The membrane anchor of penicillin-binding protein PBP2a from Streptococcus pneumoniae influences peptidoglycan chain length

Authors

  • Nordine Helassa,

    1.  CEA, Institut de Biologie Structurale, Grenoble, France
    2.  CNRS, Institut de Biologie Structurale, Grenoble, France
    3.  Université Joseph Fourier, Institut de Biologie Structurale, Grenoble, France
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  • Waldemar Vollmer,

    1.  Centre for Bacterial Cell Biology, Newcastle University, UK
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  • Eefjan Breukink,

    1.  Department of Chemical Biology and Organic Chemistry, Utrecht University, The Netherlands
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  • Thierry Vernet,

    1.  CEA, Institut de Biologie Structurale, Grenoble, France
    2.  CNRS, Institut de Biologie Structurale, Grenoble, France
    3.  Université Joseph Fourier, Institut de Biologie Structurale, Grenoble, France
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  • André Zapun

    1.  CEA, Institut de Biologie Structurale, Grenoble, France
    2.  CNRS, Institut de Biologie Structurale, Grenoble, France
    3.  Université Joseph Fourier, Institut de Biologie Structurale, Grenoble, France
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T. Vernet, Pneumococcus Group, Institut de Biologie Structurale, 41 rue Jules Horowitz, 38027 Grenoble, France
Fax: +33 4 38 78 54 94
Tel: +33 4 38 78 96 81
E-mail: thierry.vernet@ibs.fr

Abstract

The pneumococcus is an important Gram-positive pathogen, which shows increasing resistance to antibiotics, including β-lactams that target peptidoglycan assembly. Understanding cell-wall synthesis, at the molecular and cellular level, is essential for the prospect of combating drug resistance. As a first step towards reconstituting pneumococcal cell-wall assembly in vitro, we present the characterization of the glycosyltransferase activity of penicillin-binding protein (PBP)2a from Streptococcus pneumoniae. Recombinant full-length membrane-anchored PBP2a was purified by ion-exchange chromatography. The glycosyltransferase activity of this enzyme was found to differ from that of a truncated periplasmic form. The full-length protein with its cytoplasmic and transmembrane segment synthesizes longer glycan chains than the shorter form. The transpeptidase active site was functional, as shown by its reactivity towards bocillin and the catalysis of the hydrolysis of a thiol-ester substrate analogue. However, PBP2a did not cross-link the peptide stems of glycan chains in vitro. The absence of transpeptidase activity indicates that an essential component is missing from the in vitro system.

Structured digital abstract

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