Force Dependent Stepping Kinetics of Myosin-V

¹ Technical University Munich
² J. Stefan Institute

^* To whom correspondence should be addressed. E-mail: mrief{at}ph.tum.de.

Submitted on September 27, 2004
Revised on October 29, 2004
Accepted on 3 March 2005

	Abstract

Myosin-V is a processive two-headed actin-based motor protein involved in many intra-cellular transport processes. A key question for understanding myosin-V function and the communication between its two heads is its behavior under load. Since in vivo myosin-V co-localizes with other much stronger motors like kinesins its behavior under super-stall forces is especially relevant. We used optical tweezers with a long-range force feedback to study myosin-V motion under controlled external forward and backward loads over its full run length. We find the mean step size remains constant at ~36 nm over a wide range of forces from 5 pN forward to 1.5 pN backward load. We also find two force dependent transitions in the chemo-mechanical cycle. The slower ADP release is rate-limiting at low loads and depends only weakly on force. The faster rate depends stronger on force. The stronger force-dependence suggests this rate represents the diffusive search of the leading head for its binding site. In contrast to kinesin motors, myosin-V run length is essentially independent of force between 5 pN of forward to 1.5 pN of backward load. At super-stall forces of 5 pN, we observe continuous backward stepping of myosin-V indicating a force-driven reversal of the power-stroke is possible.

Key Words: mechanics, molecular motors, optical, single molecule experiments, tweezers

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