Willy Wriggers and Klaus Schulten.
Investigating a back door mechanism of actin phosphate release by
steered molecular dynamics.
PROTEINS: Structure, Function, and Genetics, 35:262-273, 1999.
WRIG99In actin-based cell motility, phosphate (Pi) release after ATP hydrolysis is an essential biochemical process, but the actual pathway of Pi dissociation from actin is not well understood. We report a series of molecular dynamics simulations that induce the unbinding of Pi from actin. After cleavage from ATP, the singly protonated phosphate (HPO4 2-) rotates about the ADP-associated Ca2+ ion, turning away from the negatively charged ADP towards the putative exit near His73. To reveal the microscopic processes underlying the unbinding of Pi, adhesion forces were measured when pulling the substrate out of its binding pocket. The results suggest that the dissociation from the divalent cation is the rate-limiting step in Pi release. Protonation of HPO4 2- to H2PO4- lowers the electrostatic barrier during Pi liberation from the ion. The simulations revealed a propensity of charged His73+ to form a salt bridge with HPO4 2-, but not with H2PO4-. His73 stabilizes HPO4 2- and, thereby, inhibits rapid Pi dissociation from actin. Arg177 remains attached to Pi along the putative back door pathway, suggesting a shuttle function which facilitates the transport of Pi to a binding site on the protein surface.
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