title: Anchoring of surface proteins to the cell wall of Staphylococcus aureus. A conserved arginine residue is required for efficient catalysis of sortase A.
abstract: Surface proteins of Staphylococcus aureus are anchored to the cell wall envelope by a mechanism requiring a C-terminal sorting signal with an LPXTG motif. Sortase A cleaves surface proteins between the threonine (T) and the glycine (G) residues of the LPXTG motif and catalyzes the formation of an amide bond between the carboxyl group of threonine at the C-terminal end of polypeptides and the amino group of pentaglycine cross-bridges of cell wall peptidoglycan. Previous work showed that Cys(184) and His(120) of sortase A are absolutely essential for catalysis; however an active site thiolateimidazolium ion pair may not be formed. The three-dimensional crystal structure of sortase A revealed that Arg(197) is located in close proximity to both the active site Cys(184) and the scissile peptide bond between threonine and glycine. We show here that substitution of Arg(197) with alanine, lysine, or histidine severely reduced sortase A function both in vivo and in vitro, whereas Asn(98), which had earlier been implicated in hydrogen bonding to His(120), was found to be dispensable for catalysis. As the structural proximity of Arg(197) and Cys(184) is conserved in sortase enzymes and as ionization of the Cys(184) sulfhydryl group seems required for sortase activity, we propose that Arg(197) may function as a base, facilitating thiolate formation during sortase-mediated cleavage and transpeptidation reactions.
authors.affiliation: Committee on Microbiology, University of Chicago, Chicago, IL 60637, USA.
authors.forename: Luciano A
authors.initials: LA
authors.lastname: Marraffini
authors.forename: Hung
authors.initials: H
authors.lastname: Ton-That
authors.forename: Yinong
authors.initials: Y
authors.lastname: Zong
authors.forename: Sthanam V L
authors.initials: SV
authors.lastname: Narayana
authors.forename: Olaf
authors.initials: O
authors.lastname: Schneewind
mesh_terms: D019881:Aminoacyltransferases / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D001483:Base Sequence; D002384:Catalysis; D002473:Cell Wall / Q000378:metabolism*; D003546:Cysteine Endopeptidases; D017931:DNA Primers; D016297:Mutagenesis, Site-Directed; D013211:Staphylococcus aureus / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D017931:DNA Primers; D019881:Aminoacyltransferases; C120542:sortase A; D003546:Cysteine Endopeptidases