Genetic constraints on protein evolution

Abstract

Evolution requires the generation and optimization of new traits ("adaptation") and involves the selection of mutations that improve cellular function. These mutations were assumed to arise by selection of neutral mutations present at all times in the population. Here we review recent evidence that indicates that deleterious mutations are more frequent in the population than previously recognized and that these mutations play a significant role in protein evolution through continuous positive selection. Positively selected mutations include adaptive mutations, i.e. mutations that directly affect enzymatic function, and compensatory mutations, which suppress the pleiotropic effects of adaptive mutations. Compensatory mutations are by far the most frequent of the two and would allow potentially adaptive but deleterious mutations to persist long enough in the population to be positively selected during episodes of adaptation. Compensatory mutations are, by definition, context-dependent and thus constrain the paths available for evolution. This provides a mechanistic basis for the examples of highly constrained evolutionary landscapes and parallel evolution reported in natural and experimental populations. The present review article describes these recent advances in the field of protein evolution and discusses their implications for understanding the genetic basis of disease and for protein engineering in vitro.

Fig. 1. Fidelity versus overall activity of active mutants of E. coli DNA Polymerase I

69 active mutants of E. coli DNA polymerase I were tested for fidelity and activity. Replication fidelity (X-axis) was established as the reversion frequency of a β-lactamase gene encoding an ochre stop codon. Polymerase activity (Y-axis) was determined by incorporation of radio labeled dTMP. Values have been normalized to wild-type values, with wild-type controls occupying the position 1,1. Error bars represent standard error p<0.05 ((Camps et al., 2003), with permission).

Fig. 2. Substitutability of the polymerase domain of E. coli DNA Polymerase I

A random library of E. coli DNA Polymerase I was selected for activity using functional complementation of JS200 cells, a strain encoding a temperature-sensitive DNA polymerase I gene (recA718 PolA12). For each position, a substitutability index was calculated based on the number of tolerated mutations. The crystal structure of the Pol I Klenow fragment (pdb file 1KLN) was colored according to a gradient, going from blue (lowest) to red (highest) tolerance for mutations. The 5′ exonuclease domain of the Klenow fragment, which did not undergo mutagenesis, is colored in grey. (Modified from (Loh et al., 2007b)).

Fig. 3. Effects of mutation fitness on protein evolution

The lifespan of mutations in the population depends on their effects on the fitness of the organism. Neutral mutations, having little effect on fitness, accumulate and persist for a long time in the population (although they may be eliminated via recombination). Deleterious mutations are rapidly eliminated by purifying selection. Adaptive mutations often have pleiotropic deleterious effects because they affect residues that are important for function. In this case, compensatory mutations rapidly arise, typically in bursts of multiple mutations, allowing adaptive mutations to persist longer in the population. The presence of compensatory mutations has an epistatic effect that generally precludes a return to the ancestral sequence. In a genome with a significant history of positive selection such as the human genome, the final contribution of positive and negative selection likely reflects a trade-off between persistence in the population (neutral mutations) and the need to generate novel traits (adaptive mutations). Some adaptive mutations with potential deleterious effects remain in the population at low frequencies because compensatory mutations delay purifying selection.