Positive selection drives a correlation between non-synonymous/synonymous divergence and functional divergence

Motivation: Functional divergence among proteins is often assumed to be strongly influenced by natural selection, as inferred from the ratio of non-synonymous nucleotide divergence (d(N)) to synonymous nucleotide divergence (d(S)). That is, the more a mutation changes protein function, the more likely it is to be either selected against or selectively favored, and because the d(N)/d(S) ratio is a measure of natural selection, this ratio can be used to predict the degree of functional divergence (d(F)). However, these hypotheses have rarely been experimentally tested. Results: I present a novel method to address this issue, and demonstrate that divergence in bacteria-killing activity among animal antimicrobial peptides is positively correlated with the log of the d(N)/d(S) ratio. The primary cause of this pattern appears to be that positively selected substitutions change protein function more than neutral substitutions do. Thus, the d(N)/d(S) ratio is an accurate estimator of adaptive functional divergence.