| Project/Area Number |
07680628
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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 |
Bioorganic chemistry
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| Research Institution | Nagoya University |
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Principal Investigator |
ICHIBA Ikuko Nagoya University, School of Agricutural Sciences, Assistant Professor, 農学部, 助手 (40247680)
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| Co-Investigator(Kenkyū-buntansha) |
ICHIKAWA Yoshiyasu Nagoya University, School of Agicutural Sciences, Associate Professor, 農学部, 助教授 (60193439)
ISOBE Minoru Nagoya University, School of Agricutural Sciences, Professor, 農学部, 教授 (00023466)
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| Project Period (FY) |
1995 – 1996
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| Project Status |
Completed (Fiscal Year 1996)
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| Budget Amount *help |
¥2,300,000 (Direct Cost: ¥2,300,000)
Fiscal Year 1996: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 1995: ¥1,100,000 (Direct Cost: ¥1,100,000)
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| Keywords | tautomycin / protein phosphatase inhibitor / firefly bioluminescence system / molecular mechanics calculation / オカダ酸 |
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| Research Abstract |
We have established a new assay method for protein phosphatases type 1 (PP1) and type 2A (PP2A) inhibitors, in which the activities are monitored by a firefly bioluminescence system. The established method successfully determined the activities of known inhibitors, i.e., okadaic acid, calyculin A,microcystin-LR,and tautomycin, using less than 10pmol of samples. Tautomycin exists in two forms as equilibrium (acid anhydride and diacid). We successfully isolated them and proved that diacid is the real active form of tautomyicin. DELTA^<21,22> and 22-deoxy-tautomycin derivatives were prepared for structure-activity relationship studies. None of them showed PP2A inhibitory activities in our assay system, therefore flexibility around the C19 to the C23 and hydroxyl group at the C22 maybe indispensable for PP22A inhibitory activity. Trans-esterification mechanism on alkali degradation of tautomycin was proposed, which proved that hydroxyl group at C22 is near to C-1' in tautomycin molecule.
Stable conformation of okadaic acid was calculated by a molecular mechanics calculation based on NMR data. Obtained conformation was superimposable on that determined by X-ray analysis. Same strategy was adapted to tautomycin diacid (active form) to give one of the stable conformations that supports the trans-esterification mechanism. Similar conformation was obtained for 22-deoxytautomycin diacid (inactive) by a combination of molecular mechanics calculation and NMR analysis. This evidence suggested that hydroxyl group on C-22 is essential for protein phosphatase inhibitory activity.
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