| Project/Area Number |
10460147
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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 |
Applied molecular and cellular biology
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| Research Institution | Nagoya University |
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Principal Investigator |
SASAKI Yukiko Graduate School of Agricultural Sciences, Nagoya University, Professor, 大学院・生命農学研究科, 教授 (00026519)
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| Project Period (FY) |
1998 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥12,300,000 (Direct Cost: ¥12,300,000)
Fiscal Year 2000: ¥2,900,000 (Direct Cost: ¥2,900,000)
Fiscal Year 1999: ¥4,700,000 (Direct Cost: ¥4,700,000)
Fiscal Year 1998: ¥4,700,000 (Direct Cost: ¥4,700,000)
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| Keywords | Fatty acid biosynthesis / Acetyl-CoA carboxylase / Light-activation / Redox regulation / Pea plants / RNAエディテング / レドックスカスケード / ビオチンカルボキシラーゼ / カルボキシトランスフェラーゼ |
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| Research Abstract |
Fatty acid synthesis in pea chloroplasts is regulated by light/dark. The regulatory enzyme, acetyl-CoA carboxylase (ACCase) which catalyzes the synthesis of malonyl-CoA, is modulated by light/dark, presumably under redox regulation. The plastidic ACCase is multienzyme complex composed of biotin carboxylase and carboxyltransferase (CT), and the latter enzyme is redox-regulated. The CT is composed of the nuclear-encoded α and the chloroplast-encoded β subunits, which contains 2 and 11 cysteine residues, respectively. To demonstrate the redox-regulation of this enzyme, we identified the cysteine residues involved in the regulation and and investigated the role of the two subunits. We expressed the recombinant CT in E.coli and found that the partly deleted CT was sensitive to redox state like the full-length CT.Site-directed mutagenesis of the deleted CT showed that replacement of a cysteine by alanine at the amino acid position 267 in the α polypeptide or at position 442 in the β polypeptide resulted in redox-insensitive CT and breaking intermolecular disulfide bond between the α and β polypeptides. Similar results were confirmed in the full-length CT.These results indicated that the two cysteines are involved in redox-regulation by intermolecular disulfide/dithiol exchange between the α and β subunits. This is the first demonstration that a covalent bond between nuclear-encoded polypeptide and chloroplast-encoded polypeptide regulates the enzyme activity.
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