MOLECULAR BASIS OF PLASMINOGEN-STREPTOKINASE INTERACTION

纤溶酶原-链激酶相互作用的分子基础

• 批准号: 6184532
• 负责人: Xuejun Cai Zhang
• 金额: $ 26.02万
• 依托单位: OKLAHOMA MEDICAL RESEARCH FOUNDATION
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-07-06 至 2002-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6184532
• 关键词: X ray crystallography; active sites; conformation; crystallization; enzyme substrate complex; intermolecular interaction; plasminogen; plasminogen activator; protein binding; protein engineering; protein structure function; site directed mutagenesis; streptokinase

Of central importance to thrombolysis is the activation of plasminogen (Plg) to form plasmin (Pm) which catalyze the degradation of fibrin clots. Streptokinase (SK), a bacterial protein, is a plasminogen activator widely used in the clinical treatment of clotting disorders, including myocardial infarction. Unlike tissue plasminogen activator and urokinase, SK is not a protease. SK and Plg form a non-covalent complex, which is proteolytically active and converts other Plg molecules to Pm leading to fibrinolysis. The detailed activation mechanism of Plg by SK remains unclear due largely to the lack of information on their three-dimensional structure. Since both proteins contain multiple domains which may be flexibly connected, the crystallization of the native proteins for structural studies has been unsuccessful so far. Using protein engineering the applicants have produced a functional core of the SK-Plg complex. An active-site mutant of a truncated Plg (muPlg*) complexed with SK has produced crystals which diffract to high resolution. The current application proposes three aims. First, the crystal structure of the SK-muPlg* complex on hand will be determined at high resolution. Second, the understanding of the SK and Plg interaction from the core complex structure will be extended to the full size proteins. This will be accomplished by studying the interaction and structure of the domain of the proteins. The third aim is to gain an understanding of how SK-binding activates Plg. This will be done by comparing the structures of the proteins in both free state and in complexes and by designed mutagenesis. The structural and functional understanding of the SK-Plg interaction may promote design of better thrombolytic agents for clinical treatment.

溶栓的核心是纤溶酶原的激活 (PLG)形成纤溶酶(PM)，催化纤维蛋白凝块的降解。 链激酶是一种细菌蛋白，是一种广泛存在的纤溶酶原激活剂。 用于临床治疗凝血障碍，包括心肌 脑梗塞。与组织型纤溶酶原激活剂和尿激活剂不同，SK不是 一种蛋白酶。SK和PLG形成非共价络合物，这是 并将其他PLG分子转化为PM，从而导致 纤溶作用。 SK对PLG的具体激活机制仍不清楚，主要原因是 缺乏关于它们的三维结构的信息。自.以来 这两种蛋白质都包含可以灵活连接的多个结构域， 用于结构研究的天然蛋白质的结晶已经被 到目前为止还没有成功。使用蛋白质工程的申请者拥有 产生了SK-PLG复合体的功能核心。的一个活性位点突变体 截断的PLG(muPlg*)与SK络合，产生了 衍射到高分辨率。目前的申请提出了三个目标。 首先，手头的SK-muPlg*络合物的晶体结构将是 在高分辨率下确定的。第二，对SK和PLG的理解 从核心复杂结构的相互作用将扩展到完全 大小的蛋白质。这将通过研究相互作用和 蛋白质结构域的结构。第三个目标是获得一个 理解SK结合是如何激活PLG的。这将通过以下方式完成 比较游离态和非游离态蛋白质的结构 并通过设计诱变。 对SK-PLG相互作用的结构和功能的理解可能 推动设计更好的溶栓剂用于临床治疗。