Structural Characterization of SARS-CoV-2 Genome Organization by NMR

该项目的目标是从结构上表征核衣壳(N)介导的SARS-CoV-2病毒基因组包装的特征，SARS-CoV-2病毒是致命的新冠肺炎大流行的病原体。拟议的调查和将要开发的一般方法框架将具有广泛的应用，因为它们将提供1)生物学见解，开启干扰病毒基因组包装的新策略，以及2)可供其他从业者(生物化学家、生物物理学家、分子生物学家)使用的基于磁共振的综合方案。因此，拟议的研究将影响几个科学领域，包括结构生物学、生物化学、物理化学、磁共振光谱学、生物技术和药理学。拟议的创新研究计划将为各级学生提供最先进的实验方法方面的独特培训。重要的是，该项目中的教育计划将包括为PI、共同PI和工业合作者的实验室成员提供机会，这将极大地加强STEM劳动力的培训。该项目的目标是通过结合实验魔角旋转(MAS)和动态核极化(DNP)核磁共振(NMR)方法，从结构上表征核衣壳(N)是如何包装SARS-CoV-2病毒基因组的。通过拟议的研究建立的这一新的方法学框架将克服目前在研究复杂的异质病毒基因组核糖核颗粒方面的挑战，这些核糖核颗粒表现出不同程度的无序和动力学。这项拟议的研究将提供关于SARS-CoV-2N蛋白结构和动力学的原子水平知识，以及N蛋白与基因组RNA组装的结构组织。使用高磁场以及快速和超快的1H和19F检测的MAS和DNP增强的MAS核磁共振将使研究非常大的病毒系统成为可能，只需要传统实验中使用的一小部分材料。将开发的一般方法框架将广泛适用于研究其他病毒的打包基因组，并可供这些方法的广泛实践者(生物化学家、生物物理学家、分子生物学家)使用。因此，计划中的研究将影响多个科学领域，包括结构生物学、生物化学、物理化学、磁共振波谱、生物技术和药理学。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

The goal of this project is to structurally characterize the nucleocapsid (N)-mediated packaging of the genome of the SARS-CoV-2 virus, the causative agent of the deadly COVID-19 pandemic. The proposed investigations and the general methodological framework to be developed will have broad applications since they will provide 1) biological insights that open new strategies for interfering with viral genome packaging, and 2) integrative magnetic resonance-based protocols that can be used by other practitioners (biochemists, biophysicists, molecular biologists). As such, the proposed research will impact several areas of science, including structural biology, biochemistry, physical chemistry, magnetic resonance spectroscopy, biotechnology and pharmacology. The proposed innovative research program will provide unique training for students at all levels in state-of-the-art experimental methods. Importantly, an educational program within this project will include embedding opportunities for members of the laboratories of the PI, Co-PI and the industrial collaborator that will greatly enhance the training of the STEM workforce. The goal of this project is to structurally characterize how the nucleocapsid (N) packages the genome in the SARS-CoV-2 virus, by integrating experimental magic angle spinning (MAS) and dynamic nuclear polarization (DNP) nuclear magnetic resonance (NMR) methods. This new methodological framework, established through the proposed research, will overcome the current challenges in studying complex heterogeneous viral genome ribonuclear particles that exhibit different degrees of disorder and dynamics. The proposed research will provide atomic-level knowledge on the structure and dynamics of the SARS-CoV-2 N protein as well as the structural organization of the N protein in assemblies with genomic RNA. Employing high magnetic fields together with fast and ultrafast 1H- and 19F- detected MAS and DNP-enhanced MAS NMR will enable studies into very large virus systems, requiring only a fraction of the material used in conventional experiments. The general methodological framework that will be developed will be broadly applicable to investigate packaged genomes of other viruses and can be used by a wide range of practitioners of these methodologies (biochemists, biophysicists, molecular biologists). As such, the planned research will impact several areas of science, including structural biology, biochemistry, physical chemistry, magnetic resonance spectroscopy, biotechnology and pharmacology.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.