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: 8155294
• 负责人: Matthew Auton
• 金额: $ 34.61万
• 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 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8155294
• 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. PUBLIC HEALTH RELEVANCE: Von Willebrand factor (VWF) is a main responder to vascular injury, and functions to sequester and adhere platelets to the subendothelium and initiate coagulation. The objective of the proposed investigation is to test a novel model for how inherited abnormalities of VWF result in clinical disorders of hemostasis. The principles learned from this research project will provide a unique perspective into the mechanisms of disease that can potentially be translated to modern medical approaches for diagnosis and treatment of cardiovascular disorders.

描述(申请人提供)：血管性血友病因子(VWF)是一种由血管内皮细胞和激活的血小板分泌的多聚体糖蛋白，其功能是隔离和黏附血小板到内皮下层并启动凝血。自从最初观察到VWF的功能受血流流变切应力的构象调控以来，主流观点一直认为，暴露于高水平的血管切应力会使VWF呈现一种激活的构象，从而增加其A1结构域与血小板GPIba表面受体的结合亲和力。当我们在VWF的“血小板GPIba结合”A1结构域中发现存在热力学上不同的展开中间体时，我们对这种构象在健康和疾病中所起的功能作用进行了大量的研究。VWF值得广泛研究，因为它与人类最常见的遗传性出血性疾病--血管性血友病(VWD)有关。2型VWD的两个亚型的特征是由于VWF A1结构域的点突变而导致的血小板-VWF相互作用增强(2B型)或缺陷(2M型)。我们的研究已经证实，这些表型相反的突变不同地影响A1结构域的热力学稳定性，A1结构域与血小板GPIba的结合亲和力，以及A1和GPIba之间的力依赖的接滑结合，这是一种随着剪切流的增加而调节VWF上血小板滚动速度的特性。这些突变对A1结构域这些性质的影响程度表明：1)结合亲和力与这种天然到中(N？I)构象平衡相连；2)A1-GPIba相互作用的力/剪应力依赖性性质是天然低亲和力构象和中高亲和力构象之间热力学联系的直接结果。这些研究产生了第一个VWF功能机制的定量工作模型，该模型形成了本申请的中心假设的基础，即A1结构域的中间构象形成了组成VWF活性状态的结构的组成部分。我们的目标是确定VWF的A1A2A3三结构域片段中的A1稳定性和结构域关联是如何在热力学上耦合的，以及这一调节如何与临床疾病中出血的严重程度有关。为了实现这一目标，我们将通过调查导致Von Willebrand病的2B和2M型突变的全面临床数据库来测试构象模型，并建立这些临床突变对A1A2A3三域内四元结构域相互作用的影响。这项工作将对VWD的临床诊断具有重要价值，因为它有可能根据突变对VWF结构和功能的构象影响对其进行分类。彻底了解由遗传突变引起的构象缺陷的广度对于发展VWF的定量结构-活性关系以及开发心血管疾病的新治疗方法至关重要。 公共卫生相关性：血管性血友病因子(VWF)是血管损伤的主要反应者，其功能是隔离和黏附血小板到内皮下层并启动凝血。这项拟议的研究的目的是测试一种新的模型，即遗传的VWF异常如何导致临床止血障碍。从这项研究项目中学到的原理将为疾病机制提供一个独特的视角，这些机制可能被转化为诊断和治疗心血管疾病的现代医学方法。