CONTENT AND ASSEMBLAGE OF PLATELET GRANULE PROTEINS

血小板颗粒蛋白的含量和组装

• 批准号: 3338936
• 负责人: Daniel A Walz
• 金额: $ 16.56万
• 依托单位: WAYNE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-04-01 至 1994-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3338936
• 关键词: antibody; chemical binding; electrophoresis; fibrinogen; fibrinolysis; fibronectins; fluorescent dye /probe; glycoproteins; granule; high performance liquid chromatography; ligands; monoclonal antibody; plasmin; plasminogen; platelet activating factor; platelet aggregation; platelets; protein sequence; synthetic peptide; thrombosis; thrombospondins

Platelets play a critical role in mediating a variety of biological processes, including cell proliferation, cell adhesion, immunoregulation, thrombosis and fibrinolysis; many of these events are directly influenced by proteins secreted from the platelet alpha-granule. The central hypothesis of this proposal is that at least two such proteins, namely thrombospondin and platelet factor 4, participate in such regulatory mechanisms through their interaction with specific ligands and, furthermore, that this interaction is the function of unique and specific peptide domains, both within a given ligand and the platelet proteins. Specifically, the requisite thrombospondin binding domains within the Aalpha- an Bbeta-fibrinogen chains will be identified. Synthetic peptide analogues to each of these domains will be synthesized and evaluated for their ability to modulate the secretion-dependent phase of platelet aggregation. Utilizing a blot-binding assay, the region of thrombospondin which binds fibrinogen will also be identified, the requisite peptide domain(s) characterized, synthesized, and assessed for their ability to regulate thrombospondin-fibrinogen interaction and platelet aggregation. The COOH-terminal 18 kDa domain of thrombospondin has been purified in quantities which will permit a direct assessment of its ability to bind to non-activated and activated platelets. These studies will be conducted using fluorescent-labeled 18 kDa domain as well as polyclonal and monoclonal antibody probes. The binding of thrombospondin to specific kringle regions of plasminogen, particularly kringle 5, will be determined. The ability of peptide analogues to this region will be probed for their potency in regulating plasminogen activators' kinetics effect on plasmin generation. The plasmin generation protocols will also be assessed when histidine rich glycoprotein, and the thrombospondin binding domain regions from that protein, are included in the trimolecular complex activation of thrombospondin-histidine rich glycoprotein-plasminogen. Further studies on the interaction of thrombospondin and the 29 kDa domain of fibronectin will also be performed. Monoclonal antibodies against thrombospondin will be utilized to define which regions within the thrombospondin molecule are "decorated" by specific protein ligands. These studies should provide a better understanding of the role of thrombospondin in platelet aggregation, fibrin formation and fibrinolysis and permit the development of synthetic peptides which prevent or attenuate these processes.

血小板在介导多种生物学过程中起关键作用， 过程，包括细胞增殖，细胞粘附，免疫调节， 血栓形成和纤维蛋白溶解;这些事件中的许多直接影响 由血小板α颗粒分泌的蛋白质引起中央 这一建议的假设是，至少有两种这样的蛋白质，即 血小板反应蛋白和血小板因子4参与这种调节 通过与特定配体的相互作用， 此外，这种相互作用是独特和具体的功能， 肽结构域，在给定的配体和血小板蛋白内。 具体地，在细胞内必需的血小板反应蛋白结合结构域是必需的。 将鉴别A α-和B β-纤维蛋白原链。合成肽 将合成这些结构域中的每一个的类似物，并评估其用于 它们调节血小板分泌依赖性阶段的能力 聚合来利用印迹结合分析，血小板反应蛋白区域 结合纤维蛋白原的肽也将被鉴定， 结构域表征、合成和评估其 调节血小板反应蛋白-纤维蛋白原相互作用和血小板聚集。 血小板反应蛋白的COOH-末端18 kDa结构域已在 这将允许直接评估其约束能力的数量 非活化和活化的血小板。这些研究将在 使用荧光标记的18 kDa结构域以及多克隆和 单克隆抗体探针血小板反应蛋白与特异性 纤溶酶原的Kringle区，特别是Kringle 5， 测定肽类似物对该区域的能力将是 探讨其调节纤溶酶原激活剂动力学的效力 对纤溶酶生成的影响。纤溶酶生成方案还将 当富含组氨酸的糖蛋白和血小板反应蛋白 来自该蛋白质的结合结构域区域包括在所述结合结构域区域中。 富凝血酶敏感蛋白-组氨酸三分子复合物活化 糖蛋白-纤溶酶原进一步研究了 血小板反应蛋白和纤连蛋白的29 kDa结构域也将被 执行。将使用抗血小板反应蛋白的单克隆抗体 为了确定血小板反应蛋白分子内的哪些区域 被特定的蛋白质配体“修饰”。这些研究将提供一个 更好地理解血小板反应蛋白在血小板中的作用 聚集，纤维蛋白形成和纤维蛋白溶解，并允许发展 合成肽来阻止或减弱这些过程。