Investigating processive molecular motor myosin-V under load using optical tweezers

Mojca Vilfan, Anabel Clemen, Johann Jaud, Matthias Rief

J. Stefan Institute, Ljubljana, Slovenia; Technische Universität München, Germany

Myosin-V is a processive two-headed motor protein associated to actin. It is involved in many transport processes in the cell. A key question for understanding myosin-V function and the communication between the 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. To gain insight into the chemo-mechanical cycle under load, we used optical tweezers with long range force feedback. This ensured controlled external forward and backward loads. We find that the mean step size remains constant at ~36 nm (corresponding to the periodicity of the actin filament) over a wide range of loads from 5 pN forward to 1.5 pN backward load. We also find two force dependent transitions in the chemo-mechanical cycle. The slower rate, previously identified as 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. At super-stall forces of 5 pN, we observe continuous backward stepping.

Email: mojca.vilfan@ijs.si

Address: Mojca Vilfan, J. Stefan Institute, Jamova 39, SI-1000 Ljubljana, Slovenia