| Project/Area Number |
61480468
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
代謝生物化学
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| Research Institution | Jichi Medical School |
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Principal Investigator |
SONE Nobuhito Dept. of Biochem, Jichi Medical School, Associate Prof.., 医学部, 助教授 (20049034)
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| Co-Investigator(Kenkyū-buntansha) |
ISHIZUKA Morio Dept. of Industrial Chem., Tyuo Univ. Associate Prof., 理工学部, 助教授 (50168241)
KAGAWA Yasuo Dept. of Biochem.,Jichi Medical School, Prof., 医学部, 教授 (30048962)
SAIGO Satoshi Dept. of Physics, Jichi Medical School, Associate Prof., 医学部, 助教授 (70049071)
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| Project Period (FY) |
1986 – 1988
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| Project Status |
Completed (Fiscal Year 1988)
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| Budget Amount *help |
¥6,000,000 (Direct Cost: ¥6,000,000)
Fiscal Year 1988: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1987: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 1986: ¥3,500,000 (Direct Cost: ¥3,500,000)
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| Keywords | Cytochrome oxidase / Thermophilic bacterium / Proton pump / Gene structure / Crystalization of membrane protein / DNA sequence / タン白質一次構造 / 耐熱性酵素 / 遺伝子構造 / アミノ酸配列 / 好熱菌 / チトクロム酸化酵素 / 膜タンパク質の結晶化 / 高速電子線照射実験による標的分子量 / 好熱菌のチトクロム酸化酵素 / 膜蛋白質の結晶化 |
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| Research Abstract |
1. The genes for subunits of cytochrome oxidase were cloned from a library in EMBL-3 of the thermophilic bacterium PS3 and sequenced. The genes formed an operon in the order of subunits II,I,III and IV. Comparison of the amino acid sequences of each subunit with those from mitochondria and paracoccus denitrificans tells us places of important amino acid residues and a rough image of three-dimensional structure.
2. Crystalization of the enzyme was tried. However, only small crystals seemed to appear, when octylglucoside and heptane triol were added to the mother liquor, which was not suitable for crystalography.
3. Target size of PS3 cytochrome oxidase in respect to both oxidase and proton pump activities were measured. Dimeric structure may be necessary for proton pumping, whereas the monomer can mediate oxidation. The enzyme exist as monomer in several non-ionic detergent solution. Dimeric enzyme may be vormed in the bed of phospholipids.
4. Several methods to measure proton pump activity at the cytochrome oxidase-level have been developed, and applied on several preparations from various bacteria. After consulting several papers on this topics, a review on respiratory complexes in bacteria generating electrochemical proton gradients have been written,
5. Introduction of an expression system will make it possible to confirm putative relationship between structure and function in cytochrome oxidase.
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