The Pox Virion Molecular Interactome

痘病毒体分子相互作用组

• 批准号: 8582931
• 负责人: Paul D Gershon
• 金额: $ 21.73万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-09 至 2015-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8582931
• 关键词: Africa; Agriculture; Animals; Antibodies; Antiviral Agents; Appearance; Applications Grants; Architecture; Area; Binding; Boxing; Capsid Proteins; Complement; Complex; Core Protein; DNA Binding; Development; Disease; Disease Outbreaks; Enzymes; Equipment and supply inventories; Family; Family Picornaviridae; Future; Genetic Transcription; Genome; Herpesviridae; Heterogeneity; Human; In Situ; Intervention; Knowledge; Lateral; Mass Spectrum Analysis; Mediating; Medical; Methodology; Molecular; Molecular Structure; Monkeypox; Normal Pressure Hydrocephalus; Nuclear Pore Complex; Nucleoproteins; Nucleosomes; Peptides; Positioning Attribute; Poxviridae; Proteins; RNA Polymerase II; RNA Viruses; Resolution; Risk; Role; Route; Shapes; Smallpox; Solubility; Staging; Structural Protein; Structure; Techniques; Testing; Textiles; Therapeutic; Therapeutic Agents; Vaccinia; Vaccinium; Virion; Virus; Virus Diseases; Yeasts; anthrax lethal factor; base; crosslink; design; multicatalytic endopeptidase complex; novel; particle; protein complex; protein crosslink; protein protein interaction; public health relevance; therapeutic vaccine; tool; transmission process; virus envelope; yeast two hybrid system

DESCRIPTION (provided by applicant): Smallpox has, historically, been one of the great killers of mankind. Although this disease is considered to have been eradicated some 35 years ago, the poxviruses, nonetheless, comprise a major family of viruses of medical, ecological and agricultural importance. For humans, aside from the possibility of smallpox re-introduction, eradication has coincided with the appearance of human monkeypox in Africa during the past 40 years and in the US during the past decade. Not knowing the lethal factor in smallpox, the full potential of such outbreaks remains uncertain. The importance of virus envelopes and capsid proteins in mediating the effects of antiviral therapeutics and vaccines is undisputed. From the picornaviruses to the herpesviruses, an understanding of virion outer molecular structures at molecular or atomic resolution, has instructed the rational design of therapeutic agents and an understanding of mechanisms that cause and thwart virus infection and disease. Due to their complexity, asymmetry and heterogeneity, the poxviruses have, however, been particularly persistent in defeating attempts to understand their virion structure at the molecular level, thus evading an important potential avenue for intervention. The P.I. hypothesizes that the complexity of the vaccinia virion may prove to be an Achilles heel. In addition, a full understanding of virion structure may be regarded as one of the last remaining black boxes in the lifecycle of the poxviruses - one which impinges upon the early transcription, genome uncoating and virion assembly stages of poxvirus replication. The major hole in our knowledge of pox virion structure lies at a level between the inventory of proteins present within the virion (which is largely known) and the basic topological and topographical features of the intact particle (also known). This intervening area may be referred to as the virion's "molecular architecture", or protein "interactome". In this R21 proposal, the P.I. has chosen a protein-protein crosslinking approach in combination with mass spectrometry. Such an approach for interactome analysis is unbiased in many respects, and has a track record of informing the molecular architectures of elaborate cellular assemblies such as the nuclear pore complex, 20S proteasome and RNA polymerase II. Aim 1 of this proposal seeks to identify directly juxtaposed proteins within the virion core via covalent protein-protein crosslinking/MS, taking "top-down" (protein-level) and "bottom-up" (peptide-level) approaches. The P.I. hypothesizes that the pox virion core wall may not be fundamentally dissimilar to the matrix protein layers of some enveloped RNA viruses, and that the classical delineation of enzymes in the deep interior with structural proteins surrounding may not be as clear cut as currently supposed.

描述（由申请人提供）：从历史上看，天花是人类最大的杀手之一。虽然这种疾病被认为在大约35年前就已被根除，但痘病毒仍然是具有医学、生态和农业重要性的病毒的主要家族。对于人类来说，除了天花重新引入的可能性之外，过去40年中，非洲和过去十年中，美国人类猴痘的出现恰逢天花的根除。由于不知道天花的致命因素，这种爆发的全部潜力仍然不确定。病毒包膜和衣壳蛋白在介导抗病毒疗法和疫苗的作用中的重要性是无可争议的。从微小核糖核酸病毒到疱疹病毒，在分子或原子分辨率上对病毒体外部分子结构的理解指导了治疗剂的合理设计以及对引起和阻止病毒感染和疾病的机制的理解。然而，由于其复杂性、不对称性和异质性，痘病毒特别坚持不懈地挫败了在分子水平上理解其病毒体结构的尝试。 这是一个重要的潜在干预渠道。私家侦探假设牛痘病毒粒子的复杂性可能是致命的弱点。此外，对病毒体结构的充分理解可以被认为是痘病毒生命周期中最后剩下的黑盒子之一，其影响痘病毒复制的早期转录、基因组脱壳和病毒体组装阶段。我们对痘病毒体结构的认识中的主要漏洞在于病毒体中存在的蛋白质的库存之间的水平 （这在很大程度上是已知的）和完整颗粒的基本拓扑和形貌特征（也是已知的）。这个中间区域可以被称为病毒体的“分子结构”或蛋白质“相互作用组”。在这个R21提案中，P.I.选择了蛋白质-蛋白质交联方法结合质谱分析。这种用于相互作用组分析的方法在许多方面是无偏的，并且具有告知精细细胞组装体（诸如核孔复合物、20 S蛋白酶体和RNA聚合酶II）的分子结构的跟踪记录。本提案的目的1旨在通过共价蛋白质-蛋白质交联/MS，采用“自上而下”（蛋白质水平）和“自下而上”（肽水平）方法，直接鉴定病毒体核心内的并列蛋白质。私家侦探假设痘病毒粒子的核心壁可能与某些包膜RNA病毒的基质蛋白层没有根本的不同，并且在结构蛋白围绕的内部深处的酶的经典描绘可能不像目前假设的那样清晰。