title: Transcriptional regulation of the agr locus and the identification of DNA binding residues of the global regulatory protein SarR in Staphylococcus aureus.
abstract: Many members of the transcriptional regulator SarA protein family are winged-helix proteins that are involved in gene regulation essential to Staphylococcus aureus pathogenesis. Investigation of the mechanism by which this family of genes acts at the molecular level will likely contribute to the understanding of the pathogenesis process and enhance the potential for the development of inhibitors capable of controlling S. aureus infections. Our previously published studies demonstrate that SarR is a repressor of sarA expression. Here, we show that SarR (a member of the SarA protein family) in part regulates agr expression by direct binding to the agr intergenic promoter region as determined by gel shift and DNase I footprinting assays. Analysis of the double sarA/sarR mutant in the early phase of growth reveals its significant role in regulating agr expression as compared with single mutants. Based on the previously reported crystal structure of SarR, we conducted site-specific mutagenesis and demonstrate that K52 residues within helix-turn-helix (HTH), K80, R82 and R88 (in the wing) and L105 (in the alpha5 helix) are important for DNA binding. Interestingly, SarR and SarA binding sites on the agr promoter are confined within the same region of DNA. Additional gel shift studies with SarR and SarA suggest that these two proteins may bind the same region of the agr promoter.
authors.affiliation: Division of Basic Biomedical Sciences, University of South Dakota School of Medicine, Vermillion, SD 57069, USA.
authors.forename: Adhar C
authors.initials: AC
authors.lastname: Manna
authors.forename: Ambrose L
authors.initials: AL
authors.lastname: Cheung
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D015964:Gene Expression Regulation, Bacterial; D006801:Humans; D008969:Molecular Sequence Data; D005810:Multigene Family; D016297:Mutagenesis, Site-Directed; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D016415:Sequence Alignment; D013211:Staphylococcus aureus / Q000235:genetics* / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic
chemical_list: C095698:Agr protein, Staphylococcus aureus; D001426:Bacterial Proteins; C492107:SarA protein, Staphylococcus aureus; D015534:Trans-Activators