An iron-regulated sortase anchors a class of surface protein during Staphylococcus aureus pathogenesis

Abstract

Sortase (SrtA), an enzyme that anchors surface proteins to the cell wall of Gram-positive bacteria, cleaves sorting signals at the LPXTG motif. We have identified a second sortase (SrtB) in the Gram-positive pathogen Staphylococcus aureus that is required for anchoring of a surface protein with a NPQTN motif. Purified SrtB cleaves NPQTN-bearing peptides in vitro, and a srtB mutant is defective in the persistence of animal infections. srtB is part of an iron-regulated locus called iron-responsive surface determinants (isd), which also contains a ferrichrome transporter and surface proteins with NPQTN and LPXTG motifs. Cell wall-anchored surface proteins and the isd locus seem involved in a novel mechanism of iron acquisition that is important for bacterial pathogenesis.

Engineering the Substrate Specificity of Staphylococcus aureus Sortase A

“…(33,39). By contrast, SrtB had no detectable activity against the LPETG substrate, even after 12 h at 37°C and an enzyme concentration of 94.8 M. This is consistent with earlier studies in which no LPETG-cleaving activity was observed for SrtB in vitro (22). SrtLS was able to hydrolyze the LPETG substrate but with a vastly reduced k cat /K m app of 3.7 Ϯ 0.6 ϫ 10 Ϫ4 M Ϫ1 s Ϫ1 (Table 2).…”

“…SrtA anchors LPXTGcontaining proteins, whereas SrtB is specific for NPQTN-containing proteins (22,26). In order to better understand the basis for this specific recognition, we used sequence analysis to look at SrtA and SrtB isoforms from various Gram-positive bacteria.…”

“…First, SrtB has a specificity profile different from that of SrtA. It selectively anchors the NPQTN-containing protein IsdC, and it does not anchor LPXTG-containing proteins (22). Second, the anchor structure for SrtB is not Lipid II (a peptidoglycan biosynthetic precursor) but is the Gly 5 cross-bridge of mature peptidoglycan from the staphylococcal cell wall (23).…”

“…Second, the anchor structure for SrtB is not Lipid II (a peptidoglycan biosynthetic precursor) but is the Gly 5 cross-bridge of mature peptidoglycan from the staphylococcal cell wall (23). In addition, the gene for SrtB is located along with its substrate in an ironresponsive operon, which is only transcribed under conditions of iron starvation (22). Since the acquisition of iron is an essential process for bacterial colonization of mammalian host tissues (24), it is likely that SrtB carries out an essential function for virulence by anchoring a protein with a specific role in iron acquisition to a unique anchor motif in the cell wall (25).…”

Sortase D Forms the Covalent Bond That Links BcpB to the Tip of Bacillus cereus Pili

“…Sortase A cleaves secreted protein at LPXTG sorting signals and recognizes the amino group of lipid II peptidoglycan precursors as a nucleophile (8,9). Sortase B cleaves protein substrates at NPQTN sorting signals (10). This enzyme immobilizes proteins within fully assembled cell walls, utilizing the cell wall cross-bridge as a nucleophile (11).…”