The occurrence of a new yellow enzyme in Gluconobacter strains
| Project/Area Number |
18580084
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Applied microbiology
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| Research Institution | Ube National College of Technology |
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Principal Investigator |
SHINAGAWA Emiko Ube National College of Technology, Department of Chemical and Biological Engineering, Professor (20116726)
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| Project Period (FY) |
2006 – 2007
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| Project Status |
Completed (Fiscal Year 2007)
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| Budget Amount *help |
¥3,740,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥240,000)
Fiscal Year 2007: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000)
Fiscal Year 2006: ¥2,700,000 (Direct Cost: ¥2,700,000)
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| Keywords | old yellow enzyme / acetic acid bacteria / oxidative fermentation / new yellow enzyme / NADH dehydrogenase / NADPH dehvdrogenase / 生体エネルギー |
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| Research Abstract |
A new yellow enzyme, distinct from an old yellow enzyme, was found in the same cytoplasmic fraction of Gluconobacter strains. A new yellow enzyme and an old yellow enzyme were purified from the cytoplasm of G. oxydans IFO 3244. These two enzymes were purified and crystallized. While an old yellow enzyme is NADPH dehydrogenase catalyzing the oxidation of NADPH to NADP, a new yellow enzyme was hind to be NADH dehydrogenase oxidizing NADH to NAD. Molecular mass of NADH dehydrogenase and NADPH dehydrogenase was estimated to be 120kDa and 50-60kDa, respectively. The former enzyme was consisted of four identical subunits with 301cDa and the molecular mass of subunit of the latter enzyme was 50kDa. The absorption spectra of NADH dehydrogenase showed two absorption maxima in the visible region at 445 and 340nm, with clear shoulders at 468 and 420nm, and NADPH dehydrogenase also showed two absorption maxima in the visible region, at 470 and 377 nm, with slight shoulders at 490 and 450nm. The ab
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sorption spectrum of these two enzymes was reduced by the addition of each substrate, NADH and NADPH, and the spectrum of NADH-or NADPH-reduced enzyme was restored to the original level after the enzyme solution was bubbled with oxygen. FAD and FMN were identified as the coenzyme of NADH dehydrogenase and NADPH dehydrogenase, respectively. The oxidation rate of NADH to NADPH with NADH dehydrogenase gave 3.0, and that of NADPH to NADH with NADPH dehydrogenase gave 2.0. The reaction rate by NADH dehydrogenase and also NADPH dehydrogenase was relatively low when molecular oxygen was used as the electron acceptor. The artificial electron acceptors such as p-benzoquinone and 2, 6-clichlorophenolindophenol were more effective electron acceptors than molecular oxygen for both enzymes. The N-terminal amino acid sequence of NADH dehydrogenase showed 100% similarity with a non-heme chloroperoxidase and that of NADPH dehydrogenase matched 100% a putative oxidoreductase containing the old yellow enzyme-like FMN-binding domains. NADH dehydrogenase might function to regenerate NAD coupling with NAD-dependent dehydrogenases in cytoplasm of Gluconobacter strains like NADPH dehydrogenase function to regenerate NADP coupling with NADP-dependent dehydrogenases. Less
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(3 results)
Research Products
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