Experimental Evolution of Ribitol Dehydrogenase

Abstract

This project began in 1968, when the impact of amino acid sequencing and protein crystallography had revealed a flood of data with great impact on evolutionary theory. These facts allowed the following important conclusions: 1. The conformation of the same protein from different species is carefully conserved throughout evolution. 2. Internal amino acid residues that clearly contribute to that conformation are generally, but not invariably, conserved. 3. Surface amino acids are more variable, except for those that clearly contribute to the catalytic activity. This encouraged the concept of “neutral mutations.” 4. Evolutionary trees can be constructed from a matrix of species differences, either overall or by translating back to DNA sequences via the known genetic code or by assuming “invariant” and “variable” regions. These evolutionary trees can be made to bear a satisfying resemblance to the known fossil record. 5. The rate of sequence variation appears to correlate with time rather than with the assumed number of generations between species. This led to the “neutral drift” theory of Kimura (1969). 6. Protein families with similar functions, such as myoglobin and hemoglobin, or the pancreatic serine proteases, also have almost superimposable conformations. Divergence from a common ancestor via gene duplication was clearly implied. 7. Sequence variations between these similar proteins in the same individual follow the same pattern as species differences for a single protein. Hence, evolutionary trees implying distance from a common ancestor could be constructed. 8. Specificity differences between chymotrypsin, trypsin, and elastase appeared to require only one or two amino acid changes, respectively, with no significant conformational change in the specificity sites (Hartley and Shotton, 1971). 9. Convergent evolution to a common enzyme mechanism from two completely different protein ancestors was obvious from the structures of chymotrypsin and subtilisin (Kraut et al., 1971).