Complement Convertase: Assembly, Function and Regulation

补体转化酶：组装、功能和调节

• 批准号: 6616462
• 负责人: DENNIS EMIL HOURCADE
• 金额: $ 22.95万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-04-01 至 2007-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6616462
• 关键词: active sites; complement; complement pathway; enzyme activity; enzyme linked immunosorbent assay; hydrolase; molecular assembly /self assembly

DESCRIPTION (provided by applicant): The complement system is a critical participant in both innate and acquired immunity. Complement marks infectious agents for immune clearance or cell lysis and it forms a focal point for inflammatory reactions. Moreover, complement facilitates antigen localization to the spleen and lymph nodes, where it lowers the threshold for the activation of specific B cells. Complement can also be a principal cause of tissue damage in human diseases such as antibody-mediated autoimmunity, immune complex deposition syndromes, ischemic reperfusion injury, and hyperacute graft rejection. Nearly all the biological consequences of complement require the enzymatic cleavage of C3, an abundant serum glycoprotein. The enzymes that cleave C3, the C3 convertases, are major players in the complement activation pathways and occur in two structurally and functionally homologous forms, the alternative pathway (AP) C3 convertase (C3bBb) and the classical pathway (CP) C3 convertase (C4bC2a). Several structurally related proteins, DAF, CR1, C4BP, and factor H, inhibit inappropriate complement activation (e.g. on self-tissue). They act to promote the irreversible dissociation of active convertases, a process known as decay acceleration. In contrast, the serum protein properdin can partially stabilize C3bBb on target surfaces. The long-range goal of this work is to create new approaches to the therapeutic control of complement. Our strategy is to elucidate the unique biochemical features of the C3 convertases, the key enzymes in complement activation, and identify promising therapeutic targets. A simple assay was developed for analyzing the assembly, stabilization, and decay acceleration of the AP C3 convertases C3bBb and C3bBbP: Microtiter wells were coated with C3b, incubated with fluid phase factor B, factor D, divalent cation and, in some cases, properdin; complexes were detected by ELISA. Employment of the ELISA-based assay with panels of factor B, DAF and CR1 mutants generated by site-directed mutagenesis has led to the identification of a number of possible active sites and the proposal of several key hypotheses. To test these hypotheses, studies are proposed with the following specific aims: 1) Develop a model for convertase assembly and the activation of its serine protease domain; 2) Determine how properdin stabilizes AP convertases; 3) Elucidate the mechanisms of decay acceleration.

描述（由申请人提供）：补体系统是先天免疫和获得性免疫的关键参与者。补体标志着免疫清除或细胞溶解的感染因子，并形成炎症反应的焦点。此外，补体促进抗原定位到脾脏和淋巴结，在那里它降低了特异性B细胞激活的门槛。补体也可能是人体疾病组织损伤的主要原因，如抗体介导的自身免疫、免疫复合物沉积综合征、缺血再灌注损伤和超急性移植排斥反应。