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Chemomechanical coupling of human mitochondrial F1-ATPase motor

Nature Chemical Biology volume 10, pages 930–936 (2014)Cite this article

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Abstract

The rotary motor enzyme F1-ATPase (F1) is a catalytic subcomplex of FoF1-ATP synthase that produces most of the ATP in respiring cells. Chemomechanical coupling has been studied extensively for bacterial F1 but very little for mitochondrial F1. Here we report ATP-driven rotation of human mitochondrial F1. A rotor-shaft γ-subunit in the stator α3β3 ring rotates 120° per ATP with three catalytic steps: ATP binding to one β-subunit at 0°, inorganic phosphate (Pi) release from another β-subunit at 65° and ATP hydrolysis on the third β-subunit at 90°. Rotation is often interrupted at 90° by persistent ADP binding and is stalled at 65° by a specific inhibitor azide. A mitochondrial endogenous inhibitor for FoF1-ATP synthase, IF1, blocks rotation at 90°. These features differ from those of bacterial F1, in which both ATP hydrolysis and Pi release occur at around 80°, demonstrating that chemomechanical coupling angles of the γ-subunit are tuned during evolution.

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Figure 1: ATP-driven rotation of human F1.
Figure 2: Catalytic events that occur in dwells.
Figure 3: Angular positions for ATP hydrolysis and Pi releasing or binding.
Figure 4: Rotation of human F1 in high [ATP] plus low [ATPγS].
Figure 5: Angular position of human F1 in inhibitory states.
Figure 6: A model of rotary catalysis of human F1 and assignment of crystal structures in the rotation scheme of human F1.

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Acknowledgements

We thank our colleagues B. Feniouk, N. Taniguchi and K. Sugawara (Japan Science and Technology Agency), S. Furuike (Osaka Medical college), K. Kinosita Jr., N. Soga and K. Adachi (Waseda University) and T. Hisabori (Tokyo Institute of Technology) for their valuable discussions. S. Furuike helped T.S. in setting up the center-shielded dark-field microscopic system. We thank K. Adachi (Waseda University) for the kind gift of the software program Center-5. The cDNA library from human lung cells was a kind gift from F. Ishikawa of Kyoto University and M. Fujikawa of the Japan Science and Technology Agency. This work was supported in part by grant-in-aid for Scientific Research from the Japan Society for the Promotion of Science (grant numbers 24570149 (T.S.) and 23227006 (M.Y.)).

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Authors and Affiliations

  1. Faculty of Science and Engineering, Waseda University, Shinjuku-ku, Tokyo, Japan

    Toshiharu Suzuki & Masasuke Yoshida

  2. ATP Synthesis Regulation Project, International Research Project (ICORP), Japan Science and Technology Corporation (JST), Miraikan, Koto-ku, Tokyo, Japan

    Toshiharu Suzuki, Kazumi Tanaka, Chiaki Wakabayashi, Ei-ichiro Saita & Masasuke Yoshida

  3. Chemical Resources Laboratory, Tokyo Institute of Technology, Nagatsuta, Yokohama, Japan

    Toshiharu Suzuki

  4. Department of Molecular Bioscience, Kyoto-Sangyo University, Kamigamomotoyama, Kyoto, Japan

    Kazumi Tanaka, Ei-ichiro Saita & Masasuke Yoshida

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  1. Toshiharu Suzuki
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Contributions

T.S. and M.Y. conceived and designed the experiments and wrote the paper. T.S. led development of the human F1 expression system in E. coli and the single-molecule observation system. E.S. gave critical suggestions for experiments and interpretations. K.T. and C.W. helped T.S. in collecting large amounts of image data.

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Correspondence to Toshiharu Suzuki or Masasuke Yoshida.

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Supplementary information

Supplementary Text and Figures

Supplementary Results and Supplementary Figures 1–7. (PDF 3445 kb)

Supplementary Video 1

Stepwise rotation of human F1. A sample of rotations of human F1 under conditions in Figure 2a (4 mM ATPγS, 50 kfps). (AVI 3004 kb)

Supplementary Video 2

Stepwise rotation of human F1. A sample of rotations of human F1 under conditions in Figure 2b (50 μM ATPγS, 50 kfps). (AVI 3004 kb)

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Suzuki, T., Tanaka, K., Wakabayashi, C. et al. Chemomechanical coupling of human mitochondrial F1-ATPase motor. Nat Chem Biol 10, 930–936 (2014). https://doi.org/10.1038/nchembio.1635

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