Addition of two new acidic residues to the NH2 terminus of yeast actin increased the Vmax and catalytic efficiency of actin-activated ATPase activity of S-1 without affecting actin sliding velocity.
The number of actin NH2-terminal negative charges determines the efficiency of converting ATP hydrolysis energy to filament sliding in vitro.
We examined the effects of yeast actin NH2-terminal mutations on actomyosin interactions and the function of actin in vivo through measurements of actin-activated ATPase activity, cosedimentation with rabbit muscle myosin subfragment 1 (S-1), in vitro motility, and invertase secretion assays. As reported earlier (Cook, R. K., Blake, W., and Rubenstein, P. A. (1992) J. Biol. Chem. 267, 9430-9436), elimination of NH2-terminal acidic residues from yeast actin results in an increased actin bundling, decreased actin-activated S-1 ATPase, and complete inhibition of actin filament sliding over myosin. Here we show that the addition of 2 new acidic residues to the NH2 terminus of yeast actin increased the Vmax value and the catalytic efficiency of the actin-activated ATPase activity of S-1. However, the binding of actin to S-1 in the presence of ATP and the velocities of actin sliding over myosin in the in vitro motility assays were not affected by this mutation. Thus, the number of actin NH2-terminal negative charges is important for actin activation of myosin S-1 ATPase activity, while only a minimum number of acidic residues is required for actin sliding over myosin in vitro. The number of actin NH2-terminal negative charges therefore appears to determine the efficiency with which the energy from ATP hydrolysis is converted to filament sliding.
Cook et al. (Mon,) reported a other. Addition of 2 new acidic residues to the NH2 terminus of yeast actin was evaluated on Actin-activated ATPase activity of S-1, binding of actin to S-1, and velocities of actin sliding over myosin. Addition of two new acidic residues to the NH2 terminus of yeast actin increased the Vmax and catalytic efficiency of actin-activated ATPase activity of S-1 without affecting actin sliding velocity.