Assembly of Picornaviral Replication Complexes

小核糖核酸病毒复制复合物的组装

基本信息

批准号：
10088366
负责人：
Olve Breien Peersen
金额：
$ 37.76万
依托单位：
COLORADO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-04-01 至 2023-02-22
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10088366
关键词：
Active Sites Acute Hepatitis Address Antiviral Agents Attenuated Binding Biophysics Catalysis Cells Child Common Cold Complement Complex Coxsackie Viruses Crystallization Data Dimerization Disease Endoplasmic Reticulum Enzymes Event Family Family Picornaviridae Fingers Funding Genome Goals Growth Hand, Foot and Mouth Disease Heart Diseases Hepatitis A Virus Human Human poliovirus In Vitro Internet Lead Life Cycle Stages Lipid Bilayers Mediating Membrane Methods Microscopy Molecular Mus Nucleotides Paralysed Pathway interactions Poliomyelitis Polymerase Polyproteins Process Protein Engineering Protein Precursors Proteins Proteolytic Processing RNA RNA Binding RNA Stability RNA Viruses RNA chemical synthesis RNA replication RNA-Directed RNA Polymerase RNA-Protein Interaction Reaction Replication-Associated Process Research Project Grants Rhinovirus Role Solid Specificity Structure Surface Translations Vesicle Viral Viral Drug Resistance Viral Genome Viral Proteins Virion Virus Virus Assembly Virus Replication base biophysical techniques cell type cost human disease member novel novel vaccines particle protein protein interaction response scaffold sensor single molecule stem structural biology viral RNA

项目摘要

The picornaviruses are a family of small positive sense single stranded RNA viruses that cause a wide range of diseases at an annual cost well into the hundreds of million dollars. Members include acute hepatitis A virus, the heart disease causing coxsackie B3 virus, rhinoviruses that cause more than half the occurrences of the common cold, and the paralyzing poliovirus. These viruses share a life cycle where RNA replication and viral assembly occurs in large membrane anchored replication complexes assembled on the surfaces of vesicles derived from the endoplasmic reticulum. The replication process is driven by a virally encoded RNA dependent RNA polymerase (3Dpol) that is responsible for the synthesis of all viral RNA. This research project is focused on the structure and assembly of the viral replication centers and on structure-function studies of the viral polymerases to understand mechanisms that control of elongation rates and replication fidelity. We have solved multiple crystal structures of 3Dpol as elongation complexes with bound RNA and these structures revealed a novel mechanism by which RdRPs close their active sites for catalysis in response to nucleotide binding. In this upcoming project period we will use biophysical and structural biology methods to further our mechanistic understanding of RNA translocation by 3Dpol and to solve the structures of viral initiation and uridylylation complexes. We will also use single molecule microscopy and particle tracking methods to directly visualize the in vitro assembly of viral replication complexes onto supported lipid bilayers.

小核糖核酸病毒是一类小型正链单链 RNA 病毒，可引起广泛的每年因疾病造成的损失高达数亿美元。成员包括急性甲型肝炎病毒、导致心脏病的柯萨奇 B3 病毒、鼻病毒，导致一半以上的心脏病发生普通感冒和麻痹性脊髓灰质炎病毒。这些病毒具有相同的生命周期，其中RNA复制和病毒组装发生在囊泡表面组装的大膜锚定复制复合物中来源于内质网。复制过程是由病毒编码的RNA依赖驱动的 RNA 聚合酶 (3Dpol) 负责合成所有病毒 RNA。该研究项目的重点是病毒复制中心的结构和组装以及病毒的结构功能研究聚合酶来了解控制延伸率和复制保真度的机制。我们有解析了 3Dpol 的多个晶体结构作为与结合 RNA 的延伸复合物以及这些结构揭示了 RdRP 响应核苷酸而关闭其催化活性位点的新机制绑定。在即将到来的项目期间，我们将使用生物物理和结构生物学方法来进一步推进我们的研究通过 3Dpol 理解 RNA 易位的机制，并解决病毒起始和传播的结构尿苷酰化复合物。我们还将使用单分子显微镜和粒子追踪方法直接可视化病毒复制复合物在支持的脂质双层上的体外组装。