| Project/Area Number |
63870094
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| Research Category |
Grant-in-Aid for Developmental Scientific Research
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| Allocation Type | Single-year Grants |
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| Research Field |
Biological pharmacy
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| Research Institution | Hokkaido University |
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Principal Investigator |
KOYAMA Jiro Hokkaido University, Faculty of Pharmaceutical Sciences, Professor, 薬学部, 教授 (10025679)
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| Co-Investigator(Kenkyū-buntansha) |
TAKAHASHI Kazuhiko Hokkaido University, Faculty of Pharmaceutical Sciences, Instructor, 薬学部, 助手 (10113581)
MATSUMOTO Shuzo Hokkaido University, Faculty of Medicine, Professor, 医学部, 教授 (80000933)
山下 俊之 北海道大学, 薬学部, 助手 (90192400)
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| Project Period (FY) |
1988 – 1989
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| Project Status |
Completed (Fiscal Year 1989)
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| Budget Amount *help |
¥6,200,000 (Direct Cost: ¥6,200,000)
Fiscal Year 1989: ¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1988: ¥4,300,000 (Direct Cost: ¥4,300,000)
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| Keywords | Chronic Granulomatous Disease / NADPH Oxidase / NADPH-Cytochrome c Reductase / Leukocyte / Cytochrome b_<558> / Superoxide Anion / チトクロ-ムb_<55g> / NADPHオキシダーゼ / NADPHーチトクロームCレダクターゼ / チトクロームb_<558> |
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| Research Abstract |
Chronic granulomatous disease (CGD) is an uncommon inherited disorder in which phagocytic cells (neutrophils and macrophages) fail to produce antimicrobial oxidants such as O^-_. This failure is supposed to be the consequence of a defect in the NADPH oxidase which catalyzes the electron transfer from NADPH to molecular oxygen. We previously proposed that the NADPH-cytochrome c reductase in the plasma membrane of polymorphonuclear leukocytes (PMN) might be a flavoprotein of the NADPH oxidase which transferred electrons from NADPH to cytochrome b_<558>, and also that the association of the reductase with cytochrome b_<558> induced by stimulation of PMN might result in elicitation of activation of the NADPH oxidase.
Along this approach, we attempted to develop a new method for quantitative determination of the reductase of normal and CGD PMN, and that for comparative examination of the abilities of cytochrome b_<558> of normal and CGD PMN to couple to the reductase. The results so far obtained are as follows.
1. The reductase was highly purified from human PMN, and its biochemical properties were studied. Based on the results obtained, a method for immunoassay of the reductase in human PMN was established. 2. When the NADPH oxidase was solubilized from the membrane fraction of phorbol-myristate acetate-stimulated PMN by the use of deoxycholate and Triton X-100, it lost rapidly its activity. However, the treatment of the deoxycholate-solubilized membrane fraction with disuccinimidyl glutarate made the NADPH oxidase markedly stable. The NADPH oxidase thus stabilized may be an excellent material for biochemical study of the NADPH oxidase. 3. The activation mechanism of the NADPH oxidase was found to vary, depending upon the sort of stimulant used.
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