ANTICOAGULANT PROTEIN COMPLEX STRUCTURE AND FUNCTION

抗凝蛋白复合物的结构和功能

• 批准号: 6232546
• 负责人: TIMOTHY A. MATHER
• 金额: $ 32.49万
• 依托单位: OKLAHOMA MEDICAL RESEARCH FOUNDATION
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-02-05 至 2004-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6232546
• 关键词: X ray crystallography; activation product; active sites; anticoagulants; cellular pathology; coagulation factor VIII; cofactor; conformation; enzyme activity; enzyme structure; enzyme substrate complex; molecular pathology; protein C; protein binding; protein structure function; proteolysis; thrombin; thrombomodulin; venous thrombosis; zymogens

DESCRIPTION (Applicant's Description Verbatim): The enzymatic phase of the anticoagulation system functions by the proteolytic digestion and down-regulation of the procoagulant cofactors factors VIIla and Va by the enzyme activated protain C(APC). Hereditary defects in this system lead to a congenital predisposition to venous thrombosis. The pathway is initiated when thrombin (T) binds to thrombomodulin (TM) on the endothelium and undergoes a cofactor dependent allosteric conformational change. In the TM bound conformation, thrombin loses its procoagulant activity, and gains an alternative substrate selectivity. The T-TM complex now recognizes the zymogen protein C as its substrate and very efficiently activates it. Thus, the major procoagulant enzyme takes on an anticoagulant activity and catalyzes its own down-regulation as a result of this cofactor induced allosteric modulation. The structural nature of this transition is completely unknown. If we could understand this specificity switch on a structural basis, we might begin to design therapeutic agents which could bind to thrombin and shift it from a procoagulant towards an anticoagulant activity. Based on the results of APC infusion in septic shock, this therapeutic approach leads to a wider window of efficacy without increased bleeding. We propose to study the structure and function of the T-TM complex with X-ray crystallography to understand the structural origin of thrombin's anticoagulant activity. We have grown diffracting crystals of one form of this complex and are currently solving its structure. We plan to build on this initial success to solve the structures of related complexes, each designed to show an additional aspect of this activity, and to apply our findings towards explaining the ability of some small molecules and thrombin point mutations to achieve partial anti-coagulant activity. Ultimately we will undertake structure based drug design to improve these partial effects.

描述（申请人的逐字描述）： 抗凝系统通过蛋白水解和消化发挥作用 促凝血辅助因子 VIIla 和 Va 的下调 酶激活蛋白C(APC)。该系统的遗传缺陷导致 先天性静脉血栓形成倾向。该路径启动时 凝血酶（T）与内皮细胞上的血栓调节蛋白（TM）结合并经历 辅因子依赖性变构构象变化。在TM界 构象，凝血酶失去其促凝血活性，并获得 替代底物选择性。 T-TM 复合物现在可以识别酶原 蛋白 C 作为其底物并非常有效地激活它。因此，主要 促凝血酶具有抗凝血活性并催化其自身 由于该辅因子诱导的变构调节而下调。这 这种转变的结构性质是完全未知的。 如果我们能够在结构基础上理解这种特异性开关，我们可能会 开始设计可以结合凝血酶并使其转移的治疗剂 从促凝血活性转变为抗凝血活性。根据结果 APC输注治疗感染性休克，这种治疗方法带来更宽的窗口 的疗效而不增加出血。我们建议研究结构和 通过 X 射线晶体学研究 T-TM 复合物的功能，以了解 凝血酶抗凝活性的结构起源。我们已经成长 这种复合体的一种形式的衍射晶体，目前正在解决它的问题 结构。我们计划在这一初步成功的基础上解决以下问题的结构： 相关的综合体，每个综合体都旨在展示该活动的另一个方面， 并应用我们的发现来解释一些小 分子和凝血酶点突变实现部分抗凝 活动。最终我们将进行基于结构的药物设计以改进 这些部分影响。