The bound nucleotide and the rates of transition between states, i.e., ATP → ADP+P i ⇌ ADP regulate actin filament assembly and aging. Experiments showed that the rate of inorganic phosphate (P i ) release post-hydrolysis is almost 300× faster at the ends of actin filaments than interior subunits. Two hypotheses have been proposed to explain the physical origin of this difference with the rate-determining step either the transient opening of the R177 back door gate or dissociation of P i from Mg 2+ in the active site. To provide a cohesive mechanistic understanding of P i release across the filament, we performed extensive molecular dynamics simulations of P i release from the pointed end, the barbed end, and the interior subunits of an ADP-Pi actin filament. Using enhanced sampling methods to calculate the transition rate from contact ion pair (CIP; Mg 2+ -P i − ) to solvent-separated ion pair (SSIP), we show how the dissociation of P i from Mg 2+ can account for the experimentally measured higher rate of P i release from filament ends. Specifically, an analysis of the subunit active sites in the three locations revealed how the weakly hydrated active site in interior subunits stabilizes the CIP, which significantly slows its dissociation compared to the filament ends. In addition, terminal subunits favor egress pathways other than the R177 backdoor gate, highlighting the complexity of the release mechanism and its dependence on subtle differences in subunit conformations. In addition, we investigated how the natural product jasplakinolide inhibits P i release to provide a comprehensive understanding of how the structure of actin subunits modulates the rate of P i release, with implications for other ATPases and GTPases.
Herman et al. (Sun,) studied this question.