| Project/Area Number |
09480167
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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 |
Functional biochemistry
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| Research Institution | NIPPON MEDICAL SCHOOL |
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Principal Investigator |
NISHINO Takeshi NIPPON MEDICAL SCHOOL, BIOCHEMISTRY AND MOLECULAR BIOLOGY, Professor, 医学部, 教授 (40094312)
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| Co-Investigator(Kenkyū-buntansha) |
OKAMOTO Ken NIPPON MEDICAL SCHOOL, BIOCHEMISTRY AND MOLECULAR BI0LOGY, Assistant, 医学部, 助手 (60267143)
MATSUMURA Tomohiro NIPPON MEDICAL SCHOOL, BIOCHEMISTRY AND MOLECULAR BIOLOGY, Assistant, 医学部, 助手 (20297930)
IWASAKI Toshio NIPPON MEDICAL SCHOOL, BIOCHEMISTRY AND MOLECULAR BIOLOGY, Lecturer, 医学部, 講師 (40277497)
堀 弘幸 日本医科大学, 医学部, 助手 (20256960)
永原 則之 日本医科大学, 医学部, 助教授 (10208043)
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| Project Period (FY) |
1997 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥12,200,000 (Direct Cost: ¥12,200,000)
Fiscal Year 2000: ¥2,800,000 (Direct Cost: ¥2,800,000)
Fiscal Year 1999: ¥2,300,000 (Direct Cost: ¥2,300,000)
Fiscal Year 1998: ¥2,400,000 (Direct Cost: ¥2,400,000)
Fiscal Year 1997: ¥4,700,000 (Direct Cost: ¥4,700,000)
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| Keywords | reactive oxygen species / xanthine dehydrogenase / xanthine oxidase / niric oxide synthase / metallo-protein / non-heme iron / peroxiredoxin / heme binding protein / NO合成酵素 / 金属フラビン酵素 / セロビオース脱水素酵素 / 電子スピン共鳴 / X線結晶構造 / フリーラジカル / EPR / nNOS / Xanthine Oxidase / Xanthine Dehydrogenase / Flavoprotein / metalloprotein / free radical / nitric oxide / super oxide anion |
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| Research Abstract |
Xanthine dehydrogenase/oxidase and nitric acid synthetase are complex flavoprotein that produce free radicals. In this study, we presented the crystal structure of the dimeric (Mr 290,000) bovine milk XDH at 2.1 Å resolution and XO at 2.5 Å resolution and describe the major changes that occur upon the proteolytic transformation of XDH to the XO form. Each molecule is composed of an N-terminal 20 kDa domain containing two iron sulfur centers, a central 40 kDa FAD domain, and a C-terminal 85 kDa molybdopterin-binding domain with the four redox centers aligned in an almost linear fashion. Cleavage of surface-exposed loops of XDH causes major structural rearrangement of another loop close to the flavin ring (Gin-423-Lys-433). This movement partially blocks access of the NAD substrate to the FAD cofactor and changes the electrostatic environment of the active site, reflecting the switch of substrate specificity observed for the two forms of this enzyme. We have also constructed several mutant enzyme of rat liver XDH and determined the character to confirm the responsible residues for conversion.
We have expressed mouse neuronal nitic acid synthase (NOS) as well as the iso-form of natural mutant and determined characters of the enzymes by several physicochemical methods. We have also studied some other related enzymes such as heme containing flavo-protein, cellobiose dehydrogenase, and heme binding protein, HBP23 has peroxidase activity.
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