Purification and characterization of sortase, the transpeptidase that cleaves surface proteins of Staphylococcus aureus at the LPXTG motif

Surface proteins of Staphylococcus aureus are linked to the bacterial cell wall by sortase, an enzyme that cleaves polypeptides at the threonine of th e LPXTG motif. Surface proteins can be released from staphylococci by treat ment with hydroxylamine, resulting in the formation of threonine hydroxamat e. Staphylococcal extracts, as well as purified sortase, catalyze the hydro xylaminolysis of peptides bearing an LPXTG motif, a reaction that can be in hibited with sulfhydryl-modifying reagents. Replacement of the single conse rved cysteine at position 184 of sortase with alanine abolishes enzyme acti vity. Thus, sortase appears to catalyze surface-protein anchoring by means of a transpeptidation reaction that captures cleaved polypeptides as thioes ter enzyme intermediates.