Molecular Interactions of Factor XI

XI 因子的分子相互作用

• 批准号: 7117391
• 负责人: PETER Newton WALSH
• 金额: $ 42.58万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-09-30 至 2007-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7117391
• 关键词: SDS polyacrylamide gel electrophoresis; activation product; aminoacid analyzer; amyloid proteins; autoradiography; binding sites; blood proteins; cell surface receptors; clinical research; coagulation factor XI; cofactor; dimer; enzyme activity; high performance liquid chromatography; human tissue; immunochemistry; intermolecular interaction; mass spectrometry; nuclear magnetic resonance spectroscopy; protease inhibitor; protein protein interaction; protein structure function; site directed mutagenesis; surface plasmon resonance; western blottings

DESCRIPTION (provided by applicant): Previous studies have focused attention upon FXI and its interactions with other plasma proteins, i.e., thrombin, FXla, FXIla, prothrombin, high molecular weight kininogen (HK), and FIX, with the platelet plasma membrane and with various inhibitory molecules, e.g., alpha-l-protease inhibitor (alpha1PI), and protease nexin II (PN2), in the initiation and regulation of blood coagulation. The long term goals of the present proposal are to elucidate the molecular mechanisms involved in the interaction of FXI/FXla with protein and cell surface ligands involved in its activation and with plasma protein and cell surface ligands involved in the expression and regulation of FXla enzymatic activity. Specifically we propose: 1) To examine the hypothesis that GPIb/IX/V comprises the platelet membrane receptor for FXI that colocalizes it with thrombin for efficient activation on the activated platelet surface, a) To investigate the hypothesis that the N-terminal globular domain of GPIbalpha is the major binding site for FXI that colocalizes it with thrombin for efficient activation on the activated platelet surface, b) To identify the FXI domain that mediates the interaction of FXI with GPIb/IX/V, to test the hypothesis that the FXI A3 domain serves this function and to determine the amino acid residues that mediate this interaction, c) To investigate the hypothesis that the FXI/GPIb complex colocalizes within lipid microdomains (membrane rafts) on the activated platelet membrane for efficient activation by thrombin. 2) To determine the structure of the A4 domain of FXI and the mechanism and structural determinants of homodimer formation by the A4 domain, a) To determine the A4 domain solution structure and the morphology of the interface mediating dimer formation, b) To carry out hydrogen/deuterium exchange experiments as an experimental approach to the analysis of interracial contacts between A4 domains, c) To carry out a mutational analysis of the mechanism and structural determinants of A4-mediated homodimer formation. 3) To determine the mechanism and physiological importance of homodimer formation in the assembly of the FXI-activation complex and the FIX-activation complex on the surface of activated platelets, a) To determine whether the dimeric structure of FXI is required for FXI activation to FXla by thrombin or by FXIIa in the presence or absence of activated platelets or glycocalicin, b) To determine whether the dimeric structure of FXla is required for FIX activation to FIXa in the presence of glycocalicin as it is in the presence of activated platelets, c) To differentiate between alternative mechanisms to account for the failure of monomeric FXla to activate FIX on the activated platelet surface.

DESCRIPTION (provided by applicant): Previous studies have focused attention upon FXI and its interactions with other plasma proteins, i.e., thrombin, FXla, FXIla, prothrombin, high molecular weight kininogen (HK), and FIX, with the platelet plasma membrane and with various inhibitory molecules, e.g., alpha-l-protease inhibitor (alpha1PI), and蛋白酶Nexin II（PN2），在血液凝血的启动和调节中。本提案的长期目标是阐明FXI/FXLA与蛋白质和细胞表面配体相互作用所涉及的分子机制，以及参与FXLA酶促活性的表达和调节的血浆蛋白和细胞表面配体。我们具体提出：1）检验以下假说：gpib/ix/v包括FXI的血小板膜受体，它们与凝血酶进行了有效激活，以在活化的血小板表面上进行有效激活，a）调查假设的假设，以下假说，即GPIBALPHA的N端域的N-末端球形构成了fxi的主要能力，可以在fxi上累及fxi的激活，而fl xi则可以激活fxi的激活位置，以弥补fxi的范围。 surface, b) To identify the FXI domain that mediates the interaction of FXI with GPIb/IX/V, to test the hypothesis that the FXI A3 domain serves this function and to determine the amino acid residues that mediate this interaction, c) To investigate the hypothesis that the FXI/GPIb complex colocalizes within lipid microdomains (membrane rafts) on the activated platelet membrane for efficient凝血酶激活。 2）确定FXI A4结构域的结构以及通过A4结构域形成同型二聚体形成的机制和结构决定因素，a）确定A4域溶液结构和界面介导二聚体形成的界面形态的形态，b）以进行氢/氘化实验，以进行氢/氘化实验，以进行实验分析，以实验分析构成对44 A4介导的同二聚体形成的结构决定因素。 3) To determine the mechanism and physiological importance of homodimer formation in the assembly of the FXI-activation complex and the FIX-activation complex on the surface of activated platelets, a) To determine whether the dimeric structure of FXI is required for FXI activation to FXla by thrombin or by FXIIa in the presence or absence of activated platelets or glycocalicin, b) To determine whether the dimeric structure of在存在激活血小板的情况下，在存在糖蛋白的情况下将FXLA固定到FIXA所必需，c）分化替代机制，以说明单体FXLA在活化的血小板表面上激活固定的失败。