1. Structural Biology and Molecular Biophysics

Structure of the AAA protein Msp1 reveals mechanism of mislocalized membrane protein extraction

  1. Lan Wang
  2. Alexander Myasnikov
  3. Xingjie Pan
  4. Peter Walter  Is a corresponding author
  1. Howard Hughes Medical Institute, University of California, San Francisco, United States
  2. University of California, San Francisco, United States
https://doi.org/10.7554/eLife.54031
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  1. Lan Wang
  2. Alexander Myasnikov
  3. Xingjie Pan
  4. Peter Walter
(2020)
Structure of the AAA protein Msp1 reveals mechanism of mislocalized membrane protein extraction
eLife 9:e54031.
https://doi.org/10.7554/eLife.54031
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Abstract

The AAA protein Msp1 extracts mislocalized tail-anchored membrane proteins and targets them for degradation, thus maintaining proper cell organization. How Msp1 selects its substrates and firmly engages them during the energetically unfavorable extraction process remains a mystery. To address this question, we solved cryo-EM structures of Msp1-substrate complexes at near-atomic resolution. Akin to other AAA proteins, Msp1 forms hexameric spirals that translocate substrates through a central pore. A singular hydrophobic substrate recruitment site is exposed at the spiral's seam, which we propose positions the substrate for entry into the pore. There, a tight web of aromatic amino acids grips the substrate in a sequence-promiscuous, hydrophobic milieu. Elements at the intersubunit interfaces coordinate ATP hydrolysis with the subunits' positions in the spiral. We present a comprehensive model of Msp1's mechanism, which follows general architectural principles established for other AAA proteins yet specializes Msp1 for its unique role in membrane protein extraction.

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All data needed to evaluate the conclusions in the paper are present in the paper and/or the supplementary materials. The atomic models were deposited in the protein data bank under the accession codes 6PDW, 6PDY and 6PE0; the associated cryo-EM maps were deposited in the electron microscopy data bank (EMDB) under the accession codes EMD-20318, EMD-20319 and EMD-20320

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Author details

  1. Lan Wang

    Department of Biochemistry and Biophysics, Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8931-7201

  2. Alexander Myasnikov

    Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States
    Competing interests
    The authors declare that no competing interests exist.

  3. Xingjie Pan

    Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, San Francisco, United States
    Competing interests
    The authors declare that no competing interests exist.

  4. Peter Walter

    Department of Biochemistry and Biophysics, Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, United States
    For correspondence
    peter@walterlab.ucsf.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-6849-708X

Funding

National Institutes of Health (R01GM032384)

  • Peter Walter

National Institutes of Health (R01GM032384)

  • Lan Wang

Howard Hughes Medical Institute

  • Peter Walter

Damon Runyon Cancer Research Foundation (DRG-2312-17)

  • Lan Wang

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

Copyright

© 2020, Wang 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. Lan Wang
  2. Alexander Myasnikov
  3. Xingjie Pan
  4. Peter Walter
(2020)
Structure of the AAA protein Msp1 reveals mechanism of mislocalized membrane protein extraction
eLife 9:e54031.
https://doi.org/10.7554/eLife.54031

Share this article

https://doi.org/10.7554/eLife.54031

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Further reading

    1. Cell Biology
    2. Structural Biology and Molecular Biophysics

    Conserved structural elements specialize ATAD1 as a membrane protein extraction machine

    Lan Wang, Hannah Toutkoushian ... Peter Walter
    Research Advance Updated

    The mitochondrial AAA (ATPase Associated with diverse cellular Activities) protein ATAD1 (in humans; Msp1 in yeast) removes mislocalized membrane proteins, as well as stuck import substrates from the mitochondrial outer membrane, facilitating their re-insertion into their cognate organelles and maintaining mitochondria’s protein import capacity. In doing so, it helps to maintain proteostasis in mitochondria. How ATAD1 tackles the energetic challenge to extract hydrophobic membrane proteins from the lipid bilayer and what structural features adapt ATAD1 for its particular function has remained a mystery. Previously, we determined the structure of Msp1 in complex with a peptide substrate (Wang et al., 2020). The structure showed that Msp1’s mechanism follows the general principle established for AAA proteins while adopting several structural features that specialize it for its function. Among these features in Msp1 was the utilization of multiple aromatic amino acids to firmly grip the substrate in the central pore. However, it was not clear whether the aromatic nature of these amino acids were required, or if they could be functionally replaced by aliphatic amino acids. In this work, we determined the cryo-EM structures of the human ATAD1 in complex with a peptide substrate at near atomic resolution. The structures show that phylogenetically conserved structural elements adapt ATAD1 for its function while generally adopting a conserved mechanism shared by many AAA proteins. We developed a microscopy-based assay reporting on protein mislocalization, with which we directly assessed ATAD1’s activity in live cells and showed that both aromatic amino acids in pore-loop 1 are required for ATAD1’s function and cannot be substituted by aliphatic amino acids. A short α-helix at the C-terminus strongly facilitates ATAD1’s oligomerization, a structural feature that distinguishes ATAD1 from its closely related proteins.