| Project/Area Number |
11440244
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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 |
動物生理・代謝
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| Research Institution | Hiroshima University |
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Principal Investigator |
MICHIBATA Hitoshi Hiroshima Univ., Graduate School of Science, Professor, 大学院・理学研究科, 教授 (00111740)
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| Co-Investigator(Kenkyū-buntansha) |
UEKI Tatsuya Hiroshima University, Graduate School of Science, Research Associate, 大学院・理学研究科, 助手 (10274705)
UYAMA Taro Hiroshima University, Graduate School of Science, Lecturer, 大学院・理学研究科, 講師 (60232914)
KANAMORI Kan Toyama University, Faculty of Science, Professor, 理学部, 教授 (00019001)
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| Project Period (FY) |
1999 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥14,000,000 (Direct Cost: ¥14,000,000)
Fiscal Year 2000: ¥5,800,000 (Direct Cost: ¥5,800,000)
Fiscal Year 1999: ¥8,200,000 (Direct Cost: ¥8,200,000)
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| Keywords | Ascidian / Vanadium / Accumulation / Reduction / cDNA cloning / Metal-binding / 金属濃縮 / 酸化還元 / 組み換えタンパク質 / 遺伝子クローニング / 金属結合タンパク質 |
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| Research Abstract |
During 2 years supported by the research grant, we have obtained the following results. Full-length of cDNAs, encoding 12.5 kDa and 15 kDa vanadium-associated proteins (designated newly as Vanabin), respectively, which might have a clue to resolve the mechanism of vanadium accumulation, was analyzed. As a result, vanabin was disclosed to consist of about 120 amino acids in which the content of cysteine residues is very high and to be a novel protein having α-helix. Recombinant vanabin was found to bind about 20 atoms of vanadium and its binding constant is 10-7M.In vacuoles of the vanadocytes we previously pointed out a close energetic correlation between the concentrations of vanadium ions and protons accumulated by vacuolar-type H^+-ATPases (V-ATPases) expressed on the vacuole membrane. As the first step to elucidate the energetics, we isolated cDNA encoding subunit C of V-ATPase which plays the role of regulating ATPase activity and confirmed that the pH-sensitive phenotype of the corresponding vma5 mutant of a budding yeast was rescued when the ascidian cDNA for subunit C was transformed. Furthermore, we obtained about 300 EST clones and extracted 50 proteins from the homogenate of vanadocytes using vanadium-affinity chromatography.
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