1. Structural Biology and Molecular Biophysics
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Mechanism of completion of peptidyltransferase centre assembly in eukaryotes

  1. Vasileios Kargas
  2. Pablo Castro-Hartmann
  3. Norberto Escudero-Urquijo
  4. Kyle Dent
  5. Christine Hilcenko
  6. Carolin Sailer
  7. Gertrude Zisser
  8. Maria J Marques-Carvalho
  9. Simone Pellegrino
  10. Leszek Wawiórka
  11. Stefan MV Freund
  12. Jane L Wagstaff
  13. Antonina Andreeva
  14. Alexandre Faille
  15. Edwin Chen
  16. Florian Stengel
  17. Helmut Bergler
  18. Alan John Warren  Is a corresponding author
  1. Cambridge Institute for Medical Research, United Kingdom
  2. University of Konstanz, Germany
  3. University of Graz, Austria
  4. MRC Laboratory of Molecular Biology, United Kingdom
  5. University of Leeds, United Kingdom
Research Article
  • Cited 24
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Cite this article as: eLife 2019;8:e44904 doi: 10.7554/eLife.44904

Abstract

During their final maturation in the cytoplasm, pre-60S ribosomal particles are converted to translation-competent large ribosomal subunits. Here, we present the mechanism of peptidyltransferase centre (PTC) completion that explains how integration of the last ribosomal proteins is coupled to release of the nuclear export adaptor Nmd3. Single-particle cryo-EM reveals that eL40 recruitment stabilizes helix 89 to form the uL16 binding site. The loading of uL16 unhooks helix 38 from Nmd3 to adopt its mature conformation. In turn, partial retraction of the L1 stalk is coupled to a conformational switch in Nmd3 that allows the uL16 P-site loop to fully accommodate into the PTC where it competes with Nmd3 for an overlapping binding site (base A2971). Our data reveal how the central functional site of the ribosome is sculpted and suggest how the formation of translation-competent 60S subunits is disrupted in leukaemia-associated ribosomopathies.

Data availability

The cryo-EM density maps have been deposited in the Electron Microscopy Data Bank with accession numbers EMD-4558, EMD-4559, EMD-4560, EMD-4636, EMD-4884 and EMD-4630. Atomic coordinates have been deposited in the Protein Data Bank, with entry codes 6QIF, 6QIJ, 6QIK, 6QTZ, 6RI5 and 6QT0.

The following data sets were generated

Article and author information

Author details

  1. Vasileios Kargas

    Cambridge Institute for Medical Research, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  2. Pablo Castro-Hartmann

    Cambridge Institute for Medical Research, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  3. Norberto Escudero-Urquijo

    Cambridge Institute for Medical Research, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8201-5884
  4. Kyle Dent

    Cambridge Institute for Medical Research, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  5. Christine Hilcenko

    Cambridge Institute for Medical Research, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  6. Carolin Sailer

    Department of Biology, University of Konstanz, Konstanz, Germany
    Competing interests
    The authors declare that no competing interests exist.
  7. Gertrude Zisser

    Institute of Molecular Bioscience, University of Graz, Graz, Austria
    Competing interests
    The authors declare that no competing interests exist.
  8. Maria J Marques-Carvalho

    Cambridge Institute for Medical Research, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  9. Simone Pellegrino

    Cambridge Institute for Medical Research, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  10. Leszek Wawiórka

    Cambridge Institute for Medical Research, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  11. Stefan MV Freund

    MRC Laboratory of Molecular Biology, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  12. Jane L Wagstaff

    MRC Laboratory of Molecular Biology, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  13. Antonina Andreeva

    MRC Laboratory of Molecular Biology, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  14. Alexandre Faille

    Cambridge Institute for Medical Research, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  15. Edwin Chen

    Faculty of Biological Sciences, University of Leeds, Leeds, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-0742-9734
  16. Florian Stengel

    Department of Biology, University of Konstanz, Konstanz, Germany
    Competing interests
    The authors declare that no competing interests exist.
  17. Helmut Bergler

    Institute of Molecular Biosciences, University of Graz, Graz, Austria
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-7724-309X
  18. Alan John Warren

    Cambridge Institute for Medical Research, Cambridge, United Kingdom
    For correspondence
    ajw1000@cam.ac.uk
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9277-4553

Funding

Medical Research Council (MC_U105161083)

  • Alan John Warren

Bloodwise (12048)

  • Alan John Warren

Wellcome (108466/Z/15/Z)

  • Edwin Chen

German Science Foundation Emmy Noether Foundation (STE 2517/1-1)

  • Florian Stengel

Collaborative Research Center (969 Project A06)

  • Florian Stengel

Austrian Science Foundation FWF Grants (P26136)

  • Helmut Bergler

Austrian Science Foundation FWF Grants (P29451)

  • Helmut Bergler

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

Reviewing Editor

  1. Nahum Sonenberg, McGill University, Canada

Publication history

  1. Received: January 5, 2019
  2. Accepted: May 20, 2019
  3. Accepted Manuscript published: May 22, 2019 (version 1)
  4. Version of Record published: June 17, 2019 (version 2)

Copyright

© 2019, Kargas 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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