Structure and Function of Protein-Membrane Interactions in Blood Clotting

描述（由申请人提供）：血液凝固的几乎每一步都需要多种蛋白质在膜表面的组装，然而血液凝固中的蛋白质-膜相互作用在分子水平上仍然知之甚少。我们正在使用新的高分辨率技术，包括魔角旋转固态核磁共振（SSNMR）来探测凝血蛋白与磷脂表面相互作用的机制，以及膜成分的变化如何调节凝血反应。我们研究的主要重点是组织因子和VIIa因子的膜结合复合物，这是一种负责触发健康和疾病中的血液凝固的双亚基酶。目的1将在原子分辨率上描述Ca2+存在下富含PS的膜结构域的结构和动力学变化，并将研究磷脂酰丝氨酸、磷脂酰胆碱和磷脂酰乙醇胺在配置膜以支持凝血因子高亲和力结合中的作用。目的2将研究磷脂-磷脂相互作用在增强膜表面血液凝固反应中的作用。目的3将描述当凝血蛋白与双分子层结合时诱导的脂质环境/构象。Aim 4将求解膜表面组织因子的结构，识别组织因子与配体相互作用时的构象变化。总之，这些研究将在原子尺度分辨率上为膜表面在血液凝固中的作用提供有价值的新见解。

DESCRIPTION (provided by applicant): Almost every step in blood coagulation requires assembly of multiple proteins on membrane surfaces, yet protein-membrane interactions in blood clotting remain poorly understood at the molecular level. We are using new, high-resolution technologies including magic-angle spinning solid-state NMR (SSNMR) to probe the mechanisms by which blood clotting proteins interact with phospholipid surfaces, and how changes in membrane composition regulate clotting reactions. The primary focus of our studies is the membrane-bound complex of tissue factor and factor VIIa, the two-subunit enzyme responsible for triggering blood clotting in health and disease. Aim 1 will delineate, at atomic resolution, the changes in structure and dynamics of PS- rich membrane domains in the presence of Ca2+, and will examine the roles of phosphatidylserine, phosphatidylcholine and phosphatidylethanolamine in configuring the membrane to support high affinity binding of clotting factors. Aim 2 will investigate the role of phospholipid-phospholipid interactions in enhancing blood clotting reactions on membrane surfaces. Aim 3 will delineate the lipid environments/conformations induced when clotting proteins bind to bilayers. Aim 4 will solve the structure of tissue factor on the membrane surface and identify conformational changes in tissue factor when it interacts with ligands. Together these studies will provide valuable new insights into the role of the membrane surface in blood clotting, at atomic-scale resolution.