MECHANISM OF ACTIVATION OF PLATELET GPIIB AND GPIIIA

本研究的长期目标是了解 血小板整合素GPIIb-IIIa的活化。GPIIb-IIIa是 血小板粘附和聚集。 了解其规则 可能对治疗许多心血管疾病很重要 包括心肌梗塞和中风。 GPIIb-IIIa存在于 处于休眠构象的休眠血小板不能结合可溶性 纤维蛋白原。 血小板刺激后，IIb-IIIa能够 结合纤维蛋白原并介导血小板聚集。 虽然 IIb-IIIa的激活是血小板功能的关键，其机制是 激活尚未解决。 该研究的一个假设是， IIb-IIIa的胞质结构域控制 整联蛋白 这一假设将通过表征 两种纯化形式的胞质结构域的结构差异 IIb-IIIa在激活状态上不同。 本研究的第二个目标是了解 活化依赖性配体与IIb-IIIa的结合。 噬菌体展示将是 用于选择优先结合休眠的配体， 整合素的活性形式。 这项研究的结果可能会 提供了一个结构-活性系列，解释了激活依赖性 配体结合 本研究的第三个目的是了解 整合素活化。 激活是由于配体的增加吗 结合速率或配体解离速率的降低？ 这些 研究将在全血小板和纯化形式的 休眠和活跃的IIb-IIIa。 最后一个目标是了解IIb上的二价离子结合位点- 影响激活事件。 将进行结合研究 Ca 2+与休眠和活性IIb-IIIa的纯化构象之间的关系。 该分析的结果应确定哪类离子结合 位点调节整联蛋白的活化。

The long term objective of this study is to understand the mechanism of activation of the platelet integrin GPIIb-IIIa. GPIIb-IIIa is key to platelet adhesion and aggregation. An understanding of its regulation is likely to be important for treating many cardiovascular diseases including myocardial infarction and stroke. GPIIb-IIIa exists on resting platelets in a dormant conformation unable to bind soluble fibrinogen. Upon platelet stimulation, IIb-IIIa becomes capable of binding fibrinogen and mediating platelet aggregation. Although activation of IIb-IIIa is key to platelet function, the mechanism its activation has not been solved. One hypothesis of the study is that proteolytic cleavage of the cytoplasmic domains of IIb-IIIa control the activation state of the integrin. This hypothesis will be tested by characterizing the structural differences in the cytoplasmic domain of two purified forms of IIb-IIIa which differ in activation state. A second goals of the study is to understand the structural basis of activation-dependent ligand binding to IIb-IIIa. Phage-display will be used to select ligands that bind preferentially to the dormant and active forms of the integrin. Results from this study are likely to provide a structure-activity series explaining activation-dependent ligand binding. A third aim of the study is to understand the kinetic aspects of integrin activation. Does activation result from an increase in ligand association rate or a decrease in ligand dissociation rate? These studies will be performed on whole platelets and with purified forms of dormant and active IIb-IIIa. A final goal is to understand how the divalent ion binding sites on IIb- IIIa influence the activation event. Binding studies will be performed between Ca2+ and purified conformers of dormant and active IIb-IIIa. Results from this analysis should determine which class of ion binding sites regulate activation of the integrin.