| Project/Area Number |
06453168
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
応用微生物学・応用生物化学
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
KUMAGAI Hidehiko Kyoto University, Faculty of Agriculture, Professor, 農学部, 教授 (70027192)
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| Co-Investigator(Kenkyū-buntansha) |
TAMAKI Hisanori Kyoto University, Faculty of Agriculture, Instructor, 農学部, 助手 (20212045)
SUZUKI Hideyuki Kyoto University, Faculty of Agriculture, Instructor, 農学部, 助手 (10202136)
YAMAMOTO Kenji Kyoto University, Faculty of Agriculture, Associate Professor, 農学部, 助教授 (70109049)
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| Project Period (FY) |
1994 – 1996
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| Project Status |
Completed (Fiscal Year 1996)
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| Budget Amount *help |
¥7,400,000 (Direct Cost: ¥7,400,000)
Fiscal Year 1996: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 1995: ¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1994: ¥4,000,000 (Direct Cost: ¥4,000,000)
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| Keywords | gamma-glutamyltranspeptidase / monoamine oxidase / tyrosine phenol-lyase / Bifidobacterium / beta-glucosidase / amine oxidase / glutathione S-transferase / モノアミン酸化酵素 / アミン酸化酵素 / グルタチオンS-トランスフェラーゼ / X線結晶解析 / γ-グルタミルトランスフェラーゼ |
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| Research Abstract |
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.
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