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Swing of the lever arm of a myosin motor at the isomerization and phosphate-release steps

Nature volume 396, pages 380–383 (1998)Cite this article

Abstract

In muscle, the myosin head (‘crossbridge’) performs the ‘working stroke’, in which ATP is hydrolysed to generate the sliding of actin and myosin filaments. The myosin head consists of a globular motor domain and a long lever-arm domain. The ‘lever-arm hypothesis’1,2,3,4,5 predicts that during the working stroke, the lever-arm domain tilts against the motor domain, which is bound to actin in a fixed orientation. To detect this working stroke in operation, we constructed fusion proteins by connecting Aequorea victoria green fluorescent protein and blue fluorescent protein6,7,8 to the amino and carboxyl termini of the motor domain of myosin II of Dictyostelium discoideum, a soil amoeba, and measured the fluorescence resonance energy transfer between the two fluorescent proteins. We show here that the carboxy-terminal fluorophore swings at the isomerization step of the ATP hydrolysis cycle, and then swings back at the subsequent step in which inorganic phosphate is released, thereby mimicking the swing of the lever arm. The swing at the phosphate-release step may correspond to the working stroke, and the swing at the isomerization step to the recovery stroke.

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Figure 1: Diagrams of the domain of chimaeras made from the incorporation of fluorescent proteins into S1dC.
Figure 2: The relationship of FRET emission spectra in chimaeric S1dC to the hydrolysis of ATP.
Figure 3: Time course of changes in the FRET-induced emission from GFP.
Figure 4: Stereo views of a complex of an actin filament and a myosin head, prepared by using program O (ref. 26).

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Acknowledgements

We thank K. Oiwa for Cy3 nucleotides, M. Miki for discussions about the FRET experiments, and M. Steward, C.Bagshaw and P. Kuhlman for critically reading the manuscript.

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Authors and Affiliations

  1. Department of Life Sciences, School of Arts and Sciences, University of Tokyo, Komaba 3-8-1, Tokyo, 153, Japan

    Yoshikazu Suzuki, Reiko Ohkura & Kazuo Sutoh

  2. Department of Physics, School of Science, University of Tokyo, Hongo 7-3-1, Tokyo, 113, Japan

    Takuo Yasunaga & Takeyuki Wakabayashi

Authors
  1. Yoshikazu Suzuki
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  2. Takuo Yasunaga
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  3. Reiko Ohkura
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  4. Takeyuki Wakabayashi
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  5. Kazuo Sutoh
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Correspondence to Kazuo Sutoh.

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Suzuki, Y., Yasunaga, T., Ohkura, R. et al. Swing of the lever arm of a myosin motor at the isomerization and phosphate-release steps. Nature 396, 380–383 (1998). https://doi.org/10.1038/24640

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