Activation of the l,d-transpeptidation peptidoglycan cross-linking pathway by a metallo-d,d-carboxypeptidase in Enterococcus faecium

Authors

  • Emmanuelle Sacco,

    1. Centre de Recherche des Cordeliers, LRMA, Equipe 12, Université Pierre et Marie Curie – Paris6, UMR S 872, Paris, F-75006 France.
    2. Université Paris Descartes, UMR S 872, Paris, F-75006 France.
    3. INSERM, U872, Paris, F-75006 France.
    Search for more papers by this author
    • These authors contributed equally to the work.

  • Jean-Emmanuel Hugonnet,

    1. Centre de Recherche des Cordeliers, LRMA, Equipe 12, Université Pierre et Marie Curie – Paris6, UMR S 872, Paris, F-75006 France.
    2. Université Paris Descartes, UMR S 872, Paris, F-75006 France.
    3. INSERM, U872, Paris, F-75006 France.
    Search for more papers by this author
    • These authors contributed equally to the work.

  • Nathalie Josseaume,

    1. Centre de Recherche des Cordeliers, LRMA, Equipe 12, Université Pierre et Marie Curie – Paris6, UMR S 872, Paris, F-75006 France.
    2. Université Paris Descartes, UMR S 872, Paris, F-75006 France.
    3. INSERM, U872, Paris, F-75006 France.
    Search for more papers by this author
  • Julie Cremniter,

    1. Centre de Recherche des Cordeliers, LRMA, Equipe 12, Université Pierre et Marie Curie – Paris6, UMR S 872, Paris, F-75006 France.
    2. Université Paris Descartes, UMR S 872, Paris, F-75006 France.
    3. INSERM, U872, Paris, F-75006 France.
    Search for more papers by this author
  • Lionel Dubost,

    1. Muséum National d'Histoire Naturelle, Plateforme de Spectrométrie de Masse et de Protéomique du Muséum, Département Régulation Développement et Diversité Moléculaire, Paris, F-75005 France.
    2. Molécules de Communication et Adaptation des Micro-Organismes, FRE 3206 CNRS, Paris, F-75005 France.
    Search for more papers by this author
  • Arul Marie,

    1. Muséum National d'Histoire Naturelle, Plateforme de Spectrométrie de Masse et de Protéomique du Muséum, Département Régulation Développement et Diversité Moléculaire, Paris, F-75005 France.
    2. Molécules de Communication et Adaptation des Micro-Organismes, FRE 3206 CNRS, Paris, F-75005 France.
    Search for more papers by this author
  • Delphine Patin,

    1. Laboratoire des Enveloppes Bactériennes et Antibiotiques, Institut de Biochimie et Biophysique Moléculaire et Cellulaire, UMR 8619 CNRS, Univ Paris-Sud, 91405 Orsay, France.
    Search for more papers by this author
  • Didier Blanot,

    1. Laboratoire des Enveloppes Bactériennes et Antibiotiques, Institut de Biochimie et Biophysique Moléculaire et Cellulaire, UMR 8619 CNRS, Univ Paris-Sud, 91405 Orsay, France.
    Search for more papers by this author
  • Louis B. Rice,

    1. Medical and Research Services, Louis Stokes Cleveland VA Medical Center, Cleveland, OH 44106, USA.
    Search for more papers by this author
  • Jean-Luc Mainardi,

    1. Centre de Recherche des Cordeliers, LRMA, Equipe 12, Université Pierre et Marie Curie – Paris6, UMR S 872, Paris, F-75006 France.
    2. Université Paris Descartes, UMR S 872, Paris, F-75006 France.
    3. INSERM, U872, Paris, F-75006 France.
    4. AP-HP, Hôpital Européen Georges Pompidou, Paris, F-75015 France.
    Search for more papers by this author
  • Michel Arthur

    Corresponding author
    1. Centre de Recherche des Cordeliers, LRMA, Equipe 12, Université Pierre et Marie Curie – Paris6, UMR S 872, Paris, F-75006 France.
    2. Université Paris Descartes, UMR S 872, Paris, F-75006 France.
    3. INSERM, U872, Paris, F-75006 France.
    Search for more papers by this author

*E-mail michel.arthur@crc.jussieu.fr; Tel. (+33) 1 43 25 00; Fax (+33) 1 43 25 68 12.

Summary

Bypass of the penicillin-binding proteins by an l,d-transpeptidase (Ldtfm) confers cross-resistance to β-lactam and glycopeptide antibiotics in mutants of Enterococcus faecium selected in vitro. Ldtfm is produced by the parental strain D344S although it insignificantly contributes to peptidoglycan cross-linking as pentapeptide stems cannot be used as acyl donors by this enzyme. Here we show that production of the tetrapeptide substrate of Ldtfm is controlled by a two-component regulatory system (DdcRS) and a metallo-d,d-carboxypeptidase (DdcY). The locus was silent in D344S and its activation was due to amino acid substitutions in DdcS or DdcR that led to production of DdcY and hydrolysis of the C-terminal d-Ala residue of the cytoplasmic peptidoglycan precursor UDP-MurNAc-pentapeptide. The T161A and T161M substitutions affected a position of DdcS known to be essential for the phosphatase activity of related sensor kinases. Complete elimination of UDP-MurNAc-pentapeptide, which was required specifically for resistance to glycopeptides, involved substitutions in DdcY that increased the catalytic efficiency of the enzyme (E127K) and affected its interaction with the cell envelope (I14N). The ddc locus displays striking similarities with portions of the van vancomycin resistance gene clusters, suggesting possible routes of emergence of cross-resistance to glycopeptides and β-lactams in natural conditions.

Ancillary