MECHANISM OF COMPLEMENT-MEDIATED MEMBRANE DAMAGE

补体(C)是构成体液的一组血清蛋白 我们免疫防御系统的手臂。它的多种功能包括裂解 某些细菌、病毒和细胞通过直接攻击细菌， 病毒膜或细胞膜。C语言的膜攻击涉及到组装 膜上C末端蛋白(C5b-9)多聚体复合体 插入到膜中以形成可渗透到离子和小分子的通道 溶质。然而，这些插入的通道的形成机制 蛋白质仍未被分解。 拟议研究的长期目标是开发一种更清晰的 对补体介导的细胞裂解机制的理解和对 控制补体抗病毒效果的调控过程 同源细胞。本项目的具体目标是：1)分析 C5b-8和C5b-9络合物的结构/功能关系 膜的渗透深度和膜与膜之间的协同性 形成功能通道的单个复合体，2)系统地 研究温度的影响和钙离子在通道中的作用 组装和功能；3)研究膜的参与 表面分子在调节插入以及因此裂解效率中的作用 同源细胞上的膜结合末端复合体。地球的深度 单个蛋白质在末端复合体中的穿透将是 使用膜受限的光活性糖脂探针进行测定 锚定在外部或内部单层中的指定位置 模型膜，将用作C攻击的底物。这个 渠道形成的动力学将通过比较以下动力学进行研究 以囊泡释放标志物的动力学为函数的插入 插入络合物的浓度。这些方法将使我们能够 评价温度和Ca+2离子对分离的具体影响 流程(例如结合、插入、C9聚合)参与通道 集合。除了影响功能的物理化学因素 末端复合体的组装，也会受到膜因素的影响 被研究。特别是，与末端蛋白相互作用的作用 膜表面蛋白将使用光敏剂进行处理 用于检测相关蛋白质的交联剂。的关联性 这些与末端复合体插入的关联将被 学习。

Complement(C) is a group of serum proteins which constitutes the humoral arm of our immune defense system. Its diverse functions include lysis of certain bacteria, viruses and cells by direct attack on the bacterial, viral or cellular membranes. Membrane attack by C involves the assembly of a multimeric complex of C terminal proteins (C5b-9) on membrane which inserts into the membranes to form channels permeable to ions and small solutes. However, the mechanism of channel formation by these inserted proteins is still unresolved. The long-range goal of the proposed research is to develop a clearer understanding of the mechanism of complement-mediated lysis of cells and of the regulatory processes controlling the efficacy of complement against homologous cells. The specific aims of this project are: 1) to analyze structure/function relationships of the C5b-8 and C5b-9 complexes in terms of depth of penetration into the membrane and cooperativity between individual complexes in forming functional channels, 2) to systematically investigate the effect of temperature and the role of Ca+2 in channel assembly and function, and 3) to investigate the involvement of membrane surface molecules in modulating insertion, and hence lytic efficiency, of membrane-bound terminal complexes on homologous cells. The depth of penetration of the individual proteins in terminal complexes will be determined using membrane-restricted, photo-reactive glycolipid probes anchored at defined positions in either the outer or the inner monolayer of model membranes which will be used as substrates for C attack. The dynamics of channel formation will be studied by comparing the kinetics of insertion with the kinetics of marker release from vesicles as a function of the concentration of inserted complexes. These methods will allow us to evaluate the specific effects of temperature and Ca+2 ions on separate processes (eg. binding, insertion, C9 polymerization) involved in channel assembly. In addition to the physicochemical factors affecting functional assembly of terminal complexes, the influence of membrane factors will also be studied. In particular, the role of terminal protein interaction with membrane surface proteins will be addressed using photosensitive crosslinking reagents to detect associated proteins. The relevance of these associations to insertion of the terminal complex will then be studied.