Herpes Simplex Virus Capsid Assembly and DNA Packaging

单纯疱疹病毒衣壳组装和 DNA 包装

• 批准号: 7534331
• 负责人: FREDERICK L HOMA
• 金额: $ 31.83万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-12-01 至 2010-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7534331
• 关键词: Affect; Affinity Chromatography; Animals; Antiviral Agents; Baculoviruses; Binding; Biochemical; Biochemical Genetics; Biology; Capsid; Capsid Proteins; Cell physiology; Cells; Complex; Cytomegalovirus; DNA; DNA Binding; DNA Packaging; DNA Sequence; Development; Disease; Genes; Genetic; Genome; Goals; Herpesviridae; Herpesvirus 1; Herpesvirus Type 3; Homologous Gene; Human; Human Herpesvirus 4; Hybrids; Immune; Immunocompromised Host; In Vitro; Individual; Infection; Left; Length; Location; Maps; Medical; Minor; Mutation; Newborn Infant; Nucleic Acids; Population; Process; Production; Protein Binding; Proteins; Proteomics; Reaction; Research; Research Personnel; Role; Series; Simplexvirus; Site; Specificity; Staging; System; Tertiary Protein Structure; Testing; Translations; Viral; Viral Proteins; Yeasts; insertion/deletion mutation; mutant; novel; prevent; programs; protein complex; protein degradation; protein function; protein protein interaction; terminase; viral DNA; yeast two hybrid system

Infections with human herpesviruses are endemic in the population with some herpesviruses causing severe disease, especially in immune impaired individuals and newborns. The increased immunocompromised population has created an unmet medical need for antivirals against herpesviruses. The goal of the proposed project is to gain a better understanding of DNA cleavage and packaging in herpes simplex virus (HSV) through the analysis of two viral proteins, UL25 and UL28, that are required for this process. The specific hypothesis behind the proposed research is that UL25 and UL28 are required for packaging DNA into HSV capsids at separate stages in the cleavage packaging reaction. The UL28 protein is required for cleavage of concatemeric DNA into unit length molecules while UL25 serves an essential function in the production of DNA containing capsids. Although, the focus will be on UL25 and UL28 the goal is to characterize the cleavage/packaging process using genetic and biochemical approaches to examine how mutations in the UL25 and UL28 genes affect the interaction of these proteins with: (i) DNA, (ii) capsid proteins, (iii) cleavage/packaging proteins and (iv)host cell proteins. Specific goals of the project are to: 1. Determine the capsid location, DNA binding specificity and protein-protein interactions of UL25. 2. Identify functional domains of UL28 that are important for capsid incorporation, binding viral DNA packaging sites and for interaction with terminase (UL15) and portal protein (UL6) by characterizing a series of HSV-1 UL28 mutants that we have isolated; and map the second site mutation for a revertent of one of the lethal UL28 linker- insertion mutants since marker rescue and DNA sequencing have demonstrated that the mutation does not map to the UL28 gene. 3. Examine the interaction of UL25 and UL28 with viral and cell proteins using proteomic and biochemical approaches in order to identify essential protein-protein interactions and protein complexes that are involved in the cleavage/packaging reaction. These studies aim to elucidate the mechanism underlying HSV DNA cleavage and packaging and may suggest novel targets for the development of antivirals.

人类疱疹病毒感染在人群中是地方性的，一些疱疹病毒会引起严重的 疾病，尤其是免疫受损的个人和新生儿。增强的免疫力损害了 人口对抗疱疹病毒药物的医疗需求尚未得到满足。建议的目标是 该项目是为了更好地了解单纯疱疹病毒(HSV)中DNA的切割和包装 通过分析这一过程所需的两种病毒蛋白UL25和UL28。具体的 这项研究背后的假设是，将DNA包装到HSV中需要UL25和UL28 裂解包装反应中不同阶段的衣壳。UL28蛋白是切割蛋白质所必需的 将DNA连接成单位长度的分子，而UL25在生产 含有衣壳的DNA。虽然重点将放在UL25和UL28上，但目标是描述 使用遗传和生化方法的切割/包装过程来检查基因突变是如何 UL25和UL28基因影响这些蛋白质与：(I)DNA，(Ii)衣壳蛋白，(Iii) 裂解/包装蛋白和(Iv)宿主细胞蛋白。该项目的具体目标是：1.确定 UL25衣壳定位、DNA结合特异性和蛋白质-蛋白质相互作用2.确定功能 UL28的结构域对于衣壳掺入、结合病毒DNA包装位点和 HSV-1 UL28突变株与终止酶(UL15)和门静脉蛋白(UL6)的相互作用 我们已经分离出；并将第二个位点突变映射为致命的UL28连接子之一的回复- 自标记挽救和DNA测序以来的插入突变体已经证明，突变不会 定位到UL28基因。3.研究UL25和UL28与病毒和细胞蛋白的相互作用 蛋白质组学和生化方法，以确定必需的蛋白质-蛋白质相互作用和蛋白质 参与切割/包装反应的复合体。这些研究旨在阐明 HSV DNA切割和包装的潜在机制，并可能提出新的靶点 抗病毒药物的开发。