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J. Biol. Chem., Vol. 275, Issue 47, 37167-37172, November 24, 2000
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From the Department of Molecular Physiology and Biophysics,
University of Vermont, Burlington, Vermont 05405
Structural data led to the proposal that the
molecular motor myosin moves actin by a swinging of the light chain
binding domain, or "neck." To test the hypothesis that the neck
functions as a mechanical lever, smooth muscle heavy meromyosin (HMM)
mutants were expressed with shorter or longer necks by either deleting or adding light chain binding sites. The mutant HMMs were characterized kinetically and mechanically, with emphasis on measurements of unitary
displacements and forces in the laser trap assay. Two shorter necked
constructs had smaller unitary step sizes and moved actin more slowly
than WT HMM in the motility assay. A longer necked construct that
contained an additional essential light chain binding site exhibited a
1.4-fold increase in the unitary step size compared with its control.
Kinetic changes were also observed with several of the constructs. The
mutant lacking a neck produced force at a somewhat reduced level, while
the force exerted by the giraffe construct was higher than control. The single molecule displacement and force data support the hypothesis that
the neck functions as a rigid lever, with the fulcrum for movement and
force located at a point within the motor domain.
The Light Chain Binding Domain of Expressed Smooth Muscle Heavy
Meromyosin Acts as a Mechanical Lever*
,
,
*
This work was supported by National Institutes of Health
Grant HL54568 (to K. T. and D. W.) and the Totman Fund for
Cerebrovascular Research (to D. W.).The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
Present address: Dept. of Biomedical Engineering, University of
Virginia, Charlottesville, VA 22908.
§
Whitehead Institute for Biomedical Research, Massachusetts
Institute of Technology, Cambridge, MA 02142.
¶
Dept. of Biology, San Diego State University, San Diego, CA 92182.
Dept. of Molecular, Cellular, and Developmental Biology, Yale
University, New Haven, CT 06511.
**
To whom correspondence should be addressed: Molecular Physiology
and Biophysics, University of Vermont, Burlington, VT 05405. Tel.:
802-656-8750; Fax: 802-656-0747; E-mail:
trybus@salus.med.uvm.edu.
Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
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