| Project/Area Number |
03044113
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| Research Category |
Grant-in-Aid for international Scientific Research
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| Allocation Type | Single-year Grants |
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| Section | Joint Research |
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| Research Institution | Faculty of Science, Kyushu University |
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Principal Investigator |
IWANAGA Sadaaki Faculty of Science, Kyushu University, Professor, 理学部, 教授 (90029942)
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| Co-Investigator(Kenkyū-buntansha) |
HIGASHI Shouichi Faculty of Science, Kyushu University JSPS Fellowships for Japanese Junior Scien, 理学部, 日本学術振興会特別研 (10275076)
NAKAGAWA Kazunori Faculty of Medicine, Kyushu University Research Associate, 医学部, 助手 (50217668)
MUTA Tatsushi Faculty of Science, Kyushu University Research Associate, 理学部, 助手 (60222337)
KAWABATA Shun-ichiro Faculty of Science, Kyushu University Research Associate, 理学部, 助手 (90183037)
SUEISHI Katsuo Faculty of Medicine, Kyushu University Professor, 医学部, 教授 (70108710)
ICHINOSE Akitada Faculty of Medicine, Yamagata University Professor, 医学部, 教授 (10241689)
FUJIKAWA Kazuo Department of Biochemistry, Washington University Research Professor, 生化学部, 研究教授
EARL W.Davie ワシントン大学, 生化学部, 教授
DAVIE Earl W Department of Biochemistry, Washington University Professor
西村 仁 北海道大学, 薬学部, 助手 (80241347)
武谷 浩之 三重大学, 医学部, 助手 (60222105)
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| Project Period (FY) |
1991 – 1993
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| Project Status |
Completed (Fiscal Year 1993)
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| Budget Amount *help |
¥9,000,000 (Direct Cost: ¥9,000,000)
Fiscal Year 1993: ¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 1992: ¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 1991: ¥3,000,000 (Direct Cost: ¥3,000,000)
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| Keywords | Factor VII / Tissue factor / Extrinsic blood coagulation / Gla-domain / Factor VIIa / Factor IX / VII-TF molecular complex / Gla-EGF1 / 外因系血液凝固反応 / Gla-EGF1 / EGF様ドメイン / 血液凝固因子 / 外因系凝固反応 / セレクチン |
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| Research Abstract |
Initiation of the extrinsic blood coagulation pathway is mediated by a complex formed between plasma-derived factor VII/VIIa and cell-derived tissue factor (TF). To identify the site(s) of interaction, zymogen VII and VIIa were enzymatically and chemically modified, and their affinities with TF were estimated by measuring their inhibitory effects on the amidolytic activity enhanced after formation of the VIIa-TF complex. We found that the VIIa-light chain(Ki=3.5 X 10^<-7>) and its fragment consisting of the Gla-domain and the first epidermal growth factor (EGF)-like domain (Gla-EGF1 peptide ; Ki=1.0 X 10^<-6>) have an affinity with TF, but their binding capacity disappeared, respectively, by conventional chymotryptic cleavage.
Therefore, one of the binding sites of VII with TF probably locates in the Gla-EGF1 region. On the other hand, a dansyl-Glu-Gly-Arg chloromethyl ketone-treated Gla-domainless VIIa(Ki=0.7 X 10^<-7>) showed a high affinity with TF, whereas the corresponding Gla-doma
… More
inless VII similarly treated showed no binding potential, thereby indicating that binding site(s) other than in the Gla-EGF1 region is present in VIIa but not in VII. Acetylation or carbamylation of alpha-amino group of NH_2-terminal Ile-153 of VIIa resulted in the loss of binding affinity with TF ; such modifications convert VIIa into a zymogen like inactive form by destroying the salt bridge between Ile-153 and Asp343 in VIIa. The carbamylation rate of VIIa in the presence of TF was low, as compared with that in the absence of TF. The protection of alpha-amino group of Ile-153 from carbamylation after complex formation seemed to be due to a salt bridge formation between Ile-153 and Asp-343 in VIIa-TF complex. Therefore, it is concluded that the binding of TF with the heavy chain of VIIa induces a specific conformational change that brings alpha-amino group of Ile-153 close to beta-carboxyl group of Asp-343 to make a stable salt bridge. This salt bridge formed only in the presence of TF is essential for the formation of the catalytic triad of VIIa. Less
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