REGULATION OF HUMAN PLATELET PROTHROMBINASE ACTIVITY

人血小板凝血酶原活性的调节

• 批准号: 6657103
• 负责人: Paula Babiarz Tracy
• 金额: $ 18.67万
• 依托单位: UNIVERSITY OF VERMONT & ST AGRIC COLLEGE
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-01 至 2003-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6657103
• 关键词: antifibrinolytic agents; binding sites; blood coagulation; clotting factor; coagulation factor V; coagulation factor X; enzyme activity; enzyme complex; fibrinogen receptors; fibrinolysis; hirudins; human subject; intermolecular interaction; membrane proteins; molecular assembly /self assembly; platelet activation; platelets; protein C; protein binding; protein protein interaction; protein structure function; prothrombin; receptor expression; thrombin

The ability of platelets to regulate thrombin generation at their membrane surface is central to their role in hemostasis, thrombosis, and atherosclerosis. Thrombin generation is effected through the assembly and function of the enzymatic complex, and atherosclerosis. Thrombin generation is effected through the assembly and function of the enzymatic complex, Prothrombinase, consisting of a I.I stoichiometric Ca2+- dependent complex of the cofactor factor Va and the serine protease factor Xa. Subsequent to platelet activation, release platelet factor Va and/or plasma-derived factor Va bind to the platelet membrane surface and in so doing form at least part of the receptor for factor Xa. Thus, several protein/protein and protein/membrane interactions participate in and regulate complex assembly. One goal of this project is to define how platelets actively participate in and regulate Prothrombinase assembly and function. The following hypotheses have been formulated and will be tested. Platelets regulate thrombin generation through 1) the expression of two discrete platelet subpopulations-both bind factor Va, only one binds both factors Va and Xa; and 3) through the agonist-induced release of platelet factor Va which is resistant to inactivation by activation by activated protein C and plasmin, and functionally and physically unique when compared to plasma-derived factor Va. Effort will be placed on isolating and characterizing these discrete platelet populations, isolating and/or expression cloning the platelet membrane receptors for factor Va and Xa, and characterizing platelet-derived factor Va, functionally and physically as related to its participation in Prothrombinase. Because thrombin once formed positively regulates Prothrombinase assembly and function through platelet activation and release of platelet factor Va, a second goal is to define how thrombin interacts through platelet activation and release of platelet factor Va, a second goal is to define how thrombin interacts with platelet membrane proteins to modulate its activity. Effort will be place don testing the hypothesis that the platelet high affinity binding site for thrombin is 1) a unique platelet membrane protein that resembles hirudin and renders the thrombin molecule transiently inactive and, 2) is distinct from glycoprotein 1b. This will be accomplished in part through expression cloning of this unique platelet membrane protein and demonstration that anti-glycoprotein 1b antibodies shown to inhibit thrombin-induced platelet activation do so through their cross reactivity with PAR1. Successful completion of these goals will demonstrate the mechanisms by which platelets actively regulate both the generation and function of thrombin at their membrane surface.

血小板调节其膜上凝血酶生成的能力 表面是它们在止血、血栓形成和 动脉粥样硬化凝血酶的产生是通过组装 和酶复合物的功能以及动脉粥样硬化。凝血酶 产生是通过酶的组装和功能实现的， 复合物，凝血酶原酶，由I.I化学计量的Ca 2 +- 辅因子Va和丝氨酸蛋白酶的依赖性复合物 因子Xa。血小板活化后，释放血小板因子Va 和/或血浆衍生因子Va与血小板膜表面结合 并在这样做时形成因子Xa受体的至少一部分。因此，在本发明中， 几种蛋白质/蛋白质和蛋白质/膜相互作用参与 并调节复杂的装配。这个项目的一个目标是定义如何 血小板积极参与和调节凝血酶原酶组装 和功能以下假设已经制定，并将 测试.血小板通过以下途径调节凝血酶的生成：1） 两个离散的血小板亚群-都结合因子Va，只有一个结合 因子Va和Xa;和3）通过激动剂诱导的 血小板因子Va，其对通过活化而失活具有抗性， 活化的蛋白C和纤溶酶，功能和物理独特 与血浆衍生因子Va相比，将努力 分离和表征这些离散的血小板群体， 和/或表达克隆因子Va的血小板膜受体 和Xa，并表征血小板衍生因子Va的功能和 物理上与其参与凝血酶原酶有关。因为 凝血酶一旦形成，就正向调节凝血酶原酶组装， 通过血小板活化和释放血小板因子Va，a 第二个目标是确定凝血酶如何通过血小板相互作用 激活和释放血小板因子Va，第二个目标是确定如何 凝血酶与血小板膜蛋白相互作用， 活动将努力检验血小板 凝血酶高亲和力结合位点是1）独特的血小板膜 一种类似水蛭素的蛋白质，使凝血酶分子 瞬时失活，2）与糖蛋白1b不同。这将是 部分是通过表达克隆这种独特的血小板 膜蛋白和证明抗糖蛋白1b抗体 显示抑制凝血酶诱导的血小板活化是通过它们的 与PAR 1交叉反应。成功实现这些目标， 证明了血小板积极调节血小板和血小板分泌的机制。 凝血酶在其膜表面的产生和功能。