title: Two DNA-binding domains of Mga are required for virulence gene activation in the group A streptococcus.
abstract: Mga is a DNA-binding protein that activates expression of several important virulence genes in the group A streptococcus (GAS), including those encoding M protein (emm), C5a peptidase (scpA) and Mga (mga). To determine the functionality of four potential helix-turn-helix DNA-binding motifs (HTH1-HTH4) identified within the amino-terminus of Mga, alanine substitutions were introduced within each domain in a MBP-Mga fusion allele and purified proteins were assayed for binding to Mga-specific promoter fragments (Pmga, PscpA and Pemm) in vitro. Although HTH-1 and HTH-2 mutations showed wild type DNA-binding activity, an altered HTH-3 domain resulted in reduced binding to the three promoters and an HTH-4 mutant was devoid of detectable binding activity. Plasmid-encoded expression of the HTH-3 and HTH-4 alleles from a constitutive promoter (Pspac) in the mga-deleted GAS strain JRS519 demonstrated that Mga-regulated emm expression correlated directly to the DNA-binding activity observed for each mutant protein in vitro. Single-copy expression of HTH-3 and HTH-4 from their native Pmga resulted in a dramatic reduction in autoregulated mga expression in both mutant strains. Thus, Mga appears to contain two DNA-binding domains (HTH-3 and HTH-4) that are required for direct activation of the Mga virulence regulon in vivo.
authors.affiliation: Department of Microbiology, University of Texas Southwestern Medical Center, Dallas, TX 75390-9048, USA. Kevin.Mciver@UTSouthwestern.edu
authors.forename: Kevin S
authors.initials: KS
authors.lastname: McIver
authors.forename: Rhonda L
authors.initials: RL
authors.lastname: Myles
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry; D015964:Gene Expression Regulation, Bacterial; D019077:Helix-Turn-Helix Motifs / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010957:Plasmids / Q000235:genetics; D013297:Streptococcus pyogenes / Q000235:genetics / Q000254:growth & development / Q000472:pathogenicity*; D013329:Structure-Activity Relationship; D015533:Transcriptional Activation; D014774:Virulence / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C068267:mry protein, Streptococcus pyogenes