Structural studies of HIV Capsid with host factors and Capsid-targeting antivirals

 描述（由申请方提供）：HIV-1衣壳蛋白（CA）在病毒生命周期的多个步骤中起关键作用。它是Gag多聚蛋白的一部分，在成熟过程中裂解产生基质、CA和核衣壳。到目前为止，已知CA结构及其对核心稳定性的影响对于脱壳、逆转录、核进入、整合位点和组装的过程是至关重要的，并且对于掩盖DNA产物免受免疫监视也是重要的。HIV-1 CA与宿主因子相互作用，包括TRIM 5a、裂解和多聚腺苷酸化特异性因子6（CPSF 6）、核孔蛋白153和358（NUP 153、NUP 358）、MxB和亲环素A。所有这些不同的功能和相互作用都是由一个 一个非常灵活的蛋白质，CA，因此可以被称为生物瑞士军刀。 尽管在过去的20年中已经解决了HIV-1 CA片段或重工程变体的几种结构，但这些结构都没有提供控制衣壳稳定性的关键六聚体间接触的完整原子细节。Sarafianos实验室最近解决了HIV-1 CA的晶体结构，包括完整的天然全长六聚体HIV-1 CA的难以捉摸的结构。这些结构揭示了以前未知的分子细节，特别是在影响核心稳定性的3倍和2倍内六聚体界面处。他们证明，六聚体间的界面是有延展性的，可以通过新发现的适应性水化层来改变，这被假设为对岩心稳定至关重要。他们揭示了一个显着的可塑性，解释了在病毒粒子中观察到的多态性。他们还验证了我们的实验系统作为最相关的CA相互作用的研究在一个本地的上下文中的文物。最后，在额外的初步研究中，该系统已被用于研究a） 结合CA-结合抗病毒剂，B）结合宿主因子肽，和c）迄今为止增加或降低核心稳定性并影响病毒学功能的三种CA突变体。与所有以前的CA结构不同，我们的结构揭示了六聚体间界面结构变化的新信息，并建立了一个全面的框架，这将有助于通过以下目标显着推进该领域：AIM 1：表征增加或减少核心稳定性的结构决定因素AIM 2：CA与宿主因子的结构相互作用AIM 3：这些研究将揭示核心稳定性的结构基础，提供关于CA结构如何控制与宿主因子和小分子效应物的相互作用的独特信息，并且可以揭示各种生物功能的脆弱的结构中间体。

 DESCRIPTION (provided by applicant): The HIV-1 capsid protein (CA) plays a critical role in multiple steps of the virus life cycle. It is part of the Gag polyprotein, which is cleaved during maturation to give matrix, CA, and nucleocapsid. So far it is known that CA structure and its effects on core stability are critical for the processes of uncoating, reverse transcription, nucler entry, sites of integration, and assembly, and it is also important for cloaking the DNA product from immune surveillance. HIV-1 CA interacts with host factors including TRIM5a, cleavage and polyadenylation specific factor 6 (CPSF6), nucleoporins 153 and 358 (NUP153, NUP358), MxB and Cyclophilin A. All these diverse functions and interactions are regulated by the structure of a single, remarkably flexible protein, CA, which can thus be termed a biological Swiss Army knife. Although several structures of HIV-1 CA fragments or heavily engineered variants have been solved over the past 20 years, none of these has provided the full atomic details of the critical inter-hexameric contacts that govern capsid stability. The Sarafianos lab recently solved crystal structures of HIV-1 CA, including the elusive structure of the intact native full length hexameric HIV-1 CA. The structures reveal previously unknown molecular details, especially at the 3-fold and 2-fold intra-hexamer interfaces that affect core stability. They demonstrate that the inter-hexamer interfaces are malleable and can change through a newly discovered adaptable hydration layer, which is hypothesized to be critical for core stabilization. They reveal a remarkable plasticity which explains the polymorphism observed in virions. They also validate our experimental system as the most relevant for the study of CA interactions in a native context free of artifacts. Finally, in additional preliminary studies this system has been used to study a) binding of a CA-binding antiviral, b) binding of a host factor peptide, and c) so far three CA mutants that increase or decrease core stability and affect virological functions. Unlike all previous CA structures our structures reveal novel information of the structural changes at the inter-hexamer interfaces and establish a comprehensive framework that will help significantly advance the field through the following Aims: AIM 1: Characterize the structural determinants of increased or decreased core stability AIM 2: Structural interactions of CA with host factors AIM 3: Understanding the mechanisms of CA biological functions using CA-targeting antiviral probes These studies will unravel the structural basis of core stability, provide unique information on how CA structure controls interactions with host factors and small molecule effectors, and may reveal vulnerable structural intermediates of various biological functions.