Assembly of Picornaviral Replication Complexes

小核糖核酸病毒复制复合物的组装

• 批准号: 6866381
• 负责人: Olve Breien Peersen
• 金额: $ 25.13万
• 依托单位: COLORADO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-04-01 至 2009-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6866381
• 关键词: DNA directed RNA polymerase; Picornaviridae; RNA biosynthesis; SDS polyacrylamide gel electrophoresis; circular dichroism; conformation; protein folding; protein structure function; spectrometry; virus assembly; virus replication

DESCRIPTION (provided by applicant): 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 common life cycle where their 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, the 3Dpol protein, that is responsible the synthesis of all viral RNA. Like all picornaviral proteins, the polymerase is generated by proteolytic cleavage of a single large viral polyprotein. There is mounting evidence in several picornaviruses that the polymerase and its immediate precursors are directly responsible for the assembly of these replication centers. The 3Dpol polymerase of poliovirus, the best studied of the picornaviruses, has been shown to assemble into large sheet structures along a protein-protein interface that was initially identified in a partial crystal structure of 3Dpol. We have solved the complete crystal structure of 3Dpol at 2.0 A resolution and discovered that the enzyme requires a free N-terminus to properly fold the active site, providing a molecular basis for the processing dependent activation of the polymerase. We are continuing our structural studies of the poliovirus proteins by further characterizing the conformational flexibility of 3Dpol and expanding into determining the role of protein-protein interfaces in the assembly of 3CDpro and other precursor proteins. The results will yield fundamental insights into the replication of this important group of pathogens.

描述（由申请人提供）：Picornaviruses是一个小型积极的单一链RNA病毒的家族，每年以每年成本良好的成本造成多种疾病。成员包括急性肝炎病毒，造成心脏病的Coxsackie B3病毒，引起普通感冒的一半以上的鼻病毒以及瘫痪的脊髓灰质炎病毒。这些病毒具有共同的生命周期，其中RNA复制和病毒组装发生在大膜锚定复制复合物中，这些复制复制复合物组装在源自内质网的囊泡表面上。复制过程是由病毒编码的RNA依赖性RNA聚合酶，3DPOL蛋白的驱动，该蛋白是所有病毒RNA的合成。像所有PICORNAVIRAL蛋白一样，聚合酶是通过单个大型病毒多蛋白的蛋白水解裂解产生的。在几个Picornavires中有越来越多的证据表明，聚合酶及其直接前体是直接负责这些复制中心的组装。脊髓灰质炎病毒最佳研究的脊髓灰质炎病毒的3DPOL聚合酶已被证明沿蛋白质 - 蛋白质界面组装成大片结构，该结构最初在3DPol的部分晶体结构中鉴定出来。我们已经在2.0 A分辨率下解决了3DPOL的完整晶体结构，并发现该酶需要自由的N端才能正确折叠活性位点，从而为聚合酶的加工依赖性激活提供了分子基础。我们正在通过进一步表征3DPol的构象柔韧性并扩展到确定蛋白质蛋白接口在3CDPRO和其他前体蛋白组装中的作用，从而继续对脊髓灰质炎病毒蛋白进行结构研究。结果将产生对这一重要病原体复制的基本见解。