Velocity of myosin-based actin sliding depends on attachment and detachment kinetics and reaches a maximum when myosin-binding sites on actin saturate

Travis J Stewart,Vidya Murthy,Sam P Dugan,Josh E Baker

doi:10.1016/j.jbc.2021.101178

Travis J Stewart, Vidya Murthy + Show 2 more

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https://doi.org/10.1016/j.jbc.2021.101178

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Abstract

Molecular motors such as kinesin and myosin often work in groups to generate the directed movements and forces critical for many biological processes. Although much is known about how individual motors generate force and movement, surprisingly, little is known about the mechanisms underlying the macroscopic mechanics generated by multiple motors. For example, the observation that a saturating number, N, of myosin heads move an actin filament at a rate that is influenced by actin–myosin attachment and detachment kinetics is accounted for neither experimentally nor theoretically. To better understand the emergent mechanics of actin–myosin mechanochemistry, we use an in vitro motility assay to measure and correlate the N-dependence of actin sliding velocities, actin-activated ATPase activity, force generation against a mechanical load, and the calcium sensitivity of thin filament velocities. Our results show that both velocity and ATPase activity are strain dependent and that velocity becomes maximized with the saturation of myosin-binding sites on actin at a value that is 40% dependent on attachment kinetics and 60% dependent on detachment kinetics. These results support a chemical thermodynamic model for ensemble motor mechanochemistry and imply molecularly explicit mechanisms within this framework, challenging the assumption of independent force generation.

Highlights

Molecular motors such as myosin and kinesin often work in groups to perform diverse biological functions such as vesicle transport, cell division, wound healing, and muscle contraction [1,2,3]
From these studies we know that the basic molecular mechanism for muscle contraction involves a discrete displacement of an actin filament generated by a myosin structural change induced by strong actin binding
In an independent force model (Fig. 1B, top) actin sliding velocities are described in terms of the kinetics and mechanics of an individual myosin head, Vmax = d⋅kdet

Summary

Introduction

Molecular motors such as myosin and kinesin often work in groups to perform diverse biological functions such as vesicle transport, cell division, wound healing, and muscle contraction [1,2,3]. Researchers have focused more on reductionist approaches using electron microscopy, X-ray diffraction, spectroscopic techniques, stopped flow kinetics, crystal structures, and single molecule mechanics measurements [22,23,24,25,26,27,28,29] to provide detailed structural, biochemical, and mechanical descriptions of the molecules involved in muscle contraction From these studies we know that the basic molecular mechanism for muscle contraction involves a discrete displacement of an actin filament generated by a myosin structural change induced by strong actin binding. A single strongly bound myosin head is assumed to prevent the working step of other myosin heads from moving actin and transmitting forces between them, and movement is limited by detachment of the resistive myosin head

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