Thermodynamics of the Conformational Activation of Von Willebrand Factor

冯维勒布兰德因子构象激活的热力学

• 批准号: 8302350
• 负责人: Matthew Auton
• 金额: $ 35.21万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-15 至 2012-11-09
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8302350
• 关键词: Affect; Affinity; Behavior; Binding; Binding Proteins; Blood Platelets; Blood Vessels; Blood flow; Cardiovascular Diseases; Clinical; Coagulation Process; Complex; Coupled; Databases; Defect; Development; Diagnosis; Disease; Equilibrium; Exposure to; Glycoproteins; Goals; Health; Hemorrhage; Hemostatic function; Human; Induced Mutation; Inherited; Injury; Investigation; Learning; Link; Literature; Mechanics; Medical; Modeling; Molecular Conformation; Mutation; Physiological; Play; Point Mutation; Polymers; Population; Prevalence; Property; Publishing; Quantitative Structure-Activity Relationship; Regulation; Research Project Grants; Role; Severities; Structure; Surface; Testing; Thermodynamics; Translating; Vascular Endothelial Cell; Work; X-Ray Crystallography; base; clinical Diagnosis; clinical effect; loss of function; man; novel; predictive modeling; receptor; shear stress; von Willebrand Disease; von Willebrand Factor

DESCRIPTION (provided by applicant): Von Willebrand factor (VWF), a multimeric polymer glycoprotein secreted from vascular endothelial cells and activated platelets, functions to sequester and adhere platelets to the subendothelium and initiate coagulation. Since the initial observation that the function of VWF is conformationally regulated by the rheological shear stress of blood flow, the prevailing idea has been that exposure to elevated vascular shear stress unfolds VWF to an activated conformation that increases its binding affinity of the A1 domain for the platelet GPIba surface receptor. When we identified the presence of a thermodynamically distinct unfolding intermediate in the "platelet GPIba binding" A1 domain of VWF, it set forth numerous studies of the functional role that this conformation plays in health and disease. VWF has merited extensive study because of its association with the most common inherited bleeding disorder in man, Von Willebrand Disease (VWD). Two subtypes of type 2 VWD are characterized by either enhanced (type 2B) or deficient (type 2M) platelet-VWF interactions due to point mutations located specifically in the A1 domain of VWF. Our studies have established that these phenotypically opposite mutations differentially affect the thermodynamic stability of the A1 domain, the binding affinity of the A1 domain for platelet GPIba, and the force-dependent catch-slip bonding between A1 and GPIba, a property that regulates platelet-rolling velocities on VWF as shear flow is increased. The degree to which these mutations affect these properties of the A1 domain have revealed that 1) the binding affinity is coupled to this native to intermediate (N?I) conformational equilibrium and 2) the force/shear stress dependent properties of the A1-GPIba interaction are a direct consequence of the thermodynamic linkage between the native low affinity and intermediate high affinity conformations. These studies have resulted in the first quantitative working model for the mechanism of VWF function that forms the basis for the central hypothesis of this application that the intermediate conformation of the A1 domain forms an integral part of the structure that comprises the active state of VWF. Our objective is to identify how A1 stability and domain association within the A1A2A3 tri-domain fragment of VWF are thermodynamically coupled and how this regulation is linked to the severity of bleeding in clinical disease. To accomplish this objective we will test the Conformational Model through the investigation of a comprehensive clinical database of type 2B and 2M mutations that cause Von Willebrand disease and establish the effects of these clinical mutations on the quaternary domain interactions within the A1A2A3 tri-domain. The proposed work will be of significant value to the clinical diagnosis of VWD as it has the potential to classify mutations according to their conformational effects on VWF structure and function. A thorough understanding of the breadth of conformational defects caused by inherited mutations will be essential for the development of quantitative structure-activity relationships in VWF as well as the development of new treatments for cardiovascular disease.

描述（由申请人提供）：冯·威尔布兰德因子（VWF），这是一种从血管内皮细胞和活化的血小板中分泌的多聚合聚合物糖蛋白，功能隔离并粘附于下皮下皮和启动凝结。由于最初观察到，VWF的功能受到血流的流变剪切应力的构象调节，因此，普遍的想法是，暴露于升高的血管剪切应力将VWF展现为活化构象，从而增加了其对Platelet Gpiba表面受体的A1结构域的结合亲和力。当我们确定了VWF的“血小板GPIBA结合” A1域中的热力学上不同的中间体的存在时，它就对这种构型在健康和疾病中起的功能作用进行了大量研究。 VWF之所以值得广泛的研究，是因为它与人类冯·威勒（Von Willebrand）疾病（VWD）中最常见的遗传出血疾病相关。 2型VWD的两个亚型的特征是增强的（类型2b）或缺乏（类型2M）血小板-VWF相互作用，这是由于位于VWF的A1域中的点突变而引起的。我们的研究已经确定，这些表型相反的突变在差异上影响了A1结构域的热力学稳定性，A1域对血小板GPIBA的结合亲和力以及A1和GPIBA之间的力依赖性的捕获滑移键合，该特性会调节VWF上的VWF上的shear plat shear plat shear Flow。这些突变影响A1结构域的这些特性的程度表明，1）结合亲和力与该本机与中间体（n？i）构象平衡和2）A1-GPIBA相互作用的力/剪切应力相互作用耦合，是天然低亲密属于高度亲和力之间的热力学连接的直接结果。这些研究为VWF函数机制提供了第一个定量工作模型，该模型构成了本应用的中心假设的基础，即A1域的中间构型构成了组成VWF活性状态的结构的组成部分。我们的目标是确定VWF的A1A2A3三域片段中的A1稳定性和域关联是热力学耦合的，以及该调节与临床疾病中出血的严重程度如何相关。为了实现这一目标，我们将通过研究导致von Willebrand疾病的2B型和2M突变的综合临床数据库测试构象模型，并确定这些临床突变对A1A2A3三域中Quaternary域相互作用的影响。拟议的工作将对VWD的临床诊断具有重要价值，因为它有可能根据突变对VWF结构和功能的构象作用进行分类。对遗传突变引起的构象缺陷的广度的彻底理解对于发展VWF的定量结构 - 活性关系以及心血管疾病的新治疗方法至关重要。