ATP-induced asymmetric pre-protein folding as a driver of protein translocation through the Sec machinery

  1. Robin A Corey
  2. Zainab Ahdash
  3. Anokhi Shah
  4. Euan Pyle
  5. William John Allen
  6. Tomas Fessl
  7. Janet E Lovett  Is a corresponding author
  8. Argyris Politis  Is a corresponding author
  9. Ian Collinson  Is a corresponding author
  1. University of Bristol, United Kingdom
  2. King's College London, United Kingdom
  3. University of St Andrews, United Kingdom
  4. University of South Bohemia in Ceske Budejovice, Czech Republic

Abstract

Transport of proteins across membranes is a fundamental process, achieved in every cell by the 'Sec' translocon. In prokaryotes, SecYEG associates with the motor ATPase SecA to carry out translocation for pre-protein secretion. Previously, we proposed a Brownian ratchet model for transport, whereby the free energy of ATP-turnover favours the directional diffusion of the polypeptide [Allen et al. eLife 2016]. Here, we show that ATP enhances this process by modulating secondary structure formation within the translocating protein. A combination of molecular simulation with hydrogen-deuterium-exchange mass spectrometry and electron paramagnetic resonance spectroscopy reveal an asymmetry across the membrane: ATP induced conformational changes in the cytosolic cavity promote unfolded pre-protein structure, while the exterior cavity favours its formation. This ability to exploit structure within a pre-protein is an unexplored area of protein transport, which may apply to other protein transporters, such as those of the endoplasmic reticulum and mitochondria.

Data availability

All data generated during this study are included in the Figures of the mansucript. EPR data is available at https://doi.org/10.17630/0fedaeec-7e27-4876-a6d1-cda2d3a6799c.

The following data sets were generated

Article and author information

Author details

  1. Robin A Corey

    School of Biochemistry, University of Bristol, Bristol, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1820-7993
  2. Zainab Ahdash

    Department of Chemistry, King's College London, London, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  3. Anokhi Shah

    School of Physics and Astronomy, University of St Andrews, St Andrews, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  4. Euan Pyle

    Department of Chemistry, King's College London, London, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4633-4917
  5. William John Allen

    School of Biochemistry, University of Bristol, Bristol, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-9513-4786
  6. Tomas Fessl

    Faculty of Sciences, University of South Bohemia in Ceske Budejovice, České Budějovice, Czech Republic
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6969-4870
  7. Janet E Lovett

    School of Physics and Astronomy, University of St Andrews, St Andrews, United Kingdom
    For correspondence
    jel20@st-andrews.ac.uk
    Competing interests
    The authors declare that no competing interests exist.
  8. Argyris Politis

    Department of Chemistry, King's College London, London, United Kingdom
    For correspondence
    argyris.politis@kcl.ac.uk
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-6658-3224
  9. Ian Collinson

    School of Biochemistry, University of Bristol, Bristol, United Kingdom
    For correspondence
    ian.collinson@bristol.ac.uk
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3931-0503

Funding

Biotechnology and Biological Sciences Research Council (BB/M003604/1)

  • Robin A Corey
  • William John Allen
  • Ian Collinson

Wellcome (104632)

  • William John Allen
  • Ian Collinson

Royal Society (University Research Fellowship)

  • Janet E Lovett

Wellcome (109854/Z/15/Z)

  • Zainab Ahdash
  • Argyris Politis

Wellcome (099149/Z/12/Z)

  • Anokhi Shah
  • Janet E Lovett

European Regional Development Fund (CZ.02.1.01/0.0/0.0/15_003/0000441)

  • Tomas Fessl

Engineering and Physical Sciences Research Council (ep/m508214/1)

  • Anokhi Shah

Biotechnology and Biological Sciences Research Council (BB/I008675/1)

  • Robin A Corey
  • William John Allen
  • Ian Collinson

Biotechnology and Biological Sciences Research Council (BB/N015126/1)

  • Robin A Corey
  • William John Allen
  • Ian Collinson

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

Reviewing Editor

  1. Yibing Shan, DE Shaw Research, United States

Version history

  1. Received: September 7, 2018
  2. Accepted: January 1, 2019
  3. Accepted Manuscript published: January 2, 2019 (version 1)
  4. Version of Record published: January 16, 2019 (version 2)
  5. Version of Record updated: July 4, 2019 (version 3)

Copyright

© 2019, Corey 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

  • 2,848
    Page views
  • 465
    Downloads
  • 25
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, Scopus, PubMed Central.

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. Robin A Corey
  2. Zainab Ahdash
  3. Anokhi Shah
  4. Euan Pyle
  5. William John Allen
  6. Tomas Fessl
  7. Janet E Lovett
  8. Argyris Politis
  9. Ian Collinson
(2019)
ATP-induced asymmetric pre-protein folding as a driver of protein translocation through the Sec machinery
eLife 8:e41803.
https://doi.org/10.7554/eLife.41803

Share this article

https://doi.org/10.7554/eLife.41803

Further reading

    1. Biochemistry and Chemical Biology
    2. Developmental Biology
    Zhi Li, Yuedi Wang ... Zeyang Zhou
    Research Article

    Imidacloprid is a global health threat that severely poisons the economically and ecologically important honeybee pollinator, Apis mellifera. However, its effects on developing bee larvae remain largely unexplored. Our pilot study showed that imidacloprid causes developmental delay in bee larvae, but the underlying toxicological mechanisms remain incompletely understood. In this study, we exposed bee larvae to imidacloprid at environmentally relevant concentrations of 0.7, 1.2, 3.1, and 377 ppb. There was a marked dose-dependent delay in larval development, characterized by reductions in body mass, width, and growth index. However, imidacloprid did not affect on larval survival and food consumption. The primary toxicological effects induced by elevated concentrations of imidacloprid (377 ppb) included inhibition of neural transmission gene expression, induction of oxidative stress, gut structural damage, and apoptosis, inhibition of developmental regulatory hormones and genes, suppression of gene expression levels involved in proteolysis, amino acid transport, protein synthesis, carbohydrate catabolism, oxidative phosphorylation, and glycolysis energy production. In addition, we found that the larvae may use antioxidant defenses and P450 detoxification mechanisms to mitigate the effects of imidacloprid. Ultimately, this study provides the first evidence that environmentally exposed imidacloprid can affect the growth and development of bee larvae by disrupting molting regulation and limiting the metabolism and utilization of dietary nutrients and energy. These findings have broader implications for studies assessing pesticide hazards in other juvenile animals.

    1. Biochemistry and Chemical Biology
    2. Cancer Biology
    Pengfei Guo, Rebecca C Lim ... Hui Zhang
    Research Article Updated

    The Polycomb Repressive Complex 2 (PRC2) methylates H3K27 to regulate development and cell fate by transcriptional silencing. Alteration of PRC2 is associated with various cancers. Here, we show that mouse Kdm1a deletion causes a dramatic reduction of PRC2 proteins, whereas mouse null mutation of L3mbtl3 or Dcaf5 results in PRC2 accumulation and increased H3K27 trimethylation. The catalytic subunit of PRC2, EZH2, is methylated at lysine 20 (K20), promoting EZH2 proteolysis by L3MBTL3 and the CLR4DCAF5 ubiquitin ligase. KDM1A (LSD1) demethylates the methylated K20 to stabilize EZH2. K20 methylation is inhibited by AKT-mediated phosphorylation of serine 21 in EZH2. Mouse Ezh2K20R/K20R mutants develop hepatosplenomegaly associated with high GFI1B expression, and Ezh2K20R/K20R mutant bone marrows expand hematopoietic stem cells and downstream hematopoietic populations. Our studies reveal that EZH2 is regulated by methylation-dependent proteolysis, which is negatively controlled by AKT-mediated S21 phosphorylation to establish a methylation-phosphorylation switch to regulate the PRC2 activity and hematopoiesis.