HARC Center: HIV Accessory and Regulatory Complexes

描述（由申请人提供）：在整个HIV生命周期中，病毒-宿主复合物在病毒的生物学中起着不可或缺的作用。组装的范围从二元蛋白质-蛋白质和蛋白质-核酸复合物到更大的多组分复合物。这些复合物的高分辨率结构为艾滋病治疗的干预策略提供了模板的潜力，但迄今为止，结构仅在少数情况下得到解决。获得这种结构信息一直受到限制，部分原因是一些HIV蛋白在生化方面表现不佳，并且通常是孤立的非结构化的。这些特征可能反映了蛋白质在整个病毒生命周期中与不同的细胞伴侣短暂结合时采用不同构象的固有特性。该项目被称为HARC中心（HIV Accessory and Regulatory Complexes），重点研究发挥重要调控和辅助功能的五种关键HIV蛋白——整合酶（IN）、Tat、Rev、Vif和Nef——及其相互作用的病毒和细胞蛋白伙伴。我们的主要目标是获得在病毒生命周期早期阶段形成的hiv -宿主细胞相互作用的全面结构图，特别是那些调节基因表达，细胞信号传导或细胞生物学的相互作用。这些病毒和宿主蛋白中的一些在与其核酸或蛋白质伴侣复合物时发生显著的构象变化，了解这种动态行为的分子细节是一个重要的结构目标。我们对调控复合体的关注将为HIV的生物学和发病机制提供新的见解，并确定新的治疗靶点来对抗HIV感染。HARC中心将利用蛋白质表达和共表达策略来表征功能复合物，使用全面的结构方法，包括质谱，x射线晶体学，核磁共振和冷冻电镜。为了克服迄今为止限制进步的障碍，我们建议大力发展技术。关键的新方法将探索优化表达结晶，分析蛋白质动力学使用晶体学和核磁共振数据，并确定小型和非均相配合物的结构通过冷冻电镜。强有力的外联和合作组成部分将吸引中心以外的研究人员，特别是那些在艾滋病毒机制、蛋白质生物化学和病毒学方面具有专业知识的研究人员。

DESCRIPTION (provided by applicant): Throughout the HIV life cycle, viral-host complexes play an integral role in the biology of the virus. Assemblies range from binary protein-protein and protein-nucleic acid complexes to much larger multicomponent complexes. High-resolution structures of these complexes offer the potential to provide templates for intervention strategies in the treatment of AIDS, but structures have so far been solved in only a few cases. Obtaining such structural information has been limited in part because several of the HIV proteins are poorly behaved biochemically and often are unstructured in isolation. These characteristics may reflect an inherent property of the proteins to adopt different conformations when bound transiently to different cellular partners throughout the viral life cycle. This project, called the HARC Center (HIV Accessory and Regulatory Complexes), focuses on five key HIV proteins that perform important regulatory and accessory functions - Integrase (IN), Tat, Rev, Vif, and Nef- and their interacting viral and cellular protein partners. Our primary aim is to achieve a comprehensive structural picture of HIV-host cell interactions formed during early phases of the viral life cycle, particularly those that regulate gene expression, cell signaling, or cell biology. Several of these viral and host proteins undergo significant conformational changes when complexed to their nucleic acid or protein partners, and understanding the molecular details of this dynamic behavior is an important structural goal. Our focus on regulatory complexes will provide new insights into the biology and pathogenesis of HIV and define new therapeutic targets to combat HIV infection. The HARC Center will harness protein expression and co-expression strategies to characterize functional complexes using a comprehensive battery of structural methods, including mass spectrometry, x- ray crystallography, NMR, and cryo-EM. To overcome the obstacles that have limited progress to date, we propose substantial efforts for technological development. Essential new methods will be explored to optimize expression-to-crystallization, analyze protein dynamics using crystallographic and NMR data, and determine structures of small and heterogeneous complexes by cryo-EM. Strong outreach and collaborative components will draw investigators from outside the Center, particularly those with expertise in HIV mechanisms, protein biochemistry, and virology. OVERALL