Genome packaging in DNA viruses

描述（由申请人提供）：基因组包装是大双链DNA （dsDNA）病毒（包括尾双链DNA噬菌体和疱疹病毒）形态发生的关键步骤，对感染性子代病毒粒子的组装至关重要。基因组包装是一种精确协调的分子协同作用，其中来自DNA串联体的一定量的病毒DNA插入到预先形成的原衣壳中，然后将额外的病毒蛋白结合到衣壳上以保留包装的DNA。这个DNA插入过程是由一个由终止酶和入口组成的强大的分子机器完成的。在衣壳的单个顶点形成一个通道，允许病毒DNA在病毒组装期间进入并在感染期间退出。末端酶复合体包含一个特异性结合病毒DNA的DNA识别亚基，以及一个为包装反应提供能量并从串联体中切割基因组长度DNA的催化亚基。由于异常复杂和缺乏高分辨率结构数据，这些大型病毒基因组包装的分子机制尚不清楚。特别是，这些基因组包装蛋白必须形成高阶分子组装才能发挥作用。门静脉蛋白在衣壳内形成环状十二聚体。然而，对于终止酶如何组装，它如何与入口组装，以及入口如何嵌入衣壳，我们知之甚少。我们的目标是通过使用结构方法分析基因组包装蛋白的组装来了解DNA病毒基因组包装的分子机制。我们特别感兴趣的一般问题是，如何大量的蛋白质和其他生物分子，每一个存在于多个副本，按层次组装成一个复杂的系统，以完成一个复杂的生物过程。在本提案中，我们力求：(i)阐明一个完整的、具有传染性的细菌病毒的高分辨率结构；（ii）阐明末端酶组成蛋白的结构；（iii）阐明终止酶dna识别蛋白的dna结合模式；（iv）探索末端酶复合物的体外组装。疱疹病毒是重要的人类病原体，可引起从水痘到各种癌症的各种疾病。这一建议有望通过确定新的抗病毒靶点来控制和预防疱疹病毒引起的感染和疾病，从而促进公共卫生。

DESCRIPTION (provided by applicant): Genome packaging is a key step in morphogenesis of large double-stranded DNA (dsDNA) viruses including tailed double-stranded DNA bacteriophages and herpesviruses, and is essential for assembly of infectious progeny virions. Genome packaging is a precisely coordinated molecular synergy, in which a certain amount of viral DNA from a DNA concatemer is inserted into a preformed procapsid, followed by binding of additional viral proteins to the capsid to retain the packaged DNA. This DNA insertion process is fulfilled by a powerful molecular machine consisting of the terminase and the portal. The portal forms a conduit at a single vertex of the capsid that allows viral DNA to enter during virus assembly and exit during infection. The terminase complex contains a DNA-recognition subunit that specifically binds to the viral DNA, and a catalytic subunit that provides the energy for the packaging reaction and cleaves the genome-length DNA from the concatemer. Molecular mechanisms of genome packaging in these large viruses are not well understood, owing to unusual complexity and lack of high resolution structural data. In particular, these genome-packaging proteins have to form high-order molecular assemblies in order to function. The portal protein forms a ring-like dodecamer embedded in the capsid. However, little is known about how the terminase assembles, how it assembles with the portal, and how the portal is embedded in the capsid. Our goal is to understand molecular mechanisms of genome packaging in DNA viruses by analyzing assemblies of genome-packaging proteins using structural approaches. We are particularly interested in the general question of how numerous proteins and other biological molecules, each present in multiple copies, assemble hierarchically into a sophisticated system to fulfill a complex biological process. In the present proposal, we seek to: (i) elucidate the high resolution structure of a complete, infectious bacterial virus; (ii) elucidate structures of component proteins of the terminase; (iii) elucidate the mode of DNA-binding for a terminase DNA-recognition protein; and (iv) explore the assembly of the terminase complex in vitro. Herpes viruses are important human pathogens that cause diseases ranging from chickenpox to various forms of cancer. This proposal is anticipated to contribute to public health by identification of novel targets of antiviral to control and prevent infection and diseases caused by herpes viruses. PUBLIC HEALTH RELEVANCE: Human herpes viruses are linked to important human diseases ranging from chickenpox to life-threatening cancer. The majority of individuals in most human populations are infected by several herpes viruses. We seek to investigate a common molecular mechanism in life cycles of herpes virus and many other large double- stranded DNA viruses, in search of new measures to control and prevent infection and diseases caused by these viral pathogens.