期刊
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
卷 111, 期 18, 页码 E1833-E1842出版社
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1320122111
关键词
actomyosin; stable single alpha-helix; optical trapping; myosin X; myosin-5a
资金
- National Natural Science Foundation of China [31270819]
- NHLBI [HL004243 12]
- Medical Research Council [U1175.70592]
- Medical Research Council [1106254] Funding Source: researchfish
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 similar to 50 nm. In vitro motility assays show that myosin10 moves actin filaments smoothly with a velocity of similar to 310 nm/s. Steady-state and transient kinetic analysis of the ATPase cycle shows that the ADP release rate (similar to 13 s(-1)) is similar to the maximum ATPase activity (similar to 12-14 s(-1)) and therefore contributes to rate limitation of the enzymatic cycle. Single molecule optical tweezers experiments show that under intermediate load (similar to 0.5 pN), myosin-10 interacts intermittently with actin and produces a power stroke of similar to 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 similar to 35-nm steps per binding interaction. We discuss the implications of this load-dependent processivity of myosin-10 as a filopodial transport motor.
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