Structural Mechanisms for the Inhibition of Thrombosis

抑制血栓形成的结构机制

• 批准号: 6654278
• 负责人: JAMES Andrew HUNTINGTON
• 金额: $ 0.64万
• 依托单位: UNIVERSITY OF CAMBRIDGE
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-12-21 至 2004-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6654278
• 关键词: X ray crystallography; activation product; antithrombins; carbohydrate structure; chemical kinetics; coagulation factor X; cofactor; conformation; enzyme activity; enzyme complex; enzyme inhibitors; enzyme mechanism; fibrin; gene mutation; heparin; ionic bond; mucopolysaccharides; protein binding; protein structure function; serine proteinases; structural biology; thrombin; thrombosis

DESCRIPTION (provided by applicant): The long-term objective of this research proposal is to understand the mechanisms by which the principal two plasma inhibitors of thrombin, antithrombin and heparin cofactor II, are activated by heparin-like glycosaminoglycans. Such an understanding will help explain dysfunction which leads to familial thrombophilia and aid in the development of novel therapeutic antithrombotic agents. This objective will largely be fulfilled through the successful completion of the specific aims which follow from the general hypotheses that: i) antithrombin and heparin cofactor II circulate in a kinetically inactive state; ii) binding to and activation by glycosaminoglycans involves a dramatic conformational change to a kinetically active state; iii) the shared serpin mechanismn of protease inhibition plays a critical role in facilitating release of the final complex from glycosaminoglycan and fibrin to allow for clearance and in situ degradation. The specific aims fall under three general questions: I) How does heparin accelerate antithrombin inhibition of thrombin? II) How is antithrombin activated towards factor Xa? and, III) How is heparin cofactor II activated towards thrombin? A combination of protein X-ray crystallographic and biochemical studies will be employed to answer these questions. The specific aims are: 1) To determine the structures of antithrombin and thrombin bound to heparin. 2) To determine the structure of the initial antithrombin-thrombin-heparin complex. 3) To determine the structure of the final antithrombin-thrombin complex. 4) To determine the structure of native and pentasaccharide-activated antithrombins. 5) To investigate the role of surface electrostatics in the allosteric activation of antithrombin. 6) To determine the structure of native and glycosaminoglycan bound heparin cofactor II. 7) To determine the structure of the initial heparin cofactor Il-thrombin complex. 8) To investigate the conformational link between glycosaminoglycan binding and release of the N-terminal tail. 9) To determine the conformational basis for cleavage of the N-terminal tail from the final heparin cofactor II-thrombin complex.

描述(申请人提供)：本研究的长期目标 建议是理解两种主要的等离子体 凝血酶、抗凝血酶和肝素辅因子II的抑制物可通过 类肝素糖胺聚糖。这样的理解将有助于解释 导致家族性血栓形成的功能障碍 新型治疗性抗血栓药物。这个目标将在很大程度上 通过成功完成后续的具体目标来实现 根据一般假设：i)抗凝血酶和肝素辅因子II 在运动不活跃的状态下循环；ii)结合并激活 糖胺多聚糖涉及一种戏剧性的构象变化 活性状态；iii)共同的丝氨酸蛋白酶抑制机制发挥作用 在促进最终复合体从 糖胺多聚糖和纤维蛋白，以允许清除和原位降解。 具体目标分为三个一般问题：i)肝素是如何 加速抗凝血酶抑制凝血酶？II)抗凝血酶效果如何 对Xa因子的激活？以及，iii)肝素辅因子II是如何被激活的 凝血酶？蛋白质X-射线结晶学和X射线衍射仪的结合 生物化学研究将被用来回答这些问题。具体的 目的是：1)确定抗凝血酶和凝血酶结合的结构 肝素。2)确定首字母的结构 抗凝血酶-凝血酶-肝素复合体。3)确定 最终的抗凝血酶-凝血酶复合体。4)确定原生生物的结构 和五糖激活的抗凝血酶。5)调查 表面静电学在抗凝血酶变构激活中的作用。6)至 天然和糖胺聚糖结合的肝素辅因子的结构测定 Ii.7)测定初始肝素辅因子IL-凝血酶的结构 很复杂。8)研究糖胺多糖之间的构象联系 N-末端尾巴的结合和释放。9)确定构象 从最终肝素辅因子中切割N-末端尾部的基础 II-凝血酶复合体。