A point mutation in the regulatory light chain reduces the step size of skeletal muscle myosin

doi:10.1073/pnas.0401699101

A point mutation in the regulatory light chain reduces the step size of skeletal muscle myosin

Department of Molecular Physiology and Biophysics, University of Vermont, Burlington, VT 05405

Edited by James A. Spudich, Stanford University School of Medicine, Stanford, CA, and approved May 26, 2004 (received for review March 10, 2004)

Abstract

Current evidence favors the theory that, when the globular motor domain of myosin attaches to actin, the light chain binding domain or “lever arm” rotates, and thereby generates movement of actin filaments. Myosin is uniquely designed for such a role in that a long α-helix (≈9 nm) extending from the C terminus of the catalytic core is stabilized by two calmodulin-like molecules, the regulatory light chain (RLC) and the essential light chain (ELC). Here, we introduce a single-point mutation into the skeletal myosin RLC, which results in a large (≈50%) reduction in actin filament velocity (V _actin) without any loss in actin-activated MgATPase activity. Single-molecule analysis of myosin by optical trapping showed a comparable 2-fold reduction in unitary displacement or step size (d), without a significant change in the duration of the strongly attached state (τ_on) after the power stroke. Assuming that V _actin ≈ d/τ_on, we can account for the change in velocity primarily by a change in the step size of the lever arm without incurring any change in the kinetic properties of the mutant myosin. These results suggest that a principal role for the many light chain isoforms in the myosin II class may be to modulate the flexural rigidity of the light chain binding domain to maximize tension development and movement during muscle contraction.

Footnotes

↵ † To whom correspondence should be addressed at: Department of Molecular Physiology and Biophysics, University of Vermont, 128 Health Science Research Facility, Burlington, VT 05405-0068. E-mail: lowey{at}physiology.med.uvm.edu.
↵ * Present address: Whitehead Institute for Biomedical Research, Cambridge, MA 02142.
This paper was submitted directly (Track II) to the PNAS office.
Abbreviations: S1, myosin subfragment-1; RLC, regulatory light chain; ELC, essential light chain; LC1 and LC3, light chain isoforms of ELC; MV, mean-variance.