STRUCTURE/FUNCTION OF TERMINAL COMPLEMENT PROTEINS

描述(改编自《调查者摘要》)：长期目标 这个项目的主要目的是为了更详细地了解 补体细胞溶解复合体人C5b-9的组装和功能 被称为“膜攻击复合体”或MAC。Mac由C5b组成 以及终端部件C6、C7、C8和C9。其中，C6、C7、 C8-α和C8-β亚基，以及C9组成了一个独特的家族 具有高度保守的基因结构、氨基酸序列和 通用模块化设计。C8-伽马亚基是不相关的，是 广泛分布的结合小分子的Lipocalin家族 疏水配体。由于其在MAC组装中的核心作用，其 多亚基结构及其多重相互作用，将是研究的重点 主要是关于人类C8的结构和功能。具体目标是： (1)确定C8a中的C8b和C9结合位点；(2)确定片段 C8a介导C8g和C8g的细胞内识别和结合 C8a-g二聚体的形成；(3)鉴定C5b-7和C8a-g结合位点 在C8b中；(4)确定C8g的晶体结构并进行延伸 其配体结合特性和功能的研究。实验将使用 重组(R)形式的人C8a-g、C8a、C8b和C8g。结合位点将 使用rC8a和rC8b的截短突变体和嵌合体进行鉴定，其中 系统地交换细分市场，并对产品进行分析 相应的功能互换。因为他们可能在 研究将在MAC组装过程中调节蛋白质-蛋白质相互作用 主要关注每种蛋白质中保守的模块。其他实验 将尝试结晶C8、C8a-g、C8b和rC8a。衍射光质量 RC8G的晶体已经生产出来，结构将完成。 C8G的功能及其天然配体的身份尚不清楚。 实验将搜索可能的配体，并研究 C8g与炎性介质结合从而抑制的可能性 促炎反应。其他实验将检验它在 细菌致死。对人类C8的拟议研究将提供新的见解 进入所有终端组件相互作用的机制，从而 促进对治疗有用的类似物的设计和开发 MAC和MAC裂解和刺激功能的调节器。因为MAC 蛋白质家族在结构、功能、信息等方面是独一无二的 这也将有助于理解蛋白质-蛋白质 一般情况下，互动。

DESCRIPTION (Adapted from Investigator's abstract): The long-term objective of this project is to gain a detailed understanding of the mechanism of assembly and function of human C5b-9, the cytolytic complex of complement referred to as the "membrane attack complex" or MAC. MAC is composed of C5b and the terminal components C6, C7, C8 and C9. Among the latter, C6, C7, the C8-alpha and C8-beta subunits, and C9 comprise a unique family of proteins with highly conserved gene structures, amino acid sequences and a common modular design. The C8-gamma subunit is unrelated and is a member of the lipocalin family of widely distributed proteins that bind small hydrophobic ligands. Because of its central role in MAC assembly, its multi-subunit structure and its multiple interactions, studies will focus primarily on the structure and function of human C8. Specific aims are to: (1) identify the C8b and C9 binding sites in C8a; (2) identify the segment of C8a that mediates intracellular recognition and binding of C8g and formation of the C8a-g dimer; (3) identify the C5b-7 and C8a-g binding sites in C8b; (4) determine the crystal structure of C8g and extend ongoing studies of its ligand-binding properties and function. Experiments will use recombinant (r) forms of human C8a-g, C8a, C8b and C8g. Binding sites will be identified using truncated mutants and chimeras of rC8a and rC8b in which segments are systematically exchanged and the products analyzed for a corresponding exchange of function. Because of their likely role in mediating protein-protein interactions during MAC assembly, studies will focus primarily on the modules conserved in each protein. Other experiments will attempt to crystallize C8, C8a-g, C8b and rC8a. Diffraction-quality crystals of rC8g have been produced and the structure will be completed. The function of C8g and identity of its natural ligand are unknown. Experiments will search for possible ligands as well as investigate the possibility that C8g binds inflammatory mediators and thus may inhibit proinflammatory responses. Other experiments will examine its role in bacterial killing. Proposed studies of human C8 will provide new insight into the mechanism by which all the terminal components interact and thereby facilitate the design and development of therapeutically useful analogues of MAC and regulators of MAC lytic and stimulatory functions. Because the MAC family of proteins is unique in terms of structure and function, information obtained will also contribute to an understanding of protein-protein interactions in general.