The conformational landscape of fold-switcher KaiB is tuned to the circadian rhythm timescale.

One can conceptualize KaiB as an on-off switch to regulate circadian rhythms in bacteria, where the "On state" is the Fold-switched state that binds KaiC and other proteins, and the "Off state" is the Ground state. Our work exemplifies how evolution tuned the kinetics of interconversion to align with the hour-long timescale of its biological function. The Ground state is dramatically destabilized at cold temperatures, and the system contains an alternate "off" conformation that exchanges with the primary "off" conformation at faster timescales than the rate-limiting step. Our findings demonstrate a simple principle for evolving a protein switch: one part of a protein domain remains stably folded to serve as a scaffold for the rest of the protein to re-fold.