Intrinsically disordered proteins (IDPs) play key roles in signaling and regulation. Many IDPs undergo folding upon binding to their targets. We have proposed that coupled folding and binding of IDPs generally follow a dock-and-coalesce mechanism, whereby a segment of the IDP, through diffusion, docks to its cognate subsite and, subsequently, the remaining segments coalesce around their subsites. Here, by a combination of experiment and computation, we determined the precise form of dock-and-coalesce operating in the association between the intrinsically disordered GTPase binding domain (GBD) of the Wiskott-Aldrich Syndrome protein (WASP) and the Cdc42 GTPase. The association rate constants (ka) were measured by stopped-flow fluorescence under various solvent conditions. ka reached 107 M-1s-1 at physiological ionic strength and had a strong salt dependence, suggesting that an electrostatically enhanced, diffusion-controlled docking step may be rate-limiting. Our computation, based on the transient-complex theory, identified the N-terminal basic region of the GBD as the docking segment. However, several other changes in solvent conditions provided strong evidence that the coalescing step also contributed to determining the magnitude of ka. Addition of glucose and TFE and an increase in temperature all produced ka values much higher than expected from the effects on the docking rate alone. Conversely, addition of urea led to ka values much lower than expected if only the docking rate was affected. These results all pointed to ka being approximately two thirds of the docking rate constant under physiological solvent conditions. The dock-and-coalesce mechanism allows WASP and other IDPs to code electrostatic complementarity into the docking segment to gain binding speed and use additional interactions formed by the coalescing segments to reinforce binding affinity.

Dock-and-coalesce mechanism for the association of a WASP disordered region with the Cdc42 GTPase

254th National Meeting (2017)

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