STRUCTURE-BASED STUDIES OF RNA BINDING PROTEINS OF HIV

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 我们正在研究HIV基因组中的RNA-蛋白质相互作用。被称为SL1的RNA茎环位于HIV-1基因组5‘端附近，介导着几种对病毒组装至关重要的功能，因此是抗病毒治疗的潜在新靶点。通过结合HIV Gag蛋白的核衣壳(NCp7)部分，SL1形成了包装信号的一部分，将基因组RNA靶向病毒粒子。一对HIV RNA中SL1环之间的接触也会启动基因组二聚化--这一过程由NCp7结合促进，是完全感染性所必需的。SL1的所有生物学活性都依赖于它的三维结构。利用异核标记和多维核磁共振，我们建议解决全长，可信的SL1的相应结构，它包含其他生物学上重要的特征。SL1二聚体是一种70ntRNA，分子量接近22kD。通过确定这些结构，我们试图了解SL1二聚的显著效率以及稳定每个连续构象的化学特征。我们还将确定HIV NCp7蛋白与每个RNA构象结合时形成的复合体的结构和动力学，揭示相关的确切分子接触以及NCp7促进SL1和其他核酸折叠的机制。此外，我们将解决NCp7与异源、单体RNA配体(适体)形成的复合体，以确定其紧密结合的特征，并可在合理的药物设计中加以利用。这些研究的结果将为发现针对SL1/NCp7相互作用的新的抗病毒药物提供基础。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. We are working on RNA-protein Interaction in HIV genome. RNA stemloop called SL1, located near the 5'-end of the HIV-1 genome, mediates several functions crucial for viral assembly, and so is a potential new target for antiviral therapy. By binding the nucleocapsid (NCp7) portion of the HIV Gag protein, SL1 forms part of the packaging signal that targets genomic RNA into virions. Contact between SL1 loop in a pair of HIV RNAs also initiates genomic dimerization- a process that is facilitated by NCp7 binding and is essential for full infectivity. All biological activities of SL1 depend on its three-dimensional structure. Using heteronuclear labeling and multidimensional NMR, we propose to solve the corresponding structures of full-length, authentic SL1, which contains additional features that are biologically important. SL1 dimer is a 70nt RNA with molecular weight nearging 22kD. By determining these structures, we seek to understand the remarkable efficiency with which SL1 dimerizes and the chemical features that stabilize each successive conformation. We will also determine the structures and dynamics of complexes formed when HIV NCp7 protein binds each RNA conformer, revealing the exact molecular contacts involved and the mechanism by which NCp7 facilitates refolding of SL1 and other nucleic acids. In addition, we will solve the complex formed by NCp7 with a heterologous, monomeric RNA ligand (aptamer) that binds with higher affinity than SL1, to identify features that account for its tight binding and could be exploited in rational drug design. Results of these studies will provide a basis for discovering new antivirals that target the SL1/NCp7 interaction.