In 2011, Ann Gauger and Douglas Axe published a paper in BIO-Complexity, "The Evolutionary Accessibility of New Enzymes Functions: A Case Study from the Biotin Pathway." They reported results of their laboratory experiments trying to convert one enzyme (Kbl2) to perform the function of a very similar enzyme (BioF2), thought to be very closely related to Kbl2. Because these proteins are both members of the GABA-aminotransferase-like (GAT) family, and are believed to be very closely related, this is the sort of evolutionary conversion that evolutionists say ought to be easily accomplished under the standard co-option model. However, after trying multiple combinations of different mutations, they found otherwise:
We infer from the mutants examined that successful functional conversion would in this case require seven or more nucleotide substitutions.
2010 paper by Axe
Evolutionary innovations requiring that many changes would be extraordinarily rare, becoming probable only on timescales much longer than the age of life on earth. Considering that Kbl2 and BioF2 are judged to be close homologs by the usual similarity measures, this result and others like it challenge the conventional practice of inferring from similarity alone that transitions to new functions occurred by Darwinian evolution.
Now in their new study, "Enzyme Families-Shared Evolutionary History or Shared Design? A Study of the GABA-Aminotransferase [GAT] Family," Reeves, Gauger, and Axe examine nine other enzymes from the same GAT family. Once again, the idea was to see if it is possible to convert them to perform the function of BioF2. They tested proteins that are closer to BioF2, or more distant from BioF2, than the enzyme they tested in their prior study (Kbl2). But all of the proteins studied are in the same family, and are thought to be closely related.
First, they sought to determine if the enzymes could be converted to perform the function of BioF2 through a single mutation. They created mutation libraries with every single possible mutation in those nine enzymes. No BioF2 function was ever detected. As they explain:
The present study has added to our previous examination of these problems in several respects. We have shown, based on sequence alignment of α-oxoamine synthases (a subset of the GAT family), that our previous use of rational design did indeed target regions of Kbl2 that are likely to be functionally significant. Furthermore we have now shown that the lack of a simple evolutionary transition to BioF2 function is not at all unique to our initial choice of Kbl2 as the starting point. Single mutations cannot convert any of eight other members of the GAT family to that function, despite the fact that all of these enzymes are regarded as close evolutionary relatives.