Extracellular Proteins of Staphylococcus Aureus and the Role of SarA and Sigma B

Staphylococcus aureus synthesizes a large number of extracellular proteins that have been postulated to play a role in bacterial virulence. The proteomic approach was used to analyse the pattern of extracellular proteins of two different S. aureus strains, RN6390 and COL. Thirty-nine protein spots were identified by N-terminal sequencing or MALDI-TOF-MS. The differences of the extracellular protein patterns between both strains are striking. Among the 18 proteins identified in S. aureus COL there are nine proteins not yet discovered in S. aureus RN6390. These are enterotoxin B, leukotoxin D, enterotoxin, serin proteases (SplA and SplC), thermonuclease, an IgG binding protein and two so far unknown proteins in S. aureus with similarities to SceD precursor in Staphylococcus carnosus and to synergohymenotropic toxin precursor in Streptococcus intermedius. In contrast, lipase as well as staphylokinase identified in S. aureus RN6390 were not detectable in S. aureus COL under the same conditions. By using a regulatory mutant of sarA (ALC136) isogenic to strain RN6390 we identified five proteins positively regulated by SarA and 12 proteins negatively regulated by SarA. Besides V8 protease (StsP) and Hlb already described to be regulated by the sar locus new putatively sarA-dependent proteins were identified, e.g. glycerolester hydrolase and autolysin both down-regulated in the sarA mutant, and aureolysin, staphylokinase, staphopain and format tetrahydrofolate lyase up-regulated in the mutant. Moreover, the role of sigma B in expression of extracellular proteins was studied. Interestingly, we found 11 proteins at an enhanced level in a sigB mutant of S. aureus COL, among them enterotoxin B, alpha and beta hemolysin, serine proteases SplA and SplB, leukotoxin D, and staphopain homologues. The sigma B-dependent repression of gene expression occurs at the transcriptional level. Only one protein, SceD, was identified whose synthesis was down-regulated in the mutant indicating that its gene belongs to the sigma B-dependent general stress regulon.