Residue-by-residue analysis of cotranslational membrane protein integration in vivo

  1. Felix Nicolaus
  2. Ane Metola
  3. Daphne Mermans
  4. Amanda Liljenström
  5. Ajda Krč
  6. Salmo Mohammed Abdullahi
  7. Matthew Zimmer
  8. Thomas F Miller III
  9. Gunnar von Heijne  Is a corresponding author
  1. Stockholm University, Sweden
  2. University of Ljubljana, Slovenia
  3. California Institute of Technology, United States

Abstract

We follow the cotranslational biosynthesis of three multi-spanning E. coli inner membrane proteins in vivo using high-resolution Force Profile Analysis. The force profiles show that the nascent chain is subjected to rapidly varying pulling forces during translation, and reveal unexpected complexities in the membrane integration process. We find that an N-terminal cytoplasmic domain can fold in the ribosome exit tunnel before membrane integration starts, that charged residues and membrane-interacting segments such as re-entrant loops and surface helices flanking a transmembrane helix (TMH) can advance or delay membrane integration, and that point mutations in an upstream TMH can affect the pulling forces generated by downstream TMHs in a highly position-dependent manner, suggestive of residue-specific interactions between TMHs during the integration process. Our results support the 'sliding' model of translocon-mediated membrane protein integration, in which hydrophobic segments are continually exposed to the lipid bilayer during their passage through the SecYEG translocon.

Data availability

All fFL values measured in this study are included as Figures Source Data.

Article and author information

Author details

  1. Felix Nicolaus

    Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden
    Competing interests
    The authors declare that no competing interests exist.
  2. Ane Metola

    Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2885-7634
  3. Daphne Mermans

    Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden
    Competing interests
    The authors declare that no competing interests exist.
  4. Amanda Liljenström

    Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden
    Competing interests
    The authors declare that no competing interests exist.
  5. Ajda Krč

    Faculty of Chemistry and Chemical Technology, University of Ljubljana, Ljubljana, Slovenia
    Competing interests
    The authors declare that no competing interests exist.
  6. Salmo Mohammed Abdullahi

    Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden
    Competing interests
    The authors declare that no competing interests exist.
  7. Matthew Zimmer

    Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. Thomas F Miller III

    Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-1882-5380
  9. Gunnar von Heijne

    Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden
    For correspondence
    Gunnar.von.Heijne@dbb.su.se
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4490-8569

Funding

Knut och Alice Wallenbergs Stiftelse (2017.0323)

  • Gunnar von Heijne

Novo Nordisk Fonden (NNF18OC0032828)

  • Gunnar von Heijne

Vetenskapsrådet (NNF18OC0032828)

  • Gunnar von Heijne

National Institutes of Health (R01GM125063)

  • Thomas F Miller III

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

Reviewing Editor

  1. Randy Schekman, University of California Berkeley, United States

Version history

  1. Received: October 23, 2020
  2. Accepted: February 5, 2021
  3. Accepted Manuscript published: February 8, 2021 (version 1)
  4. Version of Record published: February 16, 2021 (version 2)

Copyright

© 2021, Nicolaus 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. Felix Nicolaus
  2. Ane Metola
  3. Daphne Mermans
  4. Amanda Liljenström
  5. Ajda Krč
  6. Salmo Mohammed Abdullahi
  7. Matthew Zimmer
  8. Thomas F Miller III
  9. Gunnar von Heijne
(2021)
Residue-by-residue analysis of cotranslational membrane protein integration in vivo
eLife 10:e64302.
https://doi.org/10.7554/eLife.64302

Share this article

https://doi.org/10.7554/eLife.64302

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