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

Yasuharu Takagi, Rachel E. Farrow, Neil Billington, A. Nagy, Christopher Batters, Yi Yang, James R. Sellers, Justin E. Molloy

Research output: Contribution to journalArticle

20 Citations (Scopus)

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 staircaselike 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.

Original languageEnglish
JournalProceedings of the National Academy of Sciences of the United States of America
Volume111
Issue number18
DOIs
Publication statusPublished - May 6 2014

Fingerprint

Myosins
Actin Cytoskeleton
Stroke
Optical Tweezers
Adenosine Triphosphatases
Actins
Leucine Zippers
Spindle Apparatus
Pseudopodia
Dimerization
Phagocytosis
Adenosine Diphosphate
Cell Movement
Electron Microscopy
Coloring Agents
Maintenance
Amino Acids
Proteins

Keywords

  • Actomyosin
  • Myosin X
  • Myosin-5a
  • Optical trapping
  • Stable single alpha-helix

ASJC Scopus subject areas

  • General

Cite this

Myosin-10 produces its power-stroke in two phases and moves processively along a single actin filament under low load. / Takagi, Yasuharu; Farrow, Rachel E.; Billington, Neil; Nagy, A.; Batters, Christopher; Yang, Yi; Sellers, James R.; Molloy, Justin E.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 111, No. 18, 06.05.2014.

Research output: Contribution to journalArticle

Takagi, Yasuharu ; Farrow, Rachel E. ; Billington, Neil ; Nagy, A. ; Batters, Christopher ; Yang, Yi ; Sellers, James R. ; Molloy, Justin E. / Myosin-10 produces its power-stroke in two phases and moves processively along a single actin filament under low load. In: Proceedings of the National Academy of Sciences of the United States of America. 2014 ; Vol. 111, No. 18.
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