Administrative Supplement for GM100888

GM100888 行政补充

基本信息

批准号：
9902027
负责人：
Gino Cingolani
金额：
$ 6.73万
依托单位：
THOMAS JEFFERSON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-08-01 至 2021-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9902027
关键词：
ATP phosphohydrolase Adenoviruses Administrative Supplement Affinity Antiviral Agents Architecture Award Bacteriophage P22 Bacteriophages Binding Binding Sites Biochemical Biochemical Reaction Biological Assay Capsid Catalytic Domain Cell Nucleus Chemicals Cleaved cell Complex Cryoelectron Microscopy Crystallography Cytomegalovirus DNA DNA Viruses Data Dependovirus Deuterium Enzymatic Biochemistry Fill-It Funding Genetic Materials Genome Goals Grant HIV-1 Herpesviridae Human Hybrids Hydrogen Importins Individual Knowledge Link Macromolecular Complexes Maps Mass Spectrum Analysis Methods Modernization Molecular Molecular Conformation Molecular Machines Molecular Structure Motor Nature Nuclear Pore Complex Parvovirus Pathogenicity Planet Earth Poxviridae Protein Biochemistry Proteins Protomer Pump RNA Viruses Reaction Research Role Salmonella Phages Site Site-Directed Mutagenesis Structure Surface Tail Techniques Testing Tobacco Mosaic Virus Viral Viral Genome Viral Packaging Virion Virus Virus Replication Work Yeasts base biophysical analysis comparative flexibility human disease in vivo interest macromolecular assembly novel nuclease pathogen single molecule stoichiometry structural biology synthetic antibodies terminase viral DNA

项目摘要

Project Summary Viral genome packaging is a complex, non-spontaneous, multi-step enzymatic reaction that in tailed bacteriophages and herpesviruses proceeds via formation of an empty precursor capsid (or procapsid) that is filled with genetic material by the action of two proteins, known as large and small terminase. Though commonly studied as individual subunits, viral terminases bind, pump and cleave viral DNA assembled into large macromolecular complexes of poorly characterized structure, function and composition. The recent discovery of a potent antiviral agent specific to Human Cytomegalovirus small terminase subunit (pUL56) has grown further interest in viral packaging motors. In this grant, combining hybrid methods in structural biology (i.e. X-crystallography, cryo-electron microscopy, hydrogen/deuterium exchange mass spectrometry) with modern biochemical approaches (i.e. conformation- specific synthetic Fabs, site directed mutagenesis, yeast 1-hybrid), we seek to understand the principles governing viral genome packaging through the comparative analysis of motors from different DNA viruses. We are particularly interested in deciphering the atomic structure of macromolecular assemblies formed by terminase subunits during the packaging reaction and the role of S-terminase that is functionally conserved from bacterial viruses to herpesviruses. This research tries to fill a significant and growing knowledge gap between the enzymology of genome packaging, which is increasing well-understood thanks to single molecule biophysical studies, and the molecular machines catalyzing packaging. Building upon the work initiated in the previous funding cycle, we seek to: 1.) Elucidate the architecture of terminase assemblies formed during viral genome packaging; 2.) Determine the conserved architecture of Human Cytomegalovirus small terminase (pUL56) and its interaction with viral DNA and the antiviral drug letermovir.

项目摘要病毒基因组包装是一种复杂的，非自发的多步酶反应，在尾部噬菌体和疱疹病毒通过形成空的前体帽子（或procapsid）而进行通过两种蛋白质（称为大和小末端酶）的作用充满遗传物质。尽管通常以单个亚基的研究，病毒末端酶结合，泵和裂解病毒DNA聚集到特征性较差的结构，功能和组成的大型大分子复合物。最近发现针对人类巨细胞病毒小末端酶亚基（PUL56）的有效抗病毒剂的发现对病毒包装电机的兴趣进一步兴趣。在这项赠款中，结合了结构生物学中的混合方法（即X晶格学，冷冻电子显微镜，采用现代生化方法（即构象 - 特定的合成晶圆厂，定向诱变，酵母1-Hybrid），我们试图理解原理通过对不同DNA病毒的电动机进行比较分析来管理病毒基因组包装。我们对解释由大分子组件的原子结构特别感兴趣包装反应过程中的末端酶亚基和功能保守的S端酶的作用从细菌病毒到疱疹病毒。这项研究试图填补大量且不断增长的知识差距在基因组包装的酶学之间，由于单分子，它正在越来越大量理解生物物理研究和分子机催化包装。建立在发起的工作的基础以前的资金周期，我们寻求：1。）阐明病毒期间形成的末端酶组件的结构基因组包装； 2.）确定人类巨型病毒小末端酶的保守结构（PUL56）及其与病毒DNA和抗病毒药物Letermovir的相互作用。