SUMMARY
The extent to which chance and contingency shaped the sequence outcomes of protein evolution is largely unknown. To directly characterize the causes and consequences of chance and contingency, we combined directed evolution with ancestral protein reconstruction. By repeatedly selecting a phylogenetic series of ancestral proteins in the B-cell lymphoma-2 family to evolve the same protein-protein interaction specificities that existed during history, we show that contingency and chance interact to make sequence evolution almost entirely unpredictable over the timescale of metazoan evolution. At any historical moment, multiple sets of mutations can alter or maintain specificity, and chance decides which ones occur. Contingency arises because historical sequence substitutions epistatically altered which mutations are compatible with new or ancestral functions. Evolutionary trajectories launched from different ancestors therefore lead to dramatically different outcomes over phylogenetic time, with virtually no mutations occurring repeatedly in distantly related proteins, even under identical selection conditions.
Competing Interest Statement
J.P. and B.C.D. have a patent on the proximity-dependent split RNAP technology used in this work. The content is solely the responsibility of the authors and the funders had no input on the study design, analysis, or conclusions.
Footnotes
↵* email: pujy{at}uchicago.edu, joet1{at}uchicago.edu, dickinson{at}uchicago.edu