Studies on structure and function of amine and amino acid related enzymes from microbiology.

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1. (1) Lys-257 of tyrosine phenol-lyase was mutated into Ala. And the strain over expresses the mutant enzyme was constructed using E.coli and T7 phage. (2) Transcription initiation site and regulation sites of tyrosine phenol-lyase gene was analyzed.2. (1) Structure of the main chain of gamma-glutamyltranspeptidase was determined. (2) Ser-33 of gamma-glutamyltranspeptidase was mutated into Cys. Pb^<2+> derivatives of the enzyme was analyzed. Mechanism of low temperature induction of the enzyme was analyzed. The induction was regulated by transcriptional regulation.3. (1) Monoamine oxidase of E.coli was induced by Cu^<2+> ion and the induction was regulated by transcriptional regulation. (2) The enzyme was induced by tyramine and also regulated by catabolite repression. (3) Over-expression of the regulator protein results in production of yellow enzyme. The yellow enzyme was purified and characterized. Its crystal structure was also studied.4.Copper-binding His residues and active center amino acid residues of amine oxidase of fungi were mutagenized.5.Isoenzyme gene of glutathione S-transferase was cloned from yeast. Over-expressed enzyme was purified and characterized.6.beta-Glucosidase gene of bifidobacterium was cloned in E.coli. Over-expressed enzyme was purified, characterized, and crystallized.