Crystal structure of the full Swi2/Snf2 remodeler Mot1 in the resting state

  1. Agata Butryn
  2. Stephan Woike
  3. Savera Jagathpala Shetty
  4. David Thomas Auble
  5. Karl-Peter Hopfner  Is a corresponding author
  1. Ludwig-Maximilians-Universität München, Germany
  2. University of Virginia Health System, United States

Abstract

Swi2/Snf2 ATPases remodel protein:DNA complexes in all of the fundamental chromosome‑associated processes. The single‑subunit remodeler Mot1 dissociates TATA box-binding protein (TBP):DNA complexes and provides a simple model for obtaining structural insights into the action of Swi2/Snf2 ATPases. Previously we reported how the N-terminal domain of Mot1 it binds TBP, NC2 and DNA, but the location of the C-terminal ATPase domain remained unclear (Butryn et al., 2015). Here, we report the crystal structure of the near full-length Mot1 from Chaetomium thermophilum. Our data show that Mot1 adopts a ring like structure with a catalytically inactive resting state of the ATPase. Biochemical analysis suggests that TBP binding switches Mot1 into an ATP hydrolysis-competent conformation. Combined with our previous results, these data significantly improve the structural model for the complete Mot1:TBP:DNA complex and suggest a general mechanism for Mot1 action.

Data availability

The coordinates and structure factors are deposited in the Protein Data Bank under accession code 6G7E. All data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided for Figures 2 and Figure 2-figure supplement 1.

The following data sets were generated

Article and author information

Author details

  1. Agata Butryn

    Gene Center, Ludwig-Maximilians-Universität München, Munich, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5227-4770
  2. Stephan Woike

    Gene Center, Ludwig-Maximilians-Universität München, Munich, Germany
    Competing interests
    The authors declare that no competing interests exist.
  3. Savera Jagathpala Shetty

    Department of Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. David Thomas Auble

    Department of Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Karl-Peter Hopfner

    Gene Center, Ludwig-Maximilians-Universität München, Munich, Germany
    For correspondence
    hopfner@genzentrum.lmu.de
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4528-8357

Funding

National Institutes of Health (GM055763)

  • David Thomas Auble

European Commission (ERC Advanced Grant ATMMACHINE)

  • Karl-Peter Hopfner

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

Copyright

© 2018, Butryn 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

  • 1,596
    views
  • 263
    downloads
  • 5
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

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)

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

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

  1. Agata Butryn
  2. Stephan Woike
  3. Savera Jagathpala Shetty
  4. David Thomas Auble
  5. Karl-Peter Hopfner
(2018)
Crystal structure of the full Swi2/Snf2 remodeler Mot1 in the resting state
eLife 7:e37774.
https://doi.org/10.7554/eLife.37774

Share this article

https://doi.org/10.7554/eLife.37774

Further reading

    1. Structural Biology and Molecular Biophysics
    Jesse Howe, Douglas Walker ... Elisar J Barbar
    Research Article

    53BP1 is a key player in DNA repair and together with BRCA1 regulate selection of DNA double strand break repair mechanisms. Localization of DNA repair factors to sites of DNA damage by 53BP1 is controlled by its oligomerization domain (OD) and binding to LC8, a hub protein that functions to dimerize >100 clients. Here we show that 53BP1 OD is a trimer, an unusual finding for LC8 clients which are all dimers or tetramers. As a trimer, 53BP1 forms a heterogeneous mixture of complexes when bound to dimeric LC8 with the largest mass corresponding to a dimer-of-trimers bridged by 3 LC8 dimers. Analytical ultracentrifugation and isothermal titration calorimetry demonstrate that only the second of the three LC8 recognition motifs is necessary for a stable bridged complex. The stability of the bridged complex is tuned by multivalency, binding specificity of the second LC8 site, and the length of the linker separating the LC8 binding domain and OD. 53BP1 mutants deficient in bridged species fail to impact 53BP1 focus formation in human cell culture studies, suggesting that the primary role of LC8 is to bridge 53BP1 trimers which in turn promotes recruitment of 53BP1 at sites of DNA damage. We propose that the formation of higher-order oligomers of 53BP1 explains how LC8 elicits an improvement in 53BP1 foci and affects the structure and functions of 53BP1.

    1. Immunology and Inflammation
    2. Structural Biology and Molecular Biophysics
    Douwe Schulte, Marta Šiborová ... Joost Snijder
    Research Article

    Antibodies are a major component of adaptive immunity against invading pathogens. Here, we explore possibilities for an analytical approach to characterize the antigen-specific antibody repertoire directly from the secreted proteins in convalescent serum. This approach aims to perform simultaneous antibody sequencing and epitope mapping using a combination of single particle cryo-electron microscopy (cryoEM) and bottom-up proteomics techniques based on mass spectrometry (LC-MS/MS). We evaluate the performance of the deep-learning tool ModelAngelo in determining de novo antibody sequences directly from reconstructed 3D volumes of antibody-antigen complexes. We demonstrate that while map quality is a critical bottleneck, it is possible to sequence antibody variable domains from cryoEM reconstructions with accuracies of up to 80–90%. While the rate of errors exceeds the typical levels of somatic hypermutation, we show that the ModelAngelo-derived sequences can be used to assign the used V-genes. This provides a functional guide to assemble de novo peptides from LC-MS/MS data more accurately and improves the tolerance to a background of polyclonal antibody sequences. Following this proof-of-principle, we discuss the feasibility and future directions of this approach to characterize antigen-specific antibody repertoires.