Anchor Structure of Staphylococcal Surface Proteins

Abstract

Surface proteins of Staphylococcus aureus are covalently linked to the bacterial cell wall by a mechanism requiring a COOH-terminal sorting signal with a conserved LPXTG motif. Cleavage between the threonine and the glycine of the LPXTG motif liberates the carboxyl of threonine to form an amide bond with the pentaglycyl cross-bridge in the staphylococcal peptidoglycan. Here, we asked whether altered peptidoglycan cross-bridges interfere with the sorting reaction and investigated surface protein anchoring in staphylococcal fem mutants. S. aureus strains carrying mutations in the femA, femB, femAB, or the femAX genes synthesize altered cross-bridges, and each of these strains displayed decreased sorting activity. Characterization of cell wall anchor structures purified from the fem mutants revealed that surface proteins were linked to cross-bridges containing one, three, or five glycyl residues, but not to the ⑀-amino of lysyl in muropeptides without glycine. When tested in a femAB strain synthesizing cross-bridges with mono-, tri-, and pentaglycyl as well as tetraglycyl-monoseryl, surface proteins were found anchored mostly to the five-residue cross-bridges (pentaglycyl or tetraglycyl-monoseryl). Thus, although wild-type peptidoglycan appears to be the preferred substrate for the sorting reaction, altered cell wall cross-bridges can be linked to the COOH-terminal end of surface proteins.

Amide bonds assemble pili on the surface of bacilli

“…Bacilli were grown on LB agar, suspended in water, and centrifuged and pili in the supernatant were digested with CNBr (29). Reactions were dried, solubilized and peptides chromatographed on Ni-NTA Sepharose.…”

Crystal Structures of Staphylococcus aureus Sortase A and Its Substrate Complex

“…Several observations suggest that lipid II, a membrane anchored intermediate of cell wall synthesis, functions as the peptidoglycan substrate of sortase A (10,11). The product of the complete sorting reaction, surface protein tethered to lipid II, is presumed to be incorporated into the cell wall envelope via the penicillin-sensitive transpeptidation and transglycosylation reactions of peptidoglycan synthesis (10,12).…”

Activation of Inhibitors by Sortase Triggers Irreversible Modification of the Active Site

“…Nucleophilic attack of the amino group of the peptidoglycan precursor lipid II (C 55 -PP-MurNAc-(L-Ala-D-iGln-LLys(NH 2 -Gly 5 )-D-Ala-D-Ala)-GlcNAc) at the thioester intermediate resolves the acyl enzyme and forms an amide bond between the C-terminal threonine of surface protein and pentaglycine crossbridges (9). Lipid II-linked polypeptide is subsequently incorporated into the cell wall envelope of staphylococci (10). The final product of this pathway, protein linked to cell wall pentaglycine cross-bridges, is displayed on the bacterial surface and enables interactions between the pathogen and tissues of its host.…”