MECHANISMS IN PAPOVAVIRUS ASSEMBLY

乳腺病毒组装机制

• 批准号: 2837601
• 负责人: Robert L Garcea
• 金额: $ 32.55万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-07-01 至 2002-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2837601
• 关键词: Polyomavirus; Sf9 cell line; X ray crystallography; bovine papillomavirus; calcium flux; capsid; cryoelectron microscopy; human papillomavirus; intermolecular interaction; molecular chaperones; phosphorylation; protein binding; protein purification; protein structure function; recombinant proteins; simian virus 40; site directed mutagenesis; transfection /expression vector; virion; virus assembly; virus genetics; virus protein; virus virus interaction

DESCRIPTION: The atomic resolution structures of viruses provide a new framework for studying the interaction of viruses with their cellular receptors, designing anti-viral therapeutics, and understanding the bonding interactions in complex macromolecular assemblies. Although these structures represent well-resolved end-products, the process of virus assembly remains ill-defined with regard to its regulation within virus-infected cells and the properties of the viral structural proteins which determine the pathways of assembly. The study of virus assembly has direct implications for designing anti-viral therapies, but also has relevance to the general biologic problems of protein folding, protein trafficking within cells, and the interaction of eukaryotic chromosomes with structural proteins. The aims of this proposal are directed at understanding the structure and assembly of papovaviruses at the atomic, biochemical, and cell biological levels. A structural analysis of the human papillomavirus (HPV) subtype-11 L1 capsid protein will be undertaken using a recombinant L1 protein as a substrate for interactions between the papovavirus "minor" capsid proteins and the capsid will be studied using cryoelectron microscopy of virus-like particles purified after expression of polyoma capsid proteins in insect cells, and biochemical characterization of polyomavirus VP1/VP2/VP3 or HPV L1/L2 complexes. Using mutant polyoma VP1 proteins subdomains of the VP1 carboxy terminus will be analyzed in relation to their effect upon the size, shape, and curvature of capsids formed in vitro. The interaction of polyoma VP1 with cell chaperone proteins will be analyzed to determine whether chaperone binding influences association of VP1 capsomeres into capsids. The in vivo effects of calcium, protein concentration, and chaperone binding on capsid assembly will be tested using mutant VP1 proteins expressed in insect cells.

描述：病毒的原子分辨率结构提供了新的 研究病毒与细胞相互作用的框架 受体，设计抗病毒疗法，并了解粘结 复杂的大分子组件中的相互作用。 虽然这些 结构代表良好的终产物，病毒的过程 大会在其内部的监管方面仍然不明显 病毒感染的细胞和病毒结构蛋白的特性 决定组装的途径。 病毒组装的研究具有 对设计抗病毒疗法的直接影响，但也有 与蛋白质折叠，蛋白质的一般生物学问题有关 细胞内的运输以及真核染色体与 结构蛋白。 该提案的目的旨在了解结构和 原子，生化和细胞生物学的木瓜病毒组装 水平。 人乳头瘤病毒（HPV）亚型-11的结构分析-11 将使用重组L1蛋白作为A进行L1衣壳蛋白 木瓜病毒“次要”衣壳蛋白之间相互作用的底物 并将使用类似病毒的冷冻电子显微镜研究衣壳 昆虫中多瘤衣壳蛋白表达后纯化的颗粒 细胞，以及多瘤病毒VP1/VP2/VP3或HPV的生化表征 L1/L2复合物。 使用VP1的突变聚体VP1蛋白子域 羧基终端将根据其对大小的影响进行分析， 体外形成的衣壳的形状和曲率。 多瘤的相互作用 将分析带有细胞伴侣蛋白的VP1，以确定是否 伴侣结合影响VP1辣椒菌与衣壳的关联。 钙，蛋白质浓度和伴侣结合的体内影响 在衣壳组件上，将使用在以表达的突变体VP1蛋白进行测试 昆虫细胞。