1. Structural Biology and Molecular Biophysics
Download icon

Cryo-EM structures of the autoinhibited <em>E. coli</em> ATP synthase in three rotational states

  1. Meghna Sobti
  2. Callum Smits
  3. Andrew SW Wong
  4. Robert Ishmukhametov
  5. Daniela Stock
  6. Sara Sandin
  7. Alastair G Stewart  Is a corresponding author
  1. The Victor Chang Cardiac Research Institute, Australia
  2. Nanyang Technological University, Singapore
  3. University of Oxford, United Kingdom
Research Article
  • Cited 77
  • Views 5,389
  • Annotations
Cite this article as: eLife 2016;5:e21598 doi: 10.7554/eLife.21598

Abstract

A molecular model that provides a framework for interpreting the wealth of functional information obtained on the <em>E. coli</em> F-ATP synthase has been generated using cryo-electron microscopy. Three different states that relate to rotation of the enzyme were observed, with the central stalk's &epsilon; subunit in an extended autoinhibitory conformation in all three states. The Fo motor comprises of seven transmembrane helices and a decameric c-ring and invaginations on either side of the membrane indicate the entry and exit channels for protons. The proton translocating subunit contains near parallel helices inclined by ~30&ordm; to the membrane, a feature now synonymous with rotary ATPases. For the first time in this rotary ATPase subtype, the peripheral stalk is resolved over its entire length of the complex, revealing the F1 attachment points and a coiled-coil that bifurcates towards the membrane with its helices separating to embrace subunit a from two sides.

Data availability

The following data sets were generated

Article and author information

Author details

  1. Meghna Sobti

    Molecular, Structural and Computational Biology Division, The Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    Competing interests
    The authors declare that no competing interests exist.
  2. Callum Smits

    Molecular, Structural and Computational Biology Division, The Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    Competing interests
    The authors declare that no competing interests exist.
  3. Andrew SW Wong

    NTU Institute of Structural Biology, Nanyang Technological University, Singapore, Singapore
    Competing interests
    The authors declare that no competing interests exist.
  4. Robert Ishmukhametov

    Department of Physics, Clarendon Laboratory, University of Oxford, Oxford, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  5. Daniela Stock

    Molecular, Structural and Computational Biology Division, The Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    Competing interests
    The authors declare that no competing interests exist.
  6. Sara Sandin

    NTU Institute of Structural Biology, Nanyang Technological University, Singapore, Singapore
    Competing interests
    The authors declare that no competing interests exist.
  7. Alastair G Stewart

    Molecular, Structural and Computational Biology Division, The Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    For correspondence
    a.stewart@victorchang.edu.au
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2070-6030

Funding

National Health and Medical Research Council (1004620)

  • Daniela Stock

National Health and Medical Research Council (1109961)

  • Daniela Stock

National Health and Medical Research Council (1090408)

  • Alastair G Stewart

National Health and Medical Research Council (1022143)

  • Daniela Stock

National Health and Medical Research Council (1047004)

  • Daniela Stock

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Werner Kühlbrandt, Max Planck Institute of Biophysics, Germany

Publication history

  1. Received: September 19, 2016
  2. Accepted: December 15, 2016
  3. Accepted Manuscript published: December 21, 2016 (version 1)
  4. Accepted Manuscript updated: December 22, 2016 (version 2)
  5. Version of Record published: January 5, 2017 (version 3)
  6. Version of Record updated: February 10, 2017 (version 4)

Copyright

© 2016, Sobti et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 5,389
    Page views
  • 881
    Downloads
  • 77
    Citations

Article citation count generated by polling the highest count across the following sources: Scopus, Crossref, PubMed Central.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Download citations (links to download the citations from this article in formats compatible with various reference manager tools)

Open citations (links to open the citations from this article in various online reference manager services)

Further reading

    1. Cell Biology
    2. Structural Biology and Molecular Biophysics
    Carolina Franco Nitta et al.
    Research Article Updated

    Crosstalk between different receptor tyrosine kinases (RTKs) is thought to drive oncogenic signaling and allow therapeutic escape. EGFR and RON are two such RTKs from different subfamilies, which engage in crosstalk through unknown mechanisms. We combined high-resolution imaging with biochemical and mutational studies to ask how EGFR and RON communicate. EGF stimulation promotes EGFR-dependent phosphorylation of RON, but ligand stimulation of RON does not trigger EGFR phosphorylation – arguing that crosstalk is unidirectional. Nanoscale imaging reveals association of EGFR and RON in common plasma membrane microdomains. Two-color single particle tracking captured formation of complexes between RON and EGF-bound EGFR. Our results further show that RON is a substrate for EGFR kinase, and that transactivation of RON requires formation of a signaling competent EGFR dimer. These results support a role for direct EGFR/RON interactions in propagating crosstalk, such that EGF-stimulated EGFR phosphorylates RON to activate RON-directed signaling.

    1. Structural Biology and Molecular Biophysics
    Stijn van Dorp et al.
    Research Article Updated

    The dimeric ER Ca2+ sensor STIM1 controls store-operated Ca2+ entry (SOCE) through the regulated binding of its CRAC activation domain (CAD) to Orai channels in the plasma membrane. In resting cells, the STIM1 CC1 domain interacts with CAD to suppress SOCE, but the structural basis of this interaction is unclear. Using single-molecule Förster resonance energy transfer (smFRET) and protein crosslinking approaches, we show that CC1 interacts dynamically with CAD in a domain-swapped configuration with an orientation predicted to sequester its Orai-binding region adjacent to the ER membrane. Following ER Ca2+ depletion and release from CAD, cysteine crosslinking indicates that the two CC1 domains become closely paired along their entire length in the active Orai-bound state. These findings provide a structural basis for the dual roles of CC1: sequestering CAD to suppress SOCE in resting cells and propelling it toward the plasma membrane to activate Orai and SOCE after store depletion.