Coronavirus RNA synthesis by multicomponent protein machines

总结 SARS-CoV-2是COVID-19的病原体，已成为一种全球性的人类病原体，席卷我们的 社区.制定预防或治疗COVID-19的策略对于减轻疾病和限制 进一步的病毒传播。抗病毒治疗的关键靶标是病毒RNA复制复合物。SARS-CoV-2 复制复合物是具有多种辅因子、酶和宿主调节蛋白的大型多亚基机器。 这些蛋白质亚基如何组装和合作进行病毒RNA复制和转录仍然不清楚。 这项工作的首要主题是研究冠状病毒复制复合体的未充分研究的方面， 重点是表征现有抗病毒治疗的效果以及发现新的靶点， 化合物.我们感兴趣的是病毒nsp 14核酸外切酶如何有助于病毒RNA校对，减少 核苷酸错配和提供对核苷类似物药物的天然抗性。我们亦会探讨 nsp 12核苷酸转移酶（NiRAN）的功能，并将病毒蛋白网络拼凑在一起 这些相互作用负责RNA合成复合物的组装。为此，我们将使用多种方法，包括 生物化学、生物物理学、细胞生物学、化学生物学和低温电子显微镜。这些研究将提供新的 深入了解这个复杂机器的工作原理，为现有的治疗方法提供新的作用机制， 发现新的抗病毒化合物。此外，复制机械的部件保存率高 在冠状病毒家族中，这些SARS-CoV-2的研究可以应用于未来新出现的冠状病毒， 在未来的病毒大流行成为全球危机之前阻止它们。

SUMMARY SARS-CoV-2, the causative agent of COVID-19, has emerged as a global human pathogen sweeping through our communities. The development of strategies to prevent or treat COVID-19 is essential for mitigating disease and limiting further viral spread. A key target of antiviral therapeutics is the virus RNA replication complex. The SARS-CoV-2 replication complex is a large multi-subunit machine with multiple co-factors, enzymes and host modulating proteins. How these protein subunits assemble and cooperate to carryout viral RNA replication and transcription remains unclear. The overarching theme of this work is to examine understudied aspects of the coronavirus replication complex with an emphasis on characterizing the effects of existing antiviral therapeutics as well as the discovery of novel targets and compounds. We are interested in how the virus nsp14 exonuclease contributes to viral RNA proofreading, reducing nucleotide misincorporations and providing natural resistance to nucleoside analogue drugs. We will also explore the function of the enigmatic nsp12 nucleotidyltransferase (NiRAN) and will piece together the network of viral protein interactions responsible for the assembly of the RNA synthesis complex. To this we will use diverse methods including biochemistry, biophysics, cell biology, chemical biology and cryo-electron microscopy. These studies will provide new insight into the workings of this complicated machine, provide new mechanisms of action for existing therapeutics and discover novel antiviral compounds. In addition, the high conservation of components of the replication machinery across the coronavirus family allows these studies of SARS-CoV-2 to be applicable to future emerging coronaviruses to head off future viral pandemics before they become global crises.