Intelligent Design, the best explanation of Origins

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Peptide bonding of amino acids to form proteins and its origins

http://reasonandscience.catsboard.com/t2130-peptide-bonding-of-amino-acids-to-form-proteins-and-its-origins

Primary Structure: Amino Acids Are Linked by Peptide Bonds to Form Polypeptide Chains
Proteins are linear polymers formed by linking the alpha-carboxyl group of one amino acid to the a-amino group of another amino acid. This type of linkage is called alpha peptide bond or an amide bond. The formation of a dipeptide from two amino acids is accompanied by the loss of a water molecule. The equilibrium of this reaction lies on the side of hydrolysis rather than synthesis under most conditions. Hence, the biosynthesis of peptide bonds requires an input of free energy. Nonetheless, peptide bonds are quite stable kinetically because the rate of hydrolysis is extremely slow; the lifetime of a peptide bond in aqueous solution in the absence of a catalyst approaches 1000 years.

A series of amino acids joined by peptide bonds form a polypeptide chain, and each amino acid unit in a polypeptide is called a residue. A polypeptide chain has directionality because its ends are different: an alpha -amino group is present at one end and an a -carboxyl group at the other. The amino end is taken to be the beginning of a polypeptide chain; by convention, the sequence of amino acids in a polypeptide chain is written starting with the amino-terminal residue. 12

Question: By the convention of whom ??!!

Thus, in the polypeptide Tyr-Gly-Gly-Phe- Leu (YGGFL), tyrosine is the amino-terminal (N-terminal) residue and leucine is the carboxyl-terminal (C-terminal) residue . Leu- Phe-Gly-Gly-Tyr (LFGGY) is a different polypeptide, with different chemical properties.


Amino acid sequences have direction. 
This illustration of the pentapeptide Tyr-Gly-Gly-Phe-Leu (YGGFL) shows the sequence from the amino terminus to the carboxyl terminus. This pentapeptide, Leu-enkephalin, is an opioid peptide that modulates the perception of pain. The reverse pentapeptide, Leu-Phe-Gly-Gly-Tyr (LFGGY), is a different molecule and has no such effects.

A polypeptide chain consists of a regularly repeating part, called the main chain or backbone, and a variable part, comprising the distinctive side chains (Figure below).


Components of a polypeptide chain. 
A polypeptide chain consists of a constant backbone (shown in black) and variable side chains (shown in green).

The polypeptide backbone is rich in hydrogen-bonding potential. Each residue contains a carbonyl group (C = O), which is a good hydrogen-bond acceptor, and, with the exception of proline, an NH group, which is a good hydrogen-bond donor. These groups interact with each other and with functional groups from side chains to stabilize particular structures

Peptidyl transferase catalyzes peptide-bond synthesis
A molecule called the Peptidyl Transferase Center (PTC) is considered by some as having an essential role in the emergence of life, since this catalytic ability to get together amino acids is crucial for protein synthesis and thus, for the first transition from an RNA world to a Ribonucleoprotein world, as seen in modern organisms.

All known cellular organisms have the PTC conserved and the process of reading the information contained in the messenger RNA, in general, is similar in all life forms. Would the common ancestor of all life forms be a part of the largest subunit of the ribosomal RNA? When thinking about LUCA as a molecule, and more specifically, as the large subunit of the ribosome or even more specifically as the PTC, there is an extensive modification into the junction point on which all living organisms came to be. Here the nature of LUCA is changed since it places the common point of origin in a time where the RNA was the information-carrying molecule and the cellular systems were still starting to maturate. 10

The ribosome accelerates peptide bond formation by lowering the activation entropy of the reaction due to positioning the two substrates, ordering water in the active site, and providing an electrostatic network that stabilizes the reaction intermediates. Proton transfer during the reaction appears to be promoted by a concerted proton shuttle mechanism that involves ribose hydroxyl groups on the tRNA substrate. 11

Positioning, ordering, providing, stabilizing, promoting a concerted shuttle mechanism are all tasks which we can easily attribute to the action of an intelligence, but could hardly emerge without external direction by random unguided events.

Protein synthesis in the cell is performed on ribosomes, large ribonucleoprotein particles that consist of three RNA molecules and more than 50 proteins. Ribosomes are composed of two subunits, the larger of which has a sedimentation coefficient of 50S in prokaryotes (the 50S subunit) and the smaller which sediments at 30S (the 30S subunit); together they form 70S ribosomes. The ribosome is a molecular machine that selects its substrates, aminoacyl-tRNAs (aa-tRNAs) d , rapidly and accurately and catalyzes the synthesis of peptides from amino acids. The 30S subunit contains the decoding site, where base-pairing interactions between the mRNA codon and the tRNA anticodon determines the selection of the cognate aa-tRNA.

The large ribosomal subunit contains the site of catalysis—the peptidyl transferase (PT) center—which is responsible for making peptide bonds during protein elongation and for the hydrolysis of peptidyl-tRNA (pepttRNA) during the termination of protein synthesis. The ribosome has three tRNA binding sites: A, P, and E sites ( figure below )


Schematic of Peptide Bond Formation on the Ribosome
The a-amino group of aminoacyl-tRNA in the A site (red) attacks the carbonyl carbon of the pept-tRNA in the P site (blue) to produce a new, one amino acid longer pept-tRNA in the A site and a deacylated tRNA in the P site. The 50S subunit, where the PT center is located, is shown in light gray and the 30S subunit in dark gray. A, P, and E sites of the ribosome are indicated.

During the elongation cycle of protein synthesis, aa-tRNA is delivered to the A site of the ribosome in a ternary complex e with elongation factor Tu (EF-Tu) c and GTP. Following GTP hydrolysis and release from EF-Tu, aa-tRNA accommodates in the A site of the Peptidyl Transferase Center ( PT center ) and reacts with pept-tRNA bound to the P site, yielding deacylated tRNA in the P site and A site pept-tRNA that is extended by one amino acid residue. The subsequent movement of tRNAs and mRNA through the ribosome (translocation) is catalyzed by another elongation factor (EF-G in bacteria). During translocation, pept-tRNA and deacylated tRNA move to the P and E sites, respectively; a new codon is exposed in the A site for the interaction with the next aa-tRNA, and the deacylated tRNA is released from the E site.

The movement of aa-tRNA into the A site is a multistep process that requires structural rearrangements of the ribosome, EF-Tu, and aa-tRNA.

Structure of the Active Site of the Peptidyl Transferase Center (PTC)
50S subunits are composed of two rRNA molecules, 23S rRNA and 5S rRNA, and more than 30 proteins (Figure A below).


Structure of the Peptidyl Transferase Center
(A) Crystal structure of the 50S subunit from H. marismortui with a transition state analog (red) bound to the active site. Ribosomal proteins are blue, the 23S rRNA backbone is brown, the 5S rRNA backbone is olive, and
rRNA bases are pale green. 
(B) Substrate binding to the active site. Base pairs formed between cytosine residues of the tRNA analogs in the A site (yellow) and P site (orange) with 23S rRNA bases (pale green) are indicated. The a-amino group of the A site substrate (blue) is positioned for the attack on the carbonyl carbon of the ester linking the peptide moiety of the P site substrate (green). Inner shell nucleotides are omitted for clarity.

The Mechanism of Peptide Bond Formation
The combined evidence supports the idea that peptide bond formation on the ribosome is driven by a favorable entropy change. The A and P site substrates are precisely aligned in the active center by interactions of the tRNA  CCA b sequences and of the nucleophilic a-amino group with residues of 23S rRNA in the active site. The most favorable catalytic pathway involves a six-membered transition state (Figure below) in which proton shuttling occurs via the 20-OH of A76 of the P site tRNA. The reaction does not involve chemical catalysis by ribosomal groups but may be modulated by conformational changes at the active site which can be induced by protonation.


Concerted Proton Shuttle Mechanism of Peptide Bond Formation
Pept-tRNA (P site) and aminoacyl-tRNA (A site) are blue and red, respectively, ribosome residues are pale green, and ordered water molecules are gray. The attack of the a-NH2 group on the ester carbonyl carbon results in a six-membered transition state in which the 20-OH group of the A site A76 ribose moiety donates its proton to the adjacent leaving 30 oxygen and simultaneously receives a proton from the amino group. Ribosomal residues are not involved in chemical catalysis but are part of the H bond network that stabilizes the transition state.

In addition to placing the reactive groups into close proximity and precise orientation relative to each other, the ribosome appears to work by providing an electrostatic environment that reduces the free energy of forming the highly polar transition state, shielding the reaction against bulk water, helping the proton shuttle forming the leaving group or a combination of these effects. With this preorganized network, the ribosome avoids the extensive solvent reorganization that is inevitable in the corresponding reaction in solution, resulting in significantly more favorable entropy of activation of the reaction on the ribosome.

With both the P site and the A site occupied by aminoacyl-tRNA, the stage is set for the formation of a peptide bond: the formylmethionine molecule linked to the initiator tRNA will be transferred to the amino group of the amino acid in the A site. The formation of the peptide bond, one of the most important reactions in life, is a thermodynamically spontaneous reaction catalyzed by a site on the 23S rRNA of the 50S subunit called the peptidyl transferase center. This catalytic center is located deep in the 50S subunit near the tunnel that allows the nascent peptide to leave the ribosome. The ribosome, which enhances the rate of peptide bond synthesis by a factor of 10^7 over the uncatalyzed reaction, derives much of its catalytic power from catalysis by proximity and orientation. The ribosome positions and orients the two substrates so that they are situated to take advantage of the inherent reactivity of an amine group (on the aminoacyl-tRNA in the A site) with an ester (on the initiator tRNA in the P site). The amino group of the aminoacyl-tRNA in the A site, in its unprotonated state, makes a nucleophilic attack on the ester linkage between the initiator tRNA and the formylmethionine molecule in the P site (Figure A below). 


Peptide-bond formation.
(A) The amino group of the aminoacyl-tRNA attacks the carbonyl group of the ester linkage of the peptidyl-tRNA. 
(B) An eight-membered transition state is formed. Note: Not all atoms are shown and some bond lengths are exaggerated for clarity.
(C) This transition state collapses to form the peptide bond and release the deacylated tRNA.

The nature of the transition state that follows the attack is not established and several models are plausible. One model proposes roles for the 2' OH of the adenosine of the tRNA in the P site and a molecule of water at the peptidyl transferase center (Figure B above). The nucleophilic attack of the a-amino group generates an eight-membered transition state in which three protons are shuttled about in a concerted manner. The proton of the attacking amino group hydrogen bonds to the 2' oxygen of ribose of the tRNA. The hydrogen of 2' OH, in turn, interacts with the oxygen of the water molecule at the center, which then donates a proton to the carbonyl oxygen. A collapse of the transition state with the formation of the peptide bond allows protonation of the 3'OH of the now empty tRNA in the P site (Figure C above). The stage is now set for translocation and formation of the next peptide bond.

The formation of a peptide bond is followed by the GTP-driven a translocation of tRNAs and mRNA
With the formation of the peptide bond, the peptide chain is now attached to the tRNA whose anticodon is in the A site on the 30S subunit. The two subunits rotate with respect to one another, and this structural change places the CCA b end of the same tRNA and its peptide in the P site of the large subunit (Figure below). 


Mechanism of protein synthesis. 
The cycle begins with peptidyltRNA in the P site. 
(1) An aminoacyl-tRNA binds in the A site. 
(2) With both sites occupied, a new peptide bond is formed.
(3) The tRNAs and the mRNA are translocated through the action of elongation factor G, which moves the deacylated tRNA to the E site. 
(4) Once there, the tRNA is free to dissociate to complete the cycle.

Another aminoacyl-tRNA arrives and binds at the A site (1). Again, peptide bond synthesis occurs (2). However, protein synthesis cannot continue without the translocation of the mRNA and the tRNAs within the ribosome. Elongation factor G (EF-G, also called translocase) c catalyzes the movement of mRNA, at the expense of GTP hydrolysis, by a distance of three nucleotides. Now, the next codon is positioned in the A site for interaction with the incoming aminoacyl-tRNA (3). The peptidyl- tRNA moves out of the A site into the P site on the 30S subunit and at the same time, the deacylated tRNA moves out of the P site into the E site and is subsequently released from the ribosome (4). The movement of the peptidyl-tRNA into the P site shifts the mRNA by one codon, exposing the next codon to be translated in the A site.

The three-dimensional structure of the ribosome undergoes significant change during translocation, and evidence suggests that translocation may result from properties of the ribosome itself. However, EF-G accelerates the process. A possible mechanism for accelerating the translocation process is shown in Figure below.


Translocation mechanism. 
In the GTP form, EF-G binds to the A site on the 50S subunit. This binding stimulates GTP hydrolysis, inducing a conformational change in EF-G that forces the tRNAs and mRNA to move through the ribosome by a distance corresponding to one codon.

Question: How did unguided random processes select and finely tune the forces to move the tRNAs and mRNA by the right distance of one codon?

EF-G in the GTP form binds to the ribosome near the A site, interacting with the 23S rRNA of the 50S subunit. The binding of EF-G to the ribosome stimulates the GTPase activity of EF-G. On GTP hydrolysis, EF-G undergoes a conformational change that displaces the peptidyl-tRNA in the A site to the P site, which carries the mRNA and the deacylated tRNA with it. The dissociation of EF-G leaves the ribosome ready to accept the next aminoacyl-tRNA into the A site. Note that the peptide chain remains in the P site on the 50S subunit throughout this cycle, growing into the exit tunnel. This cycle is repeated, with mRNA translation taking place in the 5' ==>> 3' direction, as new aminoacyl-tRNAs move into the A site, allowing the polypeptide to be elongated until a stop signal is found.

The direction of translation has important consequences. Transcription also is in the 5' ==>> 3' direction. If the direction of translation were opposite that of transcription, only fully synthesized mRNA could be translated. In contrast, because the directions are the same, mRNA can be translated while it is being synthesized.

Question: How could natural, unguided, random processes select the right direction to be translated? Trial and error ?

In bacteria, almost no time is lost between transcription and translation. The 5' end of mRNA interacts with ribosomes very soon after it is made, well before the 3' end of the mRNA molecule is finished. An important feature of bacterial gene expression is that translation and transcription are closely coupled in space and time.

There is a huge gap that has to be filled between " modern " polypeptide formation through ribosomes, mRNA, and tRNA's, and supposed primordial amino chain formations without this advanced machinery. How could the gap be closed? Not only are prebiotic mechanisms unlikely, but the transition would require the emergence of all the complex machinery and afterward transition from one mechanism to the other. Tamura admits that fact clearly: the ultimate route to the ribosome remains unclear.   It takes a big leap of faith to believe, that could be possible in any circumstances. 

Mystery of Life's Origin 4
Experimental evidence indicates that if there are bonding preferences between amino acids, they are not the ones found in natural organisms. There are three basic requirements for a biologically functional protein.

One: It must have a specific sequence of amino acids. At best prebiotic experiments have produced only random polymers. And many of the amino acids included are not found in living organisms.

Second: An amino acid with a given chemical formula may in its structure be either “righthanded” (D-amino acids) or “left-handed” (L-amino acids). Living organisms incorporate only L-amino acids. However, in prebiotic experiments where amino acids are formed approximately equal numbers of D- and L-amino acids are found. This is an “intractable problem” for chemical evolution (p. vi).

Third: In some amino acids there are more positions than one on the molecule where the amino and carboxyl groups may join to form a peptide bond. In natural proteins only alpha-peptide bonds (designating the location of the bond) are found. In proteinoids, however, beta, gamma and epsilon peptide bonds largely predominate. Just the opposite of what one would expect if bonding preferences played a role in prebiotic evolution.

Studies of peptide bond formation in the absence of modern biological machinery can give insight into the mechanism employed by the ribosome’s active site, as well as yield important information in the prebiotic route to the first peptides in the origin of life. The formation of a peptide bond (reaction R1 shown below) is a condensation reaction, eliminating a water molecule for each peptide bond formed, and thus faces both thermodynamic and kinetic constraints in bulk aqueous solution



Amino Acids joined together through a dehydration reaction, where a water molecule is formed and removed to form a covalent bond called a peptide bond. A structure resulting from a bunch of these bonds repeating over and over is called a polypeptide. Like DNA molecules, polypeptides have a direction: they’ve got an amino acid at one end (the N-terminus) and a carboxyl group at the other (the C-terminus).

In modern biology, the condensation reactions necessary in the formation of peptide bonds are facilitated catalytically by the large subunit of the ribosome.

Fazale Rana's Cell's design: The chemical reactions that form the bonds that join amino acids together in polypeptide chains are catalyzed or assisted by ribosomes. The ribosome, mRNA, and tRNA molecules work cooperatively to produce proteins. Using an assembly-line process, protein manufacturing machinery forms the polypeptide chains (that constitute proteins) one amino acid at a time. This protein synthetic apparatus joins together three to five amino acids per second. Ribosomes, in conjunction with mRNA and tRNAs, assemble the cell's smallest proteins, about one hundred to two hundred amino acids in length, in less than one minute. The processing of proteins in the lumen (posttranslational modification) is quite extensive. Posttranslational modifications include (1) formation and reshuffling of disulfide bonds (these bonds form between the side chains of cysteine amino acid residues within a protein, stabilizing its three-dimensional structure)

Amino Acids Are Added to the C-terminal End of a Growing Polypeptide Chain
Each amino acid is first coupled to specific tRNA molecules, next is the mechanism that joins these amino acids together to form proteins. The fundamental reaction of protein synthesis is the formation of a peptide bond between the carboxyl group at the end of a growing polypeptide chain and a free amino group on an incoming amino acid. Consequently, a protein is synthesized stepwise from its N-terminal end to its C-terminal end. Throughout the entire process, the growing carboxyl end of the polypeptide chain remains activated by its covalent attachment to a tRNA molecule (forming a peptidyl-tRNA). Each addition disrupts this high-energy covalent linkage, but immediately replaces it
with an identical linkage on the most recently added amino acid



The incorporation of an amino acid into a protein. A polypeptide chain grows by the stepwise addition of amino acids to its C-terminal end. The formation of each peptide bond is energetically favorable because the growing C-terminus has been activated by the covalent attachment of a tRNA molecule. The peptidyl-tRNA linkage that activates the growing end is regenerated during each addition. The amino acid side chains have been abbreviated as R1, R2, R3, and R4; as a referencepoint, all of the atoms in the second amino acid in the polypeptide chain are shaded gray. The figure shows the addition of the fourth amino acid (red) to the growing chain.

Peptide Bond Formation: RNA's Big Bang

The genetic code may have been established gradually (Wong, 1975). 5

observe the " may have's ", by some means, might have, proposed the idea, would have,

The second law of thermodynamics indicates that peptide bond formation does not occur spontaneously. Therefore, energy must be added into the system by some means and amino acids must be "activated." Modern biological systems use the energy of the ATP hydrolysis for coupling many reactions (Lipmann, 1941). However, during the prebiotic stage, the light from the sun, geothermal energy, pressure in the thermal vent, or other similar sources may have been used in the process of activating the molecules of a system. The development of prebiotic precursors of biomolecules might have occurred in interstellar space, and were subsequently transferred to Earth by comets, asteroids, or meteorites (Oró, 1961; Chyba et al., 1990; Chyba & Sagan, 1992). Reactions on clay (Paecht-Horowitz et al., 1970) and/or dry mixtures of amino acids (Fox & Harada, 1958) may have facilitated the condensation of activated amino acids, thereby forming peptide bonds. Iron sulfate is known to cause unusual reducing reactions, especially with H2S. Wächtershäuser (1992) previously proposed the idea of an "iron-sulfur world" where low-molecular weight constituents may have originated autotrophic metabolism. In such circumstances, amino acids would have been converted into simple peptides (Huber & Wächtershäuser, 1998). In fact, it has been demonstrated that the peptide containing a thioester at the carboxyl-terminal undergoes nucleophilic attack by the side chain of the Cys residue at the amino terminal of another peptide. Moreover, the formed thioester ligation product readily undergoes a rapid intramolecular reaction at the α-amino group of the Cys to yield a product with a native peptide bond. This series of events is called "native chemical ligation" and is important in the general application of protein chemistry (Dawson et al., 1994). These possibilities should be further considered in terms of the very early mechanisms responsible for peptide bond formation. However, because we must consider the modern ribosome, we cannot avoid consideration of RNA in the evolution of biological systems.

Its remarkable how mainstream scientific papers give to their naturalistic proposals a positive connotation, but without providing compelling evidence for their assertions.

The Emergence of Information-Rich Biopolymers 1
Given an ocean full of small molecules of the types likely to be produced on a prebiological earth with the types of processes postulated by origin of life enthusiasts, we must next approach the question of polymerization. This question poses a two edged sword: we must first demonstrate that macromolecule synthesis is possible under prebiological conditions, then we must construct a rationale for generating macromolecules rich in the information necessary for usefulness in a developing precell. We shall deal with these separately.

The synthesis of proteins and nucleic acids from small molecule precursors represents one of the most difficult challenges to the model of prebiological evolution. There are many different problems confronted by any proposal. Polymerization is a reaction in which water is a product. Thus it will only be favored in the absence of water. The presence of precursors in an ocean of water favors depolymerization of any molecules that might be formed. Careful experiments done in an aqueous solution with very high concentrations of amino acids demonstrate the impossibility of significant polymerization in this environment.

Polymer formation in aqueous environments would most likely have been necessary on early Earth because the liquid ocean would have been the reservoir of amino acid precursors needed for protein synthesis. 3

A thermodynamic analysis of a mixture of protein and amino acids in an ocean containing a 1 molar solution of each amino acid (100,000,000 times higher concentration than we inferred to be present in the prebiological ocean) indicates the concentration of a protein containing just 100 peptide bonds (101 amino acids) at equilibrium would be 10-338 molar. Just to make this number meaningful, our universe may have a volume somewhere in the neighborhood of 10^85 liters. At 10-338 molar, we would need an ocean with a volume equal to 10229 universes (100, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000) just to find a single molecule of any protein with 100 peptide bonds. So we must look elsewhere for a mechanism to produce polymers. It will not happen in the ocean.

Sidney Fox, an amino acid chemist, and one of my professors in graduate school, recognized the problem and set about constructing an alternative. Since water is unfavorable to peptide bond formation, the absence of water must favor the reaction. Fox attempted to melt pure crystalline amino acids in order to promote peptide bond formation by driving off water from the mix. He discovered to his dismay that most amino acids broke down to a tarry degradation product long before they melted. After many tries, he discovered two of the 20 amino acids, aspartic and glutamic acid, would melt to a liquid at about 200oC. He further discovered that if he were to dissolve the other amino acids in the molten aspartic and glutamic acids, he could produce a melt containing up to 50% of the remaining 18 amino acids. It was no surprise then that the amber liquid, after cooking for a few hours, contained polymers of amino acids with some of the properties of proteins. He subsequently named the product proteinoids. The polymerized material can be poured into an aqueous solution, resulting in the formation of spherules of protein-like material which Fox has likened to cells. Fox has claimed nearly every conceivable property for his product, including that he had bridged the macromolecule to cell transition. He even went so far as to demonstrate a piece of lava rock could substitute for the test tube in proteinoid synthesis and claimed the process took place on the primitive earth on the flanks of volcanoes. However, his critics, as well as his own students, have stripped his credibility. Note the following problems:

1) Proteinoids are not proteins; they contain many non-peptide bonds and unnatural cross-linkages.

2) The peptide bonds they do contain are beta bonds, whereas all biological peptide bonds are alpha.

3) His starting materials are purified amino acids bearing no resemblance to the materials available in the "dilute soup." If one were to try the experiment with condensed "prebiological soup," tar would be the only product.

4) The ratio of 50% Glu and Asp necessary for success in these experiments bears no resemblance to the vastly higher ratio of Gly and Ala found in nearly all primitive earth synthesis experiments.

5) There is no evidence of information contained in the molecules.

All of his claims have failed the tests of rationality when examined carefully. As promising as his approach seemed in theory, the reality is catastrophic to the hopes of paleobiogeochemists.

A number of other approaches have been tried. The most optimistic of these is the use of clays. Clays are very thin, very highly ordered arrays of complex aluminum silicates with numerous other cations. In this environment, the basic amino groups tend to order and polymers of several dozen amino acids have been produced. While these studies have generated enthusiastic interest on the part of prebiological evolutionists, their relevance is quickly dampened by several factors.

1) While ordered amino acids joined by peptide bonds result, the product contains no meaningful information.

2) The clays exhibit a preference for basic amino acids.

3) No polymerization of amino acids results if free amino acids are used.

4) Pure activated amino acids attached to adenine must be used in order to drive the reaction toward polymerization. Adenylated amino acids are not exactly the most likely substrate to be floating about the prebiological ocean.

5) The resultant polymers are three dimensional rather than linear, as is required for biopolymers.

At least one optimistic scientist (Cairns-Smith, 1982) believes that the clay particles themselves formed the substance of the first organisms! In reality, the best one can hope for from such a scenario is a racemic polymer of proteinous and non-proteinous amino acids with no relevance to living systems.

           A final chapter has recently been opened with the discovery of autocatalytic RNA molecules. These were originally received with great excitement by the prebiological evolutionists because they gave hope of alleviating the need to make proteins in the first cell. These so-called "ribozymes" proved incapable of rising to the occasion, however, for not only are the molecules themselves very limited in what they have been shown capable of doing, but the production of the precursors of RNA by any prebiological mechanism considered thus far is a problem at least as difficult as the one ribozymes purport to solve:

1) While ribose can be produced under simulated prebiological conditions via the formose reaction, it is a rare sugar in formaldehyde polymers (the prebiological mechanism believed to have given rise to sugars). In addition the presence of nitrogenous substances such as amino acids in the reaction mixture would prevent sugar synthesis (Shapiro, 1988). Cairns-Smith (1993) has summarized the situation as follows:"Sugars are particularly trying. While it is true that they form from formaldehyde solutions, these solutions have to be far more concentrated than would have been likely in primordial oceans. And the reaction is quite spoilt in practice by just about every possible sugar being made at the same time - and much else besides. Furthermore the conditions that form sugars also go on to destroy them. Sugars quickly make their own special kind of tar - caramel - and they make still more complicated mixtures if amino acids are around."

2) When produced and condensed with a nucleotide base, a mixture of optical isomers results, only one of which is relevant to prebiological studies.

3) Polymerization of nucleotides is inhibited by the incorporation of such an enantiomorph.

4) While only 3'-5' polymers occur in biological systems, 5'-5' and 2'-5' polymers are favored in prebiological type synthetic reactions (Joyce and Orgel, 1993, but see Usher,et. al. for an interesting sidelight).

5) None of the 5 bases present in DNA/RNA are produced during HCN oligomerization in dilute solutions (the prebiological mechanism believed to give rise to nucleotide bases). And many other non-coding bases would compete during polymerization at higher concentrations of HCN.

In addition to the problems of synthesis of the precursors and the polymerization reactions, the whole scheme is dependent on the ability to synthesize an RNA molecule which is capable of making a copy of itself, a feat that so far has eluded strenuous efforts. The molecule must also perform some function vital to initiating life force. So far all of this talk of an "RNA World" remains wishful thinking best categorized as fiction. The most devastating indictment of the scheme however, is that it offers no clue as to how one gets from such a scheme to the DNA-RNA-Protein mechanism of all living cells. The fact that otherwise rational scientists would exhibit such rampant enthusiasm for this scheme so quickly reveals how little faith they have in all other scenarios for the origin of life, including the ones discussed above.

Guanosine triphosphate ( GTP) is a high energy nucleotide (not to be confused with nucleoside) found in the cytoplasm or polymerised to form the guanine base. 17 It is a result of it's complex three dimensional structure and the variety of different chemical groups which it comprises of. 
Guanosine-5'-triphosphate (GTP) is a purine nucleoside triphosphate. It is one of the building blocks needed for the synthesis of RNA during the transcription process. Its structure is similar to that of the guanine nucleobase, the only difference being that nucleotides like GTP have a ribose sugar and three phosphates, with the nucleobase attached to the 1' and the triphosphate moiety attached to the 5' carbons of the ribose. It also has the role of a source of energy or an activator of substrates in metabolic reactions, like that of ATP, but more specific. It is used as a source of energy for protein synthesis and gluconeogenesis. GTP is essential to signal transduction, in particular with G-proteins, in second-messenger mechanisms where it is converted to guanosine diphosphate (GDP) through the action of GTPases. 16



Guanosine is a purine nucleoside comprising guanine attached to a ribose (ribofuranose) ring via a β-N9-glycosidic bond. Guanosine can be phosphorylated to become guanosine monophosphate (GMP), cyclic guanosine monophosphate (cGMP), guanosine diphosphate (GDP), and guanosine triphosphate (GTP). These forms play important roles in various biochemical processes such as synthesis of nucleic acids and proteins, photosynthesis, muscle contraction, and intracellular signal transduction (cGMP). 18



For the synthesis purines, following enzymes are required:

amidophosphoribosyltransferase, 2.4.2.14
phosphoribosylamine-glycine ligase, 6.3.4.13
phosphoribosylglycinamide formyltransferase, 2.1.2.2
phosphoribosylformylglycinamidine synthase, 6.3.5.3
phosphoribosylformylglycinamidine cyclo-ligase, 6.3.3.1 20

Guanine is one of the four main nucleobases found in the nucleic acids DNA and RNA. 
For scientists attempting to understand how the building blocks of RNA originated on Earth, guanine -- the G in the four-letter code of life -- has proven to be a particular challenge. While the other three bases of RNA -- adenine (A), cytosine (C) and uracil (U) -- could be created by heating a simple precursor compound in the presence of certain naturally occurring catalysts, guanine had not been observed as a product of the same reactions.

Ribose
How could and would random events attach a phosphate group to the right position of a ribose molecule to provide the necessary chemical activity? And how would non-guided random events be able to attach the nucleic bases to the ribose?  The coupling of a ribose with a nucleotide is the first step to form RNA, and even those engrossed in prebiotic research have difficulty envisioning that process, especially for purines and pyrimidines.”
The sugar found in the backbone of both DNA and RNA, ribose, has been particularly problematic, as the most prebiotically plausible chemical reaction schemes have typically yielded only a small amount of ribose mixed with a diverse assortment of other sugar molecules. 16

Glycosidic bond
The formation of nucleosides in abiotic conditions is a major hurdle in origin-of-life studies. The formamido pyrimidine-based syntheses are high regioselective, moderately stereoselective, multi-step, only apply to purines and afford a mixture of furanosides and pyranosides. The prebiotic worth of these syntheses is inversely proportional to the procedural complexities involved, requiring numerous concentration, purification and supplementation steps, designed to specifically overcome intermediate reactions bottlenecks. 21

Guanosine monophosphate (GMP)



b CCA is a terminal sequence required for the function of all tRNAs, is added to the 3' ends of tRNA molecules for which this terminal sequence is not encoded in the DNA. The enzyme that catalyzes the addition of CCA is atypical for an RNA polymerase in that it does not use a DNA template. A third type of processing is the modification of bases and ribose units of ribosomal RNAs. 6 CCA is added by the CCA-adding enzyme (Figure below).


Transfer RNA precursor processing. 
The conversion of a yeast tRNA precursor into a mature tRNA requires the removal of a 14-nucleotide intron (yellow), the cleavage of a 59 leader (green), and the removal of UU and the attachment of CCA at the 39 end (red). In addition, several bases are modified.

Eukaryotic tRNAs are also heavily modified on base and ribose moieties; these modifications are important for function. In contrast with prokaryotic tRNAs, many eukaryotic pre-tRNAs are also spliced by an endonuclease and a ligase to remove an intron.

tRNA nucleotidyltransferase adds the invariant CCA terminus to the tRNA 30-end, a central step in tRNA maturation.7


Protein synthesis takes place in cytosolic ribosomes, mitochondria (mitoribosomes), and in plants, the plastids (chloroplast ribosomes). Each of these compartments requires a complete set of functional tRNAs to carry out protein synthesis. The production of mature tRNAs requires processing and modification steps such as the addition of a 3’-terminal cytidine-cytidine-adenosine (CCA). Since no plant tRNA genes encode this particular sequence, a tRNA nucleotidyltransferase must add this sequence post-transcriptionally and therefore is present in all three compartments. 8

c  EF-G (elongation factor G, historically known as translocase) is involved in protein translation. As a GTPase, EF-G catalyzes the movement (translocation) of transfer RNA (tRNA) and messenger RNA (mRNA) through the ribosome. EF-G is made up of 704 amino acids that form 5 domains, labeled Domain I through Domain V. 9



d The joining of an amino acid to a tRNA molecule to form an aminoacyl-tRNA is catalyzed by a specific enzyme called an Aminoacyl tRNA synthetase 14An aminoacyl-tRNA synthetase (aaRS or ARS), also called tRNA-ligase, is an enzyme that attaches the appropriate amino acid onto its tRNA. It does so by catalyzing the esterification of a specific cognate amino acid or its precursor to one of all its compatible cognate tRNAs to form an aminoacyl-tRNA. In humans, the 20 different types of aa-tRNA are made by the 20 different aminoacyl-tRNA synthetases, one for each amino acid of the genetic code. This is sometimes called "charging" or "loading" the tRNA with the amino acid. Once the tRNA is charged, a ribosome can transfer the amino acid from the tRNA onto a growing peptide, according to the genetic code. Aminoacyl tRNA therefore plays an important role in RNA translation, the expression of genes to create proteins. 13

e ternary complex is a protein complex containing three different molecules that are bound together.  15

1) http://origins.swau.edu/papers/life/chadwick/default.html
2) http://www.ncbi.nlm.nih.gov/books/NBK22364/
3) http://www.pnas.org/content/109/39/15697.full
4) http://www.cogmessenger.org/wp-content/uploads/2013/06/Mystery_of_Life_Origin.pdf
5) http://journalofcosmology.com/Abiogenesis130.html
6. Styer, Biochemistry, 8th. edition, page 870
7. http://sci-hub.tw/10.1002/bies.201500043
8. https://en.wikipedia.org/wiki/TRNA_nucleotidyltransferase
9. https://en.wikipedia.org/wiki/EF-G
10. http://sci-hub.tw/https://www.cambridge.org/core/journals/international-journal-of-astrobiology/article/buds-of-the-tree-the-highway-to-the-last-universal-common-ancestor/ED26AA7787BA5A152090913CC7C20067
11. http://sci-hub.tw/10.1016/j.molcel.2007.03.015
12. Styer, Biochemistry, 8th. edition, page 36
13. https://en.wikipedia.org/wiki/Aminoacyl_tRNA_synthetase
14. http://reasonandscience.catsboard.com/t2057-origin-of-translation-of-the-4-nucleic-acid-bases-and-the-20-amino-acids-and-the-universal-assignment-of-codons-to-amino-acids#6011
15. https://en.wikipedia.org/wiki/Ternary_complex
16. https://en.wikipedia.org/wiki/Guanosine_triphosphate
17. https://teaching.ncl.ac.uk/bms/wiki/index.php/Guanosine_triphosphate
18. https://en.wikipedia.org/wiki/Guanosine
19. http://sci-hub.tw/https://www.sciencedirect.com/science/article/pii/B0122270800005760
20. http://reasonandscience.catsboard.com/t2028-biosynthesis-of-the-dna-double-helix-evidence-of-design
21. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5677017/

more:
http://www3.uah.es/farmamol/New_Science_Press/nsp-protein-1.pdf



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27 What Is the Metabolic Fate of Ammonium? on Thu Mar 29, 2018 9:04 am

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What Is the Metabolic Fate of Ammonium?

Plants and microorganism which utilize nitrate as nitrogen source have to reduce it to the level of ammonia before incorporation into various organic compounds of their cells, because in their cellular constituents nitrogen is present in a reduced state. 7 Reduction of nitrate to the level of ammonia involves conversion of nitrogen from its highest oxidized state (+5) to the most reduced state (-3), requiring transfer of 8 electrons. This conversion is supposed to take place in 4 steps, each step consisting of a two-electron transfer reaction.



The above sequence is known as assimilatory nitrate reduction pathway which is distinguished from a dissimilatory pathway operating in nitrate respiration (also known as de-nitrification). The first step in the above pathway involves reduction of nitrate to nitrite.

The reaction is catalysed by nitrate reductase. In microorganisms, plants and fungi, nitrate reductase is a soluble cytoplasmic enzyme. In Neurospora, the enzyme is a molybdenum containing flavo-protein. The probable pathway of electron flow to nitrate is NADPH2 —> FAD Mo —> NO3–.

Molybdenum undergoes valency change from Mo5+ to Mo6+ during electron transfer for reduction of nitrate to nitrite. Reduction of nitrite is catalysed by nitrite reductase. Further electron transport for production of NH3 requires a highly electronegative reductant. In green plants, reduced ferredoxine produced by the light-reaction of photosynthesis probably acts as the terminal reductant of nitrite to ammonia. The probable path for electron flow to nitrite is ferredoxine (reduced) —> NADP —> FAD —> NO2–.

The final product of nitrate reduction, ammonia, is then incorporated into organic compounds by the several alternative routes described below:

Incorporation of Ammonia into Organic Compounds
The key entry point is the amino acid glutamate. Glutamate and glutamine are the nitrogen donors in a wide range of biosynthetic reactions. Glutamine synthetase, which catalyzes the formation of glutamine from glutamate, is a main regulatory enzyme of nitrogen metabolism. 17

Glutamate dehydrogenase, Glutamine Synthetase & Aminotransferases play central roles in amino acid biosynthesis. The combined action of the enzymes glutamate dehydrogenase, glutamine synthetase, and the aminotransferases (Figure below) converts inorganic ammonium ion into the α-amino nitrogen of amino acids.


Glutamate, the precursor of the so-called “glutamate family” of amino acids, is formed by the reductive amidation of the citric acid cycle α-ketoglutarate, a reaction catalyzed by mitochondrial glutamate dehydrogenase ( first reaction, picture above ) The reaction strongly favors glutamate synthesis, which lowers the concentration of cytotoxic ammonium ion. The amidation of glutamate to glutamine catalyzed by glutamine synthetase ( second reaction, figure above ) and involves the intermediate formation of γ-glutamyl phosphate ( third reaction, figure above ) Following the ordered binding of glutamate and ATP, glutamate attacks the γ-phosphorus of ATP, forming γ-glutamyl phosphate and ADP. NH4+ then binds, and uncharged NH3 attacks γ-glutamyl phosphate. Release of Pi and of a proton from the γ-amino group of the tetrahedral intermediate then allows release of the product, glutamine.

Ammonia is incorporated into biomolecules through Glutamate and Glutamine. Reduced nitrogen in the form of NH4+ is assimilated into amino acids and then into other nitrogen-containing biomolecules. Two amino acids, glutamate and glutamine, provide the critical entry point. These same two amino acids play central roles in the catabolism of ammonia and amino groups in amino acid oxidation. Glutamate is the source of amino groups for most other amino acids, through transamination reactions. The amide nitrogen of glutamine is a source of amino groups in a wide range of biosynthetic processes.  An Escherichia coli cell requires so much glutamate that this amino acid is one of the primary solutes in the cytosol. Its concentration is regulated not only in response to the cell’s nitrogen requirements but also to maintain an osmotic balance between the cytosol and the external medium. The biosynthetic pathways to glutamate and glutamine are simple, and all or some of the steps occur in most organisms. The most important pathway for the assimilation of NH4+ into glutamate requires two reactions. First, glutamine synthetase catalyzes the reaction of glutamate and NH4+ to yield glutamine. This reaction takes place in two steps. 



Given the prevalence of Nitrogen atoms in cellular components, it is surprising that only three enzymatic reactions introduce ammonium into organic molecules. Of these three, glutamate dehydrogenase and glutamine synthetase are responsible for most of the ammonium assimilated into carbon compounds. 

Glutamate dehydrogenase (GDH)
Glutamate dehydrogenase (GDH) catalyzes the reductive amination of a-ketoglutarate to yield glutamate. Reduced pyridine nucleotides (NADH or NADPH) provide the reducing power.
Alpha-ketoglutarate (AKG) is a key molecule in the Krebs cycle determining the overall rate of the citric acid cycle of the organism. It is a nitrogen scavenger and a source of glutamate and glutamine that stimulates protein synthesis and inhibits protein degradation in muscles. 5

Untangling the glutamate dehydrogenase allosteric nightmare
Glutamate dehydrogenase (GDH) is found in all living organisms, but only animal GDH is regulated by a large repertoire of metabolites. More than 50 years of research to better understand the mechanism and role of this allosteric network has been frustrated by its sheer complexity. However, recent studies have begun to tease out how and why this complex behavior evolved. Much of GDH regulation probably occurs by controlling a complex ballet of motion necessary for catalytic turnover and has evolved concomitantly with a long antenna-like feature of the structure of the enzyme. Ciliates, the ‘missing link’ in GDH evolution, might have created the antenna to accommodate changing organelle functions and was refined in humans to, at least in part, link amino acid catabolism with insulin secretion. 20


A model for GDH allostery. 
This figure shows that GDH allostery might be a form of exaptation. Here, the antenna and some regulation could have been created in Ciliates in response to the requirement of organelle function. These features might have then been further adapted for a different function, insulin homeostasis, in animals.

Nitrogen transporter
Another function is to combine with nitrogen released in the cell, therefore preventing nitrogen overload. α-Ketoglutarate is one of the most important nitrogen transporters in metabolic pathways. The amino groups of amino acids are attached to it (by transamination) and carried to the liver where the urea cycle takes place. α-Ketoglutarate is transaminated, along with glutamine, to form the excitatory neurotransmitter glutamate. Glutamate can then be decarboxylated (requiring vitamin B6) into the inhibitory neurotransmitter GABA. 6

To a first approximation, two regulatory controls are paramount (Figure below):



(1) ADP inhibits the activity of nitrogenase; thus, as the ATP/ADP ratio drops, nitrogen fixation is blocked. 
(2) NH4+ represses the expression of the nif genes, the genes that encode the proteins of the nitrogen-fixing system. To date, some 20 nif genes have been identified with the nitrogen fixation process. Repression of nif gene expression by ammonium, the primary product of nitrogen fixation, is an efficient and effective way of shutting down N2 fixation when its end product is not needed. In addition, in some systems, covalent modification of nitrogenase
reductase leads to its inactivation. Inactivation occurs when Arg101 of nitrogenase reductase receives an ADP-ribosyl group donated by NAD1.

This reaction provides an important interface between nitrogen metabolism and cellular pathways of carbon and energy metabolism because a-ketoglutarate is a citric acid cycle intermediate.

Glutamine synthetase (GS)
Glutamine synthetase (GS) is an enzyme that plays an essential role in the metabolism of nitrogen by catalyzing the condensation of glutamate and ammonia to form glutamine 19





Glutamine Synthetase Is a Central Control Point in Nitrogen Metabolism
Glutamine is the amino group donor in the formation of many biosynthetic products as well as being a storage form of ammonia. The control of glutamine synthetase is therefore vital for regulating nitrogen metabolism. Mammalian glutamine synthetases are activated by α-ketoglutarate, the product of glutamate’s oxidative deamination. This control presumably helps prevent the accumulation of the ammonia produced by that reaction. Bacterial glutamine synthetase has a much more elaborate control system. The enzyme, which consists of 12 identical 468- residue subunits arranged at the corners of a hexagonal prism is regulated by several allosteric effectors as well as by covalent modification. Several aspects of its control system bear note. Nine allosteric feedback inhibitors, each with its own binding site, control the activity of bacterial glutamine synthetase in a cumulative manner. Six of these effectors—histidine, tryptophan, carbamoyl phosphate (as synthesized by carbamoyl phosphate synthetase), glucosamine- 6-phosphate, AMP, and CTP—are all end products of pathways leading from glutamine. The other three—alanine, serine, and glycine—reflect the cell’s nitrogen level. 18 


X-Ray structure of glutamine synthetase from the bacterium Salmonella typhimurium. 
The enzyme consists of 12 identical subunits, here drawn in ribbon form, arranged with D6 symmetry (the symmetry of a hexagonal prism). 
(a) View along the sixfold axis of symmetry showing only the six subunits of the upper ring in different colors, with the lower right subunit colored in rainbow order from its N-terminus (blue) to its C-terminus (red). The subunits of the lower ring are roughly directly below those of the upper ring. A pair of Mn2+ ions (purple spheres) that occupy the positions of the Mg2+ ions required for enzymatic activity are bound in each active site. The ADP bound to each active site is drawn in stick form with C green, N blue, O red, and P orange. 
(b) View along one of the protein’s twofold axes (rotated 90° about the horizontal axis with respect to Part a) showing only the eight subunits nearest the viewer. The sixfold axis is vertical in this view.

E. coli glutamine synthetase is covalently modified by adenylylation (addition of an AMP group) of a specific Tyr residue. The enzyme’s susceptibility to cumulative feedback inhibition increases, and its activity therefore decreases, with its degree of adenylylation. The level of adenylylation is controlled by a complex metabolic cascade that is conceptually similar to that controlling glycogen phosphorylase . Both adenylylation and deadenylylation of glutamine synthetase are catalyzed by adenylyltransferase in complex with a tetrameric regulatory protein, PII. This complex deadenylylates glutamine synthetase when PII is uridylylated (also at a Tyr residue) and adenylylates glutamine synthetase when PII lacks UMP residues. The level of PII uridylylation, in turn, depends on the relative levels of two enzymatic activities located on the same protein: a uridylyltransferase that uridylylates PII and a uridylyl-removing enzyme that hydrolytically excises the attached UMP groups Section 5 Amino Acid Biosynthesis of PII. The uridylyltransferase is activated by α-ketoglutarate and ATP and inhibited by glutamine and Pi, whereas uridylyl-removing enzyme is insensitive to those metabolites. This intricate metabolic cascade therefore renders the activity of E. coli glutamine synthetase extremely responsive to the cell’s nitrogen requirements.

Glutamine synthetase (GS) is essential for ammonium assimilation and the biosynthesis of glutamine. 14 All organisms contain the enzymes glutamate dehydrogenase and glutamine synthetase, which convert ammonia to glutamate and glutamine, respectively. 15 The Last Universal Common Ancestor (LUCA) accessed nitrogen via nitrogenase and via glutamine synthetase. 16

What a monumental admission.  Glutamine synthetase (GS) had to be fully operational and emerged PRIOR life and self replication began. We will see short after what that means. 

Amino and amide groups from these two compounds can then be transferred to other carbon backbones by transamination and transamidation reactions to make amino acids. Interestingly, glutamine is the universal donor of amine groups for the formation of many other amino acids as well as many biosynthetic products. Glutamine is also a key metabolite for ammonia storage. All amino acids, with the exception of proline, have a primary amino group (NH2) and a carboxylic acid (COOH) group. They are distinguished from one another primarily by , appendages to the central carbon atom.

Glutamine is a major Nitrogen donor in the biosynthesis of many organic Nitrogen compounds such as purines, pyrimidines, and other amino acids, and GS activity is tightly regulated. We require a constant supply of nitrogen to build the bases in nucleic acids and the amino acids in proteins. The amide-N of glutamine provides the nitrogen atom in these biosyntheses. Glutamine is the most abundant amino acid in humans.  The nitrogen gas in the air and the nitrogen in nitrates and nitrites, although abundant, are not reactive enough for this use. Ammonia is the preferred source of nitrogen for these reactions. Unfortunately, ammonia is very toxic and cannot be stored or transported safely. Instead, ammonia is attached to the amino acid glutamate, forming glutamine. Because it is a natural amino acid, normally used to build proteins, glutamine is easily transported throughout the body in large amounts. Ammonia may then be liberated only when needed. Glutamine synthetase connects a molecule of ammonia to the amino acid glutamate. A molecule of ATP (adenosine triphosphate) is used to power the process, to ensure that the reaction is performed only in the proper direction and not in reverse, carelessly liberating poisonous ammonia. The bacterial enzyme  is a highly regulated allosteric enzyme ( allosteric regulation or control is the regulation of an enzyme by binding an effector molecule at a site other than the enzyme's active site. ) 

Covalent Regulation of Glutamine Synthetase
" Glutamine synthetase is one of the most heavily regulated enzymes because it reacts with ammonia and pneumonias toxic we need to regulate ammonia levels very very tightly and there's a lot of ways we do that "
https://www.youtube.com/watch?v=nhmj6jnjlOQ

Glutamate, Glutamine Biosynthesis
Glutamate and glutamine are both made from the TCA cycle
https://www.youtube.com/watch?v=kygtV68ff4I

So in order to have the substrate which Glutamine synthetase (GS) processes,

The Citric acid cycle, or Krebs (TCA) cycle
http://reasonandscience.catsboard.com/t1464-the-citric-acid-cycle-or-krebs-tca-cycle

A molecular Computer
Glutamine synthetase has been likened or compared to a molecular computer. With its 12 interacting subunits, arranged in two rings of six, it senses the amounts of the amino acids and nucleotides ultimately constructed from the ammonia in glutamine. Glutamine synthetase weighs the concentrations of each, computes whether there is an overall deficit or excess, and turns on or off based on the result.  12

Our cells are continually faced with a changing environment. 13 Think about what you eat. Some days you might eat a lot of protein, other days you might eat a lot of carbohydrate. Sometimes you may eat nothing but chocolate. Your body must be able to respond to these different foods, producing the proper enzymes for capturing the nutrients in each. The same is doubly true for small organisms like bacteria, which do not have as many options in choosing their diet. They must eat whatever food happens to be close by, and then mobilize the enzymes needed to use it.

The enzyme glutamine synthetase is a key enzyme controlling the use of nitrogen inside cells. Glutamine, as well as being used to build proteins, delivers nitrogen atoms to enzymes that build nitrogen-rich molecules, such as DNA bases and amino acids. So, glutamine synthetase, the enzyme that builds glutamine, must be carefully controlled. When nitrogen is needed, it must be turned on so that the cell does not starve. But when the cell has enough nitrogen, it needs to be turned off to avoid a glut.

Glutamine synthetase acts like a tiny molecular computer, monitoring the amounts of nitrogen-rich molecules. It watches levels of amino acids like glycine, alanine, histidine and tryptophan, and levels of nucleotides like AMP and CTP. If too much of one of these molecules is made, glutamine synthetase senses this and slows production slightly. But as levels of all of these nucleotides and amino acids rise, together they slow glutamine synthetase more and more. Eventually, the enzyme grinds to a halt when the supply meets the demand.

Communication Between Many Active Sites
The glutamine synthetase molecule  is composed of twelve identical subunits, each of which has an active site for the production of glutamine. When performing its reaction, the active site binds to glutamate and ammonia, and also to an ATP molecule that powers the reaction. But, the active sites also bind weakly to other amino acids and nucleotides, partially blocking the action of the enzyme. All of the many sites communicate with one another, and as the concentrations of competing molecules rise, more and more of the sites are blocked, eventually shutting down the whole enzyme. The cell has a more direct approach when it wants to shut down the enzyme. At a key tyrosine next to the active site, colored yellow here and shown by the arrow, an ADP molecule can be attached to the protein, completely blocking its action.

We make several versions of glutamine synthetase in our own cells. Most of our cells make a version similar to the bacterial one, but with eight subunits instead of twelve. Like the bacterial enzyme, it is controlled by the nitrogen-rich compounds down the synthetic pipeline. We also make a second glutamine synthetase in our brain. There, glutamate is used as a neurotransmitter, and glutamine synthetase is used when the glutamate is recycled after a nerve impulse is delivered. In the brain, glutamine synthetase is in constant action, so a highly-regulated version is not appropriate. Instead, the alternate form is active all the time, continually performing its essential duty.

Two Doors
Each of the twelve active sites of glutamine synthetase has two metal ions, either magnesium or manganese, bound at the center of a tunnel. The substrates enter from two sides of the tunnel: ATP enters on the exposed faces on the top and bottom of the enzyme (ATP is easily seen in the upper picture on the previous page) and glutamate and ammonia squeeze through an opening between the upper ring of subunits and the lower ring. This structure contains a ADP molecule bound in the ATP site, two manganese ions (which bind tighter than magnesium, but make the enzyme slightly slower), and an inhibitor that is about the same size and shape as glutamine.




Nitrogen is found everywhere on Earth, forming about three-fourths of the air. Nitrogen gas, however, is chemically inert and of little use to us. Our primary source of nitrogen is the ammonia in amino acids and nucleotides, obtained by eating other living things. But small amounts of ammonia are lost from the biosphere over time, locked up in minerals and buried out of reach. To replenish the global supply of biological nitrogen, nitrogen gas is converted into ammonia in the process of nitrogen fixation. Today, this is accomplished in three ways: about 15% is formed geologically, by lightning and ultraviolet radiation; 25% is produced industrially and distributed as fertilizer; and the remaining 60% is produced by a small class of bacteria and algae. These "diazotrophic" microorganisms fix nitrogen using nitrogenases, enzymes that rip apart the two tightly bound atoms in nitrogen gas and add hydrogen atoms to them, forming ammonia. Nitrogenases contain dozens of reactive iron atoms, as well as rarer metals such as molybdenum and vanadium. These unusual metal ions are required to apply the chemical tension that wrenches apart the stable nitrogen molecule. However, they are extremely sensitive to oxygen.

Leguminous plants, like peas and beans, have worked out a solution to this problem. In a classic example of symbiotic cooperation, legume roots build a nodule custom-made for bacteria, filled with leghemoglobin, a protein similar to the hemoglobin that carries oxygen in our blood. Leghemoglobin soaks up any oxygen that ventures near. In return for this safe haven, the bacteria release some of their fixed nitrogen for use by the plant. This abundant supply of ammonia carries a heavy price, however. Nitrogen fixation is very expensive, requiring about 16 ATP molecules per nitrogen molecule split into ammonia.


Glutamine synthetase (GS) is found in all organisms. In addition to its importance for NH4+ assimilation in bacteria, it has a central role in amino acid metabolism in mammals, converting free NH4+ which is toxic, to glutamine for transport in the blood.     It is a huge enzyme, which has in E. coli 5628 amino acids, which catalyzes a reaction that introduces reduced nitrogen into cellular metabolism, and is among the most complex regulatory enzymes known.  In bacteria and plants, glutamate is produced from glutamine in a reaction catalyzed by glutamate synthase. alpha Ketoglutarate, an intermediate of the citric acid cycle, undergoes reductive amination with glutamine as nitrogen donor:   It is regulated allosterically (with at least eight different modulators); by reversible covalent modification; and by the association of other regulatory proteins. It catalyzes glutamine synthesis from glutamate and ammonia at the expenditure of ATP. 9  by ATP-dependent amidation of the g-carboxyl group of glutamate to form glutamine  The reaction proceeds via a g-glutamyl-phosphate intermediate. Glutamine synthetase (GS) activity depends on the presence of divalent cations such as Mg2+. 


Glutamine synthetase (GS) catalyzes the ATP-dependent amidation of the g-carboxyl group of glutamate to form glutamine




An example of an energetically unfavorable biosynthetic reaction driven by ATP hydrolysis. b
(A) Schematic illustration of the formation of A–B in the condensation reaction described in the text. 
(B) The biosynthesis of the common amino acid glutamine from glutamic acid and ammonia. Glutamic acid is first converted to a high-energy phosphorylated intermediate (corresponding to the compound B–O–PO3 described in the text), which then reacts with ammonia (corresponding to A–H) to form glutamine. In this example, both steps occur on the surface of the same enzyme, glutamine synthetase. The high-energy bonds are shaded red; here, and the symbol Pi = HPO42–, and a yellow “circled P” = PO32–. 11

What Regulatory Mechanisms Act on coli Glutamine Synthetase?
Glutamine plays a pivotal role in nitrogen metabolism by donating its amide nitrogen to the biosynthesis of many important organic N compounds. Consistent with its metabolic importance, in prokaryotic cells such as E. coli, GS is
regulated at three different levels:

1. Its activity is regulated allosterically by feedback inhibition.
2. GS is interconverted between active and inactive forms by covalent modification.
3. Cellular amounts of GS are carefully controlled at the level of gene expression and protein synthesis.

The activity of glutamine synthetase is regulated in virtually all organisms—not surprising, given its central metabolic role as an entry point for reduced nitrogen. In enteric bacteria such as E. coli, the regulation is unusually
complex. The enzyme has 12 identical subunits  and is regulated both allosterically and by covalent modification. Alanine, glycine, and at least six end products of glutamine metabolism are allosteric inhibitors of the enzyme. Each
inhibitor alone produces only partial inhibition, but the effects of multiple inhibitors are more than additive, and all eight together virtually shut down the enzyme. This control mechanism provides a constant adjustment of glutamine levels to match immediate metabolic requirements.

Glutamine Synthetase Is Allosterically Regulated
Nine distinct feedback inhibitors (Gly, Ala, Ser, His, Trp, CTP, AMP, carbamoyl-P, and glucosamine-6-P) act on GS. Gly, Ala, and Ser are key indicators of amino acid metabolism in the cell; each of the other six compounds represents an end product of a biosynthetic pathway dependent on Gln ( see figure below )


The allosteric regulation of glutamine synthetase activity by feedback inhibition.

Evolution of the glutamine synthetase gene, one of the oldest existing and functioning genes 2
December 14, 1992
We performed molecular phylogenetic analyses of glutamine synthetase (GS) genes in order to investigate their evolutionary history. We suggest that GS genes are one of the oldest existing and functioning genes in the history of gene evolution and that GSI genes should also exist in eukaryotes.

The third, carbamoyl-phosphate synthetase I, is a mitochondrial enzyme that participates in the urea cycle. This reaction provides an important interface between nitrogen metabolism and cellular pathways of carbon and energy metabolism because a-ketoglutarate is a citric acid cycle intermediate. 

The amino acid and nucleotide biosynthetic pathways make repeated use of the biological cofactors

- pyridoxal phosphate
- tetrahydrofolate
- S-adenosylmethionine


Pyridoxal phosphate is required for transamination reactions involving glutamate and for other amino acid transformations. One-carbon transfers require S-adenosylmethionine and tetrahydrofolate. Glutamine amidotransferases catalyze reactions that incorporate nitrogen derived from glutamine.

a Guanosine-5'-triphosphate (GTP) is a purine nucleoside triphosphate. It is one of the building blocks needed for the synthesis of RNA during the transcription process. Its structure is similar to that of the guanine nucleobase, the only difference being that nucleotides like GTP have a ribose sugar and three phosphates, with the nucleobase attached to the 1' and the triphosphate moiety attached to the 5' carbons of the ribose.

It also has the role of a source of energy or an activator of substrates in metabolic reactions, like that of ATP, but more specific. It is used as a source of energy for protein synthesis and gluconeogenesis. 3



Hydrolysis  usually means the cleavage of chemical bonds by the addition of water. When a carbohydrate is broken into its component sugar molecules by hydrolysis 10

2. http://www.pnas.org/content/pnas/90/7/3009.full.pdf
3. https://en.wikipedia.org/wiki/Guanosine_triphosphate
4. https://en.wikipedia.org/wiki/Glutamate_synthase_(NADPH)
5. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4703346/
6. https://en.wikipedia.org/wiki/Alpha-Ketoglutaric_acid
7. http://www.biologydiscussion.com/organism/metabolism-organism/incorporation-of-ammonia-into-organic-compounds/50870
8. https://www.sciencedirect.com/science/article/pii/S000527360000136X
9. https://www.frontiersin.org/articles/10.3389/fmicb.2016.00969/full
10. https://en.wikipedia.org/wiki/Hydrolysis
11. Molecular biology of the cell, Alberts, 6th ed. page 66
12. Goodsell, Our molecular nature, page 31
13. http://pdb101.rcsb.org/motm/30
14. https://bmcevolbiol.biomedcentral.com/articles/10.1186/1471-2148-10-198
15. https://www.nature.com/scitable/topicpage/an-evolutionary-perspective-on-amino-acids-14568445
16. https://www.nature.com/articles/nmicrobiol2016116?dom=pscau&src=syn
17. Lehninger, principles of biochemistry, page 860
18. Fundamentals of biochemistry, 6th ed. page 749
19. https://en.wikipedia.org/wiki/Glutamine_synthetase
20. http://sci-hub.tw/https://www.cell.com/trends/biochemical-sciences/abstract/S0968-0004(08)00189-8



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Glutamine synthetase (GS), a incredible molecular super-computer which defies naturalistic explanations

http://reasonandscience.catsboard.com/t2590p25-origins-what-cause-explains-best-our-existence-and-why#5935

I have written extensively about the importance of nitrogen to sustain and make life possible on earth, how it is fixed by some of the most complex enzymes known, nitrogenase, which transforms nitrogen gas into ammonia. Nitrogen gas forms about 78 percent of the air. It  is chemically inert by its triple bond and requires enormous amounts of energy to be split. For that reason, nitrogenase is called a molecular sledgehammer. Only the force of lightning is able to split dinitrogen, which illustrates the forces required. Our primary source of nitrogen is ammonia in amino acids, buildingblocks to make proteins,  and nucleotides, that is RNA, and DNA, the information storage devices inside our cells.  60% of ammonia is produced by a small class of bacteria, that is, cyanobacterias, and algae. These "diazotrophic" microorganisms fix nitrogen using nitrogenases, enzymes that rip apart the two tightly bound atoms in nitrogen gas and add hydrogen atoms to them, forming ammonia. Nitrogenases contain dozens of reactive iron atoms, as well as rarer metals such as molybdenum. These unusual metal ions are required to apply the chemical tension that wrenches apart the stable nitrogen molecule. However, they are extremely sensitive to oxygen. I wrote previously about the hyper complex biosynthesis processes, which requires complex import mechanisms of Iron, Sulfur, and Molybden into the cytoplasm of the cell, and the enormously complex multistep synthesis process to make the active centers of nitrogenase, Iron - sulfur, and Iron-sulfur-Molybden clusters. 

Overview of the Nitrogenase enzyme complex
http://reasonandscience.catsboard.com/t2590-origins-what-cause-explains-best-our-existence-and-why#5867

Biosynthesis of the Cofactors of Nitrogenase  
http://reasonandscience.catsboard.com/t2429-biosynthesis-of-the-cofactors-of-nitrogenase

Molybdenum, essential for life
http://reasonandscience.catsboard.com/t2430-molybdenum-essential-for-life

Iron Uptake and Homeostasis in Cells
http://reasonandscience.catsboard.com/t2443-iron-uptake-and-homeostasis-in-prokaryotic-microorganisms

Once ammonium is made, it has to be introduced into the process for further intracellular biosynthesis. And here Glutamine synthetase (GS), come into play. They are essential for ammonium assimilation and the biosynthesis of glutamine. 14  All organisms contain the enzymes glutamate dehydrogenase and glutamine synthetase, which convert ammonia to glutamate and glutamine, respectively. 15 The Last Universal Common Ancestor (LUCA) accessed nitrogen via nitrogenase and via glutamine synthetase

My comment: This is a monumental admission.  Glutamine synthetase (GS) had to be fully operational and had to emerge PRIOR life and cellular self replication began. This is amazing - we will see in short what that means, once we understand what kind of enzyme that is, and what it is capable of.  

Glutamine is a major Nitrogen donor in the biosynthesis of many organic Nitrogen compounds such as purines, pyrimidines, and other amino acids. We require a constant supply of nitrogen to build the bases in nucleic acids and the amino acids in proteins. Ammonia is very toxic and cannot be stored or transported safely. Instead, ammonia is attached to the amino acid glutamate, forming glutamine. Because it is a natural amino acid, normally used to build proteins, glutamine is easily transported throughout the body in large amounts. Ammonia may then be liberated only when needed. Glutamine synthetase connects a molecule of ammonia to the amino acid glutamate. A molecule of ATP (adenosine triphosphate) is used to power the process, to ensure that the reaction is performed only in the proper direction and not in reverse, carelessly liberating poisonous ammonia. The bacterial enzyme  is a highly regulated  enzyme

Covalent Regulation of Glutamine Synthetase
" Glutamine synthetase is one of the most heavily regulated enzymes because it reacts with ammonia and we need to regulate ammonia levels very very tightly and there's a lot of ways we do that "
https://www.youtube.com/watch?v=nhmj6jnjlOQ

Glutamate, Glutamine Biosynthesis
Glutamate and glutamine are both made from the TCA cycle
https://www.youtube.com/watch?v=kygtV68ff4I

So in order to have the substrate which Glutamine synthetase (GS) processes, we need the product of the TCA Cycle. The origin of the TCA cycle is a unsolved origin of life problem, since a multitude of various enzymes are required to work together to produce ATP, and amino acids. 

The Citric acid cycle, or Krebs (TCA) cycle
http://reasonandscience.catsboard.com/t1464-the-citric-acid-cycle-or-krebs-tca-cycle

A molecular Computer
Glutamine synthetase has been likened or compared to a molecular computer. With its 12 interacting subunits, arranged in two rings of six, it senses the amounts of the amino acids and nucleotides ultimately constructed from the ammonia in glutamine. Glutamine synthetase weighs the concentrations of each, computes whether there is an overall deficit or excess, and turns on or off based on the result.  

Our cells are continually faced with a changing environment. Think about what you eat. Some days you might eat a lot of protein, other days you might eat a lot of carbohydrate. Sometimes you may eat nothing but chocolate. Your body must be able to respond to these different foods, producing the proper enzymes for capturing the nutrients in each. The same is doubly true for small organisms like bacteria, which do not have as many options in choosing their diet. They must eat whatever food happens to be close by, and then mobilize the enzymes needed to use it.

The enzyme glutamine synthetase is a key enzyme controlling the use of nitrogen inside cells. Glutamine, as well as being used to build proteins, delivers nitrogen atoms to enzymes that build nitrogen-rich molecules, such as DNA bases and amino acids. So, glutamine synthetase, the enzyme that builds glutamine, must be carefully controlled. When nitrogen is needed, it must be turned on so that the cell does not starve. But when the cell has enough nitrogen, it needs to be turned off to avoid a glut.

Glutamine synthetase acts like a tiny molecular computer, monitoring the amounts of nitrogen-rich molecules. It watches levels of amino acids like glycine, alanine, histidine and tryptophan, and levels of nucleotides like AMP and CTP. If too much of one of these molecules is made, glutamine synthetase senses this and slows production slightly. But as levels of all of these nucleotides and amino acids rise, together they slow glutamine synthetase more and more. Eventually, the enzyme grinds to a halt when the supply meets the demand.

Communication Between Many Active Sites
The glutamine synthetase molecule  is composed of twelve identical subunits, each of which has an active site for the production of glutamine. When performing its reaction, the active site binds to glutamate and ammonia, and also to an ATP molecule that powers the reaction. But, the active sites also bind weakly to other amino acids and nucleotides, partially blocking the action of the enzyme. All of the many sites communicate with one another, and as the concentrations of competing molecules rise, more and more of the sites are blocked, eventually shutting down the whole enzyme. The cell has a more direct approach when it wants to shut down the enzyme. At a key tyrosine next to the active site, colored yellow here and shown by the arrow, an ADP molecule can be attached to the protein, completely blocking its action.

We make several versions of glutamine synthetase in our own cells. Most of our cells make a version similar to the bacterial one, but with eight subunits instead of twelve. Like the bacterial enzyme, it is controlled by the nitrogen-rich compounds down the synthetic pipeline. We also make a second glutamine synthetase in our brain. There, glutamate is used as a neurotransmitter, and glutamine synthetase is used when the glutamate is recycled after a nerve impulse is delivered. In the brain, glutamine synthetase is in constant action, so a highly-regulated version is not appropriate. Instead, the alternate form is active all the time, continually performing its essential duty.

Glutamine synthetase is a life essential, ultracomplex molecular computer, which had to emerge fully operational. It is also one of the huge in molecular size,  it requires over 5500 amino acids, and in case of prokaryotes, twelve subunits, used in several regulation functions. The TCA cycle has no function without Glutamine synthetase, and vice versa. I would say, unless we infer a intelligent creator, its origin is a bit misterious.... 

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29 How Do Organisms Synthesize Amino Acids? on Mon Apr 09, 2018 12:33 pm

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How Do Organisms Synthesize Amino Acids?

In 1943, Gordon, Martin, and Synge used partition chromatography to separate and study constituents of proteins (Gordon, Martin, & Synge 1943), a major breakthrough that contributed to the rapid identification of the twenty amino acids used in proteins by all living organisms. After this initial burst of discovery, two additional amino acids, which are not used by all organisms, were added to the list: selenocysteine (Bock 2000) and pyrrolysine (Srinivasan et al. 2002). 2 Aside from their role in composing proteins, amino acids have many biologically important functions. They are also energy metabolites, and many of them are essential nutrients. Amino acids can often function as chemical messengers in communication between cells. For example, Arvid Carlsson discovered in 1957 that the amine 3-hydroxytyramine (dopamine) was not only a precursor for the synthesis of adrenaline from tyrosine, but is also a key neurotransmitter. Certain amino acids — such as citrulline and ornithine, which are intermediates in urea biosynthesis — are important intermediaries in various pathways involving nitrogenous metabolism. Although other amino acids are important in several pathways, S-adenosylmethionine acts as a universal methylating agent.

The pathways for the biosynthesis of these molecules are extremely ancient, going back to the last common ancestor of all living things. 4 Many of the intermediates in energy-yielding pathways play a role in biosynthesis as well. These common intermediates allow efficient interplay between energy-yielding (catabolic) and energy-requiring biosynthetic (anabolic) pathways. Thus, cells are able to balance the degradation of compounds for energy mobilization and the synthesis of starting materials for macromolecular construction.

Metabolic processes inside the cell require a delicate and finely orchestrated balance between anabolic, and catabolic processes. The right dosage or production of the various basic building blocks is essential and must be adjusted and calibrated to the cells and organisms needs. Amino acids, the basic building blocks of proteins, if there are too many in the cell, they must be degraded. Proteins which have malfunctioned must be degraded, and their turnover is a regulated process requiring complex enzyme systems and ATP energy supply. Proteasome enzymes - protein grinders,   had to emerge prior life began, to do this job.

Proteasome Garbage Grinders, evidence of luck, evolution, or design?
http://reasonandscience.catsboard.com/t1851-proteasome-garbage-grinders

The proteasome also requires ATP energy to function.

Question: Have you ever seen a garbage recycle factory emerge randomly, by a lucky accident?

Each of the 20 amino acids used in life requires specific complex degradation enzymes and coenzymes, which are all there, keen to do their job. Did prebiotic molecules lying around on early earth suddenly see the need to join together to form these enzymes, foreseeing that they would be required for amino acid degradation, one day?

Not only that. Once amino acids are degraded, there is a complex factory, keenly waiting for them for reuse: The machinery is all there to do the job: The carbon atoms of degraded amino acids are converted into pyruvate, acetyl CoA, acetoacetate, or an intermediate of the citric acid cycle.

Imagine: If these processes were not fully set up, it would result in accumulation for example of phenylalanine, and subsequently, if there were advanced animals with brains, the result would be mental retardation. I would not be here, writing these lines, and you would be unable to read, and eventually understand my write up.

But luckily, you can, and also recognize, that these ultrasophisticated processes could not be the result of a lucky accident. Thank God, these processes exist, we have discovered them, and so the hidden message between the lines. God is telling us: Hey, i made all this. Do you not want to know who i am ??

Amino Acid Precursors and Biosynthesis Pathways
In the study of metabolism, a series of biochemical reactions for compound synthesis or degradation is called a pathway. Amino acid synthesis can occur in a variety of ways. For example, amino acids can be synthesized from precursor molecules by simple steps. Alanine, aspartate, and glutamate are synthesized from keto acids called pyruvate, oxaloacetate, and alpha-ketoglutarate, respectively, after a transamination reaction step. a Similarly, asparagine and glutamine are synthesized from aspartate and glutamate, respectively, by an amidation reaction step. The synthesis of other amino acids requires more steps; between one and thirteen biochemical reactions necessary to produce the different amino acids from their precursors of the central metabolism (Figure below).


Amino acid metabolism in context
Numerous metabolism pathways are depicted: 


central metabolism (in black), 
pentose phosphate metabolism (in brown), 
nitrogen metabolism (in magenta), 
and various amino acid metabolism pathways (all other colors).


Nodes (dots) represent metabolites, and lines represent enzymes and intermediaries. The nitrogen metabolism pathway overlaps with the biosynthesis of arginine and proline, with glutamate as the shared precursor. Histidine biosynthesis branches off the pentose phosphate metabolism. Lysine (AAA) biosynthesis can be synthesized through different pathways, the aminoadipate (AAA) pathway or the diaminopimelate (DAP) pathway (shown in dark blue). There are gene homologies between different biosynthetic pathways. In the dark blue pathways, shaded rectangles represent homologies between enzymes. Similarly, the AAA pathway contains enzymes that share homologies with the branched chain amino acid (BCAA) pathways, whereas the DAP pathway contains homologies with the arginine biosynthetic pathway. In various pathways, homologous enzymes are denoted by shaded rectangles. Different shaded colors indicate different pairs of homologous enzymes.

Numerous metabolism pathways and many other connecting pathways, including amino acid metabolism pathways, are depicted as interconnected lines and nodes. The nodes are shown as colored dots that represent different metabolites. The colored lines represent different enzymes and intermediaries in the pathways. The different colors are used to differentiate the pathways, and the pathways branch off of one another at nodes. The central metabolism pathway, which includes glycolysis and the citric acid cycle, is shown on the left in black. The pentose phosphate pathway, which is shown in brown, branches off of the central metabolism pathway during glycolysis. The aminoadipate pathway, which is shown in dark blue, is labeled "AAA" and the diaminopimelate pathway, which is shown in dark purple, is labeled "DAP." Both the AAA and DAP pathways branch off of the central metabolism pathway at later steps during glycolysis or during the citric acid cycle. The nitrogen metabolism pathway is shown in magenta at the far left. Some of the nodes are labeled to indicate key metabolites.

What makes an amino acid essential?
Not all the organisms are capable of synthesizing all the amino acids, and many are synthesized by pathways that are present only in certain plants and bacteria. Mammals, for example, must obtain eight of twenty amino acids from their diets. This requirement leads to a convention that divides amino acids into two categories: essential and nonessential (given a certain metabolism). Because of particular structural features, essential amino acids cannot be synthesized by mammalian enzymes. Nonessential amino acids, therefore, can be synthesized by nearly all organisms.

Nature magazine goes on and claims:
The loss of the ability to synthesize essential amino acids likely emerged very early in evolution, because this dependence on other organisms for the source of amino acids is common among all eukaryotes, not just those of mammals.

This matter of fact goes against a naturalistic explanation. There would be no survival advantage ( rather the opposite is the case), if higher animals in taxonomy would stop producing amino acids, and depend on food ingestion to obtain them. How could a slow gradual transition of this state of affair occur? It seems to make more sense, that the interdependence was set up right from the beginning.  

How do certain amino acids become essential for a given organism? Studies in ecology and evolution give some clues. Organisms evolve under environmental constraints, which are dynamic over time. If an amino acid is available for uptake, the selective pressure to keep intact the genes responsible for that pathway might be lowered, because they would not be constantly expressing these biosynthetic genes. Without the selective pressure, the biosynthetic routes might be lost or the gene could allow mutations that would lead to a diversification of the enzyme's function. Following this logic, amino acids that are essential for certain organisms might not be essential for other organisms subjected to different selection pressures. 

Might be, could, might not be..... is there something beside baseless speculation here ??!!

The relative uses of amino acid biosynthetic pathways vary widely among species because different synthesis pathways fulfill unique metabolic needs in different organisms. Although some pathways are present in certain organisms, they are absent in others. Therefore, experimental results about amino acid metabolism that are achieved with model organisms may not always have relevance for the majority of other organisms.

Amino acids are made from intermediates of the Citric Acid Cycle and other major pathways
The pathways for the biosynthesis of amino acids are diverse. However, they have an important common feature: their carbon skeletons come from intermediates of glycolysis, the pentose phosphate pathway, or the citric acid cycle. On the basis of these starting materials, amino acids can be grouped into six biosynthetic families 1 ( see below )


Major metabolic precursors are shaded blue. Amino acids that give rise to other amino acids are shaded yellow. Essential amino acids are in boldface type.

Organisms show substantial differences in their capacity to synthesize the 20 amino acids common to proteins. Typically, plants and microorganisms can form all of their nitrogenous metabolites, including all of the amino acids, from inorganic forms of N such as ammonium NH4+ and nitrate NO3+. In these organisms, the a-amino group for all amino acids is derived from glutamate, usually via transamination of the corresponding a-keto acid analog of the amino acid. In many cases, amino acid biosynthesis is thus a matter of synthesizing the appropriate a-keto acid carbon skeleton, followed by transamination with Glu. The amino acids can be classified according to the
source of intermediates for the a-keto acid biosynthesis.

Several classes of reactions play special roles in the biosynthesis of amino acids and nucleotides
(1) transamination reactions and other rearrangements promoted by enzymes containing pyridoxal phosphate (PLP) ;
(2) transfer of one-carbon groups, with either tetrahydrofolate (usually at the —CHO and —CH2OH oxidation levels) or S-adenosylmethionine (at the —CH3 oxidation level) as cofactor; and
(3) transfer of amino groups derived from the amide nitrogen of glutamine.

Pyridoxal phosphate (PLP) - Vitamin B6
Pyridoxal phosphate (PLP)-dependent enzymes are unrivaled in the diversity of reactions that they catalyze. 7  Vitamin B6 was first identified as pyridoxine, a catalytically inactive form, in 1938 while the catalytically active aldehyde (pyridoxal) and amine (pyridoxamine) forms and their phosphorylated derivatives (pyridoxal 5’-phosphate and pyridoxamine 5’-phosphate) were discovered in the early 1940’s. They all act as vitamin B6, although pyridoxal 5’-phosphate is the enzymatically active form. 9

PLP is considered as the only active form of vitamin B6. In addition to its role in several enzymatic processes, including amino acid and fatty acid metabolism 39

Following its identification as one of the active vitamers of vitamin B6, pyridoxal 5'-phosphate (PLP) has been the subject of extensive research directed toward understanding its unequaled catalytic versatility. These enzymes are principally involved in the biosynthesis of amino acids and amino acid-derived metabolites, but they are also found in the biosynthetic pathways of amino sugars.

All aminotransferases have the same prosthetic group (co-factor) and the same reaction mechanism. The prosthetic group is pyridoxal phosphate (PLP) b, the coenzyme form of pyridoxine, or vitamin B6.  

The reaction involves three sequential steps:

(i) formation of a tetrahedral intermediate with the active site lysine and the amino substrate bonded to the PLP co-factor;
(ii) non-direct proton transfer between the amino substrate and the lysine residue; and
(iii) formation of the external aldimine after the dissociation of the lysine residue. The overall reaction is exothermic (−12.0 kcal/mol), the second step being rate-limiting, with 12.6 kcal/mol for the activation energy

The cofactor in all cases functions to stabilize negative charge development at C - alpha in the transition state that is formed after condensation of the amino acid substrate with PLP to form a Schiff base (referred to as the external aldimine c )

The majority of known structures are of Fold Type I (aspartate aminotransferase family) enzymes, a group that includes many of the best-characterized PLP enzymes. They invariably function as homodimers or higher-order oligomers, with two active sites per dimer. The active sites lie on the dimer interface, and each monomer contributes essential residues to both active sites. 

The breadth of reaction specificity enabled by PLP is illustrated to the right, using serine as an example substrate. The first and common step for all PLP-dependent enzyme catalyzed reactions is a Schiff base exchange reaction (transimination). All known PLP enzymes exist in their resting state as a Schiff base (internal aldimine) with an active site lysine residue. The incoming, amine-containing substrate displaces the lysine e-amino group from the internal aldimine, in the process forming a new aldimine with the substrate (external aldimine).

The external aldimine is the common central intermediate for all PLP catalyzed reactions, enzymatic and nonenzymatic. Divergence in reaction specificity occurs from this point. The great majority of pyridoxal phosphate catalyzed reactions depend on the formation of a carbanionic intermediate.

From the external aldimine intermediate, carbanions formed by heterolytic cleavage of any one of the bonds to Ca (except for the C-N bond) can be stabilized. Loss of CO2 gives a carbanion that is commonly reprotonated on Ca to give the corresponding amine as the product. Less commonly, for example with dialkylglycine decarboxylase, the resulting carbanion is reprotonated on C4’ of the coenzyme to give oxidized substrate and the reduced, amino form (PMP) of the coenzyme. 9



Proton abstraction is the most common forward step that external aldimines undergo since racemization, transamination, and beta-elimination, three common reaction types, all require it. Retro-aldol cleavage of serine, central to one-carbon metabolism, is initiated by abstraction of a proton from the beta-hydroxyl group followed by Ca-Cb cleavage. Other known reaction types include beta-decarboxylation of aspartate, beta-elimination and replacement, gamma-elimination and replacement, a/g-elimination, cyclopropyl ring opening, radical-based 1,2-amino migrations, and others. This extraordinarily wide range of reaction types makes PLP enzymes extraordinarily useful to cells. The enzyme commission has more than 140 EC numbers assigned to PLP enzymes, and free living prokaryotes devote ~1.5% of their open reading frames to them.

The commonly accepted mechanism for stabilization of the resulting carbanion is resonance delocalization within the extended conjugated pi system. This is illustrated to the right where the three most significant resonance forms are shown. The rightmost resonance structure is referred to as the “quinonoid” since its structure resembles that of a quinone. It has strong absorption at ~500 nm and is sometimes but not always spectroscopically observable in enzyme catalyzed reactions. This quinonoid resonance structure is commonly considered the major species responsible for the catalytic power of PLP since the electrons from Ca are neutralized by the protonated pyridine nitrogen. This simple view of PLP catalyzed reactions may not be wholly accurate.



Molecular origin of Pyridoxal phosphate enzymes
The pyridoxal-5'-phosphate (PLP)-dependent or vitamin B6-dependent enzymes that catalyze manifold reactions in the metabolism of amino acids belong to no fewer than four evolutionarily independent protein families. 6

It is remarkable that the authors of this science paper do not make a distinction, recognizing that the origin of pyridoxal phosphate enzymes ( Vitamin B6) had to be fully operational at LUCA, and when life began, and could therefore not be the result of evolution.


The multiple evolutionary origin and the essential mechanistic role of PLP in these enzymes argue for the cofactor having arrived on the evolutionary scene before the emergence of the respective apoenzymes and having played a dominant role in the molecular evolution of the B6 enzyme families.

Why would have natural occurrences on a prebiotic earth have produced co-enzymes without the respective apo-enzymes to interact with. Did they emerge, without function at all, just waiting for the respective proteins to interact with, to arrive later on the scene, to then eagerly looking hot to find them, and starting the molecular interaction? 

It has now become clear that the majority of organisms capable of producing this vitamin do so via a different route, involving precursors from glycolysis and the pentose phosphate pathway 40

BIOSYNTHESIS OF VITAMIN B6
The biosynthesis of the vitamin B6 involves two branches with seven enzymatic steps. In one branch, the sequential action of the enzymes GapA, PdxB and PdxF results in the conversion of erythrose 4-phosphate into 4-phosphohydroxyL-threonine. The latter then undergoes oxidation and decarboxylation by PdxA to form 3-hydroxy-1-aminoacetone phosphate.

In the other branch, DXP (deoxyxylulose 5- phosphate) is derived from GAP (glyceraldehyde 3-phosphate) and pyruvate by the action of DXP synthase. The products of the two branches, i.e. 3-hydroxy-1-aminoacetone phosphate and DXP, are then condensed by PdxJ to form PNP (pyridoxine 5' -phosphate) , which must undergo oxidation, catalysed by PdxH, to form the cofactor vitamer PLP. 

The occurrence of two distinct and mutually exclusive pathways for the de novo biosynthesis of vitamin B6 poses an attractive challenge regarding the rationale for the evolution of two independent pathways for the same molecule.




Diverse functionality of vitamin B6 and its involvement in bodily functions. 
The inner ring shows three of the vitamin B6 vitamers where the chemical entity at the 4position can be an aldehyde, an alcohol or an amine. R1 can either be a hydrogen or a phosphate group, thereby representing the vitamers shown or their phosphorylated derivatives respectively. The second and third rings indicate biochemical and physiological functions respectively in humans.



The three pathways of vitamin B6 biosynthesis.
Pase*, the apparently unspecific phosphatases involved in dephosphorylating the phosphorylated B6 vitamers; Tase*, transaminase. 40

Tetrahydrofolate ( THF  H4 folate ) and Vitamin B12 
One-carbon Metabolism: Basic Concepts
There is a group of biochemical reactions that have a special set of enzymes and coenzymes.   They are involved in amino acid metabolism and also play roles in nucleotide metabolism.   This group of reactions is referred to as one-carbon metabolism because what they have in common is the transfer of one-carbon groups.
One-carbon metabolism exists because one-carbon groups are too volatile and need to be attached to something while being processed. 42

Essentially, there are three ways of moving groups of atoms containing a single carbon atom using the following molecules:


  1. Tetrahydrofolate (THF) as a cofactor in enzymatic reactions.
  2. S-adenosylmethionine (SAM) as a methyl (-CH3) donor.
  3. Vitamin B12 (Cobalamin) as a co-enzyme in methylation and rearrangement reactions.


TETRAHYDROFOLATE (THF)  is the most versatile one-carbon donor in biosynthetic reactions.   THF is composed of three types of groups.   THF is derived from the vitamin folic acid (folate).   Folate is made by plants and microorganisms. The enzyme dihydrofolate reductase converts dihydrofolate into tetrahydrofolate, which is the active form that carries 1-carbon groups in a variety of reactions.  


All organisms require reduced folate cofactors for the synthesis of a variety of metabolites. Most microorganisms must synthesize folate de novo because they lack the active transport system of higher vertebrate cells that allows these organisms to use dietary folates. 38

Tetrahydrofolate (H4 folate)has fundamental importance for the biosynthesis of purines, pyrimidines, and several amino acids. The folate derivative, 5,10-methylene-tetrahydrofolate is essential for the synthesis of dTMP from dUMP and it is, therefore, crucial for DNA replication and cell division. Tetrahydrofolate is an essential substrate in the biosynthesis of amino acid, glycine. Dihydrofolate reductase enzyme replenishes tetrahydrofolate from dihydrofolate for the above mentioned biosynthetic processes. 36

Folate is necessary for the production and maintenance of new cells, for DNA synthesis and RNA synthesis through methylation, and for preventing changes to DNA, and, thus, for preventing cancer. It is especially important during periods of frequent cell division and growth, such as infancy and pregnancy. Folate is needed to carry one-carbon groups for methylation reactions and nucleic acid synthesis (the most notable one being thymine, but also purine bases).  It gets this carbon atom by sequestering formaldehyde produced in other processes. Thus, folate deficiency hinders DNA synthesis and cell division, affecting hematopoietic cells and neoplasms the most because of their greater frequency of cell division. 37
 
In the form of a series of tetrahydrofolate (THF) compounds, folate derivatives are substrates in a number of single-carbon-transfer reactions, and also are involved in the synthesis of dTMP (2′-deoxythymidine-5′-phosphate) from dUMP (2′-deoxyuridine-5′-phosphate). It is a substrate for an important reaction that involves vitamin B12 and it is necessary for the synthesis of DNA, and so required for all dividing cells

Tetrahydrofolate THF can be imagined as an arm that transfers single carbons in different reduced states from one molecule to another. 

The importance of folate compounds in metabolism has been established for over 50 years. Folate derivatives participate in a myriad of biosynthetic reactions involving transfers of groups containing a single carbon atom. For example, these functional units are essential components in the metabolism of the amino acids glycine, serine, methionine, and histidine, and the biosynthesis of purines and pyrimidines. 21  Tetrahydrofolic acid is a cofactor in many reactions, especially in the synthesis (or anabolism) of amino acids and nucleic acids.  It gets this carbon atom by sequestering formaldehyde f produced in other processes. 10

Tetrahydrofolate acts as a donor or acceptor of one-carbon unit in biosynthetic and degradative processes and has an essential role in the biosynthesis of purines, thymidylate, pantothenate, RNA and amino acids, such as methionine and glycine-to-serine conversion 18  

There is a group of biochemical reactions that have a special set of enzymes and coenzymes.   They are involved in amino acid metabolism and also play roles in nucleotide metabolism.   This group of reactions is referred to as one-carbon metabolism because what they have in common is the transfer of one-carbon groups. 11 

It means moving a carbon atom from one molecule to another. THF is the most versatile one-carbon donor in biosynthetic reactions.   THF is composed of three types of groups.   THF is derived from the vitamin folic acid (folate). Folate is made by plants and microorganisms. The folate derivative, 5,10-methylene-tetrahydrofolate is essential for the synthesis of dTMP d from dUMP and it is, therefore, crucial for DNA replication and cell division. Tetrahydrofolate is an essential substrate in the biosynthesis of amino acid, glycine. 14  


The two essential precursors of folate biosynthesis are 4-aminobenzoate (a product of shikimate biosynthesis pathway) and GTP e .  

Thymidylate cycle, a part of folate biosynthesis pathway plays important role in the generation of amino acid glycine and dTMP. It is made of 11 enzymatic steps.

Some of the carbon atoms of purines are acquired from derivatives of N10 -formyltetrahydrofolate. The methyl group of thymine, a pyrimidine, comes from N5 , N10 -methylenetetrahydrofolate. This tetrahydrofolate derivative can also donate a one-carbon unit in an alternative synthesis of glycine that starts with oxygen CO2 and ammonium NH4+ , a reaction catalyzed by glycine synthase (called the glycine cleavage enzyme when it operates in the reverse direction).   25 

It is synthesized in bacteria, consists of substituted pterin (6-methylpterin)p-aminobenzoate, and glutamate moieties.


Chemical structure of tetrahydrofolate (THF), monoglutamyl form. 
The red arrowhead marks the oxidatively labile C9–N10 bond. A polyglutamyl tail can be attached via the γ-carboxyl group of the glutamate moiety. 19


Here another image of the same molecule:


The nitrogen atoms to which one-carbon groups are attached in tetrahydrofolate are shown in blue. The one-carbon group undergoing transfer, in any of three oxidation states, is bonded to N-5 or N-10 or both.

Most forms of tetrahydrofolate are interconvertible and serve as donors of one-carbon units in a variety of metabolic reactions. The primary source of one-carbon units for tetrahydrofolate is the carbon removed in the conversion of serine to glycine, producing N5,N10-methylenetetrahydrofolate.

The oxidized form, folate, is a vitamin for mammals; it is converted in two steps to tetrahydrofolate by the enzyme dihydrofolate reductase. The one-carbon group undergoing transfer, in any of three oxidation states, is bonded to N-5 or N-10 or both. The most reduced form of the cofactor carries a methyl group, a more oxidized form carries a methylene group, and the most oxidized forms carry a methenyl, formyl, or formimino group ( see figure below )  43


Conversions of one-carbon units on tetrahydrofolate.
The different molecular species are grouped according to oxidation state, with the most reduced at the top and most oxidized at the bottom. All species within a single shaded box are at the same oxidation state. The conversion of N5,N10-methylenetetrahydrofolate to N5- methyltetrahydrofolate is effectively irreversible. The enzymatic transfer of formyl groups, as in purine synthesis and in the formation of formylmethionine in bacteria, generally uses
N10-formyltetrahydrofolate rather than N5-formyltetrahydrofolate. The latter species is significantly more stable and therefore a weaker donor of formyl groups. N5-Formyltetrahydrofolate is a minor byproduct of the cyclohydrolase reaction, and can also form spontaneously. Conversion of N5-formyltetrahydrofolate to N5,N10-methenyltetrahydrofolate requires ATP, because of an otherwise unfavorable equilibrium. Note that N5-formiminotetrahydrofolate is derived from histidine in a catabolic pathway 

Most forms of tetrahydrofolate are interconvertible and serve as donors of one-carbon units in a variety of metabolic reactions. The primary source of one-carbon units for tetrahydrofolate is the carbon removed in the conversion of serine to glycine, producing N5,N10-methylenetetrahydrofolate. Although tetrahydrofolate can carry a methyl group at N-5, the transfer potential of this methyl (adoMet) is the preferred cofactor group is insufficient for most biosynthetic reactions. SAdenosylmethionine (adoMet) is the preferred cofactor for biological methyl group transfers. It is synthesized from ATP and methionine by the action of methionine adenosyl transferase,  step 1 ( figure below )  



Synthesis of methionine and S-adenosylmethionine in an activated methyl cycle. 
The steps are described in the text. In the methionine synthase reaction (step 4 ), the methyl group is transferred to cobalamin to form methylcobalamin, which in turn is the methyl donor in the formation of methionine. S-Adenosylmethionine, which has a positively charged sulfur (and is thus a sulfonium ion), is a powerful methylating agent in several biosynthetic reactions. The methyl group acceptor (step 2 ) is designated R. 

This reaction is unusual in that the nucleophilic sulfur atom of methionine attacks the 5'carbon of the ribose moiety of ATP rather than one of the phosphorus atoms. Triphosphate is released and is cleaved to Pi and PPi on the enzyme, and the PPi is cleaved by inorganic pyrophosphatase; thus three bonds, including two bonds of high-energy phosphate groups, are broken in this reaction. The only other known reaction in which triphosphate is displaced from ATP occurs in the synthesis of coenzyme B12. S-Adenosylmethionine is a potent alkylating agent by virtue of its destabilizing sulfonium ion. The methyl group is subject to attack by nucleophiles and is about 1,000 times more reactive than the methyl group of N5-methyltetrahydrofolate. Transfer of the methyl group from S-adenosylmethionine to an acceptor yields S-adenosylhomocysteine (Figure above, step 2 ), which is subsequently broken down to homocysteine and adenosine (step 3 ). Methionine is regenerated by transfer of a methyl group to homocysteine in a reaction catalyzed by methionine synthase (step 4 ), and methionine is reconverted to S-adenosylmethionine to complete an activated methyl cycle. One form of methionine synthase common in bacteria uses N5-methyltetrahydrofolate as a methyl donor. Another form of the enzyme present in some bacteria and mammals uses N5-methyltetrahydrofolate, but the methyl group is first transferred to cobalamin, derived from coenzyme B12, to form methylcobalamin as the methyl donor in methionine formation. This reaction and the rearrangement of L-methylmalonyl-CoA to succinyl-CoA are the only known coenzyme B12–dependent reactions in mammals.

Folic acid, a B vitamin found in green plants, fresh fruits, yeast, and liver, takes its name from folium, Latin for “leaf.” Folic acid is a pterin (the 2-amino-4-oxo derivative of pteridine). Mammals cannot synthesize pterins and thus cannot make folates; they derive folates from their diet or from microorganisms in their intestines.  Folates are acceptors and donors of one-carbon units for all oxidation levels of carbon except CO2 (for which biotin is the relevant carrier). The active form is tetrahydrofolate (THF). THF is formed through two successive reductions of folate by dihydrofolate reductase. 41




The two-stage reduction of folate to THF. Both reactions are catalyzed by dihydrofolate reductase.

One-carbon units in three different oxidation states may be bound to THF at the N5 or N10 nitrogens (table below).


*Calculated by assigning valence bond electrons to the more electronegative atom and then counting the charge on the quasi ion. A carbon assigned four valence electrons would have an oxidation number of 0. The carbon in N5-methyl-THF is assigned six electrons from the three COH bonds and thus has an oxidation number of 22. †Note: All vacant bonds in the structures shown are to atoms more electronegative than C.

The one-carbon unit carried by THF can come from formate (HCOO-), the a-carbon of glycine, the b-carbon of serine, or the 3-position carbon in the imidazole ring of histidine. NADPH-dependent reactions interconvert the oxidation states of the various THF-bound one-carbon units. 

The conversion of serine to glycine is a prominent means of generating one-carbon derivatives of THF, which are so important for the biosynthesis of purines and the C-5 methyl group of thymine (a pyrimidine ), as well as the amino acid methionine. Glycine itself contributes to both purine and heme synthesis.  glycine can be synthesized by a reversal of the glycine oxidase reaction (Figure b).


Biosynthesis of glycine from serine 
(a) via serine hydroxymethyltransferase and 
(b) via glycine oxidase.

Here, glycine is formed when N5, N10-methylene-THF condenses with ammonium (NH4)+ and CO2. Via this route, the b-carbon of serine becomes part of glycine. The conversion of serine to glycine is a prominent means of generating one-carbon derivatives of THF, which are so important for the biosynthesis of purines and the C-5 methyl group of thymine.


Folate
Folate is necessary for the production and maintenance of new cells, for DNA synthesis and RNA synthesis through methylation, and for preventing changes to DNA, and, thus, for preventing cancerIt is especially important during periods of frequent cell division and growth, such as infancy and pregnancy. Folate is needed to carry one-carbon groups for methylation reactions and nucleic acid synthesis (the most notable one being thymine, but also purine bases). Thus, folate deficiency hinders DNA synthesis and cell division, affecting hematopoietic cells and neoplasms the most because of their greater frequency of cell division. RNA transcription, and subsequent protein synthesis, are less affected by folate deficiency, as the mRNA can be recycled and used again (as opposed to DNA synthesis, where a new genomic copy must be created). 34 

In the form of a series of tetrahydrofolate (THF) compounds, folate derivatives are substrates in a number of single-carbon-transfer reactions, and also are involved in the synthesis of dTMP (2′-deoxythymidine-5′-phosphate) from dUMP (2′-deoxyuridine-5′-phosphate). It is a substrate for an important reaction that involves vitamin B12 and it is necessary for the synthesis of DNA, and so required for all dividing cells 34 

Folates are essential in all living systems with the exception of methanogenic bacteria, where they are replaced by methanopterin derivatives. The various C1 moieties carried by THF-type coenzymes serve as building blocks for the biosynthesis of purines, pyrimidines, and methionine. Methenyltetrahydrofolate serves as an optical transponder in DNA photolyases that are involved in the photochemically driven repair of photodamaged DNA in numerous organisms, albeit not in mammals. 

Folate, distinct forms of which are known as folic acid, folacin, and vitamin B9, is one of the B vitamins 12 Folic acid, also known as Vitamin B9 is important to several biological functions. Folates are among the most complex pterin coenzymes. Tetrahydrofolate (THF) is the basic molecule of the folate family. It is synthesized in micro-organisms, including bacteria and lower eukaryotes, and plants, but not in animals. THF is synthesized from GTP, chorismate and glutamate 16 The folate pathway is central to any study related to DNA methylation, dTMP synthesis or purine synthesis. 17 

Folate is a designation for cofactors that consist of three moieties: 

p-aminobenzoic acid (pABA) (synthesized by chorismate pathway), 
pterin and 
glutamates and is an essential vitamin (vitamin B9) used by all cells.

Although the number of glutamates attached to PABA can vary depending on the source, in the cellular plasma, the monoglutamated and the tetra-reduced pterin ring (tetrahydrofolate) are the most predominant forms. 


a Transamination, a chemical reaction that transfers an amino group to a ketoacid to form new amino acids. This pathway is responsible for the deamination of most amino acids. This is one of the major degradation pathways which convert essential amino acids to nonessential amino acids (amino acids that can be synthesized de novo by the organism). 3
Transamination in biochemistry is accomplished by enzymes called transaminases or aminotransferases. α-ketoglutarate acts as the predominant amino-group acceptor and produces glutamate as the new amino acid.

b Pyridoxal phosphate (PLP, pyridoxal 5'-phosphate, P5P), the active form of vitamin B6, is a coenzyme in a variety of enzymatic reactions. The Enzyme commission has cataloged more than 140 PLP-dependent activities, corresponding to ~4% of all classified activities. The versatility of PLP arises from its ability to covalently bind the substrate, and then to act as an electrophilic catalyst, thereby stabilizing different types of carbanionic reaction intermediates. 5  PLP acts as a coenzyme in all transamination reactions, and in certain decarboxylation, deamination, and racemization reactions of amino acids

c In organic chemistry, an aldimine is an imine that is an analog of an aldehyde.[1] As such, aldimines have the general formula R–CH=N–R'. Aldimines are similar to ketimines, which are analogs of ketones.
An important subset of aldimines are the Schiff bases, in which the substituent on the nitrogen atom (R') is an alkyl or aryl group (i.e. not a hydrogen atom) 8

d Thymidine monophosphate (TMP), also known as thymidylic acid (conjugate base thymidylate), deoxythymidine monophosphate (dTMP), or deoxythymidylic acid (conjugate base deoxythymidylate), is a nucleotide that is used as a monomer in DNA. It is an ester of phosphoric acid with the nucleoside thymidine. dTMP consists of a phosphate group, the pentose sugar deoxyribose, and the nucleobase thymine. Unlike the other deoxyribonucleotides, thymidine monophosphate often does not contain the "deoxy" prefix in its name; nevertheless, its symbol often includes a "d" ("dTMP") 13

e Guanosine-5'-triphosphate (GTP) is a purine nucleoside triphosphate. It is one of the building blocks needed for the synthesis of RNA during the transcription process. Its structure is similar to that of the guanine nucleobase, the only difference being that nucleotides like GTP have a ribose sugar and three phosphates, with the nucleobase attached to the 1' and the triphosphate moiety attached to the 5' carbons of the ribose. 15

Formaldehyde (systematic name methanal) is a naturally occurring organic compound with the formula CH2O (H-CHO). It is the simplest of the aldehydes (R-CHO). The common name of this substance comes from its similarity and relation to formic acid. 23 Formaldehyde was the first polyatomic organic molecule detected in the interstellar medium. Since its initial detection in 1969, it has been observed in many regions of the galaxy. Because of the widespread interest in interstellar formaldehyde, it has recently been extensively studied, yielding new extragalactic sources.
" The poisonous chemical formaldehyde may have helped create the organic compounds present in the universe that gave rise to life, new research suggests. " 24

g In the chemical sciences, methylation denotes the addition of a methyl group on a substrate, or the substitution of an atom (or group) by a methyl group. Methylation is a form of alkylation, with a methyl group, rather than a larger carbon chain, replacing a hydrogen atom. These terms are commonly used in chemistry, biochemistry, soil science, and the biological sciences. 26 A methyl group is an alkyl derived from methane, containing one carbon atom bonded to three hydrogen atoms — CH3. 27 Such hydrocarbon groups occur in many organic compounds. It is a very stable group in most molecules. While the methyl group is usually part of a larger molecule, it can be found on its own in any of three forms: anion, cation or radical. The anion has eight valence electrons, the radical seven and the cation six.

 

1. Berg, Biochemistry, 5th edition, page 973
2. https://www.nature.com/scitable/topicpage/an-evolutionary-perspective-on-amino-acids-14568445#
3. https://en.wikipedia.org/wiki/Transamination
4. Biochemistry 5th edition, Styer, page 713
5. https://en.wikipedia.org/wiki/Pyridoxal_phosphate
6. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2443152/
7. https://www.annualreviews.org/doi/10.1146/annurev.biochem.73.011303.074021
8. https://en.wikipedia.org/wiki/Aldimine
9. https://sites.google.com/site/mdtoneylab/research/pyridoxal-phosphate-enzymes
10. https://en.wikipedia.org/wiki/Tetrahydrofolic_acid
11. http://www.biochem.uthscsa.edu/med/08-Amino-Acid-Metabolism/Amino-Acid-Metabolism7.html
12. https://en.wikipedia.org/wiki/Folate
13. https://en.wikipedia.org/wiki/Thymidine_monophosphate
14. http://www.llamp.net/?q=Folate%20biosynthesis
15. https://en.wikipedia.org/wiki/Guanosine_triphosphate
16. http://pubs.rsc.org/en/content/articlelanding/2007/np/b703107f#!divAbstract
17. https://www.wikipathways.org/index.php/Pathway:WP241
18. http://sci-hub.tw/https://www.future-science.com/doi/full/10.4155/fmc-2017-0168
19. http://sci-hub.tw/10.1146/annurev-arplant-042110-103819
20. https://iubmb.onlinelibrary.wiley.com/doi/pdf/10.1002/iub.1153
21. http://what-when-how.com/molecular-biology/tetrahydrofolate-molecular-biology/
22. https://en.wikipedia.org/wiki/Dihydrofolate_reductase
23. https://en.wikipedia.org/wiki/Formaldehyde
24. https://www.livescience.com/13551-formaldehyde-poison-origin-earth-life.html
25. Biochemistry, Styer , 8th ed. page 724
26. https://en.wikipedia.org/wiki/Methylation
27. https://en.wikipedia.org/wiki/Methyl_group
28. http://www.jbc.org/content/277/19/16624.full
29. http://sci-hub.tw/10.1016/j.pep.2011.05.004
30. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2643306/
31. https://en.wikipedia.org/wiki/S-adenosylmethionine_synthetase_enzyme
32. https://en.wikipedia.org/wiki/S-Adenosyl_methionine
33. Biochemistry 6th edition, Garrett, page 1063
34. https://en.wikipedia.org/wiki/Folate#DNA_and_cell_division
35. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1282579/
36. [url=http://www.llamp.net/?q=Bbov Folate recycling]http://www.llamp.net/?q=Bbov%20Folate%20recycling[/url]
37. https://en.wikipedia.org/wiki/Folate#DNA_and_cell_division
38. https://en.wikipedia.org/wiki/Dihydropteroate_synthase
39. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3650526/
40. http://sci-hub.tw/http://www.biochemj.org/content/407/1/1
41. Biochemistry 6th edition, Garrett, page 930
42. http://www.biochem.uthscsa.edu/med/08-Amino-Acid-Metabolism/Amino-Acid-Metabolism7.html
43. Biochemistry 6th edition, Garrett, page 690



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The folate biosynthesis pathway

The pathway leading to the formation of tetrahydrofolate (FH4) begins when folic acid (F) is reduced to dihydrofolate (DHF) (FH2), which is then reduced to THF. Dihydrofolate reductase catalyses the last step. Vitamin B3 in the form of NADPH is a necessary cofactor for both steps of the synthesis. Thus, hydride molecules are transferred from NADPH to the C6 position of the pteridine ring to reduce folic acid to THF 

Folate biosynthesis - Reference pathway



Tetrahydrofolate (H4 folate) MTHFR ( in red ) in the Folate cycle


The folate biosynthesis in plants and microorganisms starts with the synthesis of the pterin ring, which is catalyzed by GTP cyclohydrolase I (GTPCHI) and this reaction is followed by other five reactions catalyzed by five distinct enzymes that convert GTP into 7,8-dihydrofolate, which is reduced by dihydrofolate reductase to produce 5,6,7,8-tetrahydrofolate ( Figure B, below ) 


Folate derivatives and biosynthesis. 
(A) Chemical moieties of folates. 
(B) Overview of the folate pathway in microorganisms.

The folate biosynthesis in plants and microorganisms starts with the synthesis of the pterin ring, which is catalyzed by GTP cyclohydrolase I (GTPCHI) and this reaction is followed by other five reactions catalyzed by five distinct enzymes that convert GTP into 7,8-dihydrofolate, which is reduced by dihydrofolate reductase to produce 5,6,7,8-tetrahydrofolate 20

pABA, which is attached to the pterin moiety by dihydropteroate synthase (the forth step of the pathway), is produced by two enzymatic steps from chorismate and makes a link between the folate and the shikimate pathways. The first reaction of the folate pathway is catalyzed by GTPCHI, a homodecamer with D5-symmetry that involves intensive rearrangement, including the ring opening, an Amadori rearrangement and finally a ring closure. In this process, a molecule of GTP yields dihydroneopterin triphosphate, which is the substrate for dihydroneopterin aldolase (DHNA).DHNA is a homo-octameric enzyme that has a catalytic mechanism similar to the class I aldolases. It does not require a Schiff base, even though this feature is characteristic of this enzyme class. In addition, this enzyme is also able to catalyze the epimerization of 7,8-dihydroneopterin (DHNP) to rend 7,8-dihydromonopterin (DHMP).The next step of the pathway is catalyzed by 7,8-hydroxymethyl-7,8-dihydropterin pyrophosphokinase (HPPK), which transfers the pyrophosphate from ATP to DHMP producing 6-hydroxymethyl-7,8-dihydropterin pyrophosphate (DHPPP), that is the substrate of dihydropteroate synthase (DHPS), an enzyme that performs the condensation of this molecule with pABA to yield 7,8-dihydropteroate. Mono/dihydrofolate synthase and the bifunctional folylpoly-γ-glutamate synthetase (DHFS/FPGS) add one or more glutamates, respectively to the 7,8-dihydropteroate, producing dihydrofolate and its derivatives. The last step of the pathway is catalyzed by dihydrofolate reductase (DHFR), which together with DHPS is the most studied enzyme of the folate pathway. DHFR has a Rossmannoid fold and catalyzes the reduction of dihydrofolate to tetrahydrofolate using NADPH as a cofactor. The last two steps of the folate pathway are found in both prokaryotic and eukaryotic organisms and have been used as antimicrobial or human disease targets, respectively. Figure B above  shows all the steps of the folate pathway.
 
The proteins used in the folate biosynthesis pathway

GTP cyclohydrolase I (GTPCH)
Dihydroneopterin aldolase (DHNA)
6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase  (HPPK)
Dihydropteroate synthase (DHPS)
folylpolyglutamate synthetase (FPGS)
Dihydrofolate reductase (DHFR)





GTP cyclohydrolase I
GTPCHI was described for the first time in 1976  and is responsible for the rate-limiting step of the folate pathway, being regulated at both transcriptional and substrate/product levels. The reaction catalyzed by GTPCHI is considered the most complex of the pathway (Figure A below), in which is involved the production of  DHNP triphosphate and formic acid from GTP. GTPCHI initially breaks the guanine imidazole ring, followed by the cleavage at the N9-C8 and theN7-C8 bonds to produce theN-formyl pyrimidine intermediate and additionally releases the formate derivative from C8. The ribose moiety still undergoes an Amadori rearrangement, producing the dihydropyrazine ring, which is recyclized by a condensation reaction to provide the pteridine ring moiety of DHNP triphosphate (Figure A)




GTP cyclohydrolase I reaction

GTPCHI is conserved in bacteria, protozoa, fungi, plants, and vertebrates; however, its product is a substrate in more than one pathway among different organisms. In bacteria, fungi and plants, this enzyme catalyzes the first step
within de novo biosynthesis of folate, while invertebrates, it is associated with the biosynthesis of tetrahydrobiopterin, a key cofactor for the nitric oxide-producing enzymes, melanin and neurotransmitters.  However, the description of GTPCHI as a metalloenzyme was only established with the determination of human GTPCHI structure. A zinc ion binds near a histidine located within the active site. Posteriorly, a careful analysis of the EcGTPCHI active site also revealed an electronic density for this metal, which is coordinated by one histidine and two cysteines. The role of the zinc ion has been further proved to be essential for opening the imidazole ring by attacking the C8 of GTP.


GTP cyclohydrolase I
(A) Mechanism of catalysis of GTPCHI proposed by. 
(B) Dodecameric structure of Escherichia coli GTPCHI. Each protomer is represented by a different color. 
(C) Structure of an EcGTPCHI protomer. 
(D) Active site of EcGTPCHI. The interface of two protomers A (electrostatic surface) and B (ribbons and residues involved in the coordination of the essential zinc ion) are represented. 
(E) Dimeric structure of Neisseria gonorrhoeae GTPCH-IB, which also has the same tunneling fold of GTPCHI. EcGTPCHI: E. coli GTPCHI; GTPCHI: GTP cyclohydrolase I.

Dihydroneopterin aldolase
DHNA  catalyzes the reversible conversion of DHNP to 6-hydroxymethyl-7,8-dihydropterin (HP) and glycolaldehyde ( Figure below )




Dihydroneopterin aldolase. 
(A) Possible catalytic reactions for dihydroneopterin aldolase proposed by Czekster et al. 
(B) Octameric structure of SaDHNA. The bend black line indicates the interaction surface between the two tetramer that forms the octameric structure of DHNA. 
(C) Active site formation of dihydroneopterin aldolase by two adjacent protomers. 
(D) First series of compound  identified against SaDHNA based in a high-throughput x-ray crystallography campaign by Sanders et al. 
(E) Optimization of the 8-aminopurine analog that rendered compounds with high affinity to SaDHNA. SaDHNA: Staphylococcus aureus dihydroneopterin aldolase.

DHNA is also reported to catalyze the epimerization at the 2-carbon to produce DHMP. DHNA protomer contains a four-strand β-sheet and four α-helices wherein two are long and the other two are shorter, a feature not frequently described in all structures (Figure B & C). There are four active sites per DNHA tetramer and they are situated on the external face of the β-barrel and at the interface between two protomers of the tetramer with the contributions of amino acids from two adjacent molecules (Figure C above) Since the enzymatic catalysis does not involve cofactors, the active site is located in a narrow, deep and highly negatively charged pocket where amino acid residues are crucial for the recognition of the pterin ring substrate; however, a lysine, which acts as a general base, and a tyrosine, which has a key role in the protonation of the enol intermediate, are essential for the enzyme catalytic mechanism and they are conserved in almost all DHNA sequences. 

6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase
HPPK is an ATP-binding enzyme which transfers a pyrophosphoryl group to 6-hydroxymethyl-HP and produces hydroxymethyl-7,8-dihydropterin pyrophosphate (HPPP) and AMP (Figure A below).



DHPPK. 
(A) Catalytic mechanism of HPPK proposed by Blaszczyk et al. 
(B) Structure of EcHPPK in complex with 6-hydroxymethyl-7,8-dihydropterin and an analog of ATP (AMPCPP) indicating the position of the three essential loops for the activity of the enzyme. 
(C) Schematic representation of the dynamic movements of the essential Loop1 (yellow), -2 (pink) and -3 (green) during the binding of substrate and catalysis of HPPK. (D) Best inhibitors identified against EcHPPK and SaHPPK using different approaches. AMPCPP: Methyleneadenosine 5’-triphosphate; HPPK: 6-Hydroxymethyl-7,8-dihydropterin pyrophosphokinase; EcHPPK: Escherichia coli HPPK; SaHPPK: Staphylococcus aureus HPPK.

The structure of HPPK from E. coli (EcHPPK) revealed that this enzyme is monomeric and has a fold composed by a β-sandwich or a three-layered αβα similar to the ribosomal S6 protein  and nucleoside diphosphate kinase
(Figure B above)

Dihydropteroate synthase
This enzyme catalyzes the condensation of pABA and HPPP to produce dihydropteroate through the formation of a carbon–nitrogen bond (Figure A below)



Dihydropteroate synthase. 
(A) Catalytic mechanism proposed for dihydropteroate synthase based on the structure of Bacillus anthracis and Yersinia pestis DHPS by Yun et al. 
(B) Examples of classical dihydropteroate synthase inhibitors: prontosil and sulfamethoxazole. 
(C) Dimeric structure of Staphylococcus aureus dihydropteroate synthase. (D) Superposition of S. aureus (green) and Y. pestis dihydropteroate synthase (salmon) indicating that sulfamethoxazole binds in p-aminobenzoic acid binding site. (E) MANIC and the series of compounds identified by Zhao et al. [88] through a structure-based design campaign against B. anthracis DHPS.

Dihydrofolate synthetase/folylpolyglutamate synthetase


Dihydrofolate synthetase/folylpolyglutamate synthetase.
(A) Overall structure of monomeric Mycobacterium tuberculosis dihydrofolate synthetase/folylpolyglutamate synthetase indicating the two domains. 
(B) Surface representation of the two possible binding sites for Escherichia coli dihydrofolate synthetase/folylpolyglutamate synthetase. The ATP binding site bound with ADP and the folate binding site bound with phosphorylated dihydropteroate are shown in blue and green, respectively (PDB: 1W78). 
(C) Surface representation of open (blue) and closed (green) active site conformations of E coli folylpolyglutamate synthetase/dihydrofolate synthetase, the cavity is shown in red dashed circle.

Dihydrofolate reductase
Dihydrofolate reductase, or DHFR, is an enzyme that reduces dihydrofolic acid to tetrahydrofolic acid, using NADPH as electron donor, which can be converted to the kinds of tetrahydrofolate cofactors used in 1-carbon transfer chemistry. Dihydrofolate reductase converts dihydrofolate into tetrahydrofolate, a methyl group shuttle required for the de novo synthesis of purines, thymidylic acid, and certain amino acids. Found in all organisms, DHFR has a critical role in regulating the amount of tetrahydrofolate in the cell. Tetrahydrofolate and its derivatives are essential for purine and thymidylate synthesis, which are important for cell proliferation and cell growth. 22



Dihydrofolate reductase. 
(A) Examples of classical dihydrofolate reductase inhibitors. 
(B) Structure of Escherichia coli dihydrofolate reductase in complex with methotrexate. 
(C) Superposition of three different conformational states of E. coli dihydrofolate reductase based in the conformation of Met20 loop. 
(D) Superposition of open (orange) and closed (green) conformation of M. tuberculosis DHFR. The black arrow indicates that in the open position, the nicotinamide group is out of the active site and disordered. The green and orange arrows indicate different distances between the loop and adenosine binding subdomain, respectively in the closed and open conformations of M. tuberculosis DHFR. 
(E) Superposition between the open and closed conformations of M. tuberculosis DHFR in complex with trimethoprim indicating that the loss of the π-interaction between the nicotinamide ring of NADPH and the pyrimidine
ring of trimethoprim possible causes a change in the conformation of the ligand in the active site and also a decreasing of affinity.

S-adenosylmethionine (adoMet or SAM)
S-ADENOSYLMETHIONINE (SAM) is the most prolific donor of one-carbon groups in biosynthetic reactions.   Its formation is catalyzed by methionine adenosyltransferase.   It costs THREE high-energy phosphate bonds to make SAM.34 All known DNA methylases use S-adenosylmethionine as a methyl group donor. 36


The particularly important feature of SAM is that it donates methyl groups (we call them " active ~CH3 " to a large number of acceptors, including DNA, RNA, phospholipids, and many proteins.   Donation of these methyl groups is part of a small cycle




The step in which methionine is re-generated requires two important vitamin derivatives:


[list="color: rgb(0, 0, 0); background-color: rgb(255, 255, 255);"]
[*]Methyl-cobalamin, a derivative of vitamin B12, which donates the methyl group to homocysteine
[*]N5 -methyl-tetrahydrofolate, which donates its methyl group to cobalamin, and allows the reaction to continue.   Dietary or other deficiencies of either vitamin can cause serious problems.   Notice that all three of the one-carbon carriers are involved in this cycle.   SAM is also used to make cysteine.
[/list]

S-adenosylmethionine (SAM or AdoMet) is a conjugate of nucleotide adenosine and amino acid methionine, two ubiquitous biological compounds that almost certainly were present in the common ancestor of living cells and may have been found in the prebiotic environment on Earth, predating the origin of Life itself 35  SAM is an essential metabolic intermediate in every studied cellular life form, and each cellular organism has several SAM-utilizing enzymes. One relatively well-understood biological role of SAM is to donate methyl groups for covalent modification of different substrates – from as simple as oxidized arsenic, chloride, bromide, and iodine ions [2-4], to as complex as rRNA, tRNA, and essential proteins, whose methylation status can serve as a regulatory signal for maturation and control interactions with other macromolecules. 

S-Adenosylmethionine is a common cofactor involved in methyl group transfers, transsulfuration, and aminopropylation 32 

SAM is a substrate of methyltransferases in a variety of methyl-donor reactions, such as the formation of phosphatidylcholine from phosphatidylethanolamine, DNA methylation, and methylation of Arg and Lys residues in the
regulation of DNA: histone interactions in chromatin.  33

S-Adenosylmethionine (AdoMet) occupies a central role in the metabolism of all cells. 28 Reactions using S-adenosylmethionine (AdoMet) are among the most abundant processes taking place in any cell 29 The routes in which the AdoMet-consuming reactions are involved allow the synthesis of a large variety of compounds, as well as the control of cell function (i.e., epigenetic modifications). This wide use of AdoMet derives from the variety of groups that this molecule is able to donate, being methyl group donation the main consumer of the compound

S-adenosylmethionine (adoMet or SAM) synthesis
It is important to note the role of methionine itself in methylation reactions. The enzyme S-adenosylmethionine synthase catalyzes the reaction of methionine with ATP to form S-adenosylmethionine, or SAM (Figure below).


The synthesis of S-adenosylmethionine (SAM) and its fates.

SAM is a substrate of methyltransferases in a variety of methyl-donor reactions, such as the formation of phosphatidylcholine from phosphatidylethanolamine, DNA methylation, and methylation of Arg and Lys residues in the regulation of DNA: histone interactions in chromatin. 

S-adenosylmethionine synthetase (or methionine adenosyltransferase (MAT)) are the only enzymes known to synthesize AdoMet in a rather unusual reaction that occurs in two steps. As a methyl donor SAM allows DNA methylation. Once DNA is methylated, it switches the genes off and therefore, S-adenosylmethionine can be considered to control gene expression. 31


S-adenosylmethionine synthase 2, tetramer 

Methionine adenosyltransferases (MAT) are the family of enzymes that synthesize the main biological methyl donor, S-adenosylmethionine. 30 Methionine is a non-polar amino acid characterized by the presence of a methyl group attached to a sulfur atom located in its side chain. In addition to its role in protein synthesis, large amounts of this amino acid are used for the synthesis of S-adenosylmethionine (AdoMet) by methionine adenosyltransferases (MAT) in a reaction that is the rate-limiting step of the methionine cycle ( see figure below )


The mammalian methionine cycle and related pathways
The figure shows a scheme of the hepatic methionine cycle and some of the related pathways. Methionine is converted to S-adenosylmethionine (AdoMet) by methionine adenosyltransferases (MAT); this compound can be used by a multitude of enzymes such as methyltransferases (MTases), SAM radical proteins  and AdoMet decarboxylase ( AdoMetDC ). Polyamine synthesis occurs with methylthioadenosine (MTA) production, a compound that can be reused for methionine synthesis by the methionine salvage pathway. On the other hand, the action of MTases renders methylated products and S-adenosylhomocysteine (AdoHcy) that can be hydrolyzed by AdoHcy hydrolase (SAHH) to adenosine and homocysteine (Hcy). This reaction is reversible and favors AdoHcy synthesis. Hcy can be metabolized through the trans-sulfuration pathway by the consecutive action of cystathionine β synthase (CBS) and cystathionine γ lyase (CγL) rendering cysteine for glutathione synthesis, among other purposes. In addition, Hcy can also serve in resynthesis of methionine by two reactions catalyzed by methionine synthase (MS) and betaine homocysteine methyltransferase (BHMT). Some of these steps can be modulated by metabolites synthesized in these pathways and dashed lines indicate the most relevant.

S-adenosylmethionine synthetase enzymes also known as methionine adenosyltransferase ( MAT) catalyzes the only known route of AdoMet biosynthesis. The synthetic process occurs in a unique reaction in which the complete triphosphate chain is displaced from ATP and a sulfonium ion formed. 

MATs from various organisms contain ∼400-amino acid polypeptide chains. We have recently found that the protein sequences comprise two categories, the extensively studied eucaryal-bacterial type (encoded by a catalytic subunit denoted α ) and the archaeal type (encoded by a subunit that we denote γ ) The sequences of the two classes are widely diverged, e.g. there is only 22% identity.

This is amongst many others, one more indication that common ancestry of the 3 domains of life is a failed hypothesis.

Tetrahydrofolate can carry a methyl group on its N-5 atom, but its transfer potential is not sufficiently high for most biosynthetic methylations. Rather, the activated methyl donor is usually S- adenosylmethionine, which is synthesized by the transfer of an adenosyl group from ATP to the sulfur atom of methionine. It is important to note the role of methionine amino acids itself in methylation reactions. The enzyme S-adenosylmethionine synthase catalyzes the reaction of methionine with ATP to form S-adenosylmethionine, or SAM. 

a Transamination, a chemical reaction that transfers an amino group to a ketoacid to form new amino acids. This pathway is responsible for the deamination of most amino acids. This is one of the major degradation pathways which convert essential amino acids to nonessential amino acids (amino acids that can be synthesized de novo by the organism). 3
Transamination in biochemistry is accomplished by enzymes called transaminases or aminotransferases. α-ketoglutarate acts as the predominant amino-group acceptor and produces glutamate as the new amino acid.

b Pyridoxal phosphate (PLP, pyridoxal 5'-phosphate, P5P), the active form of vitamin B6, is a coenzyme in a variety of enzymatic reactions. The Enzyme commission has cataloged more than 140 PLP-dependent activities, corresponding to ~4% of all classified activities. The versatility of PLP arises from its ability to covalently bind the substrate, and then to act as an electrophilic catalyst, thereby stabilizing different types of carbanionic reaction intermediates. 5  PLP acts as a coenzyme in all transamination reactions, and in certain decarboxylation, deamination, and racemization reactions of amino acids

c In organic chemistry, an aldimine is an imine that is an analog of an aldehyde.[1] As such, aldimines have the general formula R–CH=N–R'. Aldimines are similar to ketimines, which are analogs of ketones.
An important subset of aldimines are the Schiff bases, in which the substituent on the nitrogen atom (R') is an alkyl or aryl group (i.e. not a hydrogen atom) 8

d Thymidine monophosphate (TMP), also known as thymidylic acid (conjugate base thymidylate), deoxythymidine monophosphate (dTMP), or deoxythymidylic acid (conjugate base deoxythymidylate), is a nucleotide that is used as a monomer in DNA. It is an ester of phosphoric acid with the nucleoside thymidine. dTMP consists of a phosphate group, the pentose sugar deoxyribose, and the nucleobase thymine. Unlike the other deoxyribonucleotides, thymidine monophosphate often does not contain the "deoxy" prefix in its name; nevertheless, its symbol often includes a "d" ("dTMP") 13

e Guanosine-5'-triphosphate (GTP) is a purine nucleoside triphosphate. It is one of the building blocks needed for the synthesis of RNA during the transcription process. Its structure is similar to that of the guanine nucleobase, the only difference being that nucleotides like GTP have a ribose sugar and three phosphates, with the nucleobase attached to the 1' and the triphosphate moiety attached to the 5' carbons of the ribose. 15

Formaldehyde (systematic name methanal) is a naturally occurring organic compound with the formula CH2O (H-CHO). It is the simplest of the aldehydes (R-CHO). The common name of this substance comes from its similarity and relation to formic acid. 23 Formaldehyde was the first polyatomic organic molecule detected in the interstellar medium. Since its initial detection in 1969, it has been observed in many regions of the galaxy. Because of the widespread interest in interstellar formaldehyde, it has recently been extensively studied, yielding new extragalactic sources.
" The poisonous chemical formaldehyde may have helped create the organic compounds present in the universe that gave rise to life, new research suggests. " 24

g In the chemical sciences, methylation denotes the addition of a methyl group on a substrate, or the substitution of an atom (or group) by a methyl group. Methylation is a form of alkylation, with a methyl group, rather than a larger carbon chain, replacing a hydrogen atom. These terms are commonly used in chemistry, biochemistry, soil science, and the biological sciences. 26 A methyl group is an alkyl derived from methane, containing one carbon atom bonded to three hydrogen atoms — CH3. 27 Such hydrocarbon groups occur in many organic compounds. It is a very stable group in most molecules. While the methyl group is usually part of a larger molecule, it can be found on its own in any of three forms: anion, cation or radical. The anion has eight valence electrons, the radical seven and the cation six.

  

1. Berg, Biochemistry, 5th edition, page 973
2. https://www.nature.com/scitable/topicpage/an-evolutionary-perspective-on-amino-acids-14568445#
3. https://en.wikipedia.org/wiki/Transamination
4. Biochemistry 5th edition, Styer, page 713
5. https://en.wikipedia.org/wiki/Pyridoxal_phosphate
6. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2443152/
7. https://www.annualreviews.org/doi/10.1146/annurev.biochem.73.011303.074021
8. https://en.wikipedia.org/wiki/Aldimine
9. https://sites.google.com/site/mdtoneylab/research/pyridoxal-phosphate-enzymes
10. https://en.wikipedia.org/wiki/Tetrahydrofolic_acid
11. http://www.biochem.uthscsa.edu/med/08-Amino-Acid-Metabolism/Amino-Acid-Metabolism7.html
12. https://en.wikipedia.org/wiki/Folate
13. https://en.wikipedia.org/wiki/Thymidine_monophosphate
14. http://www.llamp.net/?q=Folate%20biosynthesis
15. https://en.wikipedia.org/wiki/Guanosine_triphosphate
16. http://pubs.rsc.org/en/content/articlelanding/2007/np/b703107f#!divAbstract
17. https://www.wikipathways.org/index.php/Pathway:WP241
18. http://sci-hub.tw/https://www.future-science.com/doi/full/10.4155/fmc-2017-0168
19. http://sci-hub.tw/10.1146/annurev-arplant-042110-103819
20. https://iubmb.onlinelibrary.wiley.com/doi/pdf/10.1002/iub.1153
21. http://what-when-how.com/molecular-biology/tetrahydrofolate-molecular-biology/
22. https://en.wikipedia.org/wiki/Dihydrofolate_reductase
23. https://en.wikipedia.org/wiki/Formaldehyde
24. https://www.livescience.com/13551-formaldehyde-poison-origin-earth-life.html
25. Biochemistry, Styer , 8th ed. page 724
26. https://en.wikipedia.org/wiki/Methylation
27. https://en.wikipedia.org/wiki/Methyl_group
28. http://www.jbc.org/content/277/19/16624.full
29. http://sci-hub.tw/10.1016/j.pep.2011.05.004
30. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2643306/
31. https://en.wikipedia.org/wiki/S-adenosylmethionine_synthetase_enzyme
32. https://en.wikipedia.org/wiki/S-Adenosyl_methionine
33. Biochemistry 6th edition, Garrett, page 1063
34. http://www.biochem.uthscsa.edu/med/08-Amino-Acid-Metabolism/Amino-Acid-Metabolism8.html
35. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1282579/
36. Lehninger, Principles of biochemistry, 5th ed. page 292



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In Narnia, we are transported to fantasy land. I feel sometimes, that God is taking my hands, and takes me to a walk to see what he has done to make life possible, to a reality which is more bewildering than in someone’s wildest dreams. When i move forward in my investigation, I step into new territory, and what i discover, makes me speechless.

The ability of transfer of just ONE SINGLE CARBON atom is absolutely essential for the metabolism of the amino acids glycine, serine, methionine, and histidine, and the biosynthesis of purines and pyrimidines - which constitute DNA molecules, the information carriers of cells.

And in order for biological cells to achieve this transfer, they require tetrahydrofolate cofactors, consisting of three moieties. Folates are among the most complex pterin coenzymes. The folate pathway is central to any study related to DNA methylation, dTMP synthesis or purine synthesis, and as such, to the origin of life itself, since without amino acids, and DNA - no life.


Annexed below, you can see the Folate biosynthesis pathway - at each branch point, there is a ramification of a web of complex enzymes which work in a coordinated, orchestrated, and interconnected way together to produce just this Tetrahydrofolate cofactor. To make things even more complex, the two essential precursors of folate biosynthesis are 4-aminobenzoate (a product of shikimate biosynthesis pathway) and GTP. To give you an idea about the complexity of the shikimate metabolic pathway, you can have a look here:

http://www.genome.jp/kegg-bin/show_pathway?map01063

The central pathway uses six extremely complex enzymes, which I describe in detail in the article below. Moral of the story: These metabolic networks, enzymes, and co-factors are upon which life depends, and could hardly be explained with any other causal mechanism, besides a super intelligent creator.



Cheating of secular science papers, claiming of evolutionary mechanisms in place prior to life fully setup and self-replication.
The deceptive narrative of secular science papers of origins is evident between the lines when carefully analyzed.


Anyone that follows my posts over a certain time period, will observe, that I post periodically about my findings on molecular biology. It follows a logic since I am writing a book on the intracellular world.

Origins - what cause explains best our existence, and why?
Molecular biochemistry, biology, the origin of life and biodiversity, systematically analyzed from an epistemologically universal perspective
http://reasonandscience.catsboard.com/t2590-origins-what-cause-explains-best-our-existence-and-why

Slowly, I am unraveling the unexpected awe-inspiring mechanisms that cells use to produce life. Basically, all basic building blocks used in life are synthesized inside the Cell by extremely complex, irreducible, interdependent molecular machines and factories. That is six types of macromolecules. They are

amino acids,
phosphate,
glycerol,
sugars,
fatty acids,
and nucleotides.

Now I am investigating in-depth about how Cells synthesize amino acids. That has brought me in the last month in extensive length to elucidate the bewildering unfathomably complex machinery that transforms dinitrogen ( 78% of the air we breath is dinitrogen ) in the atmosphere into ammonia and nitrate, incorporated in the cells to make amino acids, used to make proteins, the molecular workhorses of the Cell. After nitrogenase does the job of transformation of Dinitrogen into ammonia by an enormously energy consuming process, 3 enzymes, amongst them Glutamate dehydrogenase, a veritable molecular Supercomputer, converts inorganic ammonium ion into the α-amino nitrogen of amino acids.

What Is the Metabolic Fate of Ammonium?
http://reasonandscience.catsboard.com/t2590p25-origins-what-cause-explains-best-our-existence-and-why#5934

The key entry point is the amino acid glutamate. Glutamate dehydrogenase (GDH) catalyzes the reductive amination of a-ketoglutarate to yield glutamate.  Alpha-ketoglutarate (AKG) is a nitrogen scavenger and a source of glutamate and glutamine - which are the nitrogen donors in a wide range of biosynthetic reactions.

The amino acid and nucleotide biosynthetic pathways make repeated use of the biological cofactor pyridoxal phosphate.

Now comes the key point of this post.

What is the origin of Pyridoxal phosphate enzymes?

The authors of following science paper claim that:
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2443152/

The pyridoxal-5'-phosphate (PLP)-dependent or vitamin B6-dependent enzymes that catalyze manifold reactions in the metabolism of amino acids belong to no fewer than four evolutionarily independent protein families.

My comment: It is remarkable that the authors do not make a distinction, recognizing that the origin of pyridoxal phosphate enzymes ( Vitamin B6) and the respective enzyme families which they interact with had to be fully operational at the Last Universal Common Ancestor ( LUCA ),  or in other words, when life began,  and could therefore not be the result of evolution. The fact that B6 interacts with four independent protein families challenges naturalistic explanations even further. Convergent emergence means that the same enzymatic reaction would have had to emerge independently and separately four times - that is extremely unlikely without guidance and direction, and teleology - or, goal oriented.

Further, the authors go and claim:
"The multiple evolutionary origins and the essential mechanistic role of PLP in these enzymes argue for the cofactor having arrived on the evolutionary scene before the emergence of the respective apoenzymes and having played a dominant role in the molecular evolution of the B6 enzyme families."

Why would natural occurrences on a prebiotic earth have produced co-factors ( in comparison of lock and key, the key alone) without the respective apo-enzymes ( the lock ) to interact with? Did they emerge, without function at all, just waiting for the respective proteins to interact with, to arrive later on the scene, to then eagerly looking to find them, and starting the molecular interaction?

Recycling, and the orchestration of anabolism and catabolism, evidence of natural forces, or design?

http://reasonandscience.catsboard.com/t2696-metabolism-and-catabolism-evidence-of-design

Recycling or reuse of used material, organized decomposition into basic building blocks, separation, and organized re-use is an exclusive activity performed by intelligence, namely by us, humans, who have figured out of know-how. And the more we practice it, the more sustainable our and less destructive our activities are for the planet where we live in.

In biological cells, recycling is a highly orchestrated, complex, and coordinated process. It is called catabolism. While in anabolism, metabolic networks construct molecules from smaller units, while in catabolism, a set of metabolic pathways breaks down molecules into smaller units that are either oxidized to release energy or used in other anabolic reactions       Interestingly, anabolism and catabolism occur simultaneously in the cell. The conflicting demands of concomitant catabolism and anabolism are managed by cells in two ways. First, the cell maintains tight and separate regulation of both catabolism and anabolism, so metabolic needs are served in an immediate and orderly fashion. Second, competing metabolic pathways are often localized within different cellular compartments. Isolating opposing activities within distinct compartments, such as separate organelles, avoids interference between them.

Question: How could unguided, random, not goal oriented processes on early earth have brought such a system into being?  regulation, order, management, organized separation, compartmentalization are known to be exclusively brought into action by intelligence. No exception is known.

A rather limited collection of simple precursor molecules is sufficient to provide for the biosynthesis of virtually any cellular constituent, be it protein, nucleic acid, lipid, or polysaccharide.Certain of the central pathways of intermediary metabolisms, such as the citric acid cycle, and many metabolites of other pathways have dual purposes—they serve in both catabolism and anabolism. Remarkably, the opposite metabolic directions is that such pathways must be independently regulated.

Question: How could such regulation have emerged in a stepwise, slow, gradual trial and error fashion, and the fact that regulation is essential? Could and would both independent regulation implementation not have had to emerge simultaneously, if considered, that the reverse cycle is slightly different and differently adjusted in order to work properly?

If catabolism and anabolism passed along the same set of metabolic tracks, equilibrium considerations would dictate that slowing the traffic in one direction by inhibiting a particular enzymatic reaction would necessarily slow traffic in the opposite direction. Independent regulation of anabolism and catabolism can be accomplished only if these two contrasting processes move along different routes or, in the case of shared pathways, the rate-limiting steps serving as the points of regulation are catalyzed by enzymes that are unique to each opposing sequence.

It is evident that in order to implement such a system that works both ways, there must be foresight and the setting of specific goals, and teleology, which is what naturalism must try to avoid to be true.

The spatial compartmentalization of metabolic pathways within cells provides important advantages, one of which is isolating competing pathways from one another. Cells and organisms also exhibit temporal compartmentalization of their metabolic pathways. That is, metabolic pathways may be turned on and off in a time-dependent and/or cyclic fashion. For example, the metabolism of many organisms—microbes, animals, and plants—is regulated in synchrony with the 24-hour cycle of day and night, a pattern called circadian rhythmicity and often referred to as the biological clock.

Question: How could such a circadian rhythm have emerged in a random manner? It could not be explained by evolution since: 

" The 24-hour circadian clock found in human cells is the same as that found in algae and dates back millions of years to early life on Earth, and is linked to DNA and gene activity " 
https://www.sciencedaily.com/releases/2011/01/110126131540.htm

and: " Because light and/or varying nutrient availability represent key signals regarding the transitory nature of the environment, and organisms have evolved and adapted to exploit the information in such signals. ".

Remarkably, there is not only a 24h circadian clock, but also a 7-day Circadian Clock, which coincides with Gods setup of six days, and the Sabbath, the rest on the seventh day:

The seven-day Circadian Rhythms: "Nature's Intricate Clockwork"
http://reasonandscience.catsboard.com/t1487-the-seven-day-circadian-rhythms-nature-s-intricate-clockwork

Metabolic networks are extraordinarily complex. The Human Metabolomics Database (www.hmdb.ca) provides data on more than over 40,000 metabolites in cells and fluids (blood, urine, and so on) of the human body. The metabolomes of plants are even more complex, with estimates suggesting hundreds of thousands of different metabolites across the plant kingdom. Metabolomic assays must be able to resolve and discriminate this extraordinary array
of small molecules.

Atheists can tell me as much and as often as they want, that I lack credulity towards their narrative, that such complex molecular networks could have emerged randomly on early Earth. I will grant them with the same return: It requires a lot of faith/credulity that such complexity could be the process of random chemical interaction on a prebiotic earth.



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Phosphate, essential for life 3

http://reasonandscience.catsboard.com/t2463-phosphate-essential-for-life

Hugh Ross: The short supply of phosphorus poses a significant problem for a naturalistic origin of life because so much of this ingredient is required to make replicator molecules. Phosphates are part of the backbone of both DNA and RNA. A phosphate molecule must accompany every nucleoside in them. Possible precursors to DNA and RNA molecules would seem to require similar phosphate richness. Without life molecules (already assembled and operating), no known natural process can harvest the amounts of phosphorus necessary for life from the environment. All the phosphate-rich deposits on Earth are produced by life.

Biological Significance of Phosphorus 

One problem regarding the prebiotic chemistry is the ―problem of phosphorus and phosphorylation

Phosphorus is a key biogenic molecule which comprises of about 1% of the dry weight of cells. It serves in all life properties such as cellular metabolism (cellular respiration, involving sugar phosphates, P containing enzymes etc.), structure (phospholipids) and a necessary part of information storing molecules (RNA and DNA). Replication and metabolism are the two main biochemical processes that mainly rely on P as it constitutes 3% of the atomic composition of RNA and about 1% of the atomic composition of the metabolomics. Most interestingly, about 44% of all metabolic compounds are biological P compounds. The phosphorylated biomolecules occurring in living organisms are classified into the following categories;

(1) Reactive organophosphates such as acetyl phosphate, phosphoenolpyruvate, phosphocreatine,
(2) stable phosphorylated biomolecules such as glycerol phosphates, ribose phosphate, and phosphoethanolamine and
(3) condensed phosphates such as adenosine-di-phosphates (ADP), and adenosine-tri-phosphates (ATP)




The chart above shows a generic scheme to show various routes that have been described so far in the prebiotic phosphorylation reactions where; OP stands for orthophosphate and TMP stands for trimetaphosphate.



Some biological phosphates that are vital for cellular activities and hence for life. These include simple sugar phosphates such as glycerol phosphate (for respiration and cell structure) and highly complex molecules such as nucleotides (for storing information), and phospholipids (for structure).

Prebiotic Phosphorylation by Phosphates

phosphorylation reaction means a dehydration reaction between an OP and organic molecule.

R-OH + HO-PO32− R-O-PO32− + H2O [2]

The reaction given above is thermodynamically unfavorable. This is because this process is trying to ―pull out water into the aqueous environment, thus making phosphorylation extremely challenging and with low efficiency. This is a major reason why the phosphorylation process has been a challenge in prebiotic chemistry. The major challenging aspects include a common route to synthesize significant biological P compounds with good yields and with conditions relevant to the Hadean Earth and in the presence of water.

Phosphorus in prebiotic chemistry 2

A variety of pathways now are available as possible solutions for what this author used to refer to as the ‘phosphate problem’. The ‘problem’ seems to have evolved gradually into the less perplexing one of having to decide which of several possible mechanisms are most likely to have operated on the primitive Earth.

Phosphorus and sulfur for energetic bonds Phosphorus and sulfur create the principal energy-carrying bonds of organic chemistry 4 The principle phosphoryl-transfer cofactors are the nucleoside triphosphates (NTPs), with ATP serving as the major energy carrier and other NTPs performing special functions that include a signaling role because of their restricted uses. The central role and high concentration of phosphorus in biochemistry is disproportionate to its availability in most geochemical settings on Earth today. There are good reasons that no other chemical group may be able to do what phosphorus does in biochemistry, but its inevitability alone does not address the problem of either obtaining it or using it as a vehicle of disequilibrium.




Some examples of biologically relevant phosphates 1


Phosphate is essential for all living systems, serving as a building block of genetic and metabolic machinery. However, it is unclear how phosphate could have assumed these central roles on primordial Earth, given its poor geochemical accessibility. While most research on the evolution of living systems has been focused on sequences and genomes, some answers to fundamental questions about the emergence of life may be hidden in the architecture of the complex biochemical reaction networks that sustain the cell.  Among the many unanswered questions on life’s origin, the enigma of how phosphate ended up playing a prominent role in cellular biochemistry has been puzzling scientists for decades. 
1. http://sci-hub.tw/https://www.cambridge.org/core/journals/quarterly-reviews-of-biophysics/article/why-nature-really-chose-phosphate/030269AA621E6EA0CADDC263E6977930
2) http://rstb.royalsocietypublishing.org/content/361/1474/1743
3. http://www.cell.com/cell/fulltext/S0092-8674(17)30133-2
4. https://books.google.com.br/books/about/The_Origin_and_Nature_of_Life_on_Earth.html?id=vi-8CwAAQBAJ&source=kp_cover&redir_esc=y



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33 Lipids and Biological Membranes on Fri Jun 08, 2018 8:56 am

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Lipids and Biological Membranes



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How are lipids synthesized?

Lipids play a life essential role, and so, in many biochemical and cell biological processes. They are involved in the formation of biological membranes and therefore are important elements of organelle biogenesis and function. 25 This is not only true for bulk lipids such as the major classes of glycerophospholipids, sterols, and sphingolipids, but also for less abundant lipid species. Lipids with only minor concentrations in cells have more subtle functions. For example, polyphosphoinositides, lysophospholipids, or ceramides are essential for signaling processes. Certain classes of lipids, such as sphingolipids, are also sensors of cellular stress (heat shock) and important components for signal transduction. These lipids provide the appropriate environment for membrane proteins to function as enzymes or transporters, or to act as positive regulators of enzymes to gain optimum activity in membranes.

Many regulatory phenomena caused by lipids, which have not been recognized before, became evident through the advance of molecular biological research. Intracellular lipid transport and distribution play an important role in the maintenance of cellular structure and function. The lipid composition of vesicles involved in membrane traffic, recently, turned out to be a most important parameter for protein targeting. For example, the presence of sterols and sphingolipids in secretory and endocytotic vesicles appears to be essential. Membrane contact, as a mechanism of translocation of components between subcellular compartments, may also depend on the lipid patterns of donor and acceptor membranes.

Lipid metabolism must not be regarded only as an isolated process, but rather as a part of total cellular metabolism, which is highly linked to other metabolic and cell biological pathways


The first step in lipid metabolism is the hydrolysis b of the lipid c in the cytoplasm to produce glycerol and fatty acids. 5

Complex lipids consist of backbone structures to which fatty acids d are covalently bound. Principal classes include the glycerolipids, for which glycerol is the backbone, and sphingolipids, which are built on a sphingosine backbone. The two major classes of glycerolipids are glycerophospholipids and triacylglycerols

Glycerophospholipids or phosphoglycerides are glycerol-based phospholipids. They are the main component of biological membranes.

The phospholipids, which include both glycerophospholipids and sphingomyelins, are crucial components of membrane structure. Different organisms possess greatly different complements of lipids and therefore invoke somewhat different lipid biosynthetic pathways. For example, sphingolipids and triacylglycerols are produced only in eukaryotes. In contrast, bacteria usually have rather simple lipid compositions. 


Glycerolipid metabolism 12

Glycerol, essential for life
Glycerol a also called glycerine or glycerin is a simple polyol j compound. It is a colorless, odorless, viscous liquid that is sweet-tasting and non-toxic. The glycerol backbone is found in all lipids known as triglycerides. 8 Glycerin is a trihydroxy sugar alcohol that is an intermediate in carbohydrate and lipid metabolism. 9

The phospholipids characteristic of contemporary membranes are composed of two fatty acids esterified to a glycerol phosphate molecule, with any of several groups also attached to the phosphate through an ester bond, such as
choline, ethanolamine, serine, and glycerol. 7 Although hydrocarbons are abundant on today’s Earth, they are primarily products of biological processes, so there must have been supposedly abiotic sources of the amphiphiles required for assembly into first membrane structures if natural membrane assembly mechanisms are presupposed. Two possibilities are geochemical synthesis under volcanic conditions and delivery during the accretion of asteroids and comets in the late stage of planet formation.

Of course, that raises the question, why would biological processes have evolved the ultracomplex protein-machine-based processes to make lipids, if they were supposedly readily available on early earth? 

Phospholipids found in cell membranes show a wide range of chemical variability. Their head groups typically consist of a phosphate group bound to a glycerol (glycerin) backbone. 1
Cell membranes basically consist of phospholipids, which are amphiphile ( water-loving) molecules, that is to say that they possess a hydrophilic ( attracted to water ) head (consisting of glycerol-phosphate) and a hydrophobic ( repelling water )  tail (which may belong fatty acids or isoprenoid derivatives) 2 The fundamental distinction (with no known exceptions) between the phospholipids of the bacteria and eukaryotes, and those of the archaea rests in the type of stereoisomer e of glycerol used  ; 

glycerol-3-phosphate in the bacteria and eukaryotes, 
glycerol-1-phosphate in the archaea. 

The pathways by which these two stereoisomers are synthesized are so different that, for certain authors, the cenancestor did not have membranes and was an acellular organism or, according to yet others, it had mineral membranes consisting of iron monosulfide.

Here we have one reason amongst many others, why common ancestry of all three domains of life is extremely unlikely. 

The idea of an ancestor without lipid membranes, however, comes up against one piece of evidence: there are membrane proteins that are universally conserved, like the ATPase. A less radical hypothesis, which had recently been supported by molecular phylogeny analyses, would be to imagine a cenancestor with a heterochiral membrane, that is to say possessing a mixture of phospholipids constructed with glycerol-1-phosphate and glycerol-3-phosphate. Additional molecular phylogenetic studies further support the idea that the cenancestor had a complete toolkit to make phospholipids of either fatty acids or isoprenoid chains. Evolution would have subsequently led to opposite stereospecificity and choice of lateral chains in the bacteria and in the archaea.

Another explanation is that common ancestry is an unlikely hypothesis, and the origin of the three domains of life is not due to a last common universal ancestor, but distinct creation events. 

The phospholipids of Bacterial and eukarya plasma membranes consist of fatty acids ester-linked f to glycerol-3-phosphate
Archaea make theirs of isoprenoids ether-linked g to glycerol- 1-phosphate


The structure of membrane phospholipids in the three domains of life. 
The type of stereo-isomer of glycerol-phosphate distinguishes, with no known exception, between the phospholipids of bacteria and eukaryotes, and those of the archaea: glycerol-3-phosphate in bacteria and eukaryotes, glycerol-1-phosphate in archaea. Other differences are observed in the hydrophobic chains and the bond between the latter and the glycerol-phosphate, but there are exceptions. Some phospholipids in archaea have chains of fatty acids, and some phospholipids in bacteria include ether bonds.


Glycerol 1-phosphate, sometimes called as D-glycerol 3-phosphate, is an enantiomer of glycerol 3-phosphate. Most organisms use 3-phosphate, or L-configuration, for glycerolipid backborn; however, 1-phosphate is specifically used in archeal ether lipids.

Glycerol 3-phosphate is a phosphoric ester of glycerol, which is a component of glycerophospholipids. 11

Glycerol 3-phosphate synthesis and metabolism

There are several routes to synthesize Glycerol 3-phosphate. 

1. Glycerol 3-phosphate is synthesized by reducing dihydroxyacetone phosphate (DHAP), a glycolysis intermediate, with glycerol-3-phosphate dehydrogenase. 11
Glycerol-3-phosphate dehydrogenase is an enzyme that catalyzes the reversible redox conversion of dihydroxyacetone phosphate  to sn-glycerol 3-phosphate. Glycerol-3-phosphate dehydrogenase serves as a major link between carbohydrate metabolism and lipid metabolism. Sn-glycerol-3-phosphate dehydrogenase (GlpD) 4 is an essential membrane enzyme, functioning at the central junction of respiration, glycolysis, and phospholipid biosynthesis. Its critical role is indicated by the multitiered regulatory mechanisms that stringently controls its expression and function. Once expressed, GlpD activity is regulated through lipid-enzyme interactions. Homologs of GlpD are found in practically all organisms, from prokaryotes to humans, with >45% consensus protein sequences, signifying that these structural results on the prokaryotic enzyme may be readily applied to the eukaryotic GlpD enzymes.

Despite the pioneering work of Hargreaves et al. in 1977 who demonstrated that the synthesis of phosphatidic acid and other lipids could be achieved abiotically, it is considered very improbable that fatty acids, glycerol, and phosphate (i.e., the standard molecular components of a phospholipid) could have been present together in high enough concentrations on the primordial Earth. 6





2. Glycerol can be a source for glycerol-3-phosphate, in which case, a phosphate from ATP is transferred to glycerol by glycerol kinase forming glycerol-3-phosphate and ADP. 22 Glycerol kinase 23 catalyzes the phosphorylation i of glycerol to form glycerol-3-phosphate, which is then acylated at both the 1- and 2-positions to yield phosphatidic acid. Glycerol is a precursor for the synthesis of phospholipids. Before glycerol can enter the pathway of glycolysis or gluconeogenesis (depending on physiological conditions), it must be converted to their intermediate glyceraldehyde 3-phosphate in the following steps:




Glycerol kinase, encoded by the gene GK, is a phosphotransferase enzyme involved in triglycerides and glycerophospholipids synthesis. 






b Biological hydrolysis is the cleavage of biomolecules where a water molecule is consumed to effect the separation of a larger molecule into component parts. 13

c In biology, a lipid is a biomolecule that is soluble in nonpolar solvents. Non-polar solvents are typically hydrocarbons used to dissolve other naturally occurring hydrocarbon lipid molecules that do not (or do not easily) dissolve in water, including fats, waxes, sterols, fat-soluble vitamins (such as vitamins A, D, E, and K), monoglycerides, diglycerides, triglycerides, and phospholipids. 14

In chemistry, particularly in biochemistry, a fatty acid is a carboxylic acid with a long aliphatic chain, which is either saturated or unsaturated. Most naturally occurring fatty acids have an unbranched chain of an even number of carbon atoms, from 4 to 28. 15

e In stereochemistry, stereoisomers are isomeric molecules that have the same molecular formula and sequence of bonded atoms (constitution), but differ in the three-dimensional orientations of their atoms in space 16

f In chemistry, an ester is a chemical compound derived from an acid (organic or inorganic) in which at least one –OH (hydroxyl) group is replaced by an –O–alkyl (alkoxy) group 17

g In an organic chemistry general sense, an ether lipid implies an ether bridge between an alkyl group (a lipid) and an unspecified alkyl or aryl group, not necessarily glycerol. 18

h  Isoprenoids are any of a class of organic compounds composed of two or more units of hydrocarbons, with each unit consisting of five carbon atoms arranged in a specific pattern. These compounds are derived from five-carbon isoprene units and are biosynthesized from a common intermediate known as mevalonic acid, which is itself synthesized from acetyl-CoA. These lipids are considered to be the largest group of natural products, playing a wide variety of roles in physiological processes of plants and animals. 19

In chemistry, phosphorylation of a molecule is the attachment of a phosphoryl group. Together with its counterpart, dephosphorylation, it is critical for many cellular processes in biology. 21

polyol is an organic compound containing multiple hydroxyl groups. 24

1. Origins of life : biblical and evolutionary models face off / Fazale Rana & Hugh Ross., page 102
2. Young Sun, Early Earth and the Origins of Life , page 122
3. https://ipfs.io/ipfs/QmXoypizjW3WknFiJnKLwHCnL72vedxjQkDDP1mXWo6uco/wiki/Glycerol_3-phosphate.html
4. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2265192/
5. http://chemistry.elmhurst.edu/vchembook/622overview.html
6. Prebiotic Systems Chemistry: New Perspectives for the Origins of Life : http://sci-hub.tw/https://pubs.acs.org/doi/10.1021/cr2004844
7. http://sci-hub.tw/10.1016/j.resmic.2009.06.004
8. https://en.wikipedia.org/wiki/Glycerol
9. https://pubchem.ncbi.nlm.nih.gov/compound/glycerol
10. https://en.wikipedia.org/wiki/Glycerophospholipid
11. https://en.wikipedia.org/wiki/Glycerol_3-phosphate
12. https://www.genome.jp/kegg-bin/show_pathway?map00561
13. https://en.wikipedia.org/wiki/Hydrolysis
14. https://en.wikipedia.org/wiki/Lipid
15. https://en.wikipedia.org/wiki/Fatty_acid
16. https://en.wikipedia.org/wiki/Stereoisomerism
17. https://en.wikipedia.org/wiki/Ester
18. https://en.wikipedia.org/wiki/Ether_lipid
19. https://en.wikibooks.org/wiki/Structural_Biochemistry/Lipids/Isoprenoids
20. https://en.wikipedia.org/wiki/Phosphatidic_acid
21. https://en.wikipedia.org/wiki/Phosphorylation
22. http://reactome.org/content/detail/R-HSA-75887
23. https://en.wikipedia.org/wiki/Glycerol_kinase
24. https://en.wikipedia.org/wiki/Polyol
25. Lipid Metabolism and Membrane biogenesis, page 1

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