1988 Fiscal Year Final Research Report Summary
Cloning of UbiA gene coding an ubiquinone synthesis key enzyme and its expression in E. coli
| Project/Area Number |
62560106
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
発酵・醸造
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| Research Institution | Shimane University |
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Principal Investigator |
MATSUDA Hideyuki Professor, Shimane Univ., Faculty of Agriculture, 農学部, 教授 (50032595)
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| Co-Investigator(Kenkyū-buntansha) |
OCHIAI Hideo Professor, Shimane Univ., Faculty of Agriculture, 農学部, 教授 (10032971)
MORI Tadahiro Professor, Shimane Univ., Faculty of Agriculture, 農学部, 教授 (20166359)
KAWAMUKAI Makoto Associate Professor, Shimane Univ., Faculty of Agriculture, 農学部, 助教授 (70186138)
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| Project Period (FY) |
1987 – 1988
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| Keywords | Ubiquinone-10 / ubiA gene / p-Hydroxybenzoate-polyprenylpyrophosphate / Ubiquinone synthesis key enzyme / 光合成細菌のユビキノン |
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| Research Abstract |
1. The ubiA gene fragment coding a key enzyme in ubiquinone synthesis, p-hydroxybenzoate-polyprenylpyrophosphate(pHB-PPP) transferase, was purified from phage DNA 12B4 containing the ubiA gene region of E. coli genomic library. Recombinant plasmid consisting of the obtained ubiA fragment and E. coli cloning vector pUC18 was constructed and then cloned in E. coli JM109.
2. The recombinant plasmid, pUBA, was transformed into E. coli AN385 (ubiA mutant) which was unable to synthesize ubiquinone. The obtained transformant cell synthesized apparently ubiquinone-8. The results show that the ubiA gene in plasmid pUBA is expressed in E. coli AN385.
3. In order to transform pUBA into Photosynthetic bacteria, a broad-host-range cloning vector, RSF1010(Sm, Su ), was transformed into Rhodo-pseudomonas sphaeroides KA38 by the method of Fornari and Kaplan. Furthermore, a new plasmid was constructed from RSF1010 and pUBA, and then transformed into R. sphaeroides.
4.Transformation frequencies of the plasmids into E. coli and R. sphaeroides were increased by electroinjection method. The expression of ubiA gene in R. sphaeroides and a possibility of the over production of ubiquinone-10 were discussed.
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