In Vitro Analysis of HIV Gag Protein Interactions

HIV Gag 蛋白相互作用的体外分析

• 批准号: 6943960
• 负责人: ERIC W BARKLIS
• 金额: $ 29.2万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-09-01 至 2007-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6943960
• 关键词: binding sites; capsid; cryoelectron microscopy; gag protein; human immunodeficiency virus; intermolecular interaction; physical model; protein engineering; protein protein interaction; protein structure; transmission electron microscopy; virus assembly

DESCRIPTION (provided by applicant): The ultimate objective of our investigations is the rational design of antivirals targeted against human immunodeficiency virus (HIV) structural (Gag) proteins. These proteins are synthesized as domains of the precursor Gag protein, Pr55Gag, which can direct the assembly of immature virus particles. However, normal processing of Pr55Gag yields the mature HIV Gag proteins matrix (MA), capsid (CA), p2, nucleocapsid (NC), pl, and p6, which organize mature virions. The availability of atomic models for matrix, nucleocapsid, and the N-terminal domain (NTD) and C-terminal domain (CTD) of capsid potentially allows the design of ligands which could interfere with the functions of the Gag proteins. However, for this approach to be successful, it is essential to characterize molecular interfaces of the Gag proteins used to choreograph the assembly and maturation of infectious viruses. The focus of our studies is HIV CA which, along with p2 and NC, constitutes the engine of virus particle assembly. We have developed methods for the dissection of HIV capsid interactions, and propose to characterize capsid interfaces in in vitro assemblies, and examine putative capsid targets. The results we obtain will provide crucial insights into mechanisms of assembly and morphogenesis, that should serve as a foundation to attack HIV at its viral core. Our specific aims are as follows: 1. Determination of capsid protein arrangements in particle assemblies: Protein arrangements in in vitro assemblies will be characterized to examine models for how capsid proteins associate in hexamer arrays. The effects of MA and NC domains will be analyzed, and resulting models will be tested in vivo. 2. Evaluation of capsid protein assembly targets: In vitro assembly assays will be employed to test structural models, and to identify accessible capsid residues. These data should prove instrumental in choosing appropriate capsid antiviral targets.

描述(由申请人提供)： 我们研究的最终目标是针对人类免疫缺陷病毒(HIV)结构蛋白(GAG)的抗病毒药物的合理设计。这些蛋白质被合成为前体Gag蛋白Pr55Gag的结构域，可以指导未成熟病毒颗粒的组装。然而，Pr55Gag的正常处理产生了成熟的HIV Gag蛋白基质(MA)、衣壳(CA)、p2、核衣壳(NC)、p1和p6，它们组成了成熟的病毒粒子。基质、核衣壳以及衣壳的N-末端结构域(NTD)和C-末端结构域(CTD)的原子模型的可用性可能会使配体的设计成为可能，从而干扰Gag蛋白的功能。然而，对于这种方法的成功，至关重要的是表征用于编排传染性病毒的组装和成熟的Gag蛋白的分子界面。我们研究的重点是HIV CA，它与p2和NC一起构成了病毒颗粒组装的引擎。我们已经开发了剖析HIV衣壳相互作用的方法，并建议在体外组装中表征衣壳界面，并检查推测的衣壳靶标。我们获得的结果将为组装和形态发生机制提供关键的见解，这些机制应该成为从病毒核心攻击艾滋病毒的基础。我们的具体目标如下：1.确定颗粒组装中的衣壳蛋白排列：体外组装中的蛋白质排列将被表征，以检验衣壳蛋白如何结合在六聚体阵列中的模型。将分析MA和NC结构域的影响，并将在体内测试由此产生的模型。2.衣壳蛋白组装靶标的评估：将使用体外组装试验来测试结构模型，并确定可获得的衣壳残基。这些数据应该有助于选择合适的衣壳抗病毒靶点。