Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

Subnanometre-resolution structure of the intact Thermus thermophilus H+-driven ATP synthase

Nature volume 481, pages 214–218 (2012)Cite this article

Subjects

This article has been updated

Abstract

Ion-translocating rotary ATPases serve either as ATP synthases, using energy from a transmembrane ion motive force to create the cell’s supply of ATP, or as transmembrane ion pumps that are powered by ATP hydrolysis1. The members of this family of enzymes each contain two rotary motors: one that couples ion translocation to rotation and one that couples rotation to ATP synthesis or hydrolysis. During ATP synthesis, ion translocation through the membrane-bound region of the complex causes rotation of a central rotor that drives conformational changes and ATP synthesis in the catalytic region of the complex. There are no structural models available for the intact membrane region of any ion-translocating rotary ATPase. Here we present a 9.7 Å resolution map of the H+-driven ATP synthase from Thermus thermophilus obtained by electron cryomicroscopy of single particles in ice. The 600-kilodalton complex has an overall subunit composition of A3B3CDE2FG2IL12. The membrane-bound motor consists of a ring of L subunits and the carboxy-terminal region of subunit I, which are equivalent to the c and a subunits of most other rotary ATPases, respectively. The map shows that the ring contains 12 L subunits2 and that the I subunit has eight transmembrane helices3. The L12 ring and I subunit have a surprisingly small contact area in the middle of the membrane, with helices from the I subunit making contacts with two different L subunits. The transmembrane helices of subunit I form bundles that could serve as half-channels across the membrane, with the first half-channel conducting protons from the periplasm to the L12 ring and the second half-channel conducting protons from the L12 ring to the cytoplasm. This structure therefore suggests the mechanism by which a transmembrane proton motive force is converted to rotation in rotary ATPases.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Three-dimensional map of the T. thermophilus ATP synthase.
Figure 2: Fitting of the C subunit crystal structure.
Figure 3: The membrane-bound region of the enzyme.
Figure 4: Model for proton translocation.

Similar content being viewed by others

Accession codes

Primary accessions

Protein Data Bank

Data deposits

The cryo-EM map of the T. thermophilus H+-driven ATP synthase is deposited in Electron Microscopy Data Bank under accession code EMD-5335; the docked atomic models are deposited in Protein Data Bank under accession number 3J0J.

Change history

  • 11 January 2012

    The placement of an arrow was corrected in Fig. 4b.

References

  1. Muench, S. P., Trinick, J. & Harrison, M. A. Structural divergence of the rotary ATPases. Q. Rev. Biophys. 44, 311–356 (2011)

    Article  CAS  Google Scholar 

  2. Toei, M. et al. Dodecamer rotor ring defines H+/ATP ratio for ATP synthesis of prokaryotic V-ATPase from Thermus thermophilus. Proc. Natl Acad. Sci. USA 104, 20256–20261 (2007)

    Article  ADS  CAS  Google Scholar 

  3. Toei, M., Toei, S. & Forgac, M. Definition of membrane topology and identification of residues important for transport in subunit a of the vacuolar ATPase. J. Biol. Chem. 286, 35176–35186 (2011)

    Article  CAS  Google Scholar 

  4. Numoto, N., Hasegawa, Y., Takeda, K. & Miki, K. Inter-subunit interaction and quaternary rearrangement defined by the central stalk of prokaryotic V1-ATPase. EMBO Rep. 10, 1228–1234 (2009)

    Article  CAS  Google Scholar 

  5. Lau, W. C. & Rubinstein, J. L. Structure of intact Thermus thermophilus V-ATPase by cryo-EM reveals organization of the membrane-bound V(O) motor. Proc. Natl Acad. Sci. USA 107, 1367–1372 (2010)

    Article  ADS  CAS  Google Scholar 

  6. Abrahams, J. P., Leslie, A. G., Lutter, R. & Walker, J. E. Structure at 2.8 A resolution of F1-ATPase from bovine heart mitochondria. Nature 370, 621–628 (1994)

    Article  ADS  CAS  Google Scholar 

  7. Lee, L. K. et al. The structure of the peripheral stalk of Thermus thermophilus H+-ATPase/synthase. Nature Struct. Mol. Biol. 17, 373–378 (2010)

    Article  CAS  Google Scholar 

  8. Iwata, M. et al. Crystal structure of a central stalk subunit C and reversible association/dissociation of vacuole-type ATPase. Proc. Natl Acad. Sci. USA 101, 59–64 (2004)

    Article  ADS  CAS  Google Scholar 

  9. Srinivasan, S., Vyas, N. K., Baker, M. L. & Quiocho, F. A. Crystal structure of the cytoplasmic N-terminal domain of subunit I, a homolog of subunit a, of V-ATPase. J. Mol. Biol. 412, 14–21 (2011)

    Article  CAS  Google Scholar 

  10. Murata, T. et al. Structure of the rotor of the V-type Na+-ATPase from Enterococcus hirae. Science 308, 654–659 (2005)

    Article  ADS  CAS  Google Scholar 

  11. Stock, D., Leslie, A. G. & Walker, J. E. Molecular architecture of the rotary motor in ATP synthase. Science 286, 1700–1705 (1999)

    Article  CAS  Google Scholar 

  12. Meier, T. et al. Structure of the rotor ring of F-type Na+-ATPase from Ilyobacter tartaricus. Science 308, 659–662 (2005)

    Article  ADS  CAS  Google Scholar 

  13. Pogoryelov, D., Yildiz, O., Faraldo-Gomez, J. D. & Meier, T. High-resolution structure of the rotor ring of a proton-dependent ATP synthase. Nature Struct. Mol. Biol. 16, 1068–1073 (2009)

    Article  CAS  Google Scholar 

  14. Watt, I. N. et al. Bioenergetic cost of making an adenosine triphosphate molecule in animal mitochondria. Proc. Natl Acad. Sci. USA 107, 16823–16827 (2010)

    Article  ADS  CAS  Google Scholar 

  15. Preiss, L. et al. A new type of proton coordination in an F1F0-ATP synthase rotor ring. PLoS Biol. 8, e1000443 (2010)

    Article  Google Scholar 

  16. Meier, T. et al. The central plug in the reconstituted undecameric c cylinder of a bacterial ATP synthase consists of phospholipids. FEBS Lett. 505, 353–356 (2001)

    Article  CAS  Google Scholar 

  17. Fillingame, R. H., Angevine, C. M. & Dmitriev, O. Y. Mechanics of coupling proton movements to c-ring rotation in ATP synthase. FEBS Lett. 555, 29–34 (2003)

    Article  CAS  Google Scholar 

  18. Junge, W., Lill, H. & Engelbrecht, S. ATP synthase: an electrochemical transducer with rotatory mechanics. Trends Biochem. Sci. 22, 420–423 (1997)

    Article  CAS  Google Scholar 

  19. Stouffer, A. L. et al. Structural basis for the function and inhibition of an influenza virus proton channel. Nature 451, 596–599 (2008)

    Article  ADS  CAS  Google Scholar 

  20. Gonzales, E. B., Kawate, T. & Gouaux, E. Pore architecture and ion sites in acid-sensing ion channels and P2X receptors. Nature 460, 599–604 (2009)

    Article  ADS  CAS  Google Scholar 

  21. Cain, B. D. & Simoni, R. D. Proton translocation by the F1F0ATPase of Escherichia coli. Mutagenic analysis of the a subunit. J. Biol. Chem. 264, 3292–3300 (1989)

    CAS  PubMed  Google Scholar 

  22. Kawasaki-Nishi, S., Nishi, T. & Forgac, M. Arg-735 of the 100-kDa subunit a of the yeast V-ATPase is essential for proton translocation. Proc. Natl Acad. Sci. USA 98, 12397–12402 (2001)

    Article  ADS  CAS  Google Scholar 

  23. Pogoryelov, D. et al. Microscopic rotary mechanism of ion translocation in the Fo complex of ATP synthases. Nature Chem. Biol. 6, 891–899 (2010)

    Article  CAS  Google Scholar 

  24. Steed, P. R. & Fillingame, R. H. Aqueous accessibility to the transmembrane regions of subunit c of the Escherichia coli F1F0 ATP synthase. J. Biol. Chem. 284, 23243–23250 (2009)

    Article  CAS  Google Scholar 

  25. Long, J. C., Wang, S. & Vik, S. B. Membrane topology of subunit a of the F1F0 ATP synthase as determined by labeling of unique cysteine residues. J. Biol. Chem. 273, 16235–16240 (1998)

    Article  CAS  Google Scholar 

  26. Valiyaveetil, F. I. & Fillingame, R. H. Transmembrane topography of subunit a in the Escherichia coli F1F0 ATP synthase. J. Biol. Chem. 273, 16241–16247 (1998)

    Article  CAS  Google Scholar 

  27. Baker, L. A., Smith, E. A., Bueler, S. A. & Rubinstein, J. L. The resolution dependence of optimal exposures in liquid nitrogen temperature electron cryomicroscopy of catalase crystals. J. Struct. Biol. 169, 431–437 (2010)

    Article  CAS  Google Scholar 

  28. Grigorieff, N. FREALIGN: high-resolution refinement of single particle structures. J. Struct. Biol. 157, 117–125 (2007)

    Article  CAS  Google Scholar 

  29. Mindell, J. A. & Grigorieff, N. Accurate determination of local defocus and specimen tilt in electron microscopy. J. Struct. Biol. 142, 334–347 (2003)

    Article  Google Scholar 

  30. Crowther, R. A., Henderson, R. & Smith, J. M. MRC image processing programs. J. Struct. Biol. 116, 9–16 (1996)

    Article  CAS  Google Scholar 

  31. Nelder, J. A. & Mead, R. A simplex method for function minimization. Comput. J. 7, 308–313 (1965)

    Article  MathSciNet  Google Scholar 

  32. Press, W. H., Teukolsky, S. A., Vetterlin, W. T. & Flannery, B. P. Numerical Recipes in Fortran 77 2nd edn (Cambridge Univ. Press, 2003)

    Google Scholar 

  33. Sousa, D. & Grigorieff, N. Ab initio resolution measurement for single particle structures. J. Struct. Biol. 157, 201–210 (2007)

    Article  CAS  Google Scholar 

  34. Rosenthal, P. B., Crowther, R. A. & Henderson, R. An objective criterion for resolution assessment in single-particle electron microscopy (appendix). J. Mol. Biol. 333, 743–745 (2003)

    Article  Google Scholar 

  35. Rosenthal, P. B. & Henderson, R. Optimal determination of particle orientation, absolute hand, and contrast loss in single-particle electron cryomicroscopy. J. Mol. Biol. 333, 721–745 (2003)

    Article  CAS  Google Scholar 

  36. Pintilie, G. D. et al. Quantitative analysis of cryo-EM density map segmentation by watershed and scale-space filtering, and fitting of structures by alignment to regions. J. Struct. Biol. 170, 427–438 (2010)

    Article  CAS  Google Scholar 

  37. Baker, L. A. & Rubinstein, J. L. Edged watershed segmentation: a semi-interactive algorithm for segmentation of low-resolution maps from electron cryomicroscopy. J. Struct. Biol. 176, 127–132 (2011)

    Article  Google Scholar 

  38. Ludtke, S. J., Baldwin, P. R. & Chiu, W. EMAN: semiautomated software for high-resolution single-particle reconstructions. J. Struct. Biol. 128, 82–97 (1999)

    Article  CAS  Google Scholar 

  39. Kelley, L. A. & Sternberg, M. J. Protein structure prediction on the Web: a case study using the Phyre server. Nature Protocols 4, 363–371 (2009)

    Article  CAS  Google Scholar 

  40. Chacon, P. & Wriggers, W. Multi-resolution contour-based fitting of macromolecular structures. J. Mol. Biol. 317, 375–384 (2002)

    Article  CAS  Google Scholar 

  41. Goddard, T. D., Huang, C. C. & Ferrin, T. E. Visualizing density maps with UCSF Chimera. J. Struct. Biol. 157, 281–287 (2007)

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank V. Kanelis, F. Sicheri, P. Rosenthal, L. Kay and R. Henderson for discussions and reading this manuscript, and J. Walker and R. Pomès for discussions. Computations were performed on the general-purpose cluster supercomputer at the SciNet HPC Consortium. W.C.Y.L. was supported by an Ontario Graduate Scholarship. J.L.R. was supported by a New Investigator Award from the Canadian Institutes of Health Research and an Early Researcher Award from the Ontario Ministry of Research and Innovation. This research was funded by operating grant MOP 81294 from the Canadian Institutes of Health Research.

Author information

Authors and Affiliations

  1. Molecular Structure and Function Program, The Hospital for Sick Children Research Institute, 555 University Avenue, Toronto, Ontario M5G 1X8, Canada,

    Wilson C. Y. Lau & John L. Rubinstein

  2. Department of Biochemistry, The University of Toronto, Ontario M5S 1A8, Canada,

    Wilson C. Y. Lau & John L. Rubinstein

  3. Department of Medical Biophysics, The University of Toronto, Ontario M5G 2M9, Canada,

    John L. Rubinstein

Authors
  1. Wilson C. Y. Lau
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. John L. Rubinstein
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

J.L.R. and W.C.Y.L. designed the experiments and J.L.R. supervised the research. W.C.Y.L. performed protein purification and cryo-EM. J.L.R. wrote new computer programs. W.C.Y.L. and J.L.R. performed the image analysis, interpreted the data and wrote the manuscript.

Corresponding author

Correspondence to John L. Rubinstein.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

The file contains Supplementary Figures 1-4 with legends, Supplementary Table 1 and additional references. (PDF 1639 kb)

Supplementary Movie 1

The movie shows a 3-D map with docked crystals structures of subunits A (yellow), B (red), C (cyan), D (blue), E (purple), F (orange), and G (beige) subunits and segments for density from subunits I (green), L (magenta), and the missing density from subunit D (blue). The scale bar corresponds to 25 Å . (MOV 20549 kb)

PowerPoint slides

Rights and permissions

Reprints and permissions

About this article

Cite this article

Lau, W., Rubinstein, J. Subnanometre-resolution structure of the intact Thermus thermophilus H+-driven ATP synthase. Nature 481, 214–218 (2012). https://doi.org/10.1038/nature10699

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature10699

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing