1. Structural Biology and Molecular Biophysics
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Dynamic action of the Sec machinery during initiation, protein translocation and termination

  1. Tomas Fessl
  2. Daniel Watkins
  3. Peter Oatley
  4. William John Allen
  5. Robin Adam Corey
  6. Jim Horne
  7. Steve A Baldwin
  8. Sheena E Radford
  9. Ian Collinson  Is a corresponding author
  10. Roman Tuma  Is a corresponding author
  1. University of Leeds, United Kingdom
  2. University of Bristol, United Kingdom
Research Article
  • Cited 29
  • Views 2,345
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Cite this article as: eLife 2018;7:e35112 doi: 10.7554/eLife.35112

Abstract

Protein translocation across cell membranes is a ubiquitous process required for protein secretion and membrane protein insertion. In bacteria, this is mostly mediated by the conserved SecYEG complex, driven through rounds of ATP hydrolysis by the cytoplasmic SecA, and the trans-membrane proton motive force. We have used single molecule techniques to explore SecY pore dynamics on multiple timescales in order to dissect the complex reaction pathway. The results show that SecA, both the signal sequence and mature components of the pre-protein, and ATP hydrolysis each have important and specific roles in channel unlocking, opening and priming for transport. After channel opening, translocation proceeds in two phases: a slow phase independent of substrate length, and a length-dependent transport phase with an intrinsic translocation rate of ~40 amino acids per second for the proOmpA substrate. Broad translocation rate distributions reflect the stochastic nature of polypeptide transport.

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Compressed data are available together with the relevant scripts as Supplementary Source Data and Code

Article and author information

Author details

  1. Tomas Fessl

    Astbury Centre for Structural Molecular Biology, University of Leeds, Leeds, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  2. Daniel Watkins

    School of Biochemistry, University of Bristol, Bristol, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  3. Peter Oatley

    Astbury Centre for Structural Molecular Biology, University of Leeds, Leeds, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  4. William John Allen

    School of Biochemistry, University of Bristol, Bristol, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-9513-4786
  5. Robin Adam Corey

    School of Biochemistry, University of Bristol, Bristol, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  6. Jim Horne

    Astbury Centre for Structural Molecular Biology, University of Leeds, Leeds, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  7. Steve A Baldwin

    Astbury Centre for Structural Molecular Biology, University of Leeds, Leeds, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  8. Sheena E Radford

    Astbury Centre for Structural Molecular Biology, University of Leeds, Leeds, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3079-8039
  9. Ian Collinson

    School of Biochemistry, University of Bristol, Bristol, United Kingdom
    For correspondence
    ian.collinson@bristol.ac.uk
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3931-0503
  10. Roman Tuma

    Astbury Centre for Structural Molecular Biology, University of Leeds, Leeds, United Kingdom
    For correspondence
    r.tuma@leeds.ac.uk
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-0047-0013

Funding

Biotechnology and Biological Sciences Research Council (BB/N017307/1)

  • Tomas Fessl
  • Sheena E Radford
  • Roman Tuma

Biotechnology and Biological Sciences Research Council (BB/I008675/1)

  • Daniel Watkins

Biotechnology and Biological Sciences Research Council (BB/M003604/I)

  • Robin Adam Corey

Wellcome (104632)

  • William John Allen
  • Ian Collinson

Seventh Framework Programme (32240)

  • Sheena E Radford

European Regional Development Fund (CZ.02.1.01/0.0/0.0/15_003/0000441)

  • Tomas Fessl
  • Roman Tuma

Biotechnology and Biological Sciences Research Council (BB/N015126/1)

  • Daniel Watkins
  • Ian Collinson

Biotechnology and Biological Sciences Research Council (BB/I008675/1)

  • Peter Oatley
  • Steve A Baldwin
  • Sheena E Radford
  • Roman Tuma

Biotechnology and Biological Sciences Research Council (BB/M011151/1)

  • Jim Horne

Biotechnology and Biological Sciences Research Council (BB/I006737/1)

  • William John Allen
  • Ian Collinson

Biotechnology and Biological Sciences Research Council (BBSRC South West Bioscience Doctoral Training Partnership)

  • Robin Adam Corey

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

Reviewing Editor

  1. Taekjip Ha, Johns Hopkins University School of Medicine, United States

Publication history

  1. Received: January 16, 2018
  2. Accepted: June 5, 2018
  3. Accepted Manuscript published: June 7, 2018 (version 1)
  4. Version of Record published: June 27, 2018 (version 2)

Copyright

© 2018, Fessl 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.

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