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Myosin VI walks hand-over-hand along actin

Nature Structural & Molecular Biology volume 11, pages 884–887 (2004)Cite this article

Abstract

Myosin VI is a molecular motor that can walk processively on actin filaments with a 36-nm step size. The walking mechanism of myosin VI is controversial because it takes very large steps without an apparent lever arm of required length. Therefore, myosin VI is argued to be the first exception to the widely established lever arm theory. It is therefore critical to directly demonstrate whether this motor walks hand-over-hand along actin despite its short lever arm. Here, we follow the displacement of a single myosin VI head during the stepping process. A single head is displaced 72 nm during stepping, whereas the center of mass previously has been shown to move 36 nm. The most likely explanation for this result is a hand-over-hand walking mechanism. We hypothesize the existence of a flexible element that would allow the motor to bridge the observed 72-nm distance.

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Figure 1: Proposed hand-over-hand mechanism for myosin VI.
Figure 2: Run-length distribution of single fluorescently labeled myosin VI molecules in the single molecule assays.
Figure 3: Staircases of three different fluorescently labeled myosin VI molecules.
Figure 4: Step-size histogram of single fluorescently labeled myosin VI molecules.

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Acknowledgements

We thank B. Spink for assistance in the creation of the construct and H.L. Sweeney for providing the plasmid for myosin VI expression. We thank F. Müller-Planitz for assistance in MatLab programming, helpful discussions and comments on the manuscript. We also thank D.A. Altman and A. Dunn for helpful discussions and comments on the manuscript. This work was supported by Boehringer Ingelheim Fonds to Z.Ö., US National Institutes of Health grant GM33289 to J.A.S. and a Burroughs Wellcome Career Award at the Scientific Interface to R.S.R.

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Authors and Affiliations

  1. Stanford University School of Medicine, Beckman Center, 279 Campus Drive, Stanford, 94305-5307, California, USA

    Zeynep Ökten, L Stirling Churchman, Ronald S Rock & James A Spudich

  2. Department of Biology, Chemistry and Pharmacy, Freie Universität Berlin, Taku Strasse 3, Institut für Chemie, Berlin, 14195, Germany

    Zeynep Ökten

  3. Department of Physics, 382 Via Pueblo Mall, Stanford University, Stanford, 94305-4060, California, USA

    L Stirling Churchman

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  1. Zeynep Ökten
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Correspondence to James A Spudich.

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Supplementary information

Supplementary Fig. 1

Inchworm model. (PDF 181 kb)

Supplementary Fig. 2

Titration curve. (PDF 20 kb)

Supplementary Fig. 3

Labeling specificity. (PDF 26 kb)

Supplementary Fig. 4

Single photon localization. (PDF 143 kb)

Supplementary Video 1

An example of a myosin VI motor that is walking close to the evanescent field can be seen at the top of the image. This molecule appears bright as opposed to the molecules walking on fully suspended filaments (see Supplementary Video 2). Most molecules do not walk this close to the surface. Superimposed and artificially colored images of Cy5 labeled Myosin VI (green) and actin (red). (AVI 1733 kb)

Supplementary Video 2

An example of a myosin VI motor that is walking on a fully suspended filament can be seen at the top right hand corner. Most Cy5 labeled myosin VI molecules on the fully suspended filaments far away from the evanescent field appear too dim to yield the required resolution for measuring the step size. (AVI 1608 kb)

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Ökten, Z., Churchman, L., Rock, R. et al. Myosin VI walks hand-over-hand along actin. Nat Struct Mol Biol 11, 884–887 (2004). https://doi.org/10.1038/nsmb815

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