Crystal structure of the SarR protein from Staphylococcus aureus

Abstract

The expression of virulence determinants in Staphylococcus aureus is controlled by global regulatory loci (e.g., sarA and agr). The sar (Staphylococcus accessory regulator) locus is composed of three overlapping transcripts (sarA P1, P3, and P2, transcripts initiated from the P1, P3, and P2 promoters, respectively), all encoding the 124-aa SarA protein. The level of SarA, the major regulatory protein, is partially controlled by the differential activation of the sarA promoters. We previously partially purified a 13.6-kDa protein, designated SarR, that binds to the sarA promoter region to down-modulate sarA transcription from the P1 promoter and subsequently SarA expression. SarR shares sequence similarity to SarA, and another SarA homolog, SarS. Here we report the 2.3 Å-resolution x-ray crystal structure of the dimeric SarR-MBP (maltose binding protein) fusion protein. The structure reveals that the SarR protein not only has a classic helix-turn-helix module for DNA binding at the major grooves, but also has an additional loop region involved in DNA recognition at the minor grooves. This interaction mode could represent a new functional class of the ''winged helix'' family. The dimeric SarR structure could accommodate an unusually long stretch of Ϸ27 nucleotides with two or four bending points along the course, which could lead to the bending of DNA by 90°or more, similar to that seen in the catabolite activator protein (CAP)-DNA complex. The structure also demonstrates the molecular basis for the stable dimerization of the SarR monomers and possible motifs for interaction with other proteins.

Identification of amino acid residues of Salmonella SlyA that are critical for transcriptional regulation

“…The reported structures of MarR family proteins show that the dimerization domain includes a-helices in the Nterminal and C-terminal regions of each monomer (Alekshun et al, 2001;De Silva et al, 2005;Hong et al, 2005;Lim et al, 2002;Liu et al, 2001;Wu et al, 2003). The protein-protein interaction is primarily mediated by a number of hydrophobic interactions, and is further stabilized by hydrogen bonds and salt-bridge pairs.…”

“…Although the amino acid sequence similarity of this SlyA is relatively low, we demonstrated that the structure of this SlyA is very similar to the structures of the other MarR family proteins. The crystal structures of this protein family (which includes MarR, MexR, SarR, SlyA-Ef, AphA and OhrR) revealed that the protein forms a homodimer, with each subunit possessing a winged-helix DNA binding motif, which is required for DNA binding (Alekshun et al, 2001;De Silva et al, 2005;Hong et al, 2005;Lim et al, 2002;Liu et al, 2001;Wu et al, 2003). Furthermore, winged-helix proteins characteristically bind to DNA sequences containing a palindromic or pseudopalindromic structure.…”

“…These regulatory proteins control the expression of resistance to multiple antibiotics, organic solvents, agents that induce oxidative stress, and virulence factors (Alekshun & Levy, 1997). This diverse group of MarR homologues is very similar with respect to protein structure; the protein forms a dimer and contains a wingedhelix DNA-binding motif (Alekshun et al, 2001;De Silva et al, 2005;Hong et al, 2005;Lim et al, 2002;Liu et al, 2001;Wu et al, 2003). In these regulatory proteins, the helix-turn-helix motif is followed by a wing composed of two antiparallel b-strands.…”

SarA ofStaphylococcus aureusBinds to thesarAPromoter To Regulate Gene Expression

“…Cognizant of our recent structural data showing that SarA is a dimeric winged helix protein (13), it is likely the binding site of SarA is greater than 17 bp in size and contains either a palindrome or an inverted repeat (based on our unpublished gel shift data with various sizes of the duplex DNA fragments and purified SarA protein), which is typical of most winged helix proteins with two winged helix motifs (e.g., OhrR, MecI, and BlaI), with each motif capable of binding to the major groove and minor groove of target DNA (11,12,21,22). Accordingly, the data presented here are consistent with a larger binding site for two winged helix motifs.…”

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

“…Based on the reported structures, they proposed a ''shortening'' effect of DNA as a result of binding by the dimeric SarA protein, leading to gene activation (17). Independently, we reported the dimeric SarR structure to be a typical ''winged-helix DNAbinding protein'' in the absence DNA, with the helix-turn-helix (HTH) and the winged regions interacting with the major and minor grooves of target promoter DNA, respectively (18). 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).…”