NMR studies of viral proteins

在病毒出芽和病毒成熟过程中，蛋白质-蛋白质和蛋白质-脂质相互作用被提出进行结构和生化研究。重点是正链RNA病毒，它们要么引起人类疾病，要么与引起人类疾病的病毒密切相关。利用核磁共振等方法研究甲病毒、丙型肝炎病毒、风疹病毒、黄热病病毒等包膜最简单的病毒的出芽过程和核衣壳稳定性。罗斯肉瘤病毒（RSV）是一种结构更复杂的逆转录病毒，其结构特征对衣壳稳定和成熟至关重要。具体目标是通过确定各种RSV蛋白结构的三维溶液结构来了解RSV Gag蛋白的衣壳-核衣壳区域的重要性，并表征衣壳（CA）蛋白、核衣壳（NC）蛋白、衣壳-核衣壳核心颗粒和跨膜E2-糖蛋白的c端结构域的相互作用，这些已经提出并将通过生化和核磁共振实验进行测试。此外，还提出利用核磁共振方法结合诱变结果进一步研究黄热病病毒（YFV）、丙型肝炎病毒（HCV）和风疹病毒（RUBV）的核衣壳稳定性。本项目中提出的研究结果，加上项目中其他地方提出的分子遗传学和晶体学结果，将增强我们对病毒组装的结构和物理基础的理解。

Structural and biochemical studies are proposed to examine protein- protein and protein-lipid interactions involved in viral budding and viral maturation. The focus is on the plus-strand RNA viruses, which either cause human disease or are closely related to viruses which cause human disease. The budding process and nucleocapsid stabilization will be investigated by NMR and other methods for alphavirus, hepatitis C virus, rubella virus, and yellow fever virus, the most simple enveloped viruses. Structural features important for capsid stabilization and maturation will be investigated by NMR for Rous sarcoma virus (RSV), a structurally more complex retrovirus. Specific objectives are to understand the importance of the capsid-nucleocapsid region of the RSV Gag protein by determining 3-dimensional solution structures of various RSV protein constructs, and characterizing interdomain interactions for the C-terminal domain of capsid (CA) protein, the nucleocapsid (NC) protein, and the capsid-nucleocapsid core particles and the transmembrane E2- glycoprotein has been proposed and will be tested by biochemical and NMR experiments. Further studies to investigate nucleocapsid stability are also proposed for yellow fever virus (YFV), hepatitis C virus (HCV), and rubella virus (RUBV) using NMR methods combined with mutagenesis results. Results from the studies proposed in this project, coupled with results from molecular genetics and crystallography proposed elsewhere in the Program Project, will enhance our understanding of the structural and physical basis underlying viral assembly.