| Summary: | 碩士 === 國立成功大學 === 微生物及免疫學研究所 === 98 === Streptococcus pyogenes, also known as group A Streptococcus (GAS), is a Gram-positive pathogen, which causes a variety of human diseases. In this study, the crystal structure of a stress response regulator, PerR, from GAS has been determined. PerR is a repressor to block transcription initiation by binding to promoter region. In GAS, metals and hydrogen peroxide stresses abolish the DNA-binding ability of PerR. Recombinant GAS PerR protein can be successfully expressed in Escherichia coli and purified by Ni2+-NTA affinity chromatography. GAS PerR is characterized to a dimeric protein which displays the ability to bind dpr promoter DNA containing a consensus sequence, Per box. However, the crystal structure shows that the GAS PerR is a metalloprotein and is captured in an inactive conformation containing two zinc ions per monomer. One of the zinc is located at the conserved zinc-finger motif which is composed by Cys104, Cys107, Cys144, and Cys147; the other zinc coordinates in a pseudo octahedral geometry composed by six residues including His4, His6, Asn15, His19, His97, and His99. Both sites of the zinc coordination are critical for PerR biological function. Mutations at any residues involving in metal-binding sites lead to abolish the DNA-binding ability. Zinc binds to regulatory site and locks the conformation of GAS PerR in inactive form. Hydrogen peroxide also causes the dissociation of PerR from dimer to monomer. To summarize, in this study we have solved the crystal structure of PerR and discovered that the regulation of DNA-binding activity of GAS PerR are different from other homologous PerR and Fur family proteins. We propose that GAS PerR possesses dual sensing mechanisms to respond to metal ion and hydrogen peroxide stresses and regulate oxidative stress response.
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