项目名称： 金属蛋白酶ADAMTS13构象及其与血管性血友病因子VWF结合的力学调控机制的研究

项目编号： No.31500759

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 生物科学

项目作者： 林蒋国

作者单位： 华南理工大学

项目金额： 20万元

中文摘要： 血小板粘附到血管壁受损处是止血和血栓形成的重要因素，是一个多步骤、受不同分子及剪切应力调控的过程。血管性血友病因子VWF，参与调控血小板在血管壁上的初始粘附及滚动。VWF的粘附性与其分子大小有关。金属蛋白酶ADAMTS13酶切VWF A2结构域的Tyr1605-Met1606肽键，调节其大小，防止血栓的形成。因此，ADAMTS13与VWF的结合及酶切的研究一直以来均为该领域的热点。然而，此前的研究存在着不足，例如，两者的反应时间过长，偏离生理条件；ADAMTS13 C－端结构域的功能尚存争议等。为此，本项目拟采用原子力显微镜及流动腔技术在单分子水平，在更接近生（病）理的条件下，研究ADAMTS13构象的稳定性，及其与VWF结合，并提取反应动力学常数，以揭示ADAMTS13构象及其与VWF结合的分子基础，及应力调控机制，为进一步认识止血和血栓形成的过程，及相关疾病的治疗提供新的视角和思路。

中文关键词： VWF/ADAMTS13；单分子力学；原子力显微镜；流动腔；反应动力学

英文摘要： The adhesion of platelets to injury site at blood vessel wall is a critical step in hemostasis and formation of thrombosis, which is a multi-step process meditated by various molecules and shear stress. Von Willebrand Factor (VWF) mediates platelets tethering to and rolling on blood vessel wall. The adhesive of VWF relates to its molecular size. A metalloprotease ADAMTS13 cleaves the Tyr1605-Met1606 peptide bond in VWF A2 domain to regulate its size and prevent thrombosis. The binding and proteolysis of ADAMTS13 to VWF therefore have been being the hot topic in the field. However, some disadvantages exist in previous studies, such as the long incubation time far beyond physiological condition, the discrepancy of the function of ADAMTS13 c-termini, et al. Therefore, in this project Atomic Force Microscopy (AFM) and flow chamber techniques will be applied, at single molecular level, to examine the stability of ADAMTS13 conformation, its binding to VWF and extract the kinetic constants in the conditions closer to physiological/pathological scenarios. This project aims to reveal the molecular basis and regulation mechanisms of shear stress on ADAMTS13 conformation and its binding to VWF, in order to further understand to the process of hemostasis and thrombosis, and provide new insight and method in therapeutics.

英文关键词： VWF/ADAMTS13;Single Molecular Mechanics;Atomic Force Microscopy;Flow Chamber;Reaction Kinetics