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Chemomechanical coupling in F1-ATPase revealed by simultaneous observation of nucleotide kinetics and rotation

Nature Structural & Molecular Biology volume 11, pages 142–148 (2004)Cite this article

Abstract

F1-ATPase is a rotary molecular motor in which unidirectional rotation of the central γ subunit is powered by ATP hydrolysis in three catalytic sites arranged 120° apart around γ. To study how hydrolysis reactions produce mechanical rotation, we observed rotation under an optical microscope to see which of the three sites bound and released a fluorescent ATP analog. Assuming that the analog mimics authentic ATP, the following scheme emerges: (i) in the ATP-waiting state, one site, dictated by the orientation of γ, is empty, whereas the other two bind a nucleotide; (ii) ATP binding to the empty site drives an ∼80° rotation of γ; (iii) this triggers a reaction(s), hydrolysis and/or phosphate release, but not ADP release in the site that bound ATP one step earlier; (iv) completion of this reaction induces further ∼40° rotation.

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Figure 1: Experimental system.
Figure 4: F1 rotation driven by Cy3-ATP.
Figure 2: Simultaneous observation of F1 rotation and Cy3-ATP binding.
Figure 3: Timing between a rotational step of the γ subunit (upper panels) and binding or release of Cy3-nucleotide (lower panels).
Figure 5: Mechanism of F1 rotation.

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Acknowledgements

We thank R. Yasuda, K. Adachi and H. Itoh for technical assistance and critical discussion; R. Nakamori and Y. Funamoto for technical assistance; D.R. Trentham for initiation of our collaboration; T. Masaike, B. Brenner and H. Kojima for critical discussion; M. Shio, K. Abe and I. Sase for the microscope techniques; and M. Uno and H. Umezawa for management of laboratories and collaboration. This work was supported in part by grants-in-aid from the Ministry of Education, Culture, Sports, Science and Technology of Japan, and by a Core Research for Evolutional Science and Technology (CREST) grant.

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Author notes

  1. Takayuki Nishizaka

    Present address: Department of Physics, Gakushuin University, Mejiro 1-5-1, Toshima-ku, Tokyo, 171-8588, Japan

Authors and Affiliations

  1. Kansai Advanced Research Center, Protein Biophysics Group, Iwaoka 588-2, Nishi-ku, 651-2492, Kobe, Japan

    Takayuki Nishizaka, Kazuhiro Oiwa & Shigeki Kimura

  2. Precursory Research for Embryonic Science and Technology (PRESTO), Japan Science and Technology Agency, Honcho 4-1-8, Kawaguchi, 332-0012, Saitama, Japan

    Takayuki Nishizaka

  3. Institute of Industrial Science, University of Tokyo, Tokyo, 153-8505, Japan

    Hiroyuki Noji

  4. Chemical Resources Laboratory, Tokyo Institute of Technology, Yokohama, 226-8503, Japan

    Eiro Muneyuki & Masasuke Yoshida

  5. ATP system, Exploratory Research for Advanced Technology (ERATO), Japan Science and Technology Agency, Yokohama, 226-0026, Japan

    Masasuke Yoshida

  6. Center for Integrative Bioscience, Okazaki National Research Institutes, Myodaiji, 444-8585, Okazaki, Japan

    Kazuhiko Kinosita Jr

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Correspondence to Takayuki Nishizaka.

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Nishizaka, T., Oiwa, K., Noji, H. et al. Chemomechanical coupling in F1-ATPase revealed by simultaneous observation of nucleotide kinetics and rotation. Nat Struct Mol Biol 11, 142–148 (2004). https://doi.org/10.1038/nsmb721

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