title: Anchoring of surface proteins to the cell wall of Staphylococcus aureus. III. Lipid II is an in vivo peptidoglycan substrate for sortase-catalyzed surface protein anchoring.
abstract: Surface proteins of Staphylococcus aureus are anchored to the cell wall peptidoglycan by a mechanism requiring a C-terminal sorting signal with an LPXTG motif. Surface proteins are first synthesized in the bacterial cytoplasm and then transported across the cytoplasmic membrane. Cleavage of the N-terminal signal peptide of the cytoplasmic surface protein P1 precursor generates the extracellular P2 species, which is the substrate for the cell wall anchoring reaction. Sortase, a membrane-anchored transpeptidase, cleaves P2 between the threonine (T) and the glycine (G) of the LPXTG motif and catalyzes the formation of an amide bond between the carboxyl group of threonine and the amino group of cell wall cross-bridges. We have used metabolic labeling of staphylococcal cultures with [(32)P]phosphoric acid to reveal a P3 intermediate. The (32)P-label of immunoprecipitated surface protein is removed by treatment with lysostaphin, a glycyl-glycine endopeptidase that separates the cell wall anchor structure. Furthermore, the appearance of P3 is prevented in the absence of sortase or by the inhibition of cell wall synthesis. (32)P-Labeled cell wall anchor species bind to nisin, an antibiotic that is known to form a complex with lipid II. Thus, it appears that the P3 intermediate represents surface protein linked to the lipid II peptidoglycan precursor. The data support a model whereby lipid II-linked polypeptides are incorporated into the growing peptidoglycan via the transpeptidation and transglycosylation reactions of cell wall synthesis, generating mature cell wall-linked surface protein.
authors.affiliation: Committee on Microbiology, University of Chicago, Chicago, Illinois 60637, USA.
authors.forename: Adrienne M
authors.initials: AM
authors.lastname: Perry
authors.forename: Hung
authors.initials: H
authors.lastname: Ton-That
authors.forename: Sarkis K
authors.initials: SK
authors.lastname: Mazmanian
authors.forename: Olaf
authors.initials: O
authors.lastname: Schneewind
mesh_terms: D000595:Amino Acid Sequence; D019881:Aminoacyltransferases / Q000378:metabolism*; D001414:Bacitracin / Q000494:pharmacology; D001426:Bacterial Proteins; D001665:Binding Sites; D003546:Cysteine Endopeptidases; D007700:Kinetics; D008565:Membrane Proteins / Q000378:metabolism*; D010457:Peptidoglycan / Q000378:metabolism*; D010710:Phosphates / Q000378:metabolism; D010957:Plasmids; D013205:Staphylococcal Protein A / Q000378:metabolism; D013211:Staphylococcus aureus / Q000187:drug effects / Q000378:metabolism*; D013379:Substrate Specificity; D014415:Tunicamycin / Q000494:pharmacology; D014538:Uridine Diphosphate N-Acetylmuramic Acid / Q000031:analogs & derivatives* / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D008565:Membrane Proteins; D010457:Peptidoglycan; D010710:Phosphates; D013205:Staphylococcal Protein A; D014538:Uridine Diphosphate N-Acetylmuramic Acid; C069249:muramyl-NAc-(pentapeptide)pyrophosphoryl-undecaprenol; D014415:Tunicamycin; D001414:Bacitracin; D019881:Aminoacyltransferases; C120542:sortase A; D003546:Cysteine Endopeptidases