The Chemical Mechanism of Myosin-I: Implications for Actin-based Motility and the Evolution of the Myosin Family of Motor Proteins

Abstract

The Acanthamoeba myosin-IA and myosin-IB molecular motors bind to membranes, so they may produce the force to move organdies and membranes along actin filaments. We have determined the rate constants for the actin-activated myosin-I ATPase by pre-steady state kinetic analysis. ATP binds rapidly to myosin-I and dissociates the enzyme from actin filaments at a rate >500 s^-1. Myosin-I hydrolyzes ATP to ADP and inorganic phosphate (Pi) at 20-50 s^-1. Phosphate dissociation is the rate limiting step in the ATPase cycle, 0.01 s^-1 for myosin-I alone and at 10 s^-1 when myosin-I is bound to actin filaments. ADP dissociation is rapid. Phosphorylation controls the ATPase cycle by increasing the rate of phosphate release from myosin-I bound to actin. At steady state the major species are myosin-ATP and myosin-ADP-P_i, which rapidly bind to and dissociate from actin filaments. During the ATPase cycle myosin-I binds so weakly to actin filaments that it cannot support processive movement like kinesin, unless several motors cluster together on a membrane or actin filament. These properties of the enzyme emphasize the importance of characterizing mechanisms that promote the self-association of myosin-I isoforms at specific binding sites in cells.