Structures of the ATP-fueled ClpXP proteolytic machine bound to protein substrate

  1. Xue Fei
  2. Tristan A Bell
  3. Simon Jenni
  4. Benjamin M Stinson
  5. Tania A Baker
  6. Stephen C Harrison
  7. Robert T Sauer  Is a corresponding author
  1. Massachusetts Institute of Technology, United States
  2. Harvard Medical School, United States
  3. Howard Hughes Medical Institute, Harvard Medical School, United States

Abstract

ClpXP is an ATP-dependent protease in which the ClpX AAA+ motor binds, unfolds, and translocates specific protein substrates into the degradation chamber of ClpP. We present cryo-EM studies of the E. coli enzyme that show how asymmetric hexameric rings of ClpX bind symmetric heptameric rings of ClpP and interact with protein substrates. Subunits in the ClpX hexamer assume a spiral conformation and interact with two-residue segments of substrate in the axial channel, as observed for other AAA+ proteases and protein-remodeling machines. Strictly sequential models of ATP hydrolysis and a power stroke that moves two residues of the substrate per translocation step have been inferred from these structural features for other AAA+ unfoldases, but biochemical and single-molecule biophysical studies indicate that ClpXP operates by a probabilistic mechanism in which five to eight residues are translocated for each ATP hydrolyzed. We propose structure-based models that could account for the functional results.

Data availability

PDB files for the structures determined here have been deposited in the PDB under accession codes 6PPE, 6PP8, 6PP7, 6PP6, 6PP5, 6POS, 6POD, 6PO3, and 6PO1.

The following data sets were generated
    1. Fei et al
    (2020) 6PPE
    RCSB Protein Data Bank, 6PPE.
    1. Fei et al
    (2020) 6PP8
    RCSB Protein Data Bank, 6PP8.
    1. Fei et al
    (2020) 6PP7
    RCSB Protein Data Bank, 6PP7.
    1. Fei et al
    (2020) 6PP6
    RCSB Protein Data Bank, 6PP6.
    1. Fei et al
    (2020) 6PP5
    RCSB Protein Data Bank, 6PP5.
    1. Fei et al
    (2020) 6POS
    RCSB Protein Data Bank, 6POS.
    1. Fei et al
    (2020) 6POD
    RCSB Protein Data Bank, 6POD.
    1. Fei et al
    (2020) 6PO3
    RCSB Protein Data Bank, 6PO3.
    1. Fei et al
    (2020) 6PO1
    RCSB Protein Data Bank, 6PO1.

Article and author information

Author details

  1. Xue Fei

    Department of Biology, Massachusetts Institute of Technology, Cambridge, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Tristan A Bell

    Department of Biology, Massachusetts Institute of Technology, Cambridge, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3668-8412
  3. Simon Jenni

    Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Benjamin M Stinson

    Department of Biology, Massachusetts Institute of Technology, Cambridge, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Tania A Baker

    Department of Biology, Massachusetts Institute of Technology, Cambridge, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0737-3411
  6. Stephen C Harrison

    Howard Hughes Medical Institute, Harvard Medical School, Boston, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7215-9393
  7. Robert T Sauer

    Department of Biology, Massachusetts Institute of Technology, Cambridge, United States
    For correspondence
    bobsauer@mit.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-1719-5399

Funding

National Institutes of Health (GM-101988)

  • Robert T Sauer

Howard Hughes Medical Institute

  • Tania A Baker
  • Stephen C Harrison

National Institutes of Health (5T32GM-007287)

  • Tristan A Bell

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

Reviewing Editor

  1. James M Berger, Johns Hopkins University School of Medicine, United States

Publication history

  1. Received: October 16, 2019
  2. Accepted: February 27, 2020
  3. Accepted Manuscript published: February 28, 2020 (version 1)
  4. Version of Record published: April 1, 2020 (version 2)

Copyright

© 2020, Fei 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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  1. Xue Fei
  2. Tristan A Bell
  3. Simon Jenni
  4. Benjamin M Stinson
  5. Tania A Baker
  6. Stephen C Harrison
  7. Robert T Sauer
(2020)
Structures of the ATP-fueled ClpXP proteolytic machine bound to protein substrate
eLife 9:e52774.
https://doi.org/10.7554/eLife.52774
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