Neutral genetic drift can alter promiscuous protein functions, potentially aiding functional evolution

<jats:title>Abstract</jats:title> <jats:sec> <jats:title>Background</jats:title> <jats:p>Many of the mutations accumulated by naturally evolving proteins are neutral in the sense that they do not significantly alter a protein's ability to perform its primary biological function. However, new protein functions evolve when selection begins to favor other, "promiscuous" functions that are incidental to a protein's original biological role. If mutations that are neutral with respect to a protein's primary biological function cause substantial changes in promiscuous functions, these mutations could enable future functional evolution.</jats:p> </jats:sec> <jats:sec> <jats:title>Results</jats:title> <jats:p>Here we investigate this possibility experimentally by examining how cytochrome P450 enzymes that have evolved neutrally with respect to activity on a single substrate have changed in their abilities to catalyze reactions on five other substrates. We find that the enzymes have sometimes changed as much as four-fold in the promiscuous activities. The changes in promiscuous activities tend to increase with the number of mutations, and can be largely rationalized in terms of the chemical structures of the substrates. The activities on chemically similar substrates tend to change in a coordinated fashion, potentially providing a route for systematically predicting the change in one activity based on the measurement of several others.</jats:p> </jats:sec> <jats:sec> <jats:title>Conclusion</jats:title> <jats:p>Our work suggests that initially neutral genetic drift can lead to substantial changes in protein functions that are not currently under selection, in effect poising the proteins to more readily undergo functional evolution should selection favor new functions in the future.</jats:p> </jats:sec> <jats:sec> <jats:title>Reviewers</jats:title> <jats:p>This article was reviewed by Martijn Huynen, Fyodor Kondrashov, and Dan Tawfik (nominated by Christoph Adami).</jats:p> </jats:sec>