Getting proteins where they need to go: Signals in Staphylococcus aureus surface proteins

Surface proteins of Gram-positive bacteria fulfill many important roles during the pathogenesis of human infections. Surface proteins are covalently linked to the cell wall envelope by sortases through a mechanism requiring an N-terminal signal peptide and a C-terminal LPXTG motif sorting signal (CWS). The CWS is composed of a 5 amino acid motif region, the site of sortase cleavage, followed by a hydrophobic stretch of residues and a tail of mostly positively charged amino acids. Herein we address the sufficiency of the CWS pentapeptide motif for S. aureus SrtA and SrtB substrate specificity and examine the contributions of the hydrophobic and charged tail regions to in vivo substrate selection and protein anchoring. Our results suggest a specific role for the hydrophobic domain in vivo. We also show that the two staphylococcal sortases vary in their stringency for accepting substrates.;In addition we investigate the mechanisms whereby the surface proteins are distributed on the staphylococcal surface. We show here that protein A is unevenly distributed over the staphylococcal surface. Upon removal with trypsin, newly synthesized polypeptide is deposited at 2-4 discrete foci. During subsequent growth, protein A appears to be slowly distributed from these sites. This pattern of distribution requires the LPXTG sorting signal of protein A as well as sortase A, A. model is presented whereby protein A deposition at discrete sites coupled with cell wall synthesis enable its distribution on the staphylococcal surface.;Finally, we show that surface proteins of Staphylococcus aureus arrive at two distinct destinations in the bacterial envelope, either distributed as a ring surrounding each cell or as discrete assembly sites. Proteins with ring-like distribution (ClfA, Spa, FnbB, SdrC and SdrD) harbor signal peptides with a YSIRK/GS motif, whereas proteins directed to discrete assembly sites (SasA, SasD, SasF and SasK) do not. Reciprocal exchange of signal peptides between surface proteins with (ClfA) or without YSIRK/GS motif (SasF) directs recombinant products to the alternate destination. Thus, S. aureus distinguishes between signal peptides in order to address proteins to their proper destination in the envelope.