Abstract
The cell wall envelope of staphylococci and other Gram-positive pathogens is coated with surface proteins that interact with human host tissues. Surface proteins of Staphylococcus aureus are covalently linked to the cell wall envelope by a mechanism requiring C-terminal sorting signals with an LPXTG motif. Sortase (SrtA) cleaves surface proteins between the threonine (T) and the glycine (G) of the LPXTG motif and catalyzes the formation of an amide bond between threonine at the C-terminal end of polypeptides and cell wall cross-bridges. The active site architecture and catalytic mechanism of sortase A has hitherto not been revealed. Here we present the crystal structures of native SrtA, of an active site mutant of SrtA, and of the mutant SrtA complexed with its substrate LPETG peptide and describe the substrate binding pocket of the enzyme. Highly conserved proline (P) and threonine (T) residues of the LPXTG motif are held in position by hydrophobic contacts, whereas the glutamic acid residue (E) at the X position points out into the solvent. The scissile T-G peptide bond is positioned between the active site Cys(184) and Arg(197) residues and at a greater distance from the imidazolium side chain of His(120). All three residues, His(120), Cys(184), and Arg(197), are conserved in sortase enzymes from Gram-positive bacteria. Comparison of the active sites of S. aureus sortase A and sortase B provides insight into substrate specificity and suggests a universal sortase-catalyzed mechanism of bacterial surface protein anchoring in Gram-positive bacteria.
Highlights
Many bacteria adhere to host extracellular matrix proteins as an essential first step toward the pathogenesis of infection [1]
With a novel but identical structural fold compared with all other cysteine proteases, it is interesting to note that S. aureus SrtA and SrtB recognize almost identical, but subtly different, sorting motifs LPETG and NPQTN, respectively
With the help of the SrtA⌬N59Cys184Ala ϩ LPETG peptide complex crystal structure, we provide the supporting structural evidence for the recent observation regarding the absence of the thiolate-imidazolium ion pair S. aureus sortase A by Connollay et al [32], and reveal that the conserved His residue in the active site of sortases is not suitably positioned to play the conventional protonator/deprotonator role in a peptide transfer reaction
Summary
Surface proteins of Staphylococcus aureus are covalently linked to the cell wall envelope by a mechanism requiring C-terminal sorting signals with an LPXTG motif. Sortase (SrtA) cleaves surface proteins between the threonine (T) and the glycine (G) of the LPXTG motif and catalyzes the formation of an amide bond between threonine at the C-terminal end of polypeptides and cell wall crossbridges. In Gram-positive bacteria, the cell wall envelope serves as a surface organelle with immobilized surface proteins that are responsible for mediating adhesion to host tissues [2] Many of these surface proteins are covalently linked to the cell wall peptidoglycan by a mechanism requiring a C-terminal sorting signal with the conserved LPXTG motif (where X represents any amino acid) [3]. Previous determination of the three-dimensional structure of SrtA⌬N59 using NMR revealed three residues, Cys184 together with
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
