MINIPROTEIN MIMETICS

描述(由申请人提供)：本项目的中心目标是使用微型蛋白质 与gp120KD4复合体的结构一起模拟设计HIV-1与人类细胞相互作用的拮抗剂。HIV-1是艾滋病细胞感染的主要途径之一，T细胞对接和进入是由HIV包膜蛋白gp120对T细胞表面蛋白CD4和趋化因子辅助受体的特异性识别所推动的。CD4的晶体结构是已知的，它与gp120和辅助受体替代抗体17b的复合体也是已知的。揭示了GP120与CD4和辅受体结合的结构性质和相互关系。病毒-细胞识别中蛋白质参与者的高分辨率结构理解的进步，加上先进的模拟设计技术，现在使得将微型蛋白质工程与生物物理和生物分析相结合，以获得对CD4-gp120相互作用过程的结构基础的高级机制理解成为可能。这种机械性的理解将有助于指导设计新的拮抗剂 艾滋病毒感染。其具体目的是：(1)利用合成和互补噬菌体随机化方法构建新型的HIV-1包膜微蛋白模拟物，并优化其与HIV-1外膜的亲和力；(2)确定控制HIV-1包膜共受体结合位点激活的CD4微蛋白模拟物中的关键结构元件，并形成有利于与Env结合且辅受体结合位点激活减少的微蛋白变体；(3)确定HIV-1外膜三聚体与单体形式(包括胞外区和膜相关形式)的结构和相互作用性质，并完善HIV-1细胞附着和附着拮抗剂所需关键结构元件的拓扑图。总体而言，该项目将对CD4-GP 120复合体界面上的关键结构元件进行高级定义，这些结构元件可触发HIV-1共受体和随后的细胞感染的包膜亲和力增加；提高对驱动HIV-1附着和宿主细胞感染的HIV-1包膜-辅受体分子机制的理解；以及为抗病毒治疗的新拮抗剂候选药物提供分子线索。所得到的微小蛋白模拟策略将有助于确定其他蛋白质-蛋白质相互作用的机制，并为其设计拮抗剂，例如gp41-gp120和趋化因子受体。

DESCRIPTION (provided by applicant): The central objective of this Project is to use miniprotein mimetics together with the structure of the gp120KD4 complex to design antagonists of the interaction of HIV-1 with human cells. T-cell docking and entry by HIV-1, a major route of cell infection in AIDS, is driven by specific recognition of T-cell surface protein CD4 and chemokine coreceptor by HIV envelope protein gp120. The crystallographic structure of CD4 is known, as is its complex with gp120 and the coreceptor surrogate antibody 17b. The structural nature and interrelationship of gp 120 binding sites for CD4 and coreceptor have been revealed. The advancing high-resolution structural understanding of the protein participants in virus-cell recognition, together with the advancing technology of mimetics design, now make it possible to use miniprotein engineering coordinately with biophysical and biologicalanalysis to obtain an advanced mechanistic understanding of the structural basis of the CD4-gp120interaction process. This mechanistic understanding will help guide the design of new antagonists for HIV infection. The specific aims are: (1) Construct novel miniprotein mimetics of CD4 using synthetic and complementary phage randomization methodologies and optimize their affinities for HIV- 1 envelope; (2) Identify key structural elements in miniprotein mimetics of CD4 that control activation of the coreceptor binding site of HIV-1 Env, and form miniprotein variants which favor Env binding with diminished coreceptor site activation; (3) Determine structural and interaction properties of miniprotein mimetics with trimeric vs monomeric forms of HIV- 1 envelopes, including ectodomain and membrane-associated forms, and refine the topological map of key structural elements needed for HIV-1-cell attachment and attachment antagonists. Overall, this project will yield an advanced definition of key structural elements at the interface of the CD4-gp 120 complex that trigger increased HIV- 1 envelope affinity for co-receptor and consequent cell infection; improved understanding of the CD4-viral envelope-coreceptor molecular machine that drives HIV- 1 attachment and infection of host cells; and molecular leads for new antagonist candidates for antiviral therapy. The miniprotein mimetics strategies derived will be useful to determine the mechanism of, and design antagonists for, other protein-protein interactions, such as the gp41 -gp 120 and chemokine receptor.