PROTEOMICS ANALYSIS UNRAVELS THE FUNCTIONAL REPERTOIRE OF CORONAVIRUS NSP3

蛋白质组学分析揭示了冠状病毒 NSP3 的功能库

• 批准号: 8171341
• 负责人: PETER KUHN
• 金额: $ 0.08万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-01 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8171341
• 关键词: Architecture; Bioinformatics; Cells; Computer Retrieval of Information on Scientific Projects Database; Coronaviridae; Coronavirus; Coronavirus Infections; Cysteine; Data; Enzymes; Escherichia coli; Funding; Grant; Growth; Infection; Institution; Mass Spectrum Analysis; Metal Ion Binding; Molecular Chaperones; Nonstructural Protein; Nucleic Acids; Paint; Papain; Peptide Hydrolases; Phosphotransferases; Process; Proteins; Proteomics; Publishing; RNA Recognition Motif; Recombinant Proteins; Research; Research Personnel; Resolution; Resources; Role; Severe Acute Respiratory Syndrome; Signal Transduction; Source; Staging; Surveys; Techniques; Tertiary Protein Structure; United States National Institutes of Health; Viral; Viral Matrix Proteins; Viral Proteins; Virion; Virus Assembly; base; novel; viral RNA; Architecture; Bioinformatics; Cells; Computer Retrieval of Information on Scientific Projects Database; Coronaviridae; Coronavirus; Coronavirus Infections; Cysteine; Data; Enzymes; Escherichia coli; Funding; Grant; Growth; Infection; Institution; Mass Spectrum Analysis; Metal Ion Binding; Molecular Chaperones; Nonstructural Protein; Nucleic Acids; Paint; Papain; Peptide Hydrolases; Phosphotransferases; Process; Proteins; Proteomics; Publishing; RNA Recognition Motif; Recombinant Proteins; Research; Research Personnel; Resolution; Resources; Role; Severe Acute Respiratory Syndrome; Signal Transduction; Source; Staging; Surveys; Techniques; Tertiary Protein Structure; United States National Institutes of Health; Viral; Viral Matrix Proteins; Viral Proteins; Virion; Virus Assembly; base; novel; viral RNA

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Severe acute respiratory syndrome (SARS) coronavirus infection and growth are dependent on initiating signaling and enzyme actions upon viral entry into the host cell. Proteins packaged during virus assembly may subsequently form the first line of attack and host manipulation upon infection. A complete characterization of virion components is therefore important to understanding the dynamics of early stages of infection. Mass spectrometry and kinase profiling techniques identified nearly 200 incorporated host and viral proteins. We used published interaction data to identify hubs of connectivity with potential significance for virion formation. Surprisingly, the hub with the most potential connections was not the viral M protein but the nonstructural protein 3 (nsp3), which is one of the novel virion components identified by mass spectrometry. Based on new experimental data and a bioinformatics analysis across the Coronaviridae, we propose a higher-resolution functional domain architecture for nsp3 that determines the interaction capacity of this protein. Using recombinant protein domains expressed in Escherichia coli, we identified two additional RNA-binding domains of nsp3. One of these domains is located within the previously described SARS-unique domain, and there is a nucleic acid chaperone-like domain located immediately downstream of the papain-like proteinase domain. We also identified a novel cysteine-coordinated metal ion-binding domain. Analyses of interdomain interactions and provisional functional annotation of the remaining, so-far-uncharacterized domains are presented. Overall, the ensemble of data surveyed here paint a more complete picture of nsp3 as a conserved component of the viral protein processing machinery, which is intimately associated with viral RNA in its role as a virion component.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这不一定是调查员的机构。 严重的急性呼吸综合征（SARS）冠状病毒感染和生长取决于在病毒进入宿主细胞时引发信号传导和酶作用。在病毒组装过程中包装的蛋白质随后可能会在感染后形成第一线攻击和宿主操纵。因此，病毒素成分的完整表征对于理解感染早期阶段的动态很重要。质谱和激酶分析技术确定了近200个掺入宿主和病毒蛋白。我们使用已发布的交互数据来识别与形成病毒体形成潜在意义的连通性的枢纽。令人惊讶的是，具有最潜在连接的枢纽不是病毒M蛋白，而是非结构蛋白3（NSP3），这是通过质谱鉴定的新型病毒粒子成分之一。基于新的实验数据和整个冠状病毒的生物信息学分析，我们提出了NSP3的高分辨率功能结构域结构，以决定该蛋白质的相互作用能力。使用大肠杆菌中表达的重组蛋白结构域，我们确定了NSP3的另外两个RNA结合结构域。这些结构域之一位于先前描述的SARS唯一结构域内，并且有一个核酸伴侣样结构域位于木瓜蛋白酶样蛋白酶结构域的下游。我们还确定了一个新型半胱氨酸协调的金属离子结合域。介绍了剩余的，纯净的域的临时相互作用和临时功能注释的分析。总体而言，此处调查的数据集合将NSP3的更完整图片描绘为病毒蛋白加工机制的保守组成部分，该组件与病毒RNA与病毒RNA密切相关。