Myosin-10 produces its power-stroke in two phases and moves processively along a single actin filament under low load.

ABSTRACT: Myosin-10 is an actin-based molecular motor that participates in essential intracellular processes such as filopodia formation/extension, phagocytosis, cell migration, and mitotic spindle maintenance. To study this motor protein's mechano-chemical properties, we used a recombinant, truncated form of myosin-10 consisting of the first 936 amino acids, followed by a GCN4 leucine zipper motif, to force dimerization. Negative-stain electron microscopy reveals that the majority of molecules are dimeric with a head-to-head contour distance of ?50 nm. In vitro motility assays show that myosin-10 moves actin filaments smoothly with a velocity of ?310 nm/s. Steady-state and transient kinetic analysis of the ATPase cycle shows that the ADP release rate (?13 s(-1)) is similar to the maximum ATPase activity (?12-14 s(-1)) and therefore contributes to rate limitation of the enzymatic cycle. Single molecule optical tweezers experiments show that under intermediate load (?0.5 pN), myosin-10 interacts intermittently with actin and produces a power stroke of ?17 nm, composed of an initial 15-nm and subsequent 2-nm movement. At low optical trap loads, we observed staircase-like processive movements of myosin-10 interacting with the actin filament, consisting of up to six ?35-nm steps per binding interaction. We discuss the implications of this load-dependent processivity of myosin-10 as a filopodial transport motor.

SUBMITTER: Takagi Y 

PROVIDER: S-EPMC4020102 | biostudies-literature | 2014 May

REPOSITORIES: biostudies-literature