Heavy isotope labeling and mass spectrometry reveal unexpected remodeling of bacterial cell wall expansion in response to drugs

  1. Heiner Atze
  2. Yucheng Liang
  3. Jean-Emmanuel Hugonnet
  4. Arnaud Gutierrez
  5. Filippo Rusconi  Is a corresponding author
  6. Michel Arthur  Is a corresponding author
  1. INSERM, UMR-S 1138, Centre de Recherche des Cordeliers, France
  2. Sorbonne Université-INSERM, France
  3. PAPPSO, Universite Paris-Saclay, INRAE, CNRS, France

Abstract

Antibiotics of the β-lactam (penicillin) family inactivate target enzymes called D,D-transpeptidases or penicillin-binding proteins (PBPs) that catalyze the last cross-linking step of peptidoglycan synthesis. The resulting net-like macromolecule is the essential component of bacterial cell walls that sustains the osmotic pressure of the cytoplasm. In Escherichia coli, bypass of PBPs by the YcbB L,D-transpeptidase leads to resistance to these drugs. We developed a new method based on heavy isotope labeling and mass spectrometry to elucidate PBP- and YcbB-mediated peptidoglycan polymerization. PBPs and YcbB similarly participated in single-strand insertion of glycan chains into the expanding bacterial side wall. This absence of any transpeptidase-specific signature suggests that the peptidoglycan expansion mode is determined by other components of polymerization complexes. YcbB did mediate β-lactam resistance by insertion of multiple strands that were exclusively cross-linked to existing tripeptide-containing acceptors. We propose that this undocumented mode of polymerization depends upon accumulation of linear glycan chains due to PBP inactivation, formation of tripeptides due to cleavage of existing cross-links by a β-lactam-insensitive endopeptidase, and concerted cross-linking by YcbB.

Data availability

MS/MS spectra have been provided in Supplementary data file.The software developments required to predict and analyze the labeled/unlabeled muropeptide ions isotopic clusters either in MS or MS/MS experiments are hosted at https://gitlab.com/kantundpeterpan/masseltof and published under a Free Software license.

Article and author information

Author details

  1. Heiner Atze

    INSERM, UMR-S 1138, Centre de Recherche des Cordeliers, Paris, France
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1497-6373
  2. Yucheng Liang

    Sorbonne Université-INSERM, PARIS, France
    Competing interests
    The authors declare that no competing interests exist.
  3. Jean-Emmanuel Hugonnet

    INSERM, UMR-S 1138, Centre de Recherche des Cordeliers, Paris, France
    Competing interests
    The authors declare that no competing interests exist.
  4. Arnaud Gutierrez

    INSERM, UMR-S 1138, Centre de Recherche des Cordeliers, Paris, France
    Competing interests
    The authors declare that no competing interests exist.
  5. Filippo Rusconi

    PAPPSO, Universite Paris-Saclay, INRAE, CNRS, Paris, France
    For correspondence
    filippo.rusconi@universite-paris-saclay.fr
    Competing interests
    The authors declare that no competing interests exist.
  6. Michel Arthur

    INSERM, UMR-S 1138, Centre de Recherche des Cordeliers, Paris, France
    For correspondence
    michel.arthur@crc.jussieu.fr
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1007-636X

Funding

Agence Nationale de la Recherche (ANR-16-CE11-0030-12)

  • Heiner Atze
  • Michel Arthur

National Institute of Allergy and Infectious Diseases (R56AI045626)

  • Yucheng Liang

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

Reviewing Editor

  1. Bavesh D Kana, University of the Witwatersrand, South Africa

Version history

  1. Preprint posted: August 6, 2021 (view preprint)
  2. Received: August 6, 2021
  3. Accepted: June 9, 2022
  4. Accepted Manuscript published: June 9, 2022 (version 1)
  5. Version of Record published: July 1, 2022 (version 2)

Copyright

© 2022, Atze 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. Heiner Atze
  2. Yucheng Liang
  3. Jean-Emmanuel Hugonnet
  4. Arnaud Gutierrez
  5. Filippo Rusconi
  6. Michel Arthur
(2022)
Heavy isotope labeling and mass spectrometry reveal unexpected remodeling of bacterial cell wall expansion in response to drugs
eLife 11:e72863.
https://doi.org/10.7554/eLife.72863

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https://doi.org/10.7554/eLife.72863

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