长庚大学生物医学研究所微生物组唾液链球菌尿素酶基因组表现之.PDF

長庚大學生物醫學研究所微生物組 唾液鏈球菌尿素酶基因組表現之調控分析 Molecular analysis of the urease regulatory circuit in Streptococcus salivarius 指導教授：陳怡原副教授 研 究 生：黃司權 0 Abstract Background: Streptococcus salivarius is a highly ureolytic oral bacterium. Urease expression in S. salivarius 57.I is derepressed at acidic growth pH, and the expression is further enhanced by excess amount of carbohydrate and fast growth rate. A consensus sequence for the global regulator CodY was recently identified 14-bases 5’ to the -35 sequence of pureI. The activity of CodY is modulated by the presence of GTP and branched chain amino acid in low G+C Gram positive bacteria, and the regulation by CodY ensures an accurate sensing of intracellular energy pool that allows for an optimal adaptation of the bacteria to various growth conditions. The goal of this study was to investigate the impact of CodY in the differential expression of ure operon. Methods: Both the wild-type and CodY knockout strains were grown in chemostat under glucose-excess (100 mM) and -limiting (20 mM) conditions, at pH 7.0 and 5.5. The expression levels of pureI and urease were evaluated by RT-PCR and urease assay, respectively. The impact of CodY in ure operon expression was further confirmed in the CodY-complementation strain. The direct interaction between CodY and pureI was demonstrated by EMSA and ChIP assay. Results: Both the pureI expression and urease activity were up-regulated in the CodY-deficient strain compared to the wild-type strain under all growth conditions used, and the