1. Structural Biology and Molecular Biophysics
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Allosteric communication in DNA polymerase clamp loaders relies on a critical hydrogen-bonded junction

  1. Subu Subramanian
  2. Kent Gorday
  3. Kendra Marcus
  4. Matthew R Orellana
  5. Peter Ren
  6. Xiao Ran Luo
  7. Michael E O'Donnell
  8. John Kuriyan  Is a corresponding author
  1. University of California, Berkeley, United States
  2. Howard Hughes Medical Institute, Rockefeller University, United States
Research Article
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Cite this article as: eLife 2021;10:e66181 doi: 10.7554/eLife.66181
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Abstract

Clamp loaders are AAA+ ATPases that load sliding clamps onto DNA. We mapped the mutational sensitivity of the T4 bacteriophage sliding clamp and clamp loader by deep mutagenesis, and found that residues not involved in catalysis or binding display remarkable tolerance to mutation. An exception is a glutamine residue in the AAA+ module (Gln 118) that is not located at a catalytic or interfacial site. Gln 118 forms a hydrogen-bonded junction in a helical unit that we term the central coupler, because it connects the catalytic centers to DNA and the sliding clamp. A suppressor mutation indicates that hydrogen bonding in the junction is important, and molecular dynamics simulations reveal that it maintains rigidity in the central coupler. The glutamine-mediated junction is preserved in diverse AAA+ ATPases, suggesting that a connected network of hydrogen bonds that links ATP molecules is an essential aspect of allosteric communication in these proteins.

Data availability

All data generated or analyzed during this study are included in the manuscript and supporting files. Source data files with raw counts and relative fitness scores are included.

Article and author information

Author details

  1. Subu Subramanian

    Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6095-7021
  2. Kent Gorday

    Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States
    Competing interests
    No competing interests declared.
  3. Kendra Marcus

    Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States
    Competing interests
    No competing interests declared.
  4. Matthew R Orellana

    Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States
    Competing interests
    No competing interests declared.
  5. Peter Ren

    Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States
    Competing interests
    No competing interests declared.
  6. Xiao Ran Luo

    Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States
    Competing interests
    No competing interests declared.
  7. Michael E O'Donnell

    Howard Hughes Medical Institute, Rockefeller University, New York, United States
    Competing interests
    No competing interests declared.
  8. John Kuriyan

    Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States
    For correspondence
    jkuriyan@mac.com
    Competing interests
    John Kuriyan, Senior editor, eLife.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4414-5477

Funding

Howard Hughes Medical Institute

  • Subu Subramanian
  • Kent Gorday
  • Peter Ren
  • Xiao Ran Luo
  • John Kuriyan

Amgen Foundation

  • Matthew R Orellana

National Institutes of Health (5T32AI100829-07)

  • Kent Gorday

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: January 4, 2021
  2. Accepted: April 3, 2021
  3. Accepted Manuscript published: April 13, 2021 (version 1)

Copyright

© 2021, Subramanian 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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