Crystal Structure of the SarS Protein from Staphylococcus aureus

Abstract

The expression of virulence determinants in Staphylococcus aureus is controlled by global regulatory loci (e.g., sarA and agr). One of these determinants, protein A (spa), is activated by sarS, which encodes a 250-residue DNA-binding protein. Genetic analysis indicated that the agr locus likely mediates spa repression by suppressing the transcription of sarS. Contrary to SarA and SarR, which require homodimer formation for proper function, SarS is unusual within the SarA protein family in that it contains two homologous halves, with each half sharing sequence similarity to SarA and SarR. Here we report the 2.2 Å resolution X-ray crystal structure of the SarS protein. SarS has folds similar to those of SarR and, quite plausibly, the native SarA structure. Two typical winged-helix DNA-binding domains are connected by a well-ordered loop. The interactions between the two domains are extensive and conserved. The putative DNA-binding surface is highly positively charged. In contrast, negatively charged patches are located opposite to the DNA-binding surface. Furthermore, sequence alignment and structural comparison revealed that MarR has folds similar to those of SarR and SarS. Members of the MarR protein family have previously been implicated in the negative regulation of an efflux pump involved in multiple antibiotic resistance in many gram-negative species. We propose that MarR also belongs to the winged-helix protein family and has a similar mode of DNA binding as SarR and SarS and possibly the entire SarA protein family member. Based on the structural differences of SarR, SarS, and MarR, we further classified these winged-helix proteins to three subfamilies, SarA, SarS, and MarR. Finally, a possible transcription regulation mechanism is proposed.

Structural and function analyses of the global regulatory protein SarA from Staphylococcus aureus

“…Third, the HTH motif and ␤-hairpin (wing) showed that the SarA protein family belongs to the very well characterized superfamily of transcription factors called the ''winged-helix'' proteins. Based on potential interactions between the elongated ␤-hairpin and minor groove of their target DNAs, SarA and its homologs likely belong to a unique subfamily of winged-helix proteins (18)(19). Finally, we discovered the same structure of SarA by using different crystallization condition (see below).…”

“…We also proposed the two conserved acidic patches on the convex side of the SarR structure as possible functional motifs, whereas the concave surface, containing a tract of basic residues, is likely to be the DNA-binding side (18). Based on sequence alignment, we speculated SarR, SarA, SarS, and possibly other MarR homologs to have similar folds, which are strongly supported by the structure of SarS (13,19); however, these structures diverge significantly from the reported SarA structure with a completely different topology. Additionally, despite low sequence homology, the reported MarR structure has similar topological folds as the SarR and SarS proteins, consistent with our structural and sequence alignment predictions (20).…”

Analysis of RovA, a Transcriptional Regulator of Yersinia pseudotuberculosis Virulence That Acts through Antirepression and Direct Transcriptional Activation

“…How MarR-like proteins interact with DNA is unknown, but sequence recognition by other winged-helix proteins is generally achieved by interaction of the HTH recognition helix with DNA. In addition, the wings can contact the DNA in the minor groove and/or the phosphate backbone (20,36,37). The isolated amino acid changes in the central region of RovA locate in the postulated recognition helix ␣ 4 or in the stabilization helix ␣ 3 of the HTH motif (Fig.…”

Regulatory Elements of theStaphylococcus aureusProtein A (Spa) Promoter

“…SarS has been reported to be a DNA binding protein, although its binding site on the spa promoter has not been specifically identified (8,25,42). When the results of the study of the effect of Agr on the promoter constructs (Fig.…”