Evolution of protein specificity: insights from ancestral protein reconstruction

Abstract

Specific interactions between proteins and their molecular partners drive most biological processes, so understanding how these interactions evolve is an important question for biochemists and evolutionary biologists alike. It is often thought that ancestral proteins were systematically more “promiscuous” than modern proteins, and specificity often does evolve by partitioning and refining the activities of multifunctional ancestors after gene duplication. However, recent studies using ancestral protein reconstruction (APR) have found that ligand-specific functions in some modern protein families evolved de novo from ancestors that did not already have those functions. Further, the new specific interactions evolved by simple mechanisms, with just a few mutations changing classically recognized biochemical determinants of specificity, such as steric and electrostatic complementarity. Acquiring new specific interactions during evolution therefore appears to be neither difficult nor rare. Rather, it is likely that proteins continually gain and new lose activities over evolutionary time as mutations cause subtle but consequential changes in the shape and electrostatics of interaction interfaces. Only a lucky few of these, however, are incorporated into the biological processes that contribute to fitness before they are lost to the ravages of further mutation.

Development and retention of a primordial moonlighting pathway of protein modification in the absence of selection presents a puzzle

“…In this scenario proteins continually gain and lose new activities, only a few of which contribute to fitness and are retained. Indeed, examples are known in which one or two residue changes can alter enzyme specificity and protein-protein interactions (12). This view may provide an explanation for the diverse activities of the five annotated A. aeolicus GcvH proteins.…”

“…It has been argued that during evolution, acquisition of specific new interactions is neither difficult nor rare (12). In this scenario proteins continually gain and lose new activities, only a few of which contribute to fitness and are retained.…”

Higher-order epistatic networks underlie the evolutionary fitness landscape of a xenobiotic-degrading enzyme

“…Previous studies using ASR examined functions that evolved millions or billions of years ago [2][3][4][31][32][33][34][35] . Our study demonstrates that this technique, combined with biochemical and mutational assays, can effectively uncover the molecular mechanisms underlying recently evolved functional novelty.…”

“…Our observations of the mechanisms underlying the evolution of MPH are consistent with the conclusions of several previous studies on protein evolution. Specifically, efficient MPH enzymes emerged rapidly, via a handful of genetic changes, by optimizing a promiscuous activity present in its ancestral state 3,4,27,35,42,43 . At the same time, epistasis between key adaptive mutations is prevalent, and acts to constrain the evolutionary pathways that were available 10,14,15,17,18,21,22 ; early mutation(s) played a permissive role by epistatically generating or enhancing the positive effect of later mutations.…”

“…Understanding the molecular mechanisms that underlie the evolution of new enzyme functions requires 1) the identification of a minimal set of genotypic changes that are necessary and sufficient to cause the emergence and optimization of a new function, and 2) an understanding of the biophysical molecular changes caused by these mutations that resulted in a novel function. Recent advances in experimental evolution and phylogenetically informed ancestral sequence reconstruction (ASR) aid in such endeavours by unveiling a set of adaptive mutations that allow substantial functional transitions [1][2][3][4][5][6][7][8][9] . Further biochemical, biophysical and structural characterization of enzyme variants enables us to uncover the molecular basis of the functional transition.…”