used to encode proteins 
Gayle K. Philip (2011): The last universal common ancestor of contemporary biology (LUCA) used a precise set of 20 amino acids as a standard alphabet with which to build genetically encoded protein polymers. Many alternatives were also available, which highlights the question: what factors led biological evolution on our planet to define its standard alphabet? Here, we demonstrate unambiguous support that the standard set of 20 amino acids represents the possible spectra of size, charge, and hydrophobicity more broadly and more evenly than can be explained by chance alone. 2

We know that conscious intelligent agents with foresight are able to conceptualize and visualize apriori, a system of building blocks, like Lego bricks, that have a set of properties that optimally perform a specific function or/and task, that is intended to be achieved, and subsequently, we know that intelligent agents can physically instantiate the physical 3D object previously conceptualized. 

Lego bricks in their present form were launched in 1958. The interlocking principle with its tubes makes it unique and offers unlimited building possibilities. It's just a matter of getting the imagination going – and letting a wealth of creative ideas emerge through play. 8

Amino acids are analogous to Lego bricks. Bricks to build a house are made with the right stability, size, materials, and capacity of isolation for maintaining adequate narrow-range temperatures inside a house. Glass is made with transparency to serve as windows.  (Rare earth) Metals, plastic, rubber, etc. are made to serve as building blocks of complex machines. A mix of atoms will never by itself organize to become the building blocks of a higher-order complex integrated system based on functional, well-integrated, and matching sub-parts. But that is precisely what nature needs in order to complexify into the integrated systems-level organization of cells and multicellularity. We know about the limited range of unguided random processes. And we know the infinite range of engineering solutions that capable intelligent agents can instantiate. 

Gayle K. Philip continues: We performed three specific tests: we compared (in terms of coverage) (i) the full set of 20 genetically encoded amino acids for size, charge, and hydrophobicity with equivalent values calculated for a sample of 1 million alternative sets (each also comprising 20 members)  results showed that the standard alphabet exhibits better coverage (i.e., greater breadth and greater evenness) than any random set for each of size, charge, and hydrophobicity, and for all combinations thereof. Results indicate that life genetically encodes a highly unusual subset of amino acids relative to any random sample of what was prebiotically plausible. A maximum of 0.03% random sets out-performed the standard amino acid alphabet in two properties, while no single random set exhibited greater coverage in all three properties simultaneously. These results combine to present a strong indication that the standard amino acid alphabet, taken as a set, exhibits strongly nonrandom properties. Random chance would be highly unlikely to represent the chemical space of possible amino acids with such breadth and evenness in charge, size, and hydrophobicity (properties that define what protein structures and functions can be built). It is remarkable that such a simple starting point for analysis yields such clear results.

If the set does exhibit nonrandom properties, and random chance is highly unlikely, where does that optimality come from? It cannot be due to physical necessity. Matter has not the necessity to instantiate, to sort out a set of building blocks for distant goals. Evolution and natural selection is a hopelessly inadequate mechanism that was not at play at that stage. The only option left is intelligent design.

Melissa Ilardo (2015) : We compared the encoded amino acid alphabet to random sets of amino acids. We drew 10^8 random sets of 20 amino acids from our library of 1913 structures and compared their coverage of three chemical properties: size, charge, and hydrophobicity, to the standard amino acid alphabet. We measured how often the random sets demonstrated better coverage of chemistry space in one or more, two or more, or all three properties. In doing so, we found that better sets were extremely rare. In fact, when examining all three properties simultaneously, we detected only six sets with better coverage out of the 10^8 possibilities tested. Sets that cover chemistry space better than the genetically encoded alphabet are extremely rare and energetically costly. The amino acids used for constructing coded proteins may represent a largely global optimum, such that any aqueous biochemistry would use a very similar set. 9

That's pretty impressive and remarkable. That means, that only one in 16 million sets is better suited for the task. The most recent paper to be mentioned was written by Andrew J. Doig in 2016. He wrote:

Why the particular 20 amino acids were selected to be encoded by the Genetic Code remains a puzzle. They were selected to enable the formation of soluble structures with close-packed cores, allowing the presence of ordered binding pockets. Factors to take into account when assessing why a particular amino acid might be used include its component atoms, functional groups, biosynthetic cost, use in a protein core or on the surface, solubility and stability. Applying these criteria to the 20 standard amino acids, and considering some other simple alternatives that are not used, we find that there are excellent reasons for the selection of every amino acid. Rather than being a frozen accident, the set of amino acids selected appears to be near ideal.10 The last sentence is noteworthy: "the set of amino acids selected appears to be near ideal." It remains a puzzle as so many other things in biology that find no answer by the ones that build their inferences on a constraint set of possible explanations, where an intelligent causal agency is excluded a priori. Selecting things for specific goals is a conscious process, that requires intelligence. Attributes, that chance alone lacks, but an intelligent creator can employ to create life.

Biosynthetic cost: Protein synthesis takes a major share of the energy resources of a cell. Leu costs only 1 ATP, but its isomer Ile costs 11. Why would life ever, therefore, use Ile instead of Leu, if they have the same properties? Larger is not necessarily more expensive; Asn and Asp cost more in ATP than their larger alternatives Gln and Glu, and large Tyr costs only two ATP, compared to 15 for small Cys. The high cost of sulfur-containing amino acids is notable.

This is indeed completely counterintuitive and does not conform with naturalistic predictions.

Burial and surface: Proteins have close-packed cores with the same density as organic solids and side chains fixed into a single conformation. A solid core is essential to stabilize proteins and to form a rigid structure with well-defined binding sites. Nonpolar side chains have therefore been selected to stabilize close-packed hydrophobic cores. Conversely, proteins are dissolved in water, so other side chains are used on a protein surface to keep them soluble in an aqueous environment.

The problem here is that molecules and an arrangement of correctly selected varieties of amino acids would bear no function until life began. Functional subunits of proteins, or even fully operating proteins on their own would only have a function after life began, and the cells intrinsic operations were on the go. It is as if molecules had the inherent drive to contribute to life to have a first go, which of course is absurd. The only rational alternative is that a powerful creator had the foresight, and knew which arrangement and selection of amino acids would fit and work to make life possible.

Which amino acids came first? It is plausible that the first proteins used a subset of the 20 and a simplified Genetic Code, with the first amino acids acquired from the environment.

Why is plausible? It is not only not plausible, but plain and clearly impossible. The genetic code could not emerge gradually, and there is no known explanation for how it emerged. The author also ignores that the whole process of protein synthesis requires all parts in the process fully operational right from the beginning. A gradual development by evolutionary selective forces is highly unlikely.

Energetics of protein folding: Folded proteins are stabilized by hydrogen bonding, removal of nonpolar groups from water (hydrophobic effect), van der Waals forces, salt bridges, and disulfide bonds. Folding is opposed by loss of conformational entropy, where rotation around bonds is restricted, and introduction of strain. These forces are well balanced so that the overall free energy changes for all the steps in protein folding are close to zero.

Foresight and superior knowledge would be required to know how to get a protein fold that bears function, and where the forces are outbalanced naturally to get an overall energy homeostatic state close to zero. In the most recent  Christopher Mayer-Bacon (2021): Three fundamental physicochemical properties of size, charge, and hydrophobicity have received the most attention to date in identifying how the standard amino acid alphabet appears most clearly unusual. The standard amino acid alphabet appears more evenly distributed across a broader range of values than can reasonably be explained by chance. This model indicates a probability of approximately one in two million that an amino acid set would exhibit better coverage by chance 13

How is ammonium introduced to synthesize amino acids?

Unsolved issues about the origin of amino acids on early earth:
How did unguided nondesigned coincidence select the right amino acids amongst over 300 ( known, but the number is theoretically limitless ) that occur naturally on earth? All life on Earth uses the same 20 ( in some cases, 22 genetically encoded) amino acids to construct its proteins even though this represents a small subset of the amino acids available in nature?
How would twenty amino acids be selected (+2)  and not more or less to make proteins?
How was the concomitant synthesis of undesired or irrelevant by-products avoided?
How were bifunctional monomers, that is, molecules with two functional groups so they combine with two others selected, and unfunctional monomers (with only one functional group) sorted out?
How were β, γ, δ… amino acids sorted out?
How did a prebiotic synthesis of biological amino acids avoid the concomitant synthesis of undesired or irrelevant by-products?
How could achiral precursors of amino acids have produced and concentrated only left-handed amino acids? ( The homochirality problem )?
How did the transition from prebiotic enantiomer selection to the enzymatic reaction of transamination occur that had to be extant when cellular self-replication and life began?
How did ammonia (NH3), the precursor for amino acid synthesis, accumulate on prebiotic earth, if the lifetime of ammonia would be short because of its photochemical dissociation?
How could prebiotic events have delivered organosulfur compounds required in a few amino acids used in life, if in nature sulfur exists only in its most oxidized form (sulfate or SO4), and only some unique groups of procaryotes mediate the reduction of SO4 to its most reduced state (sulfide or H2S)?
How did a prebiotic synthesis of biological amino acids avoid the concomitant synthesis of undesired or irrelevant by-products?
How did the transition from prebiotic enantiomer selection to the enzymatic reaction of transamination occur that had to be extant when cellular self-replication and life began?
How did natural events have foreknowledge that the selected amino acids are best suited to enable the formation of soluble structures with close-packed cores, allowing the presence of ordered binding pockets inside proteins?
How did nature select the set of amino acids which appears to be near-optimal in regard to size, charge, and hydrophobicity more broadly and more evenly than in 16 million alternative sets?
How did natural events have foreknowledge that the selected amino acids are best suited to enable the formation of soluble structures with close-packed cores, allowing the presence of ordered binding pockets inside proteins?
How did Amino acid synthesis regulation emerge? Biosynthetic pathways are often highly regulated such that building blocks are synthesized only when supplies are low.
How did the transition from prebiotic synthesis to the synthesis through metabolic pathways of amino acids occur? A minimum of 112 enzymes is required to synthesize the 20 (+2) amino acids used in proteins.

1. S L Miller: Reasons for the occurrence of the twenty coded protein amino acids 1981 
2. Gayle K. Philip: Did evolution select a nonrandom 2011 Mar 24 
3. Chiara Cabrele: Peptides Containing β-Amino Acid Patterns: Challenges and Successes in Medicinal Chemistry September 10, 2014 
4. Pre-Life Building Blocks Spontaneously Align in Evolutionary Experiment 
5. Areski Flissi: Norine: update of the nonribosomal peptide resource 
6. Stuart A. Kauffman: Theory of chemical evolution of molecule compositions in the universe, in the Miller-Urey experiment and the mass distribution of interstellar and intergalactic molecules  30 Nov 2019
7. LibreTexts: Amino Acids
8. https://web.archive.org/web/20150905173143/http://www.lego.com/en-us/aboutus/lego-group/the_lego_history
9. Melissa Ilardo: Extraordinarily Adaptive Properties of the Genetically Encoded Amino Acids 24 March 2015 
10. Andrew J. Doig: Frozen, but no accident – why the 20 standard amino acids were selected 2 December 2016
11. Joongoo Lee: Ribosome-mediated polymerization of long chain carbon and cyclic amino acids into peptides in vitro 27 August 2020 
12. Jan Mrazek: Polyribosomes Are Molecular 3D Nanoprinters That Orchestrate the Assembly of Vault Particles 2014 Oct 30 
13. Christopher Mayer-Bacon: Evolution as a Guide to Designing xeno Amino Acid Alphabets 10 March 2021 
14. Quantum chemistry solves mystery why there are these 20 amino acids in the genetic code February 1, 2018 
15. Miller–Urey experiment 
16. John Maynard Smith: The Major Transitions in Evolution 1997
17. Soo Yeon Jeong: Crystal structure of L-aspartate aminotransferase from Schizosaccharomyces pombe August 29, 2019 
18. https://proteopedia.org/wiki/index.php/Aspartate_Aminotransferase#cite_note-AST_Structure-3
19. Benner SA  Life, the universe and the scientific method. (2009)

The problem of the prebiotic origin of amino acids
1. Amino acids are of a very specific complex functional composition and made by cells in extremely sophisticated orchestrated metabolic pathways, which were not extant on the early earth. If abiogenesis were true, these biomolecules had to be prebiotically available and naturally occurring ( in non-enzyme-catalyzed ways by natural means ) and then somehow join in an organized way.  Twelve of the proteinogenic amino acids were never produced in sufficient concentrations in any lab experiment. There was no selection process extant to sort out those amino acids best suited and used in life, amongst those that were not useful. There was potentially an unlimited number of different possible amino acid compositions extant prebiotically. (The amino acids alphabet used in life is more optimal and robust than 2 million tested alternative amino acid "alphabets")  
2. There was no concentration process to collect the amino acids at one specific assembly site. There was no enantiomer selection process ( the homochirality problem). Amino acids would have disintegrated, rather than complexified There was no process to purify them.
3. Taken together, all these problems make an unguided origin of Amino Acids extremely unlikely. Making things for a specific purpose, for a distant goal, requires goal-directedness. We know that a) unguided random purposeless events are unlikely to the extreme to make specific purposeful elementary components to build large integrated macromolecular systems, and b) intelligence has goal-directedness. Bricks do not form from clay by themselves, and then line up to make walls. Someone made them.

Serine Synthesis:

Phosphoserine phosphatase (EC 3.1.3.3)
Phosphoserine aminotransferase (EC 2.6.1.52)
Glycine Synthesis:
3. Serine hydroxymethyltransferase (EC 2.1.2.1)

Cysteine Metabolism:
4. Serine O-acetyltransferase (EC 2.3.1.30)

Cysteine synthase (EC 2.5.1.47)
Methionine adenosyltransferase (EC 2.5.1.6)
S-Adenosylhomocysteine hydrolase (EC 3.3.1.1)
Cystathionine gamma-synthase (EC 2.5.1.48)
Alanine Metabolism:
9. Aspartate 4-decarboxylase (EC 4.1.1.12)

Alanine transaminase (EC 2.6.1.2)
Alanine-glyoxylate transaminase (EC 2.6.1.44)
Alanine dehydrogenase (EC 1.4.1.1)
Alanine racemase (EC 5.1.1.1)
Valine biosynthesis:
14. Acetolactate synthase (EC 2.2.1.6)

Acetohydroxy acid isomeroreductase (EC 1.1.1.86)
Dihydroxyacid dehydratase (EC 4.2.1.9)
Branched-chain amino acid aminotransferase (EC 2.6.1.42)

Leucine Biosynthesis in Bacteria:

Acetolactate synthase (EC 2.2.1.6)
Acetohydroxy acid isomeroreductase (EC 1.1.1.86)
Dihydroxy-acid dehydratase (EC 4.2.1.9)
3-isopropylmalate synthase (EC 2.3.3.13)
3-isopropylmalate dehydratase (EC 4.2.1.33)
3-isopropylmalate dehydrogenase (EC 1.1.1.85)
Branched-chain amino acid aminotransferase (EC 2.6.1.42)
Isoleucine Metabolism:
8. Threonine deaminase (EC 4.3.1.19)

3-methyl-2-oxobutanoate hydroxymethyltransferase (EC 2.1.2.11)
3-isopropylmalate dehydratase (EC 4.2.1.33) (Note: This enzyme is listed twice since it's also involved in leucine biosynthesis)
3-isopropylmalate dehydrogenase (EC 1.1.1.85) (Note: This enzyme is also involved in leucine biosynthesis)
Histidine Synthesis:
12. Phosphoribosylamine--glycine ligase (EC 6.3.4.13)

Phosphoribosylformylglycinamidine synthase (EC 6.3.5.3)
Phosphoribosylformylglycinamidine cyclo-ligase (EC 6.3.3.1)
AIR carboxylase (EC 4.1.1.21)
Phosphoribosylformimino-5-amino-1-(5-phosphoribosyl)imidazolecarboxamide isomerase (EC 5.3.1.16)
Imidazoleglycerol-phosphate synthase (EC 4.1.3.15)
Imidazoleglycerol-phosphate hydrolase (EC 3.13.1.5)
Histidinol-phosphate aminotransferase (EC 2.6.1.9)
Histidinol-phosphate phosphatase (EC 3.1.3.15)
Histidinol dehydrogenase (EC 1.1.1.23)
Histidine ammonia-lyase (EC 4.3.1.3)
Phenylalanine/Tyrosine Synthesis:
23. Chorismate mutase (EC 5.4.99.5)

For Tyrosine synthesis:
24. Prephenate dehydrogenase (EC 1.3.1.12)

4-Hydroxyphenylpyruvate dioxygenase (EC 1.13.11.27)
Homogentisate 1,2-dioxygenase (EC 1.13.11.5) (Noted for completeness)
For Phenylalanine synthesis:
27. Prephenate aminotransferase (EC 2.6.1.78)

Arogenate dehydratase (EC 4.2.1.91)

Tryptophan Synthesis:

Chorismate pyruvate-lyase (EC 4.2.99.21)
Anthranilate phosphoribosyltransferase (EC 2.4.2.18)
Phosphoribosylanthranilate isomerase (EC 5.3.1.24)
Indole-3-glycerol-phosphate synthase (EC 4.1.1.48)
Tryptophan synthase (EC 4.2.1.20)
Aspartate Metabolism:
6. Aspartate transaminase (EC 2.6.1.1)

Aspartate carbamoyltransferase (EC 2.1.3.2)
Aspartokinase (EC 2.7.2.4)
Adenylosuccinate synthase (EC 6.3.4.4)
Asparagine Metabolism:
10. Asparagine synthetase (EC 6.3.5.4)

Asparaginase (EC 3.5.1.1)
Asparagine aminotransferase (EC 2.6.1.14)
Methionine Metabolism:
13. Homoserine dehydrogenase (EC 1.1.1.3)

O-succinylhomoserine (thiol)-lyase (EC 2.5.1.48)
Cystathionine beta-lyase (EC 4.4.1.
Methionine synthase (EC 2.1.1.13)
Methylthiotransferase (EC 2.8.4.4)
Lysine Biosynthesis:
18. Dihydrodipicolinate synthase (EC 4.2.1.52)

Dihydrodipicolinate reductase (EC 1.3.1.26)
2,3,4,5-tetrahydropyridine-2,6-dicarboxylate N-succinyltransferase (EC 2.3.1.117)
2,3,4,5-tetrahydropyridine-2,6-dicarboxylate N-acetyltransferase (EC 2.3.1.89)
Diaminopimelate reductase (EC 1.3.1.26)
Diaminopimelate epimerase (EC 5.1.1.7)
Diaminopimelate decarboxylase (EC 4.1.1.20)

Threonine Biosynthesis (from Aspartate):

Aspartokinase (EC 2.7.2.4)
Aspartate-semialdehyde dehydrogenase (EC 1.2.1.11)
Homoserine dehydrogenase (EC 1.1.1.3)
Homoserine kinase (EC 2.7.1.39)
Threonine synthase (EC 4.2.3.1)
Glutamine/Glutamate Synthesis:

Glutamate dehydrogenase (NAD+) (EC 1.4.1.2)
Glutamate dehydrogenase (NADP+) (EC 1.4.1.4)
Glutamate 5-kinase (EC 2.7.2.11)
Glutamine synthetase (EC 6.3.1.2)
Glutamine-dependent NAD+ synthetase (EC 6.3.5.1)
Arginine/Ornithine Synthesis:

N-acetylglutamate synthase (EC 2.3.1.1)
N-acetylglutamate kinase (EC 2.7.2.
N-acetyl-gamma-glutamyl-phosphate reductase (EC 1.2.1.38 )
Acetylornithine aminotransferase (EC 2.6.1.11)
Ornithine carbamoyltransferase (EC 2.1.3.3)
Argininosuccinate synthase (EC 6.3.4.5)
Argininosuccinate lyase (EC 4.3.2.1)
Arginine and Proline Metabolism in Prokaryotes:

Ornithine carbamoyltransferase (EC 2.1.3.3)
Ornithine decarboxylase (EC 4.1.1.17)
Acetylornithine deacetylase (EC 3.5.1.16)
Proline dehydrogenase (EC 1.5.5.2)
Pyrroline-5-carboxylate reductase (EC 1.5.1.2)

The list you provided is a comprehensive overview of various enzymes involved in amino acid metabolism. However, when considering the Last Universal Common Ancestor (LUCA) or simple bacteria resembling LUCA, it's important to understand that our understanding of LUCA is largely hypothetical. LUCA is not a specific organism we've observed, but rather an inferred ancestral entity reconstructed from comparative genomics and molecular phylogenetics.

Serine, Glycine, and Cysteine Metabolism: The enzymes you listed for the biosynthesis of serine, glycine, and cysteine are indeed ancient. For instance, serine is synthesized from 3-phosphoglycerate, an intermediate in glycolysis, highlighting its antiquity.

Alanine Metabolism: The enzymes for alanine metabolism, including transaminases, are expected to be ancient, as alanine is a simple amino acid and its synthesis and degradation tie into central metabolism.

Branched-chain Amino Acids (Valine, Leucine, Isoleucine): The synthesis pathways for these amino acids are interconnected, and while they involve several steps, they tie back to pyruvate and other central metabolites.

Histidine Synthesis: The synthesis of histidine is complex and whether LUCA had the full pathway is a matter of debate. However, it's plausible that some of the pathway, if not all, existed in early organisms.

Phenylalanine/Tyrosine Synthesis: The synthesis of these aromatic amino acids is tied to the shikimate pathway, which is ancient and leads to the synthesis of chorismate, a precursor for both amino acids.

Tryptophan Synthesis: Like phenylalanine and tyrosine, tryptophan synthesis also starts with chorismate.

Aspartate Family (Aspartate, Asparagine, Methionine, Threonine, Lysine): These amino acids are derived from aspartate, which is directly formed from oxaloacetate, an intermediate in the TCA cycle. This family is believed to be ancient.

Glutamate Family (Glutamate, Glutamine, Arginine, Proline): Glutamate is synthesized from alpha-ketoglutarate, another TCA cycle intermediate, making its synthesis pathway ancient.

It's worth noting that while the pathways themselves may be ancient, some specific enzymes or isoforms might be later evolutionary additions or innovations. Additionally, the actual presence of these pathways in simple bacteria resembling LUCA depends on the specific bacterium in question.

Overall, while the list you provided seems generally appropriate for ancient metabolic pathways, pinning down the exact biochemistry of LUCA is a challenging endeavor due to the deep time scales involved and the lack of direct evidence.

Serine Synthesis (3 enzymes):
Phosphoserine phosphatase (EC 3.1.3.3)
Phosphoserine aminotransferase (EC 2.6.1.52)
Serine hydroxymethyltransferase (EC 2.1.2.1)

Cysteine Metabolism (6 enzymes):
Serine O-acetyltransferase (EC 2.3.1.30)
Cysteine synthase (EC 2.5.1.47)
Methionine adenosyltransferase (EC 2.5.1.6)
S-Adenosylhomocysteine hydrolase (EC 3.3.1.1)
Cystathionine gamma-synthase (EC 2.5.1.48)

Alanine Metabolism (5 enzymes):
Aspartate 4-decarboxylase (EC 4.1.1.12)
Alanine transaminase (EC 2.6.1.2)
Alanine-glyoxylate transaminase (EC 2.6.1.44)
Alanine dehydrogenase (EC 1.4.1.1)
Alanine racemase (EC 5.1.1.1)

Valine biosynthesis (4 enzymes):
Acetolactate synthase (EC 2.2.1.6)
Acetohydroxy acid isomeroreductase (EC 1.1.1.86)
Dihydroxyacid dehydratase (EC 4.2.1.9)
Branched-chain amino acid aminotransferase (EC 2.6.1.42)

Leucine Biosynthesis in Bacteria (7 enzymes):
Acetolactate synthase (EC 2.2.1.6)
Acetohydroxy acid isomeroreductase (EC 1.1.1.86)
Dihydroxy-acid dehydratase (EC 4.2.1.9)
3-isopropylmalate synthase (EC 2.3.3.13)
3-isopropylmalate dehydratase (EC 4.2.1.33)
3-isopropylmalate dehydrogenase (EC 1.1.1.85)
Branched-chain amino acid aminotransferase (EC 2.6.1.42)

Isoleucine Metabolism (6 enzymes):
Threonine deaminase (EC 4.3.1.19)
3-methyl-2-oxobutanoate hydroxymethyltransferase (EC 2.1.2.11)
3-isopropylmalate dehydratase (EC 4.2.1.33)
3-isopropylmalate dehydrogenase (EC 1.1.1.85)

Histidine Synthesis (10 enzymes):
Phosphoribosylamine--glycine ligase (EC 6.3.4.13)
Phosphoribosylformylglycinamidine synthase (EC 6.3.5.3)
Phosphoribosylformylglycinamidine cyclo-ligase (EC 6.3.3.1)
AIR carboxylase (EC 4.1.1.21)
Phosphoribosylformimino-5-amino-1-(5-phosphoribosyl)imidazolecarboxamide isomerase (EC 5.3.1.16)
Imidazoleglycerol-phosphate synthase (EC 4.1.3.15)
Imidazoleglycerol-phosphate hydrolase (EC 3.13.1.5)
Histidinol-phosphate aminotransferase (EC 2.6.1.9)
Histidinol-phosphate phosphatase (EC 3.1.3.15)
Histidinol dehydrogenase (EC 1.1.1.23)

Phenylalanine/Tyrosine Synthesis (6 enzymes):
Chorismate mutase (EC 5.4.99.5)
Prephenate dehydrogenase (EC 1.3.1.12)
4-Hydroxyphenylpyruvate dioxygenase (EC 1.13.11.27)
Prephenate aminotransferase (EC 2.6.1.78)
Arogenate dehydratase (EC 4.2.1.91)
Homogentisate 1,2-dioxygenase (EC 1.13.11.5)

Tryptophan Synthesis (5 enzymes):
Chorismate pyruvate-lyase (EC 4.2.99.21)
Anthranilate phosphoribosyltransferase (EC 2.4.2.18)
Phosphoribosylanthranilate isomerase (EC 5.3.1.24)
Indole-3-glycerol-phosphate synthase (EC 4.1.1.48)
Tryptophan synthase (EC 4.2.1.20)

Aspartate Metabolism (4 enzymes):
Aspartate transaminase (EC 2.6.1.1)
Aspartate carbamoyltransferase (EC 2.1.3.2)
Aspartokinase (EC 2.7.2.4)
Adenylosuccinate synthase (EC 6.3.4.4)

Asparagine Metabolism (3 enzymes):
Asparagine synthetase (EC 6.3.5.4)
Asparaginase (EC 3.5.1.1)
Asparagine aminotransferase (EC 2.6.1.14)

Methionine Metabolism (5 enzymes):
Homoserine dehydrogenase (EC 1.1.1.3)
O-succinylhomoserine (thiol)-lyase (EC 2.5.1.48 )
Cystathionine beta-lyase (EC 4.4.1.8 )
Methionine synthase (EC 2.1.1.13)
Methylthiotransferase (EC 2.8.4.4)

Cysteine Metabolism (3 enzymes):
Serine acetyltransferase (EC 2.3.1.30)
O-acetylserine(thiol)-lyase (EC 2.5.1.47)
Cystathionine beta-synthase (EC 4.2.1.22)

Threonine Metabolism (3 enzymes):
Threonine dehydratase (EC 4.3.1.19)
Homoserine kinase (EC 2.7.1.39)
Threonine synthase (EC 4.2.3.1)

Isoleucine/Valine Synthesis (6 enzymes):
Acetolactate synthase (EC 2.2.1.6)
Keto-acid reductoisomerase (EC 1.1.1.86)
Dihydroxy-acid dehydratase (EC 4.2.1.9)
Alpha-isopropylmalate synthase (EC 2.3.3.13)
Alpha-isopropylmalate isomerase (EC 4.2.1.33)
Branched-chain amino acid aminotransferase (EC 2.6.1.42)

Lysine Synthesis (9 enzymes):
Aspartokinase (EC 2.7.2.4)
Aspartate-semialdehyde dehydrogenase (EC 1.2.1.11)
Dihydrodipicolinate synthase (EC 4.2.1.52)
Dihydrodipicolinate reductase (EC 1.3.1.26)
Tetrahydrodipicolinate acylase (EC 3.5.1.18)
N-succinyldiaminopimelate aminotransferase (EC 2.6.1.17)
N-succinyldiaminopimelate desuccinylase (EC 3.5.1.18)
Diaminopimelate decarboxylase (EC 4.1.1.20)
Diaminopimelate epimerase (EC 5.1.1.7)

Proline Synthesis (3 enzymes):
Glutamate 5-kinase (EC 2.7.2.11)
Glutamyl-phosphate reductase (EC 1.2.1.41)
Pyrroline-5-carboxylate reductase (EC 1.5.1.2)

Arginine Synthesis (8 enzymes):
Argininosuccinate synthase (EC 6.3.4.5)
Argininosuccinate lyase (EC 4.3.2.1)
Arginine decarboxylase (EC 4.1.1.19)
Agmatinase (EC 3.5.3.11)
SpeB (EC 3.5.3.10)
Ornithine carbamoyltransferase (EC 2.1.3.3)
Arginase (EC 3.5.3.1)
N-acetylglutamate synthase (EC 2.3.1.1)

Alanine Metabolism (2 enzymes):
Alanine transaminase (EC 2.6.1.2)
Alanine-glyoxylate transaminase (EC 2.6.1.44)

Serine Metabolism (3 enzymes):
Phosphoglycerate dehydrogenase (EC 1.1.1.95)
Phosphoserine aminotransferase (EC 2.6.1.52)
Phosphoserine phosphatase (EC 3.1.3.3)

Glycine Metabolism (4 enzymes):
Glycine hydroxymethyltransferase (EC 2.1.2.1)
Glycine cleavage system (EC 1.4.4.2, 1.8.1.4, 2.1.2.10)
Serine hydroxymethyltransferase (EC 2.1.2.1)
Glycine N-methyltransferase (EC 2.1.1.20)

Glutamate Metabolism (4 enzymes):
Glutamate dehydrogenase (EC 1.4.1.3)
Glutamine synthetase (EC 6.3.1.2)
Glutamate synthase (EC 1.4.1.13)
Glutaminase (EC 3.5.1.2)

Glutamine Metabolism (2 enzymes):
Glutamine synthetase (EC 6.3.1.2)
Glutaminase (EC 3.5.1.2)

Lost City’ seabed rocks hold clues to Earth’s first amino acids

‘James Urquhart 2018-11-14

The amino acid tryptophan has been detected deep beneath the seafloor. Its discoverers believe it wasn’t life that created this chemical but geochemical processes. The finding supports the idea that alkaline deep-sea hydrothermal systems could have provided ideal conditions for producing the organic molecules necessary to kick-start life on Earth.

Amino acids are integral to life as the building blocks of all proteins, as well as enabling crucial biological functions in their own right. However, this throws up a ‘chicken and egg’ conundrum: if amino acids are required for – and made by – life then how did life get started without them?

Amino acids have been discovered on meteorites suggesting an extra-terrestrial abiotic source of the compounds. Meanwhile, experiments and theoretical calculations have shown that amino acids could have been synthesised abiotically on Earth under the right conditions. But no one has ever found a naturally occurring non-living mechanism for their production.

Source: 2018, Springer Nature Limited Optical microscopy image of a sample of serpentinite where the abiotically produce tryptophan was discovered. Image also show olivine (Ol) and magnetite (Mag), with the green arrow showing orientation of sample

Now, Bénédicte Ménez at Paris Diderot University, France and her colleagues have identified the amino acid tryptophan – and other organic molecules – preserved in rock samples taken from almost 175m below the mid-Atlanic ocean floor at the Lost City hydrothermal field. Ruling out biological sources and contamination, the researchers suggest these chemicals formed from interactions between seawater and mantle-derived rocks called serpentinites, which would have been abundant in the prebiotic Earth’s crust.

‘The first building blocks of life must first have been produced abiotically, so it is very exciting to have evidence for the production of amino acids in serpentinised peridotites,’ comments Andrew McCaig, who investigates hydrothermal systems at the University of Leeds, UK, and was not involved in the study.

To analyse the core samples, the team combined three high resolution imaging techniques that all pointed to tryptophan inside a network of clay nanopores. Although amino acids have been detected in fluids at the Lost City site before, their source was unknown. The team are confident that the tryptophan they identified was abiotic in origin because they found no evidence of biomarkers or other amino acids which would suggest a biological source.

Deep-sea synthesis

The researchers propose that the tryptophan could have been synthesised in the nanopores of the iron-rich clay when it formed during hydrothermal alteration of serpentinites. The team also detected indole, which is an intermediate organic compound in the synthesis of tryptophan via Friedel–Crafts-type reactions, and indicates the nanopore network may have acted as a confined microreactor to promote this reaction, while the iron-rich clay helped catalyse it and subsequently aided its preservation.

‘Hydrothermal alteration of mantle rocks never ceases to amaze me,’ says Frieder Klein, a geochemist at Woods Hole Oceanographic Institution, US. ‘This remarkable study corroborates the idea that hydrothermal systems, which likely existed throughout most of Earth’s history and possibly elsewhere in the solar system, have the potential for the abiotic synthesis of amino acids and possibly even more complex organic compounds.’

However, not everyone is convinced. ‘The spectroscopic evidence for the detection of tryptophan in this work is not terribly strong and so it seems very unlikely to me,’ says Jim Cleaves, who investigates geochemistry and the origin of life at the Tokyo Institute of Technology, Japan. He explains that some of the fluorescence spectra peaks do not match up very well, suggesting that the data could similarly fit other compounds. ‘Even if tryptophan was conclusively detected, it seems plausible there has been fluid flow within these samples providing an avenue for biological contamination.’

Jeffrey Bada, who investigates the geochemistry of amino acids at the Scripps Institution of Oceanography, US, agrees. ‘The supposed tryptophan smells like an artefact to me,’ he says. What’s more he questions its relevance to the origin of life. ‘Tryptophan is not an amino acid commonly thought to be part of the prebiotic compound inventory on the early Earth and elsewhere. It has only one codon in DNA, which has been interpreted as indicating it was a late addition.’

However, although tryptophan may not have been one of the original amino acids required for life to emerge, Ménez and colleagues argue that amino acids can act as biochemical precursors that catalyse the synthesis of sugars, aldehydes and nucleotide intermediates.


Challenges in Explaining Life's Origin from Seabed Amino Acids

1. Single Amino Acid Detection: Only tryptophan was identified:
- How representative is this of prebiotic amino acid synthesis?
- What about the other 19 proteinogenic amino acids?

2. Spectroscopic Evidence Concerns: Some experts question the data's reliability:
- How conclusive is the spectroscopic evidence for tryptophan?
- Could the data fit other compounds?

3. Contamination Possibility: Biological contamination hasn't been ruled out:
- How can we be certain the tryptophan isn't from fluid flow contamination?
- What methods could definitively prove abiotic origin?

4. Evolutionary Relevance: Tryptophan may be a late addition to life:
- How relevant is tryptophan to early life if it was incorporated later?
- What explains its single codon in DNA if it was an early amino acid?

5. Prebiotic Plausibility: Tryptophan isn't typically considered prebiotic:
- Why focus on tryptophan instead of simpler, more likely prebiotic amino acids?
- How does this align with other prebiotic chemistry theories?

6. Reaction Specificity: The proposed synthesis is complex:
- How likely are Friedel-Crafts reactions in this environment?
- What about side reactions or unwanted products?

7. Catalytic Role of Clay: Clay's role in synthesis and preservation is proposed:
- How specific is this catalysis to tryptophan?
- Could it produce a variety of organic compounds, complicating the picture?

8. Quantity and Concentration: Amount of tryptophan produced is unclear:
- Is the quantity sufficient to be relevant for prebiotic chemistry?
- How could it accumulate to useful concentrations?

9. Environmental Conditions: Deep-sea vents differ from early Earth surface:
- How would these molecules reach the surface?
- Are deep-sea conditions relevant to origin of life theories?

10. From Precursors to Life: Leap from amino acids to life remains unexplained:
- How would tryptophan lead to more complex biomolecules?
- What about the development of genetic code and metabolism?

While this discovery is intriguing, it doesn't resolve fundamental questions about life's origin. The leap from abiotic amino acid synthesis to self-replicating systems remains a significant challenge. Further research is needed to address these issues and strengthen the connection between deep-sea chemistry and the emergence of life.

 
 
 
 
 
 
Keywords
#amino acids, #origin of life, #abiogenesis, #chemical evolution
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

1.2. Simple organic molecules
1.3. Origin of the organic compounds on the prebiotic earth
1.4. Prebiotic Amino Acid Synthesis
1.4.1. Availability and challenges associated with major atoms required to synthesize amino acids
1.4.2. Availability of Chemical Precursors: Fundamental Obstacles
1.5. Quantity and Concentration: Challenges in Prebiotic Amino Acid Availability
1.6. Stability and Reactivity: The Prebiotic Amino Acid Paradox
1.7. Selection of the 20 proteinogenic amino acids on early earth
1.8. Optimality of the amino acid set used to encode proteins
1.9. The Requirement of Chiral Amino Acids: Unraveling the Mystery of Homochirality
1.10. Amplification of Enantiomeric Excess
1.11. The racemization of amino acids and polypeptides under natural conditions
1.12. Prebiotic Peptide Bond Formation
1.13. Thermodynamic and Kinetic Barriers to Polymerization
1.14. Thermodynamic and Kinetic Barriers to Prebiotic Polypeptide Formation
1.15. Sequence and Structure Formation in Prebiotic Protein Emergence: A Critical Analysis
1.16. Protein Folding and Chaperones
1.17. Metabolic Integration




1.3. Prebiotic Amino Acid Synthesis: Open Questions in a Naturalistic Origin

Amino acids are organic compounds that serve as the fundamental building blocks of proteins in living systems. These molecules consist of a central carbon atom bonded to an amino group (-NH2), a carboxyl group (-COOH), a hydrogen atom, and a variable side chain (R group) that gives each amino acid its unique properties. In living organisms, amino acids play following essential roles:

1. Protein synthesis: They are the monomers that link together to form the long chains of proteins, which are essential for virtually all biological processes.
2. Metabolic functions: Some amino acids serve as precursors for important biomolecules like neurotransmitters, pigments, and hormones.
3. Energy source: When carbohydrates are scarce, amino acids can be broken down to provide energy.

The importance of amino acids in life cannot be overstated. They are integral to the structure and function of enzymes, which catalyze biochemical reactions. They also contribute to cellular signaling, immune responses, and the transport of key molecules throughout organisms. While there are hundreds of amino acids found in nature, life predominantly uses a set of 20 standard amino acids to build proteins. This specific set is thought to have been evolutionarily selected for its ability to create a diverse array of protein structures and functions while maintaining a balance between complexity and efficiency. These 20 amino acids provide a wide range of chemical properties - including hydrophobic, hydrophilic, acidic, and basic characteristics - allowing for the creation of proteins with highly specialized structures and functions. The reasons for the selection of these particular 20 amino acids are still debated in the scientific community. Hypotheses range from their availability in the prebiotic world to their ability to form a complete and efficient "chemical toolkit" for life. Understanding the origin and selection of these 20 amino acids remains an active area of research in the fields of biochemistry and the origin of life studies.


Image source: Link 

The synthesis of amino acids under hypothesized prebiotic conditions faces substantial and unresolved challenges. A thorough exploration of these challenges reveals deep conceptual issues with current naturalistic explanations for the origin of life. This narrative seeks to examine these problems in a coherent and detailed manner, relying on current scientific evidence and avoiding any unwarranted assumptions of naturalistic mechanisms.

1.3.1. Availability and challenges associated with major atoms required to synthesize amino acids

Carbon (C)
a) Abundance: Carbon is relatively abundant, ranking 15th in the Earth's crust (about 0.025% by weight).
b) Availability: In the prebiotic world, carbon would have been available mainly as CO2 in the atmosphere and dissolved in water.
c) Challenges: 
   - Reducing CO2 to organic compounds requires energy and catalysts.
   - Forming complex carbon skeletons of amino acids from simple precursors.

Hydrogen (H)
a) Abundance: Hydrogen is the most abundant element in the universe but less common on Earth (0.14% of the Earth's crust by weight).
b) Availability: Mainly present in water (H2O) and in reduced form in the early Earth's atmosphere (H2, CH4, NH3).
c) Challenges:
   - Maintaining a reducing environment for amino acid synthesis.
   - Balancing hydrogen availability between water and organic compounds.

Oxygen (O)
a) Abundance: Oxygen is the most abundant element in the Earth's crust (46.6% by weight).
b) Availability: Mainly present in water (H2O) and minerals. Free oxygen was scarce in the early Earth's atmosphere.
c) Challenges:
   - Controlled incorporation of oxygen into amino acids without excessive oxidation.
   - Balancing the need for oxygen in amino acids with the potentially damaging effects of oxidation on other prebiotic molecules.

Nitrogen (N)
a) Abundance: Nitrogen is relatively scarce in the Earth's crust (0.002% by weight) but abundant in the atmosphere (78% by volume today).
b) Availability: In the prebiotic world, likely present as N2 in the atmosphere and as NH3 in solution.
c) Challenges:
   - Breaking the strong triple bond in N2 requires significant energy.
   - Incorporating nitrogen into complex organic molecules like amino acids.
   - Maintaining a sufficient concentration of reactive nitrogen species.

Sulfur (S)
a) Abundance: Sulfur is moderately abundant in the Earth's crust (0.042% by weight).
b) Availability: Likely present in volcanic emissions as H2S and in various mineral forms.
c) Challenges:
   - Incorporating sulfur into specific amino acids (cysteine and methionine).
   - Balancing the reactivity of sulfur compounds with their incorporation into stable organic molecules.

Availability of Chemical Precursors

The origin of life poses a fundamental challenge: understanding how the essential building blocks of life, such as RNA, amino acids, lipids, and carbohydrates, were assembled prebiotically. On modern Earth, these molecules are synthesized by complex metabolic networks within cells, but on prebiotic Earth, no such cellular machinery existed. The first challenge in explaining the origin of life is identifying where these building blocks came from and how they were selected for the emergence of life.

Robert M. Hazen (2012): In *Fundamentals of Geobiology*, Hazen presented a hypothesis about the emergence of natural selection on a molecular level. He suggested that molecular selection, where key molecules earned their roles in life’s origins, proceeded in many ways. Some molecules were inherently unstable or highly reactive, while others dissolved in oceans or bonded to unhelpful minerals. Over time, competition among molecules in different geochemical environments maintained a molecular equilibrium, but no evolution occurred. Hazen posited that in self-replicating chemical systems, competition eventually drove the emergence of natural selection, leading to increasingly complex autocatalytic networks. This ongoing process gave rise to new chemical pathways, overlaying the old, and driving the evolution of life. Link.

While Hazen’s explanation provides a plausible account of molecular competition, critics argue that it is speculative and lacks empirical evidence. Each of life’s building blocks—nucleotides, amino acids, phospholipids, and carbohydrates—is complex and requires sophisticated, integrated cellular machinery for synthesis in modern cells. Scientific investigations have so far failed to demonstrate that random chemical interactions could assemble functional biomolecules without some form of guidance.

Craig Venter (2008): Venter emphasized the importance of selection in biology, stating that no matter what we do in synthetic biology or genomics, selection is always part of the process. While Venter acknowledged that synthetic selection is a form of intelligent design, he pointed out that biology is one hundred percent dependent on selection. Link.

This insight highlights a significant problem for naturalistic explanations of the origin of life: natural mechanisms lack goal-directedness. On prebiotic Earth, a potentially unlimited variety of molecules existed. Without a mechanism for competition and selection among them, it is difficult to explain how the molecules essential for life were selected while others were discarded. The idea that competition and selection alone could generate life’s complexity without guidance is seen by some as an inadequate explanation for the emergence and organization of life.

Graham Cairns-Smith (1988): In his book *Genetic Takeover*, Cairns-Smith expressed skepticism about the gradual emergence of life through chemical evolution. He argued that even if a primordial system had developed the ability to convert carbon dioxide into D-glucose, for example, it would not have been a significant step toward life. Without the ability to pass on the secret of its success to offspring, such a system would be short-lived and insignificant. Cairns-Smith suggested that life’s emergence was not inevitable from the evolution of the cosmos through chemical evolution. Link.

Cairns-Smith’s perspective aligns with those who view chemical evolution as insufficient to explain the complex coordination and replication needed for life to persist. Random mixtures of chemicals in a prebiotic soup have never been shown to purify, select, and accumulate the specific building blocks necessary for life in an organized and purpose-driven process.

The origin of life remains an open question. While theories like those proposed by Hazen and Venter attempt to explain the role of molecular competition and selection in life’s emergence, they fall short of providing concrete mechanisms. The complexity and specificity of life’s building blocks suggest that a guiding force—whether through natural processes we do not yet understand or through intelligent design—may be necessary to explain the transition from non-living chemistry to living organisms. Cairns-Smith’s critique adds further weight to the argument that life’s origins are unlikely to be the result of unguided chemical evolution.

Availability of Chemical Precursors: Fundamental Obstacles

Amino acid synthesis requires the availability of specific chemical precursors, including fixed nitrogen, carbon sources, and organosulfur compounds. The scarcity of these elements under prebiotic conditions presents a major hurdle. Current evidence suggests that the availability of these precursors was inconsistent and insufficient to support widespread amino acid synthesis. 

The limited sources of fixed nitrogen, such as through abiotic nitrogen fixation by lightning or volcanic activity, were highly sporadic and inefficient. This leads to the fundamental question: how could early Earth consistently supply enough nitrogen to sustain the necessary prebiotic reactions? Similarly, carbon, in the form of CO₂ or CH₄, requires highly specific conditions to be converted into reactive organic molecules, but such conditions appear to have been rare.

Conceptual problem: Scarcity and Instability of Precursors
- Lack of consistent, widespread nitrogen and carbon sources under early Earth conditions
- Abiotic nitrogen fixation processes too sporadic to sustain necessary reactions

2. Fixed Nitrogen and Carbon: Insufficient Supply Chains
The availability of nitrogen in bioavailable forms (e.g., ammonia or nitrate) is critical for amino acid synthesis. However, nitrogen fixation on early Earth would have been limited to non-biological processes, such as sporadic lightning strikes or occasional volcanic activity. These events are inconsistent, making it improbable that sufficient amounts of fixed nitrogen could have been produced to fuel large-scale amino acid synthesis.

Furthermore, carbon must be in a reactive form to participate in organic synthesis. The challenge lies in how CO₂ or CH₄ would be consistently converted into useful organic molecules under prebiotic conditions. Without specific catalysts and environmental settings, this conversion process lacks the efficiency needed for sustained reactions.

Conceptual problem: Sporadic Nature of Key Fixation Processes
- Non-biological nitrogen fixation events too rare to support widespread synthesis
- Lack of evidence for continuous and efficient carbon conversion pathways

3. Organosulfur Compounds: Unresolved Challenges in Sulfur Incorporation
Certain amino acids, such as cysteine and methionine, require sulfur in reduced forms. On early Earth, sulfur primarily existed as sulfate (SO₄²⁻), an oxidized and unreactive form. For sulfur to be incorporated into amino acids, it would need to exist in a reduced state, such as hydrogen sulfide (H₂S). The processes required to reduce sulfur compounds in a prebiotic setting are complex and poorly understood.

Furthermore, there is little empirical evidence supporting the large-scale presence of reduced sulfur compounds necessary for amino acid synthesis. Without a reliable mechanism for sulfur reduction, the synthesis of organosulfur-containing amino acids remains a critical open question.

Conceptual problem: Lack of Mechanism for Sulfur Reduction
- No clear pathway for the reduction of oxidized sulfur compounds into reactive forms
- Difficulty explaining the availability of reduced sulfur under plausible early Earth conditions

4. Ammonia Stability: The Problem of Photochemical Decomposition
Ammonia (NH₃) serves as a crucial nitrogen source in prebiotic chemistry. However, ammonia is highly susceptible to photochemical dissociation under ultraviolet radiation, which was prevalent on early Earth. This process breaks ammonia down into nitrogen and hydrogen, rapidly depleting any available supply.

Without continuous replenishment, ammonia's instability under prebiotic conditions poses a significant obstacle to maintaining a nitrogen source sufficient for amino acid synthesis. The question remains: how could early Earth sustain stable concentrations of ammonia in the face of rapid photodecomposition?

Conceptual problem: Instability of Key Nitrogen Sources
- Ammonia dissociates quickly under UV radiation, reducing its availability
- No known mechanism to continuously replenish ammonia at the necessary rates

5. Specific Requirements for Amino Acid Synthesis: Environmental and Chemical Barriers
For amino acids to form naturally under prebiotic conditions, a set of precise requirements must be met:
- A consistent source of fixed nitrogen and carbon
- The availability of reduced sulfur compounds
- Continuous replenishment of ammonia to counteract photodecomposition
- Localized concentrations of precursors to facilitate efficient reactions
- Environmental conditions that simultaneously support the stability and reactivity of all necessary precursors

These requirements present significant barriers to natural, unguided synthesis. Early Earth would need to provide highly localized, specific environments capable of overcoming the inherent instabilities and scarcities of critical precursors. However, no plausible natural setting has yet been identified that meets these conditions, leaving an unresolved gap in the understanding of prebiotic chemistry.

Conceptual problem: Contradictions in Required Environmental Conditions
- Simultaneously meeting all the necessary conditions for amino acid synthesis seems implausible
- No identified natural environment can account for the complex, localized conditions required for precursor stability and reactivity

6. Implications for Prebiotic Chemistry: An Unsolved Mystery
The challenges outlined above point to deep conceptual issues in the naturalistic origin of amino acids. The scarcity and instability of precursors, combined with the highly specific environmental requirements, raise significant doubts about the feasibility of spontaneous amino acid synthesis under prebiotic conditions. Without a guided process or alternative explanation, the current naturalistic frameworks face critical gaps that remain unresolved by contemporary scientific research.

Open questions:
- How could early Earth environments consistently provide the necessary chemical precursors?
- What natural processes could account for the reduction of sulfur compounds and the stabilization of ammonia?
- How can prebiotic chemistry explain the complex, specific conditions required for amino acid formation?

These unresolved issues challenge the naturalistic narrative of life's origins and require deeper investigation into alternative mechanisms or processes that could have driven the emergence of life's building blocks.

1.4. Challenges of Prebiotic Peptide Bond Formation

The challenges of prebiotic peptide bond formation are multifaceted, as highlighted by recent empirical data and simulations ⁴. The thermodynamic and kinetic barriers present significant hurdles, with equilibrium concentrations of even short peptides like nonapeptides calculated to be exceedingly low under prebiotic conditions ⁵. These findings critically challenge current origin-of-life models that rely on the spontaneous formation of polypeptides in aqueous environments, especially considering the rapid racemization of amino acids that impedes the formation of homochiral peptides essential for functional biology ^[5]. To naturally form peptide bonds, numerous simultaneous requirements must be met, including high amino acid concentrations, energetically favorable conditions, homochirality, selective activation, catalytic surfaces, protection from hydrolysis, sequential polymerization, stable intermediate structures, environmental stability, and efficient concentration mechanisms ⁶. However, many of these requirements are contradictory or mutually exclusive under prebiotic conditions, posing significant challenges to the spontaneous formation of functional peptides essential for the emergence of life ⁷.

1.4.1. Quantitative Findings Challenging Conventional Theories

A critical examination of the formation of peptide bonds reveals significant thermodynamic and kinetic barriers. Recent empirical data and computer simulations illustrate these challenges starkly. For instance, the equilibrium concentration of a nonapeptide (nine amino acids long) such as glycine ([Gly]₉) in water at temperatures between 25°C and 37°C is calculated to be less than 10^-50 M. This implies that under prebiotic conditions, not even a single molecule of [Gly]₉ would likely exist, let alone the much larger polypeptides required for primitive life forms.

1.4.2. Implications for Current Scientific Models

These findings pose a critical challenge to the current origin-of-life (OoL) models, which often rely on the spontaneous formation of polypeptides in aqueous environments. The extremely low equilibrium concentrations of even short peptides significantly undermine the plausibility of these models. Furthermore, the rapid racemization of amino acids under natural conditions exacerbates the problem, as it would prevent the formation of homochiral peptides necessary for functional biology.

1.4.3. Specific Requirements for Naturalistic Peptide Formation

For peptide bond formation to occur naturally under prebiotic conditions, the following requirements must be met simultaneously:

1. High Concentration of Amino Acids
A significant accumulation of amino acids in a localized environment is crucial for peptide bond formation. In dilute conditions, the probability of amino acids colliding and reacting is minimal. Natural mechanisms that could lead to high concentrations include evaporation in shallow pools, adsorption onto mineral surfaces, and encapsulation within lipid vesicles or micelles. These processes concentrate amino acids, increasing the likelihood of interactions that lead to peptide bond formation.

2. Energetically Favorable Conditions
Peptide bond formation is thermodynamically unfavorable under standard conditions because it requires energy input to form the bond and release a water molecule (condensation reaction). Natural energy sources such as heat from geothermal vents, ultraviolet (UV) radiation from the sun, or electrical energy from lightning could provide the necessary activation energy. Additionally, cyclical processes like wet-dry cycles can shift the equilibrium toward peptide formation by removing water during the drying phase.

3. Homochirality
Life on Earth predominantly uses L-amino acids, and the incorporation of these exclusively is essential for the proper folding and function of peptides. In a prebiotic world, amino acids would likely be present in a racemic mixture (equal amounts of L- and D- forms). Mechanisms that could lead to homochirality include asymmetric synthesis influenced by chiral mineral surfaces, circularly polarized light favoring one enantiomer over the other, or selective degradation of one form, resulting in an excess of the other.

4. Selective Activation
Amino acids need to be activated to form peptide bonds selectively without engaging in unwanted side reactions. Activation could occur through natural catalysts or by forming energy-rich intermediates like amino acid adenylates or phosphates. For example, coupling agents such as cyanamide or imidazole could facilitate the activation. The challenge is to achieve activation under mild conditions that prevent side reactions like cyclization or decomposition.

5. Catalytic Surfaces
Mineral surfaces can act as catalysts by providing sites that facilitate the orientation and proximity of amino acids, thus enhancing peptide bond formation. Clays like montmorillonite have layered structures that can adsorb organic molecules. Metal sulfides present in hydrothermal vents could also serve as catalytic surfaces, providing electrons or facilitating redox reactions that drive peptide synthesis.

6. Protection from Hydrolysis
In aqueous environments, peptides are prone to hydrolysis, which breaks peptide bonds and reverts peptides back to amino acids. Protection mechanisms might include the formation of peptides in microenvironments with low water activity, such as salt crusts or ice matrices. Alternatively, encapsulation within lipid bilayers or binding to mineral surfaces could shield peptides from water molecules, reducing the rate of hydrolysis.

7. Sequential Polymerization
Functional peptides require a specific sequence of amino acids. Random polymerization is unlikely to yield biologically useful peptides. Template-directed synthesis is one possible mechanism, where existing polymers or mineral surfaces guide the addition of amino acids in a particular order. Specific environmental conditions might also favor the incorporation of certain amino acids over others, leading to non-random sequences.

8. Stable Intermediate Structures
Intermediate compounds formed during peptide synthesis must be stable enough to participate in further reactions without decomposing. Stability can be influenced by environmental factors such as pH, temperature, and the presence of stabilizing agents like metal ions. For example, metal ion coordination can protect intermediates by forming complexes that prevent decomposition.

9. Environmental Stability
A stable environment is necessary to maintain the delicate balance required for peptide formation. Frequent fluctuations in temperature, pH, or other conditions can disrupt the process. Environments like deep-sea hydrothermal vents or sheltered tidal pools may offer the necessary stability. Consistent conditions over extended periods increase the chances of successful peptide synthesis.

10. Efficient Concentration Mechanisms
Beyond initial concentration, mechanisms are needed to continually gather reactants and prevent the dilution of products. Physical processes such as evaporation, freezing, or the formation of lipid vesicles can concentrate amino acids and peptides. Microenvironments like porous rocks or clay matrices can trap molecules, effectively increasing their local concentrations and facilitating ongoing reactions.

1.4.4. Contradictions and Mutually Exclusive Conditions

Many of these requirements are mutually exclusive or contradictory under prebiotic conditions. For example, the need for high temperatures to drive peptide formation (Requirement #2) conflicts with the necessity to prevent racemization (Requirement #3), as higher temperatures accelerate racemization rates. Similarly, the need for an aqueous environment to provide a medium for reactions (Requirement #1) contradicts the requirement to protect peptides from hydrolysis (Requirement #6).

1.4.5. Illustrative Examples

Hydrothermal Vents: While hydrothermal vents provide the high temperatures and mineral surfaces that could facilitate peptide bond formation, the harsh conditions also lead to rapid hydrolysis and racemization of peptides.
Drying Lagoon Hypothesis: The theory that peptides could form in drying lagoons where water evaporates and concentrates amino acids faces the challenge of maintaining homochirality and preventing hydrolysis during subsequent wet-dry cycles.

Current naturalistic explanations for peptide bond formation under prebiotic conditions face significant challenges. The quantitative data indicating extremely low peptide concentrations, coupled with the rapid racemization of amino acids, strongly suggest that these processes are highly improbable without additional, yet-to-be-discovered mechanisms. The simultaneous fulfillment of all necessary conditions under naturalistic scenarios appears implausible given our current understanding.

Unresolved Challenges in Prebiotic Peptide Bond Formation

1. Thermodynamic and Kinetic Barriers
Recent empirical data and simulations reveal significant thermodynamic and kinetic obstacles to prebiotic peptide bond formation. The equilibrium concentrations of even short peptides like nonapeptides are calculated to be exceedingly low under prebiotic conditions. For instance, the equilibrium concentration of a glycine nonapeptide ([Gly]₉) in water at 25-37°C is less than 10^-50 M, effectively meaning not a single molecule would likely exist in a prebiotic setting.

Conceptual problems:
- Spontaneous formation of peptides is thermodynamically unfavorable in aqueous environments
- Kinetic barriers further impede the reaction, even if energy input is available
- No known prebiotic mechanism to overcome these fundamental physical constraints

2. Amino Acid Concentration and Stability
Prebiotic peptide formation requires high concentrations of amino acids in localized areas. However, maintaining such concentrations in primitive Earth environments poses significant challenges. Additionally, amino acids are prone to decomposition and side reactions under various conditions.

Conceptual problems:
- No clear mechanism for concentrating amino acids to levels required for peptide formation
- Difficulty in explaining the stability of amino acids over long periods in prebiotic environments
- Competing reactions that could deplete amino acid pools before peptide formation occurs

3. Chirality and Homochirality
Life as we know it utilizes exclusively L-amino acids. However, prebiotic synthesis would produce racemic mixtures of D- and L-amino acids. The rapid racemization of amino acids under natural conditions further complicates the formation of homochiral peptides necessary for functional biology.

Conceptual problems:
- No known prebiotic mechanism for selecting only L-amino acids
- Racemization occurs rapidly under many prebiotic conditions, working against homochirality
- The origin of biological homochirality remains unexplained by unguided processes

4. Selective Activation and Sequential Polymerization
Forming functional peptides requires not just the formation of peptide bonds, but the creation of specific sequences. This necessitates selective activation of amino acids and a mechanism for controlled, sequential polymerization.

Conceptual problems:
- No known prebiotic mechanism for selectively activating specific amino acids
- Difficulty in explaining how unguided processes could produce specific sequences required for functionality
- Lack of a plausible explanation for the origin of the genetic code linking amino acid sequences to nucleic acids

5. Protection from Hydrolysis
Peptide bonds are susceptible to hydrolysis, especially in aqueous environments likely present on the early Earth. For peptides to accumulate, they must be protected from this breakdown.

Conceptual problems:
- Hydrolysis is thermodynamically favored in water, working against peptide formation and stability
- No clear mechanism for protecting nascent peptides from hydrolysis in a prebiotic aqueous environment
- Difficulty reconciling the need for water as a reaction medium with its detrimental effects on peptide stability

6. Catalytic Surfaces and Mineral Interfaces
Some theories propose that mineral surfaces could have catalyzed peptide bond formation. However, experimental evidence for efficient, long-chain peptide synthesis on mineral surfaces under prebiotic conditions is lacking.

Conceptual problems:
- Limited evidence for efficient peptide synthesis on mineral surfaces under realistic prebiotic conditions
- Difficulty in explaining how mineral-catalyzed reactions could produce the diverse range of peptides required for life
- Lack of a clear mechanism for the transition from mineral-surface reactions to free-solution biochemistry

7. Energy Sources and Coupling
Peptide bond formation is endergonic and requires an energy source. In modern biology, this is typically provided by ATP, but the origin of such sophisticated energy coupling systems in a prebiotic context is problematic.

Conceptual problems:
- No clear prebiotic analog for the high-energy phosphate bonds used in modern biochemistry
- Difficulty in coupling available energy sources to peptide bond formation without sophisticated enzymes
- Lack of a plausible explanation for the origin of complex energy transduction systems

8. Environmental Stability and Cycles
The formation of complex peptides likely required stable environmental conditions over long periods. However, the early Earth was characterized by fluctuating and often extreme conditions.

Conceptual problems:
- Difficulty in reconciling the need for stable conditions with the dynamic nature of the early Earth
- No clear mechanism for maintaining consistent chemical environments conducive to peptide formation over geological timescales
- Lack of explanation for how primitive peptide-based systems could have survived environmental fluctuations

9. Functional Thresholds and Minimal Complexity
For peptides to contribute to the origin of life, they must reach a threshold of functional complexity. However, the minimal complexity required for life-supporting peptides is far greater than what can be reasonably expected from unguided prebiotic processes.

Conceptual problems:
- No clear pathway from simple, randomly formed peptides to the complex, functional proteins required for life
- Difficulty in explaining the origin of enzyme-like catalytic activity without invoking highly improbable chance events
- Lack of a plausible model for the emergence of the intricate protein folding and structure-function relationships observed in even the simplest living systems

10. Integration with Other Prebiotic Systems
The origin of life requires not just peptides, but their integration with other key components such as nucleic acids and lipids. Explaining how these distinct systems could have coemerged and become interdependent without guidance poses significant challenges.

Conceptual problems:
- No clear mechanism for the simultaneous emergence of peptides, nucleic acids, and lipids in a coordinated manner
- Difficulty in explaining the origin of the complex interdependencies observed in even the simplest living systems
- Lack of a plausible model for the emergence of the genetic code linking peptide sequences to nucleic acid information

In conclusion, the formation of peptides under prebiotic conditions faces numerous, interconnected challenges that remain unresolved. These issues span from basic chemical and physical constraints to the complex requirements of functional biological systems. Current scientific understanding lacks plausible, empirically supported explanations for how these challenges could be overcome through unguided processes alone. The cumulative improbability of simultaneously meeting all the necessary conditions for prebiotic peptide formation and their subsequent organization into functional biological systems presents a significant conceptual hurdle for naturalistic origin-of-life scenarios.

1.5. Quantity and Concentration: Challenges in Prebiotic Amino Acid Availability

The challenges in prebiotic amino acid availability, as outlined in recent scientific literature, highlight the significant quantitative and qualitative obstacles faced by current abiogenesis models. Computational models suggest the need for concentrations in the millimolar range, far exceeding known prebiotic synthesis capabilities ¹⁰. Experimental studies indicate low yields in peptide formation, necessitating initial amino acid concentrations orders of magnitude higher than achievable through current methods ⁸. The absence of eight "never-observed" proteinogenic amino acids in prebiotic synthesis experiments raises fundamental questions about the completeness of origin-of-life models ¹¹. Proposed concentration mechanisms like thermophoresis or mineral surface adsorption face challenges in selectivity and efficiency, emphasizing the complexity of achieving the required molecular densities for polymerization ⁹. Addressing these quantitative and qualitative requirements is crucial for advancing our understanding of the origin of life and refining abiogenesis hypotheses.

1.5.1. Quantitative Challenges

Recent computational models suggest that the formation of even the simplest self-replicating systems would require a minimum of 10^9 to 10^12 amino acid molecules (Lancet et al., 2018). This translates to local concentrations in the millimolar range, far exceeding those achievable through known prebiotic synthesis routes. Furthermore, studies on mineral-catalyzed peptide formation indicate that yields rarely exceed 1% under optimal laboratory conditions, implying that initial amino acid concentrations would need to be orders of magnitude higher to compensate for inefficient polymerization. These quantitative constraints severely limit the plausibility of "primordial soup" hypotheses. Most prebiotic synthesis experiments produce amino acids in micromolar concentrations at best, falling short of the required levels by several orders of magnitude. This discrepancy undermines the assumption that simple chemical processes could lead to the spontaneous emergence of complex biomolecules.

1.5.2. Requirements for Natural Occurrence

For the prebiotic synthesis and concentration of amino acids to occur naturally, the following conditions must be simultaneously met:

1. Presence of all 20 proteinogenic amino acids in sufficient quantities
2. Protection mechanisms against UV radiation and hydrolysis
3. Chirality selection to produce only L-amino acids
4. Concentration mechanisms to achieve millimolar levels
5. Absence of interfering molecules that could disrupt synthesis or polymerization
6. Stable pH and temperature conditions conducive to amino acid stability
7. Energy sources for synthesis and concentration processes
8. Selective surfaces or environments for amino acid accumulation
9. Mechanisms to prevent the preferential concentration of simpler, competing molecules
10. Pathways for the synthesis of the eight "never-observed" proteinogenic amino acids

These requirements must coexist in a prebiotic environment, presenting a formidable challenge to naturalistic explanations. Several of these conditions are mutually exclusive or contradictory. For instance, the energy sources required for synthesis (point 7) often lead to the breakdown of complex molecules, conflicting with the need for protection mechanisms (point 2). The "never-observed" amino acids present a particular challenge. Despite decades of prebiotic chemistry research, eight of the 20 proteinogenic amino acids have never been synthesized under plausible prebiotic conditions. These include arginine, lysine, histidine, tryptophan, methionine, asparagine, glutamine, and phenylalanine. Their absence in prebiotic synthesis experiments raises fundamental questions about the completeness of current origin-of-life models. Moreover, the concentration problem extends beyond mere quantity. Amino acids would need to accumulate at specific assembly sites to facilitate polymerization. Proposed mechanisms like thermophoresis or mineral surface adsorption face significant limitations in selectivity and efficiency (Baaske et al., 2007). The quantitative and qualitative requirements for prebiotic amino acid availability present substantial challenges to current naturalistic explanations for the origin of life. 

Unresolved Challenges in Prebiotic Amino Acid Availability

1. Quantitative Requirements and Concentration Dilemma
Recent computational models suggest that even the simplest self-replicating systems would require local amino acid concentrations in the millimolar range, far exceeding known prebiotic synthesis capabilities. Experimental studies on mineral-catalyzed peptide formation show yields rarely exceeding 1% under optimal laboratory conditions.

Conceptual problems:
- No known prebiotic mechanism can produce amino acid concentrations sufficient for life's emergence
- Vast discrepancy between required concentrations (millimolar) and those achievable through prebiotic synthesis (micromolar at best)
- Lack of plausible explanation for achieving the molecular densities necessary for polymerization without guided processes

2. Qualitative Completeness of Amino Acid Set
The absence of eight "never-observed" proteinogenic amino acids in prebiotic synthesis experiments poses a significant challenge. These include arginine, lysine, histidine, tryptophan, methionine, asparagine, glutamine, and phenylalanine.

Conceptual problems:
- No known prebiotic pathway for synthesizing all 20 proteinogenic amino acids
- Inability to explain the origin of the complete set of amino acids required for life
- Lack of a plausible mechanism for the coemergence of the missing amino acids with those more easily synthesized

3. Protection from Degradation
Amino acids are susceptible to degradation by UV radiation and hydrolysis in aqueous environments, likely present on the early Earth.

Conceptual problems:
- No clear mechanism for protecting amino acids from UV radiation in a prebiotic environment lacking an ozone layer
- Difficulty in reconciling the need for water as a reaction medium with its detrimental effects on amino acid stability
- Lack of explanation for how amino acids could accumulate over time without sophisticated protection mechanisms

4. Chirality Selection
Life exclusively uses L-amino acids, but prebiotic synthesis would produce racemic mixtures of D- and L-amino acids.

Conceptual problems:
- No known prebiotic mechanism for selecting only L-amino acids on a global scale
- Difficulty in explaining how a chiral preference could be maintained over time without biological systems
- Lack of plausible explanation for the origin of homochirality in prebiotic environments

5. Interference from Competing Molecules
Prebiotic environments likely contained a complex mixture of organic compounds, many of which could interfere with amino acid synthesis or polymerization.

Conceptual problems:
- No clear mechanism for selectively concentrating amino acids while excluding interfering molecules
- Difficulty in explaining how amino acids could outcompete simpler, more abundant molecules in prebiotic reactions
- Lack of a plausible model for the emergence of chemical selectivity without sophisticated biological machinery

6. Environmental Stability
The formation and accumulation of amino acids likely required stable pH and temperature conditions over long periods.

Conceptual problems:
- Difficulty in reconciling the need for stable conditions with the dynamic and often extreme nature of the early Earth
- No clear mechanism for maintaining consistent chemical environments conducive to amino acid stability over geological timescales
- Lack of explanation for how primitive amino acid-based systems could have survived environmental fluctuations

7. Energy Sources and Coupling
The synthesis and concentration of amino acids require energy input, but coupling this energy to specific chemical processes without enzymes is problematic.

Conceptual problems:
- No clear prebiotic analog for the sophisticated energy coupling systems observed in modern biochemistry
- Difficulty in explaining how available energy sources could drive amino acid synthesis and concentration without harmful side reactions
- Lack of a plausible model for the emergence of selective energy transduction in prebiotic systems

8. Selective Surfaces and Environments
Some theories propose that mineral surfaces or specific microenvironments could have concentrated amino acids. However, evidence for efficient, selective accumulation under prebiotic conditions is lacking.

Conceptual problems:
- Limited evidence for selective amino acid concentration on mineral surfaces under realistic prebiotic conditions
- Difficulty in explaining how surface-based concentration could transition to the solution-phase chemistry of life
- Lack of a clear mechanism for the coemergence of selective surfaces and the amino acids they supposedly concentrate

9. Concentration Mechanism Limitations
Proposed concentration mechanisms like thermophoresis or mineral surface adsorption face significant challenges in selectivity and efficiency.

Conceptual problems:
- No known prebiotic mechanism can achieve the degree of concentration required for amino acid polymerization
- Difficulty in explaining how concentration mechanisms could operate selectively on amino acids versus other organic molecules
- Lack of plausible explanation for the origin of the sophisticated concentration mechanisms observed in modern cells

10. Integration with Other Prebiotic Systems
The emergence of life requires not just amino acids, but their integration with other key components such as nucleic acids and lipids.

Conceptual problems:
- No clear mechanism for the simultaneous concentration and organization of diverse prebiotic molecules
- Difficulty in explaining the origin of the complex interdependencies between amino acids and other biomolecules
- Lack of a plausible model for the coemergence of the various molecular systems required for life

In conclusion, the availability of amino acids in prebiotic environments faces numerous interconnected challenges that remain unresolved. These issues span from basic chemical and physical constraints to the complex requirements of emerging biological systems. Current scientific understanding lacks plausible, empirically supported explanations for how these challenges could be overcome through unguided processes alone. The quantitative requirements for amino acid concentrations, coupled with the qualitative need for a complete set of proteinogenic amino acids, present formidable obstacles to naturalistic origin-of-life scenarios. The cumulative improbability of simultaneously meeting all the necessary conditions for prebiotic amino acid availability and their subsequent organization into functional biological systems presents a significant conceptual hurdle for abiogenesis hypotheses.

1.6. Stability and Reactivity: The Prebiotic Amino Acid Paradox

The origin of life theories faces a significant challenge in explaining how amino acids could have remained stable enough to accumulate in prebiotic environments while simultaneously being reactive enough to form peptides without enzymatic assistance. This analysis examines the stability-reactivity paradox and its implications for naturalistic explanations of abiogenesis. The stability-reactivity paradox concerning the prebiotic amino acid environment is a crucial aspect in understanding abiogenesis. Research has shown that amino acids exhibit varying stability in aqueous solutions at different temperatures, with half-lives ranging from a few days to several years, depending on the specific amino acid and environmental factors ¹². Additionally, the formation of peptides without enzymatic assistance is a significant challenge, as dehydration to form amide bonds is highly unfavorable in water ¹³. However, recent studies have demonstrated unique reactivity of free amino acids at the air-water interface, leading to the rapid formation of peptide isomers on a millisecond scale under ambient conditions, showcasing the potential for abiotic peptide synthesis in aqueous environments ¹³. These findings shed light on the delicate balance between stability and reactivity that must have existed in the prebiotic world to enable the accumulation of amino acids and the formation of essential biomolecules.

1.6.1. Quantitative Challenges

Studies on amino acid stability in aqueous solutions at various temperatures reveal a half-life ranging from a few days to several years, depending on the specific amino acid and environmental conditions (Radzicka & Wolfenden, 1996). For instance, at 25°C and neutral pH, the half-life of aspartic acid is approximately 253 days, while that of tryptophan is about 74 days. However, these half-lives decrease dramatically at higher temperatures, which are often invoked in prebiotic scenarios. At 100°C, most amino acids have half-lives of less than a day.

Conversely, the rate of spontaneous peptide bond formation between amino acids in aqueous solutions is extremely slow. Experimental studies have shown that the half-time for dipeptide formation at 25°C and pH 7 is on the order of 10^2 to 10^3 years (Martin et al., 2007). This presents a significant kinetic barrier to the formation of even short peptides under prebiotic conditions.

1.6.2. Implications for Current Models

These quantitative findings challenge the plausibility of current models for prebiotic peptide formation. The disparity between the rates of amino acid decomposition and peptide bond formation suggests that in most prebiotic scenarios, amino acids would degrade faster than they

could polymerize into functionally relevant peptides. This stability-reactivity paradox undermines the assumption that simple accumulation of amino acids in a primordial soup could lead to the spontaneous emergence of proto-proteins.

1.6.3. Requirements for Natural Occurrence

For the stability and reactivity of prebiotic amino acids to support the emergence of life, the following conditions must be simultaneously met:

1. Protection mechanisms against hydrolysis and thermal decomposition
2. Sufficient reactivity to form peptide bonds without enzymatic catalysis
3. Selective polymerization to form functional peptide sequences
4. Prevention of side reactions leading to unusable byproducts
5. Maintenance of a pH range that balances stability and reactivity (typically pH 7-9)
6. Temperature conditions that allow for both stability and reactivity
7. Presence of activating agents to facilitate peptide bond formation
8. Absence of competing molecules that could interfere with polymerization
9. Mechanisms to remove water, driving peptide bond formation
10. Recycling processes to regenerate degraded amino acids

These requirements must coexist in a prebiotic environment, presenting a formidable challenge to naturalistic explanations. Several of these conditions are mutually exclusive or contradictory. For example, the need for protection against hydrolysis (point 1) conflicts with the requirement for sufficient reactivity (point 2). Similarly, the presence of activating agents (point 7) often leads to increased rates of side reactions (conflicting with point 4).

The stability-reactivity paradox is further illustrated by the "aspartic acid problem." Aspartic acid, a crucial amino acid in many proteins, is particularly prone to cyclization reactions, forming unreactive succinimide derivatives. Studies have shown that at pH 7 and 37°C, about 4% of aspartic acid residues in a peptide chain will convert to succinimides within 24 hours (Geiger & Clarke, 1987). This cyclization not only removes aspartic acid from the pool of available monomers but also disrupts the integrity of any formed peptides.

The requirement for water removal to drive peptide bond formation (point 9) contradicts the aqueous environment typically assumed in prebiotic scenarios. Proposed solutions, such as wet-dry cycles or mineral surface catalysis, introduce additional complexities and limitations.

The stability and reactivity requirements for prebiotic amino acids present substantial challenges to current naturalistic explanations for the origin of life. Future discussions on this topic should focus on:
1. Developing more realistic models that account for the stability-reactivity paradox.
2. Investigating novel mechanisms that could simultaneously protect and activate amino acids.
3. Exploring the potential role of non-aqueous environments in early peptide formation.
4. Addressing the mutual exclusivity of certain required conditions in prebiotic scenarios.
5. Critically examining the assumptions underlying current abiogenesis hypotheses in light of these kinetic and thermodynamic challenges.

By rigorously addressing these points, the scientific community can work towards a more comprehensive and evidence-based understanding of the chemical processes that could have led to the emergence of life.

1.7. Thermodynamic and Kinetic Barriers to Polymerization

The challenges of polymerization in water, especially for polypeptides like [Gly]n, are well-documented due to both thermodynamic and kinetic barriers, leading to equilibrium concentrations as low as < 10^-50 M at temperatures of 25° - 37°, making the existence of even short polypeptides like [Gly]9 highly improbable ^14 15. Recent studies by Dr. Royal Truman, Dr. Charles McCombs, and Dr. Change Tan further emphasize the difficulties by outlining nine additional requirements for OoL-relevant polypeptides, including the need for specific sequences, three-dimensional structures, continuous production, and self-replication, all of which pose significant challenges under natural conditions ^[14]. These stringent requirements, such as the need for about 300 amino acids to form proteins and the exclusion of nonbiological amino acids, highlight the complex interplay of factors that must be simultaneously satisfied for peptides/proteins to be relevant in origin-of-life scenarios, presenting a formidable obstacle for OoL discussions ^[14].

Polypeptides do not form in water at any temperature for thermodynamic and kinetic reasons.
Detailed quantitative analysis shows extremely low equilibrium concentrations of even short polypeptides.
The concentration of [Gly]9 would converge to < 10^-50 M at equilibrium in water at temperatures of 25° - 37°.
Nine additional requirements for OoL-relevant polypeptides are outlined, all of which violate fundamental chemical and statistical principles under unguided, natural conditions.

In two recent ground-breaking reports, senior scientists Dr. Royal Truman, Dr. Charles McCombs, and Dr. Change Tan examined the polymerization of amino acids in water, using kinetic and thermodynamic empirical data along with computer simulations. A detailed quantitative understanding was provided for the first time of how the concentrations of polypeptides decrease with length, using mostly the best-studied amino acid, glycine (Gly):
[Gly]_n << [Gly]_n-1 << [Gly]_n-2 << [Gly]_n-3 << [Gly]_n-4 …
The quantitative analysis showed that the concentration of [Gly]₉ would converge to < 10^-50 M at equilibrium in water at temperatures of 25° - 37°. In other words, not even one Gly₉ would have existed on prebiotic earth, far less the necessary huge concentrations of much larger polypeptides required by origin of life (OoL) theories.
This is a devastating conclusion for the OoL community! To make matters even worse, if that were possible, the authors provided a table with nine more requirements polypeptides must all fulfill to be relevant for OoL purposes, all of which violate fundamental chemical and statistical principles under unguided, natural conditions.
To permit structured and productive OoL discussions the authors recommend beginning with this table, which applies also to RNA and DNA polymers, to decide which dilemma to discuss.

1. Many amino acids must be linked together, about 300 on average for proteins.
2. Only enantiomers of L-amino acids should be included.
3. Only linear polymers should form; that is, the side chains of the amino acids must not react.
4. Precise sequences of amino acid residues must be formed to perform useful functions.
5. Long chains must adopt a suitable three-dimensional structure.
6. Large numbers of peptide copies must be produced continuously for millions of years.
7. The correct proportion of peptides with a specific sequence must be colocalized.
8. Other molecules, including nonbiological amino acids, should be avoided in peptides.
9. The entire system or organism must self-replicate, including all necessary peptide copies.
10. The polymers and the three-dimensional structure must be formed under relevant conditions.

These 10 requirements must be met simultaneously for peptides/proteins to be relevant in origin-of-life scenarios, but there are contradictory trade-offs between many of these requirements. For example, raising the temperature to facilitate a Gly adding to Gly_n to form Gly_n+1 (requirement #1) would have the effect of accelerating the rate of racemization L-Gly ⇆ D-Gly (requirement #2).

Unresolved Challenges in Prebiotic Amino Acid Stability and Reactivity

1. The Stability-Reactivity Paradox
Amino acids must be stable enough to accumulate in prebiotic environments while simultaneously being reactive enough to form peptides without enzymatic assistance. Studies show amino acid half-lives ranging from days to years, while spontaneous peptide bond formation has half-times of 10^2 to 10^3 years at 25°C and pH 7.

Conceptual problems:
- No known prebiotic mechanism can balance the conflicting requirements of stability and reactivity
- Difficulty in explaining how amino acids could accumulate without degrading faster than they polymerize
- Lack of plausible explanation for overcoming the kinetic barriers to peptide bond formation without enzymes

2. Temperature Dependence
Amino acid stability decreases dramatically at higher temperatures, often invoked in prebiotic scenarios. At 100°C, most amino acids have half-lives of less than a day.

Conceptual problems:
- No clear mechanism for protecting amino acids in high-temperature prebiotic environments
- Difficulty in reconciling the need for higher temperatures to drive reactions with the rapid degradation of amino acids
- Lack of explanation for how amino acids could have accumulated in the dynamic thermal conditions of the early Earth

3. Aqueous Environment Challenges
Peptide bond formation is thermodynamically unfavorable in water, yet water is typically assumed to be the medium for prebiotic chemistry.

Conceptual problems:
- No known prebiotic mechanism for efficient peptide bond formation in aqueous environments
- Difficulty in explaining how water could be removed to drive peptide formation while maintaining an aqueous reaction medium
- Lack of plausible model for the emergence of non-aqueous microenvironments conducive to peptide synthesis

4. Specific Amino Acid Vulnerabilities
Certain amino acids, like aspartic acid, are particularly prone to side reactions. Aspartic acid can form unreactive succinimide derivatives, with about 4% converting within 24 hours at pH 7 and 37°C.

Conceptual problems:
- No clear mechanism for preventing or mitigating these side reactions in a prebiotic setting
- Difficulty in explaining how vulnerable amino acids could have participated in early protein formation
- Lack of plausible explanation for the selection of stable amino acid sequences in the face of these chemical vulnerabilities

5. Polymerization Thermodynamics
Recent studies show that the equilibrium concentration of even short polypeptides like [Gly]₉ would be less than 10^-50 M at 25-37°C, making their existence highly improbable.

Conceptual problems:
- No known prebiotic mechanism can overcome these unfavorable thermodynamics
- Difficulty in explaining how polypeptides of sufficient length for biological function could have formed
- Lack of plausible model for shifting the equilibrium towards longer peptides without sophisticated biological machinery

6. Sequence Specificity
Functional proteins require specific amino acid sequences, yet prebiotic peptide formation would be random.

Conceptual problems:
- No known prebiotic mechanism for selecting specific amino acid sequences
- Difficulty in explaining how functional sequences could emerge from random polymerization
- Lack of plausible explanation for the origin of the genetic code linking amino acid sequences to nucleic acids

7. Structural Requirements
Proteins must adopt specific three-dimensional structures to function, but this requires precise sequences and folding conditions.

Conceptual problems:
- No clear mechanism for the emergence of complex protein structures in a prebiotic environment
- Difficulty in explaining how specific folding conditions could be maintained without cellular machinery
- Lack of plausible model for the coemergence of protein sequence and structure specificity

8. Continuous Production and Self-Replication
Origin of life scenarios require the continuous production of specific peptides and their self-replication.

Conceptual problems:
- No known prebiotic mechanism for the continuous, targeted production of specific peptides
- Difficulty in explaining how early peptide-based systems could self-replicate without modern cellular machinery
- Lack of plausible explanation for the origin of the complex interdependencies required for self-replication

9. Exclusion of Non-Biological Amino Acids
Functional proteins use only a specific set of amino acids, yet prebiotic synthesis would produce a wider variety.

Conceptual problems:
- No clear mechanism for selecting only the 20 canonical amino acids from a complex prebiotic mixture
- Difficulty in explaining how non-biological amino acids could be excluded from early peptide synthesis
- Lack of plausible model for the emergence of the specific amino acid set used in modern proteins

10. Simultaneous Fulfillment of Multiple Requirements
The emergence of functional peptides requires the simultaneous fulfillment of multiple, often contradictory, conditions.

Conceptual problems:
- No known prebiotic scenario can satisfy all necessary conditions simultaneously
- Difficulty in explaining how trade-offs between conflicting requirements could be navigated without guidance
- Lack of plausible explanation for the coemergence of the various systems required to meet all conditions

In conclusion, the stability and reactivity requirements for prebiotic amino acids present formidable challenges to naturalistic explanations of the origin of life. The quantitative analysis of polypeptide formation, coupled with the multiple specific requirements for biologically relevant peptides, reveals a series of hurdles that appear insurmountable through unguided processes alone. The stability-reactivity paradox, unfavorable polymerization thermodynamics, and the need for specific sequences and structures collectively present a multi-faceted problem that current scientific understanding cannot resolve without invoking highly improbable chance events or unknown chemical processes. These challenges call for a critical re-examination of the assumptions underlying abiogenesis hypotheses and highlight the need for new, evidence-based approaches to understanding the chemical origins of life.

1.8. Thermodynamic and Kinetic Barriers to Prebiotic Polypeptide Formation

The spontaneous formation of polypeptides in aqueous prebiotic environments encounters significant thermodynamic and kinetic barriers, challenging current naturalistic explanations for the origin of life. Thermodynamic calculations indicate that peptide bond formation in water is energetically unfavorable, with a standard Gibbs free energy change of approximately 3.5 kcal/mol at 25°C and pH 7 ¹⁶. Computational exploration of organic molecule formation from water and hydrogen cyanide reveals diverse reactivity landscapes and lower activation energies for biologically relevant molecules, impacting the interpretation of network kinetics ¹⁷. In fluctuating silica environments, the presence of water activity enhances peptide formation through hydration steps, resulting in the formation of self-assembled peptide aggregates with defined secondary structures ¹⁸. Additionally, a new abiotic route demonstrates peptide chain growth from protonated glycine dimers in a cold gaseous atmosphere without the need for a solid catalytic substrate ¹⁹. Experimental simulations under hydrothermal and extraterrestrial ice crystal environments show the formation of small functional peptides, shedding light on potential prebiotic pathways for catalytically active peptides ²⁰.

1.8.1. Quantitative Challenges

Thermodynamic calculations reveal that the formation of peptide bonds in aqueous solutions is energetically unfavorable. The standard Gibbs free energy change (ΔG°) for peptide bond formation is approximately +3.5 kcal/mol at 25°C and pH 7 (Jakubke & Jeschkeit, 1977). This positive value indicates that the reaction is non-spontaneous under standard conditions.

Kinetic studies further compound this challenge. The rate constant for uncatalyzed peptide bond formation in water at 25°C is estimated to be around 10^-4 M^-1 year^-1 (Sievers & von Kiedrowski, 1994). In contrast, the rate constant for peptide bond hydrolysis under the same conditions is approximately 10^-9 to 10^-11 s^-1 (Radzicka & Wolfenden, 1996). These values translate to a half-life of peptide bond formation on the order of thousands of years, while the half-life for hydrolysis is typically days to months.

1.8.2. Implications for Current Models

These quantitative findings present severe challenges to current models of prebiotic polypeptide formation. The unfavorable thermodynamics imply that even if peptides were to form, they would be thermodynamically driven to hydrolyze back into amino acids. The slow kinetics of formation coupled with the relatively rapid hydrolysis suggests that maintaining any significant concentration of polypeptides in a prebiotic aqueous environment is highly improbable.

1.8.3. Requirements for Natural Occurrence

For the spontaneous formation and persistence of polypeptides in a prebiotic setting, the following conditions must be simultaneously met:

1. Energy input to overcome the unfavorable thermodynamics of peptide bond formation
2. Mechanisms to dramatically accelerate the rate of peptide bond formation
3. Protection against hydrolysis to maintain formed peptides
4. Concentration mechanisms to achieve sufficiently high local amino acid densities
5. Selective polymerization to form functional peptide sequences
6. Removal of water to drive the condensation reaction forward
7. pH conditions that balance peptide bond formation and stability (typically pH 2-5 for formation, pH 5-8 for stability)
8. Temperature regime that allows for both formation and stability of peptides
9. Absence of competing side reactions that could deplete the amino acid pool
10. Recycling mechanisms to regenerate hydrolyzed amino acids

These requirements must coexist in a prebiotic environment, presenting a formidable challenge to naturalistic explanations. Several of these conditions are mutually exclusive or contradictory. For instance, the need for water removal (point 6) conflicts with the aqueous environment typically assumed in prebiotic scenarios. Similarly, the pH conditions favorable for peptide bond formation (point 7) are not optimal for peptide stability.

The challenges are illustrated by the "alanine problem." Alanine, one of the simplest amino acids, forms peptides extremely slowly in aqueous solutions. Experiments have shown that at 25°C and pH 7, the equilibrium concentration of the alanine dipeptide is only about 10^-4 M when starting from a 1 M solution of alanine (Danger et al., 2012). This low yield highlights the thermodynamic barriers to even the simplest peptide formations.

Moreover, the requirement for energy input (point 1) often leads to increased rates of side reactions and decomposition, conflicting with the need for selective polymerization (point 5) and protection against hydrolysis (point 3).

Unresolved Challenges in Prebiotic Protein Formation

1. Thermodynamic Unfavorability
Peptide bond formation in water is energetically unfavorable, with a standard Gibbs free energy change of approximately +3.5 kcal/mol at 25°C and pH 7.

Conceptual problems:
- No known prebiotic mechanism can consistently overcome this thermodynamic barrier
- Difficulty in explaining how peptides could form and persist in aqueous environments
- Lack of plausible explanation for the accumulation of polypeptides against thermodynamic gradients

2. Kinetic Barriers
The rate constant for uncatalyzed peptide bond formation in water at 25°C is estimated to be around 10^-4 M^-1 year^-1, while hydrolysis occurs much faster.

Conceptual problems:
- No clear mechanism for accelerating peptide bond formation without sophisticated catalysts
- Difficulty in explaining how peptides could form faster than they hydrolyze in prebiotic conditions
- Lack of plausible model for the emergence of kinetically favored peptide synthesis pathways

3. Hydrolysis Susceptibility
Formed peptides are susceptible to hydrolysis, with half-lives typically ranging from days to months in aqueous environments.

Conceptual problems:
- No known prebiotic mechanism for protecting formed peptides from rapid hydrolysis
- Difficulty in explaining how early peptides could have persisted long enough to serve functional roles
- Lack of plausible explanation for the accumulation of long peptides in the face of constant hydrolytic pressure

4. Concentration Requirements
High local concentrations of amino acids are required for significant peptide formation, yet prebiotic environments likely had dilute conditions.

Conceptual problems:
- No clear mechanism for achieving sufficiently high amino acid concentrations in prebiotic settings
- Difficulty in explaining how localized high concentrations could be maintained without cellular compartmentalization
- Lack of plausible model for the coemergence of concentration mechanisms and peptide synthesis

5. Sequence Specificity
Functional proteins require specific amino acid sequences, yet prebiotic peptide formation would be largely random.

Conceptual problems:
- No known prebiotic mechanism for selecting specific amino acid sequences
- Difficulty in explaining how functional sequences could emerge from random polymerization
- Lack of plausible explanation for the origin of the genetic code linking amino acid sequences to nucleic acids

6. Water Paradox
Water is necessary as a solvent but its presence makes peptide bond formation thermodynamically unfavorable.

Conceptual problems:
- No clear mechanism for removing water to drive peptide formation while maintaining an aqueous environment
- Difficulty in explaining how early life could have emerged in water while requiring water's absence for key chemical steps
- Lack of plausible model for the emergence of micro-environments with controlled water activity

7. pH and Temperature Constraints
Optimal conditions for peptide bond formation (pH 2-5) differ from those for peptide stability (pH 5-8 ), and temperature affects both formation and stability.

Conceptual problems:
- No known prebiotic mechanism for maintaining optimal pH and temperature conditions for both formation and stability
- Difficulty in explaining how early peptides could have formed and persisted in fluctuating prebiotic environments
- Lack of plausible explanation for the emergence of pH and temperature regulation mechanisms

8. Competing Side Reactions
Prebiotic environments likely contained a complex mixture of organic compounds that could interfere with peptide formation.

Conceptual problems:
- No clear mechanism for selectively promoting peptide bond formation over competing reactions
- Difficulty in explaining how amino acids could have preferentially reacted with each other rather than with other abundant molecules
- Lack of plausible model for the emergence of chemical selectivity without sophisticated catalysts

9. Recycling and Regeneration
Continuous peptide formation would require mechanisms to recycle hydrolyzed amino acids and regenerate reactive species.

Conceptual problems:
- No known prebiotic mechanism for efficiently recycling amino acids from hydrolyzed peptides
- Difficulty in explaining how a continuous supply of reactive amino acids could be maintained
- Lack of plausible explanation for the emergence of complex recycling systems in prebiotic settings

10. Energy Input and Management
Overcoming thermodynamic barriers requires energy input, but managing this energy without cellular machinery is problematic.

Conceptual problems:
- No clear mechanism for coupling available energy sources to peptide bond formation without harmful side effects
- Difficulty in explaining how energy could be harnessed for specific chemical reactions in a prebiotic setting
- Lack of plausible model for the emergence of sophisticated energy management systems

In conclusion, the formation of proteins in prebiotic environments faces numerous interconnected challenges that remain unresolved. The thermodynamic unfavorability of peptide bond formation in water, coupled with slow kinetics of formation and rapid hydrolysis, presents a formidable barrier to the spontaneous emergence of polypeptides. The requirements for specific sequences, protection against hydrolysis, and the need for high local concentrations further compound these difficulties. Current scientific understanding lacks plausible, empirically supported explanations for how these challenges could be overcome through unguided processes alone. The simultaneous fulfillment of multiple, often contradictory conditions necessary for prebiotic protein formation presents a significant conceptual hurdle for naturalistic origin-of-life scenarios. These unresolved issues call for a critical re-evaluation of current abiogenesis hypotheses and highlight the need for new, evidence-based approaches to understanding the chemical origins of life.

References

1.3.1. Availability and challenges associated with major atoms required to synthesize amino acids

Hazen, R. M. (2012). Fundamentals of Geobiology. Link. (This book explores geobiology, highlighting the intersection of geology and biology in the emergence of life.)

Venter, C., et al. (2008). Life: What A Concept! Link. (Craig Venter explores the foundational aspects of life and synthetic biology’s role in redefining life’s origins.)

Cairns-Smith, A. G. (1988). Genetic Takeover: And the Mineral Origins of Life. Link. (This book discusses the mineral origins hypothesis, suggesting that early life may have begun on mineral surfaces.)

1.4.  Challenges in the Availability of Precursors for Prebiotic Amino Acid Synthesis

1. Nogal, N., Sanz-Sánchez, M., Vela-Gallego, S., Ruiz-Mirazo, K., & de la Escosura, A. (2023). The protometabolic nature of prebiotic chemistry. Chemical Society Reviews, 52(17), 7229-7248. Link. (This review explores the concept of protometabolism in prebiotic chemistry and its implications for the origin of life.)

2. Tran, Q.P., Yi, R., & Fahrenbach, A.C. (2023). Towards a prebiotic chemoton – nucleotide precursor synthesis driven by the autocatalytic formose reaction. Chemical Science, 14(25), 6999-7008. Link. (This study investigates the synthesis of nucleotide precursors using the formose reaction in a prebiotic context.)

3. Peters, S., Semenov, D., Hochleitner, R., & Trapp, O.E. (2023). Synthesis of prebiotic organics from CO2 by catalysis with meteoritic and volcanic particles. Scientific Reports, 13(1), 7054. Link. (This research examines the synthesis of organic compounds from CO2 using meteoritic and volcanic particles as catalysts under prebiotic conditions.)

Further references:
Stuart, A.H., Rammu, H., & Lane, N. (2023). Prebiotic Synthesis of Aspartate Using Life's Metabolism as a Guide. Reproductive and developmental Biology, 13(5), 1177. Link. (This study investigates the prebiotic synthesis of aspartate using metabolic pathways found in modern life as a guide.)

Magrino, T., Pietrucci, F., & Saitta, A.M. (2021). Step by Step Strecker Amino Acid Synthesis from Ab Initio Prebiotic Chemistry. Journal of Physical Chemistry Letters, 12(9), 2376-2382. Link. (This work uses ab initio simulations to model a step-by-step Strecker synthesis of amino acids under prebiotic conditions.)

Ashe, K. (2018). Studies towards the prebiotic synthesis of nucleotides and amino acids. Doctoral thesis, University of Cambridge. Link. (This thesis explores various routes for the prebiotic synthesis of both nucleotides and amino acids.)

McDonald, G.D., & Storrie-Lombardi, M.C. (2010). Biochemical constraints in a protobiotic earth devoid of basic amino acids: the "BAA(-) world". Astrobiology, 10(10), 989-1000. Link. (This paper proposes a "BAA(-) world" hypothesis, exploring biochemical constraints in a protobiotic Earth lacking basic amino acids.)

Engel, M.H., & Perry, R.S. (2008). The origins of amino acids in ancient terrestrial and extraterrestrial materials. Proceedings of SPIE, 7097, 70970O. Link. (This review examines evidence for amino acid origins in ancient terrestrial and extraterrestrial materials.)

1.4. Challenges of Prebiotic Peptide Bond Formation

4. Nogal, N., Sanz-Sánchez, M., Vela-Gallego, S., Ruiz-Mirazo, K., & de la Escosura, A. (2023). The protometabolic nature of prebiotic chemistry. Chemical Society Reviews, 52(17), 7229-7248. Link. (This review explores the concept of protometabolism in prebiotic chemistry and its implications for the origin of life.)

5. Diederich, P., Geisberger, T., Yan, Y., Seitz, C., Ruf, A., Huber, C., Hertkorn, N., & Schmitt-Kopplin, P. (2023). Formation, stabilization and fate of acetaldehyde and higher aldehydes in an autonomously changing prebiotic system emerging from acetylene. Communications Chemistry, 6(1), 69. Link. (This study investigates the formation and behavior of aldehydes in a prebiotic system derived from acetylene.)

6. Zhang, W. (2023). The formation and stability of homochiral peptides in aqueous prebiological environment in the Earth's crust. arXiv preprint. Link. (This preprint examines the formation and stability of homochiral peptides in prebiotic aqueous environments within the Earth's crust.)

7. Chi, Y., Li, X.Y., Chen, Y., Zhang, Y., Liu, Y., Gao, X., & Zhao, Y. (2022). Prebiotic formation of catalytically active dipeptides via trimetaphosphate activation. Chemistry - An Asian Journal, 17(23), e202200926. Link. (This research demonstrates the prebiotic formation of catalytically active dipeptides using trimetaphosphate activation.)

Further references:

Szilagyi, R.K. (2023). Peptide condensation and hydrolysis mechanisms from a proton-transfer network perspective. Organic and Biomolecular Chemistry, 21(21), 3974-3987. Link. (This study explores peptide formation and breakdown mechanisms from a proton-transfer perspective.)

Sydow, C., Sauer, F., Siegle, A.F., & Trapp, O. (2022). Iron‐mediated peptide formation in water and liquid sulfur dioxide under prebiotically plausible conditions. ChemSystemsChem, 4(5), e202200034. Link. (This work investigates iron-mediated peptide formation under prebiotic conditions.)
El Samrout, O., Berlier, G., Lambert, J.F., & Martra, G. (2023). Polypeptide Chain Growth Mechanisms and Secondary Structure Formation in Glycine Gas-Phase Deposition on Silica Surfaces. Journal of Physical Chemistry B, 127(13), 3017-3028. Link. (This study examines polypeptide formation on silica surfaces through gas-phase deposition.)

Trapp, O., Sauer, F., Haas, M., Sydow, C., Siegle, A.F., & Lauer, C. (2021). Peptide formation as on the early Earth: from amino acid mixtures to peptides in sulphur dioxide. Research Square. Link. (This preprint explores peptide formation in sulfur dioxide as a model for early Earth conditions.)

Stolar, T., Grubešić, S., Cindro, N., Meštrović, E., Užarević, K., & Hernández, J.G. (2021). Mechanochemical Prebiotic Peptide Bond Formation. Angewandte Chemie, 133(22), 12678-12682. Link. (This paper investigates mechanochemical methods for prebiotic peptide bond formation.)

Comte, D., Lavy, L., Bertier, P., Calvo, F., Daniel, I., Farizon, B., Farizon, M., & Märk, T.D. (2023). Glycine Peptide Chain Formation in the Gas Phase via Unimolecular Reactions. Journal of Physical Chemistry A, 127(8 ), 1768-1776. Link. (This study examines glycine peptide chain formation through gas-phase unimolecular reactions.)

Rousseau, P., Piekarski, D.G., Capron, M., Domaracka, A., Adoui, L., Martín, F., Alcamí, M., Díaz-Tendero, S., & Huber, B.A. (2020). Polypeptide formation in clusters of β-alanine amino acids by single ion impact. Nature Communications, 11(1), 3818. Link. (This work demonstrates polypeptide formation in β-alanine clusters through single ion impact.)

1.5.  Quantity and Concentration: Challenges in Prebiotic Amino Acid Availability

8.Rolf, J., Handke, J., Burzinski, F., Luetz, S., & Rosenthal, K. (2023). Amino acid balancing for the prediction and evaluation of protein concentrations in cell-free protein synthesis systems. Biotechnology Progress, 39(5), e3373. Link. (This study investigates amino acid balancing for optimizing protein synthesis in cell-free systems.)

9. (2023). Amino acid balancing for the prediction and evaluation of protein concentrations in cell-free protein synthesis systems. arXiv preprint. Link. (This preprint discusses amino acid balancing techniques for cell-free protein synthesis systems.)

10. (2023). Geochemical and Photochemical Constraints on S[IV] Concentrations in Natural Waters on Prebiotic Earth. ESSOAr. Link. (This study examines the constraints on sulfur concentrations in prebiotic Earth's natural waters.)

11. Gómez Ortega, J., Raubenheimer, D., Tyagi, S., Mirth, C.K., & Piper, M.D.W. (2023). Biosynthetic constraints on amino acid synthesis at the base of the food chain may determine their use in higher-order consumer genomes. PLOS Genetics, 19(5), e1010635. Link. (This research explores how biosynthetic constraints on amino acids at lower troph

ic levels may influence their use in higher-order organisms' genomes.)

1.6.  Stability and Reactivity: The Prebiotic Amino Acid Paradox

12. Stuart, A.H., Rammu, H., & Lane, N. (2023). Prebiotic Synthesis of Aspartate Using Life's Metabolism as a Guide. Reproductive and developmental Biology, 13(5), 1177. Link. (This study investigates the prebiotic synthesis of aspartate using metabolic pathways found in modern life as a guide.)

13. Holden, D.T., Morato, N.M., & Cooks, R.G. (2022). Aqueous microdroplets enable abiotic synthesis and chain extension of unique peptide isomers from free amino acids. Proceedings of the National Academy of Sciences of the United States of America, 119(44), e2212642119. Link. (This research demonstrates the abiotic synthesis and chain extension of peptide isomers in aqueous microdroplets, providing insights into potential prebiotic peptide formation mechanisms.)

1.7. Thermodynamic and Kinetic Barriers to Polymerization

14. Vaida, V., & Deal, A.M. (2022). Peptide synthesis in aqueous microdroplets. Proceedings of the National Academy of Sciences of the United States of America, 119(50), e2216015119. Link. (This study investigates the synthesis of peptides in aqueous microdroplets, providing insights into potential prebiotic chemistry mechanisms.)

15. Carvalho-Silva, V.H., Coutinho, N.D., & Aquilanti, V. (2020). From the Kinetic Theory of Gases to the Kinetics of Rate Processes: On the Verge of the Thermodynamic and Kinetic Limits. Molecules, 25(9), 2098. Link. (This review explores the connections between kinetic theory of gases and the kinetics of rate processes, discussing thermodynamic and kinetic limits relevant to chemical reactions.)

Further references:
Royal Truman and Charles McCombs, Negligible concentrations of peptides form in water: part 1 - using high temperatures or high pH, ​​J. Creation 38(1):126‒135, 2024.
Royal Truman, Change Tan, and Charles McCombs, Insignificant concentrations of peptides form in water: part 2-using moderate temperatures, J. Creation 38(1):136‒149, 2024.
Chemical evolution of amino acids and proteins? Impossible!!
https://reasonandscience.catsboard.com/t2887-chemical-evolution-of-amino-acids-and-proteins-impossible

1.8. Thermodynamic and Kinetic Barriers to Prebiotic Polypeptide Formation

16. Harold, S.E., Warf, S.L., & Shields, G.C. (2023). Prebiotic dimer and trimer peptide formation in gas-phase atmospheric nanoclusters of water. Physical Chemistry Chemical Physics, 25(31), 20890-20901. Link. (This study investigates the formation of small peptides in atmospheric water nanoclusters, providing insights into potential prebiotic chemistry mechanisms.)

17. Zhao, Q., Garimella, S.S., & Savoie, B.M. (2023). Thermally Accessible Prebiotic Pathways for Forming Ribonucleic Acid and Protein Precursors from Aqueous Hydrogen Cyanide. Journal of the American Chemical Society, 145(10), 5735-5745. Link. (This research explores thermally accessible pathways for the formation of RNA and protein precursors from hydrogen cyanide in aqueous environments.)

18. El Samrout, O., Berlier, G., Lambert, J.F., & Martra, G. (2023). Polypeptide Chain Growth Mechanisms and Secondary Structure Formation in Glycine Gas-Phase Deposition on Silica Surfaces. Journal of Physical Chemistry B, 127(13), 3017-3028. Link. (This study examines polypeptide formation on silica surfaces through gas-phase deposition of glycine.)

19. Comte, D., Lavy, L., Bertier, P., Calvo, F., Daniel, I., Farizon, B., Farizon, M., & Märk, T.D. (2023). Glycine Peptide Chain Formation in the Gas Phase via Unimolecular Reactions. Journal of Physical Chemistry A, 127(8 ), 1768-1776. Link. (This study examines glycine peptide chain formation through gas-phase unimolecular reactions.)

20. Chi, Y., Li, X.Y., Chen, Y., Zhang, Y., Liu, Y., Gao, X., & Zhao, Y. (2022). Prebiotic formation of catalytically active dipeptides via trimetaphosphate activation. Chemistry - An Asian Journal, 17(23), e202200926. Link. (This research demonstrates the prebiotic formation of catalytically active dipeptides using trimetaphosphate activation.)

7.7.9. Critical Examination

Current models often invoke "self-organization" or "emergent complexity" to bridge the gap between simple chemical systems and self-replicating protocells. However, these concepts lack specificity and often amount to restatements of the problem rather than solutions. The transition from non-living to living systems represents a staggering increase in functional information content, which is not adequately explained by known physical or chemical principles.

7.7.10. Conclusion and Future Discussions

Future discussions on the origin of self-replicating systems should:
1. Quantify the minimal functional requirements for self-replication rigorously
2. Address each requirement explicitly, providing plausible prebiotic mechanisms
3. Propose testable hypotheses for the coordinated emergence of replication, metabolism, and containment
4. Consider alternative models that do not rely solely on chance assembly or gradual accumulation of features
5. Investigate potential non-aqueous environments or unique geological settings that might facilitate more rapid exploration of chemical space
6. Explore the concept of "functional information" and its origins in prebiotic systems

By structuring the debate around these points, we can more accurately assess the viability of current theories and guide future research into the origins of biological complexity. The field must grapple with the enormous gulf between simple chemical reactions and the sophisticated, information-rich systems characteristic of even the simplest known life forms.

7.8. Amplification of Enantiomeric Excess

The amplification of enantiomeric excess (ee) from a small initial value to 100% L-amino acids has been a topic of extensive research and debate. Literature experiments have not supported the idea that small excesses of L-amino acids can be amplified to complete homochirality, with proposed mechanisms often requiring unrealistic experimental conditions. Studies have explored various scenarios like partial sublimation, crystal separation, and chiral catalysts but have faced significant limitations in achieving and maintaining high ee values. Research has shown that natural processes alone may not be sufficient to drive the amplification of ee to complete homochirality, highlighting the complexity of this phenomenon ^{1 2 3 4 5}.

7.8.1. Literature experiments do not corroborate that a small excess of L-amino acid could be amplified to form 100% L-amino acids

In a series of remarkable papers, senior chemists from several firms, Dr. Royal Truman, Dr. Chris Basel, and Dr. Stephen Grocott did an extensive analysis of the key literature on amplification experiments of small excesses of L-amino acids. The evolutionary experiments reviewed had been designed to find special conditions to preferentially extract excess L-amino acids from mixtures and separate a portion having a higher proportion of L-amino acid (aa).
Their conclusions are very bad news for the origin of life (OoL) community, demonstrating that implausible experimental conditions had to be used. Objective evaluation of the results showed that the attempts to find relevant amplification scenarios had failed badly.
To illustrate, a hypothetical astronomical source of right-circularly polarized UV light (r-CPL) is the preferred evolutionary theory for the origin of homochiral amino acids. However, astronomers have been unable to find polarized UV light anywhere in the relevant region of space.
We encourage you to read the papers covering the topics of interest. Here are some bullet points extracted from this series of papers.

1. Claims of significant enantiomeric excess produced by a hypothetical astronomical circularly polarized light (CPL) source are misleading:
- Astronomers have not found polarized UV light in a relevant region of space
- The theory required very specific conditions and laboratory conditions untypical in space
- The theory requires almost 100% photodestruction of all amino acids before an excess could result (but 100% destruction would serve no purpose!).
2. Different amino acids absorb left-handed and right-handed CPL differently at various UV wavelengths. Therefore, the expected result would be an averaging out with little or no survival of one enantiomer.
3. Experimental approaches focused on the very exceptional amino acids with the highest anisotropy factors and used optimal wavelengths designed to produce the researcher’s goal instead of real-world outcomes.
4. Even if a small enantiomeric excess were produced in space, it would likely be further diluted upon reaching Earth.
5. The major literature examples like alkylation of Soai and polymerization of cyclobutene have no relevance to OoL. In many cases such as selective adsorption on minerals such as kaolinite and montmorillonite clay, even the irrelevant examples published have been disproven when the experiments were repeated.
6. Adsorption on chiral calcite and quartz would produce equal amounts of D and L enantiomers overall.
7. Extracting only L-amino acids from glycine crystals required pure D-leucine, an unlikely natural scenario.
8. The formation of enantiomer-specific crystalline islands required laboratory processes not found in nature.
9. There is no credible reason why enantiomers of separate amino acids would not remix in natural environments.
10. Any enantiomeric excess produced would racemize in water over time.
11. Proposed mechanisms required carefully planned and executed laboratory conditions that are unlikely in nature.
12. None of the proposals known could have occurred naturally without intelligent guidance.
13. Different amino acid precursors behave differently with the same sugar catalysts, making it impossible to generalize about sugar-induced chirality.
14. The used D-lyxose to favor production of L-amino acids did not occur when tested using alanine.
15. On longer time scales relevant to prebiotic scenarios, racemic mixtures would result regardless of the initial sugar-induced excess.
16. Impurities have been tested to enhance the crystallization of a particular enantiomer.
- This only produced conglomerate crystals, but not separated enantiomers.
- The supersaturated pure solutions used to form crystals used would not have existed naturally.
- Conditions to favor one L-amino acid would have increased the amount of D- enantiomer of other amino acids, making matters worse for OoL purposes.
17. Excess of non-biological L-α-methyl amino acids have been claimed in meteorites. Experiments showed that mixing L-α-methylamine with racemic L- and D amino acid produced more of the wrong D-amino acids. Even had the desired outcome been obtained, equilibration L ⇆ D occurred rapidly, especially at elevated temperature.
18. Extensive experiments with L-α-methyl amino acids and many catalysts showed the desired outcome when using copper, but at plausible concentrations, the enantiomeric effect was negligible and racemization would have occurred in the presence of such a catalyst rapidly.
19. Simulations of wet-dry cycles with L-amino acids and L-isovaline in montmorillonite clay:
- Led to rapid racemization of amino acids, very bad news for OoL purposes.
- Showed that chirality could not be effectively transferred from L-isovaline to produce L-amino acids.
20. Instant sublimation experiments at ~ (430°C) followed by instant cooling at sub-freezing temperatures (!?) to avoid thermal degradation experiments have no relevance for OoL purposes: aa degrade at much lower temperatures.
21. Sublimation experiments using serine relied on unrealistic, optimized laboratory conditions (and did not work with other aa):
- High temperatures around exactly 205°C with short heating times (2-18 hours)
- Rapid cooling with dry ice and N₂ gas flow to quickly remove sublimate
- Avoiding serine racemization and decomposition at high temperatures.
- The maximum enantiomeric excess achieved was too low for biological purposes.
Worse, starting with an L-enantiomeric excess of serine actually produced a sublimate with a lower excess!
22. Sublimation experiments using mixtures of Asn, Thr, Asp, Glu, and Ser cleverly mixed with volatile racemic Ala, Leu, Pro, or Val required carefully optimized laboratory conditions to obtain the intended goal:
- Low pressure (0.3-0.7 mbar), controlled temperature (100-105°C), and duration of 14 hours
- Use pure L enantiomers, and prevent remixing of sublimate and residue
- Use of an icy cold finger to trap the sublimate.
The results were less than encouraging. Using L-enantiomers of the less sublimated aa produced sublimates enriched in D-enantiomers of the volatile aa, the opposite needed for biology!
23. Only two biological amino acids (threonine and asparagine) naturally crystallize as conglomerates of distinct D and L crystals, but under conditions not relevant for OoL.
24. Most biological aa form racemic crystals (equal amounts of D and L enantiomers), preventing crystalline excesses. Any excess in solution would simply racemize over time.
25. Random temperature variation would prevent the precise control needed to take advantage of the eutectic point for some aa to separate crystals with an excess of an enantiomer.
26. Laboratory conditions were used to extract enantiomer excesses already present, including saturated solutions of a pure aa, controlled temperatures, and constant agitation.
27. Natural processes such as rainwater, seawater, and groundwater would have diluted any enantiomeric excess and led to remixing.
28. If hypothetically an excess would exist in solution that could crystallize preferentially into L-crystals, eventually, the excess would be depleted and all the aa would then crystallize out of solution, contaminating the first batches.
29. In any scenario of excess in liquid or crystal phase remixing would occur, racemization over time, and contamination with racemic mixtures in the environments.
30. Some experiments used complex catalysts not found naturally to increase racemization, of the wrong D-amino acids, but they would have also racemized all the L-amino acids indiscriminately.
31. Techniques like Preferential Enrichment and CIAT rely on an initial excess of L-enantiomer, organic solvents like aspartic acid and acetic acid, and typically salicylaldehyde as catalyst at 90-160°C with agitation in a special container. None of these are relevant for OoL purposes.
32. Attempts to use chiral minerals like quartz as selective catalysts have shown only very small enantiomeric excesses, typically less than 1%.
33. Proposed autocatalytic reactions like the Soai reaction require highly specific precursor molecules and conditions not plausible in prebiotic environments.
34. Theoretical models of amplification often rely on unrealistic assumptions about reaction kinetics and equilibrium conditions.
35. Experiments using temperature gradients to separate enantiomers produce only transient and localized excesses that quickly dissipate.
36. Proposed mechanisms involving chiral light or spin-polarized electrons lack a demonstrated source in early Earth environments.
37. Attempts to use amino acid precursors like α-methyl amino acids as chiral catalysts have shown limited effectiveness and selectivity.
38. Proposed amplification via polymerization faces issues of reversibility and lack of selectivity for homochiral products.
39. Scenarios involving partial crystallization require precise control of supersaturation, nucleation, and growth conditions unlikely in nature.
40. Attempts to exploit slight solubility differences between enantiomers have produced only marginal enrichment.

41. Proposed chiral amplification via asymmetric autocatalysis faces issues of product inhibition and side reactions.

The overall picture reinforces the significant hurdles facing naturalistic explanations for the origin of biological homochirality.

7.8.2. Additional Considerations - Optimal Set of Amino Acids

Recent studies by Philip and Freeland (2011) and Ilardo et al. (2015) have highlighted the exceptional optimality of the standard 20 amino acid alphabet in life, showcasing high coverage of crucial chemical properties like size, charge, and hydrophobicity that outperform vast alternative alphabets. These findings challenge conventional theories of chemical evolution, indicating a level of selection or foresight that contradicts undirected processes. To naturally achieve such an optimal amino acid set, prebiotic conditions must simultaneously provide a diverse amino acid pool, a sophisticated selection mechanism, discernment of subtle chemical differences, balance simplicity with functional diversity, compatibility with translation machinery, stability under prebiotic conditions, reactivity for peptide bond formation, and rapid selection before other biochemical systems emerge, presenting significant contradictions in the origin of life hypotheses ^{1 2}.

7.8.3. Quantitative Findings Challenging Conventional Theories

A study by Philip and Freeland (2011) compared the standard 20 amino acid alphabet to random sets of amino acids chosen from a larger pool of 50 plausible prebiotic amino acids. They found that the standard alphabet exhibits unusually high coverage of three key chemical properties: size, charge, and hydrophobicity. Out of 10^19 possible alternative alphabets, only one in a million matched or exceeded the standard alphabet's coverage of these properties.

Another study by Ilardo et al. (2015) used a computational model to assess the designability and folding stability of proteins made from various amino acid alphabets. They found that the standard 20 amino acid set outperformed most alternative sets, including those with more amino acids, in producing stable, well-folded proteins.

7.8.4. Implications for Current Scientific Models

These findings pose significant challenges to current models of chemical evolution. Conventional theories typically assume that the set of amino acids used in life was determined by availability in the prebiotic environment or by chance. However, the observed optimality suggests a level of "foresight" or selection that is difficult to reconcile with undirected processes.

7.8.5. Requirements and Conditions

For the optimal set of amino acids to arise naturally, the following conditions must be met simultaneously under prebiotic conditions:

1. A diverse pool of amino acids must be available in the prebiotic environment.
2. A mechanism must exist to select amino acids based on their functional properties rather than just their abundance.
3. The selection process must be able to distinguish between subtle differences in chemical properties among similar amino acids.
4. The chosen set must provide a balance between simplicity (fewer amino acids) and functional diversity.
5. The selection process must occur before the establishment of the genetic code, as the code itself would constrain further changes to the amino acid alphabet.
6. The selected set must be compatible with the emerging translation machinery, including tRNA and aminoacyl-tRNA synthetases.
7. The chosen amino acids must be stable under prebiotic conditions yet reactive enough to form peptide bonds.
8. The selection process must occur rapidly enough to establish the optimal set before other, potentially incompatible biochemical systems emerge.

These requirements present several contradictions:
- The need for a diverse initial pool conflicts with the selective pressures that would limit the variety of compounds produced abiotically.
- The requirement for a sophisticated selection mechanism conflicts with the presumed simplicity of prebiotic chemical systems.
- The need for rapid selection conflicts with the gradual nature of evolutionary processes.

7.8.6. Relevant Scientific Terminology

Proteinogenic amino acids, chemical evolution, prebiotic chemistry, abiogenesis, protein folding, hydrophobicity, designability, genetic code, tRNA, aminoacyl-tRNA synthetases, peptide bond formation.

7.8.7. Illustrative Examples

Consider the case of lysine and arginine, two positively charged amino acids in the standard set. Both could plausibly form in prebiotic conditions, but arginine is more complex and less likely to arise spontaneously. However, arginine's guanidinium group provides unique properties for protein function. A purely abundance-based selection would likely have chosen lysine alone, missing the functional advantages of including both.

7.8.8. Critical Examination of Current Theories

Current theories of chemical evolution struggle to explain the observed optimality of the amino acid alphabet. Models based on prebiotic availability fail to account for the inclusion of less common amino acids like tryptophan or the exclusion of simpler, more abundant ones like norvaline. Scenarios invoking serial selection of amino acids face the challenge of explaining how early choices could anticipate future functional needs.

7.8.9.  Further Discussion

Future discussions on this topic should focus on developing testable hypotheses that can explain the apparent optimality of the amino acid set without invoking teleological mechanisms. This might include exploring potential feedback loops between amino acid availability and early metabolic cycles, or investigating whether alternative optimal sets exist that might have been discoverable through plausible chemical evolution scenarios. In conclusion, the near-optimal nature of the 20 proteinogenic amino acids presents a significant challenge to naturalistic explanations for the origin of life. While not insurmountable, this challenge requires careful consideration and may necessitate revisions to current models of chemical evolution and abiogenesis.

7.9. Protein Folding and Chaperones

Recent studies highlight that a substantial portion of newly synthesized proteins in eukaryotic and prokaryotic cells rely on molecular chaperones for proper folding, challenging conventional theories of early protein evolution. The intricate process of protein folding, with vast conformational possibilities, occurs rapidly due to the energy landscape and chaperone assistance. These findings raise significant questions about the evolution of functional proteins without pre-existing chaperone systems, presenting a "chicken and egg" dilemma. Early protein evolution faces contradictions regarding the necessity of complex regulatory mechanisms, specific environmental conditions, and the availability of energy sources for chaperone-assisted folding. The GroEL/GroES chaperonin system exemplifies the complexity

 of chaperones, challenging the idea of their evolution in the absence of functional proteins. Addressing these challenges requires exploring primitive folding mechanisms and potential evolutionary starting points for protein folds, urging a reevaluation of current models of early protein evolution ^[35].

7.9.1. Quantitative Findings Challenging Conventional Theories

Recent studies have shown that approximately 30-50% of newly synthesized proteins in eukaryotic cells require assistance from molecular chaperones to achieve their native, functional states (Balchin et al., 2016). In prokaryotes, this percentage is lower but still significant, with about 10-20% of proteins needing chaperone assistance (Hartl et al., 2011).

The folding process itself is extremely complex. For a small protein of 100 amino acids, there are approximately 10^30 possible conformations. Yet, proteins typically fold into their native states on timescales of milliseconds to seconds (Dill and MacCallum, 2012). This speed is possible only because of the energy landscape of protein folding and the assistance of chaperones.

7.9.2. Implications for Current Scientific Models

These findings pose significant challenges to current models of early protein evolution. The high percentage of proteins requiring chaperones for proper folding suggests that early functional proteins would have faced severe limitations without a pre-existing chaperone system. This creates a "chicken and egg" problem: how could complex, functional proteins evolve if they required equally complex chaperone systems to fold correctly?

7.9.3. Requirements and Conditions

For early proteins to fold correctly and function in a prebiotic environment, the following conditions must be met simultaneously:

1. Amino acids must spontaneously form peptide bonds in the correct sequence.
2. The resulting polypeptides must be able to fold into stable, functional conformations.
3. The prebiotic environment must provide conditions conducive to protein folding (appropriate pH, temperature, and ionic concentrations).
4. Mechanisms must exist to prevent protein aggregation and misfolding.
5. For proteins requiring chaperones, a primitive chaperone system must already be in place.
6. This primitive chaperone system must itself be composed of properly folded proteins.
7. Energy sources (e.g., ATP) must be available to power chaperone-assisted folding.
8. Feedback mechanisms must exist to regulate chaperone activity and prevent over-assistance.
9. A system must be in place to degrade misfolded proteins that escape chaperone assistance.

These requirements present several contradictions:
- The need for a pre-existing chaperone system conflicts with the assumption that early proteins evolved in its absence.
- The requirement for complex regulatory mechanisms contradicts the presumed simplicity of early biological systems.
- The need for specific environmental conditions conflicts with the variable and often extreme conditions of the prebiotic Earth.

7.9.4. Relevant Scientific Terminology

Protein folding, molecular chaperones, native state, energy landscape, aggregation, misfolding, ATP-dependent chaperones, chaperonins, heat shock proteins (HSPs), protein quality control, proteostasis.

7.9.5. Illustrative Examples

Consider the GroEL/GroES chaperonin system in E. coli. This complex molecular machine encapsulates unfolded proteins in a hydrophilic chamber, allowing them to fold without interference. The system requires 14 identical 57 kDa GroEL subunits and 7 identical 10 kDa GroES subunits, arranged in a highly specific structure. It's challenging to envision how such a complex system could have evolved in the absence of already functional proteins.

7.9.6. Critical Examination of Current Theories

Current theories of early protein evolution often overlook or underestimate the challenges posed by protein folding. Models that propose the gradual evolution of protein function fail to account for the complex folding requirements of even relatively simple proteins. Scenarios invoking short peptides as early functional molecules face the challenge of explaining how these could have evolved into complex, chaperone-dependent proteins.

The RNA World hypothesis, which proposes RNA as the original self-replicating molecule, also faces challenges in explaining the transition to a protein-based metabolism. The complexity of the translation machinery and the need for already-folded proteins in this process create significant hurdles for this model.

7.9.7. Suggestion for Further Discussion

Future discussions on this topic should focus on developing testable hypotheses for primitive folding mechanisms that could have operated in the absence of modern chaperone systems. This might include exploring the potential role of mineral surfaces or simple organic molecules in facilitating early protein folding, or investigating whether certain protein folds are inherently more likely to form spontaneously and could have served as evolutionary starting points. In conclusion, the complexity of protein folding and the widespread requirement for chaperones in modern cells present significant challenges to naturalistic explanations for the origin of life. These challenges necessitate a reevaluation of current models and may require new, innovative approaches to understanding early protein evolution.

7.10 Metabolic Integration

The integration of synthesized proteins into functional metabolic pathways presents significant challenges to current naturalistic explanations for the origin of life. This analysis will focus on the complexities of metabolic integration, particularly in the context of amino acid biosynthesis, and the implications for early cellular evolution.

7.10.1. Quantitative Findings Challenging Conventional Theories

Recent studies have shown that a minimum of 112 enzymes is required to synthesize the 20 standard proteinogenic amino acids plus selenocysteine and pyrrolysine (Fujishima et al., 2018). This number represents a significant increase from earlier estimates and highlights the complexity of even the most basic cellular metabolic processes. Furthermore, these 112 enzymes are involved in a network of interdependent reactions. A study by Ravasz et al. (2002) on the metabolic network of E. coli revealed a hierarchical organization with a scale-free topology, characterized by a few highly connected metabolic hubs. This structure implies that the removal of even a small number of key enzymes could lead to catastrophic system-wide failures.

7.10.2. Implications for Current Scientific Models

These findings pose significant challenges to current models of early cellular evolution. The high number of enzymes required for amino acid biosynthesis suggests that early cells would have needed a remarkably complex metabolic system from the outset. This complexity is difficult to reconcile with the idea of a gradual evolution of metabolic pathways from simpler precursors. The interdependence of these enzymes also creates a "chicken and egg" problem: how could such a complex system of protein-based enzymes evolve when proteins themselves require this system to be synthesized?

7.10.3. Requirements and Conditions

For metabolic integration to occur naturally in a prebiotic environment, the following conditions must be met simultaneously:

1. A diverse pool of amino acids must be available in sufficient quantities.
2. Mechanisms for forming peptide bonds must exist to create functional enzymes.
3. Each of the 112+ enzymes required for amino acid biosynthesis must be present and functional.
4. These enzymes must be produced in the correct ratios to maintain metabolic balance.
5. Cofactors and coenzymes necessary for enzyme function must be available.
6. Energy sources (e.g., ATP) must be present to drive unfavorable reactions.
7. Cellular compartmentalization must exist to concentrate reactants and products.
8. Regulatory mechanisms must be in place to control enzyme activity and metabolic flux.
9. Transport systems must exist to move substrates and products between compartments.
10. A system for maintaining genomic information encoding these enzymes must be present.

These requirements present several contradictions:
- The need for a complex, interdependent enzyme system conflicts with the assumption of simpler precursor systems.
- The requirement for specific regulatory mechanisms contradicts the presumed lack of sophisticated control systems in early cells.
- The need for compartmentalization conflicts with models proposing metabolism-first scenarios in open prebiotic environments.