STRUCTURE BASED STUDIES OF RNA BINDING PROTEINS FROM HIV

（改编自申请人摘要）一个35个碱基的RNA茎环，称为 SL-1位于HIV基因组的5 '端附近，介导几种HIV-1的转录。 对于病毒组装至关重要，因此是一个潜在的靶点， 抗病毒治疗通过结合HIV Gag蛋白，SL 1形成了HIV Gag蛋白的一部分。 将基因组RNA靶向到病毒体中的包装信号。之间的接触 一对HIV RNA中的SL 1元件（通过最初的“吻环”） 然后转化为线性双链体的复合物）也启动基因组 二聚化--一个由Gag促进的过程， 完全传染性。SL 1的所有生物活性取决于其三个- 三维结构，其确切性质是未知的。PI实验室 已经使用NMR光谱来检查SL 1的截短形式， 吻环和线性双链构象。他们获得了优秀的 RNA的核磁共振谱。他们建议解决 这两种RNA的单体和二聚体形式，从而获得第一个高- 逆转录病毒二聚体界面的分辨率图像。时遵循这些 结构，他们将系统地诱变SL 1，以绘制分子图谱， 接触需要稳定每个构象状态。他们也建议 使用实时NMR动力学研究来研究 SL 1二聚化。对杂原子标记的RNA的多维NMR将 用于解决全长SL 1类似物的结构， 能够进行Gag结合以及二聚化。这种RNA在 与未标记的Gag蛋白的复合物将揭示 Gag识别和Gag促进SL 1介导的细胞凋亡的机制 二聚化。这些研究的结果可能为合理的 设计靶向SL 1/Gag相互作用的新抗病毒药物。

(Adapted from the applicant's abstract) A 35-base RNA stem-loop called SL-1, located near the 5 ' end of the HIV genome, mediates several functions crucial for viral assembly, and so is a potential target for antiviral therapy. By binding the HIV Gag protein, SL1 forms part of the packaging signal that targets genomic RNA into virions. Contact between SL1 elements in a pair of HIV RNAs (through an initial "kissing-loop" complex which then converts to a linear duplex) also initiates genomic dimerization--a process that is facilitated by Gag and is essential for full infectivity. All biological activities of SL1 depend upon its three- dimensional structure, whose exact nature is unknown. The PIs laboratory has used NMR spectroscopy to examine a truncated form of SL1 in its kissing-loop and linear duplex conformations. They have obtained excellent NMR spectra on the RNA. They propose to solve the structures of the monomer and both dimer forms of this RNA, and so to obtain the first high- resolution images of a retroviral dimer interface. Guided by these structures, they will systematically mutagenize SL1 to map the molecular contacts needed to stabilize each conformational state. They also propose to use real-time NMR kinetic studies to study the sequence of events in SL1 dimerization. Multidimensional NMR on heteronuclear labeled RNA will be used to solve the structure of a full-length SL1 analogue that is fully competent for Gag binding as well as dimerization. Studies of this RNA in complex with unlabeled Gag protein will reveal the structural basis for Gag recognition and the mechanism by which Gag facilitates SL1-mediated dimerization. Results of these studies may provide a basis for rational design of new antivirals that target the SL1/Gag interaction.