ADP markedly inhibited the actin-activated MgATPase activity and in vitro motility of mouse myosin V, with an ADP dissociation rate of ~11.5 s(-1) indicating it is close to the rate-limiting step.
The high affinity for actin and slow rate of ADP release allow the myosin V head to remain attached to actin for a large fraction of its ATPase cycle, enabling actin filament movement by only a few molecules.
Mouse myosin V is a two-headed unconventional myosin with an extended neck that binds six calmodulins. Double-headed (heavy meromyosin-like) and single-headed (subfragment 1-like) fragments of mouse myosin V were expressed in Sf9 cells, and intact myosin V was purified from mouse brain. The actin-activated MgATPase of the tissue-purified myosin V, and its expressed fragments had a high V(max) and a low K(ATPase). Calcium regulated the MgATPase of intact myosin V but not of the fragments. Both the MgATPase activity and the in vitro motility were remarkably insensitive to ionic strength. Myosin V and its fragments translocated actin at very low myosin surface densities. ADP markedly inhibited the actin-activated MgATPase activity and the in vitro motility. ADP dissociated from myosin V subfragment 1 at a rate of about 11.5 s(-1) under conditions where the V(max) was 3.3 s(-1), indicating that, although not totally rate-limiting, ADP dissociation was close to the rate-limiting step. The high affinity for actin and the slow rate of ADP release helps the myosin head to remain attached to actin for a large fraction of each ATPase cycle and allows actin filaments to be moved by only a few myosin V molecules in vitro.
Wang et al. (Tue,) reported a other. ADP and Ionic Strength was evaluated on Kinetic and motile properties (MgATPase activity, in vitro motility, ADP dissociation rate). ADP markedly inhibited the actin-activated MgATPase activity and in vitro motility of mouse myosin V, with an ADP dissociation rate of ~11.5 s(-1) indicating it is close to the rate-limiting step.