Browsing by Author "Cai, Tao"
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- Regulatory targets of quorum sensing in Vibrio cholerae: evidence for two distinct HapR-binding motifsTsou, Amy M.; Cai, Tao; Liu, Zhi; Zhu, Jun; Kulkarni, Rahul V. (2009-05)The quorum-sensing pathway in Vibrio cholerae controls the expression of the master regulator HapR, which in turn regulates several important processes such as virulence factor production and biofilm formation. While HapR is known to control several important phenotypes, there are only a few target genes known to be transcriptionally regulated by HapR. In this work, we combine bioinformatic analysis with experimental validation to discover a set of novel direct targets of HapR. Our results provide evidence for two distinct binding motifs for HapR-regulated genes in V. cholerae. The first binding motif is similar to the motifs recently discovered for orthologs of HapR in V. harveyi and V. vulnificus. However, our results demonstrate that this binding motif can be of variable length in V. cholerae. The second binding motif shares common elements with the first motif, but is of fixed length and lacks dyad symmetry at the ends. The contributions of different bases to HapR binding for this second motif were demonstrated using systematic mutagenesis experiments. The current analysis presents an approach for systematically expanding our knowledge of the quorum-sensing regulon in V. cholerae and other related bacteria.
- Thermal stability, pH dependence and inhibition of four murine kynurenine aminotransferasesHan, Qian; Cai, Tao; Tagle, Danilo A.; Li, Jianyong (2010-05-19)Background Kynurenine aminotransferase (KAT) catalyzes the transamination of kynunrenine to kynurenic acid (KYNA). KYNA is a neuroactive compound and functions as an antagonist of alpha7-nicotinic acetylcholine receptors and is the only known endogenous antagonist of N-methyl-D-aspartate receptors. Four KAT enzymes, KAT I/glutamine transaminase K/cysteine conjugate beta-lyase 1, KAT II/aminoadipate aminotransferase, KAT III/cysteine conjugate beta-lyase 2, and KAT IV/glutamic-oxaloacetic transaminase 2/mitochondrial aspartate aminotransferase, have been reported in mammalian brains. Because of the substrate overlap of the four KAT enzymes, it is difficult to assay the specific activity of each KAT in animal brains. Results This study concerns the functional expression and comparative characterization of KAT I, II, III, and IV from mice. At the applied test conditions, equimolar tryptophan with kynurenine significantly inhibited only mouse KAT I and IV, equimolar methionine inhibited only mouse KAT III and equimolar aspartate inhibited only mouse KAT IV. The activity of mouse KAT II was not significantly inhibited by any proteinogenic amino acids at equimolar concentrations. pH optima, temperature preferences of four KATs were also tested in this study. Midpoint temperatures of the protein melting, half life values at 65°C, and pKa values of mouse KAT I, II, III, and IV were 69.8, 65.9, 64.8 and 66.5°C; 69.7, 27.4, 3.9 and 6.5 min; pH 7.6, 5.7, 8.7 and 6.9, respectively. Conclusion The characteristics reported here could be used to develop specific assay methods for each of the four murine KATs. These specific assays could be used to identify which KAT is affected in mouse models for research and to develop small molecule drugs for prevention and treatment of KAT-involved human diseases.