MAS NMR Studies of the NIV-1 Gag Polyprotein Assembled into Virus-Like Particles

 描述（由申请人提供）：由人类免疫缺陷病毒（HIV）引起的获得性免疫缺陷综合征（AIDS）是一个重大的全球公共卫生问题。虽然存在治疗艾滋病患者的有效疗法，但目前还没有治愈方法。此外，艾滋病毒可以迅速对现有治疗产生耐药性。Gag（组特异性抗原）是HIV病毒体的关键结构蛋白，约占整个HIV病毒体质量的50%。Gag在HIV病毒成熟中的作用包括驱动未成熟病毒颗粒的组装和将病毒RNA基因组包装到出芽病毒体中。迄今为止，组装成病毒样颗粒（VLP）的Gag的结构研究仅限于较低分辨率的电子显微镜和生物化学研究。魔角自旋核磁共振（MAS NMR）是表征Gag的原子水平、位点特异性相互作用的最佳生物物理技术，允许在非低温温度下对Gag进行高分辨率研究，没有尺寸或溶解度限制。利用MAS NMR，本研究项目将表征组装成VLP的Gag的结构和动力学特征，以及蛋白质与RNA的相互作用。所有具体目标的初步结果表明，所提出的目标将得以实现。目的1，共振分配和二级结构的测定组装在VLP中的Gag，是任何生物分子的NMR研究中至关重要的第一步，并将解决有关蛋白质的各个结构域的二级结构的开放性问题。关于目标2的研究将表征组装成VLP的Gag多聚蛋白的位点特异性动力学;定量蛋白中动态过程的时间尺度将有助于深入了解蛋白的特定残基和区域在VLP中的作用。 尊重功能。RNA与组装成VLP的Gag的原子水平相互作用的表征将在目标3中完成，这是多蛋白在病毒成熟期间经历的关键构象变化。该研究项目解决了对HIV病毒成熟和复制的理解中的一个关键空白：Gag在分子水平上的结构和动态特征及其与RNA病毒基因组相互作用的细节。从这项研究中获得的见解可能对开发治疗艾滋病的新疗法具有潜在的重要意义。

 DESCRIPTION (provided by applicant): Acquired Immunodeficiency Syndome (AIDS), caused by the Human Immunodeficiency Virus (HIV), is a significant global public health issue. While effective therapeutics exist to treat AIDS patients, at present there is no cure. Furthermore, the HIV virus can rapidly develop resistance to existing treatments. Gag (group specific antigen) is a key structural protein of the HIV virion, comprising approximately 50% of the entire HIV virion mass. The roles of Gag in HIV viral maturation include driving assembly of the immature viral particle and packaging the viral RNA genome into the budding virion. To date, structural studies of Gag assembled into virus-like particles (VLPs) have been limited to lower resolution electron microscopy and biochemical studies. Magic angle spinning nuclear magnetic resonance (MAS NMR) is the optimal biophysical technique to characterize atomic-level, site-specific interactions of Gag, allowing for high-resolution studies of Gag at non- cryogenic temperatures, with no size or solubility restrictions. With MAS NMR, this research project will characterize structural and dynamic characteristics of Gag assembled into VLPs, as well as the protein's interactions with RNA. Preliminary results in all specific aims demonstrate that the proposed aims will be accomplished. Aim 1, resonance assignments and secondary structure determination of Gag assembled in VLPs, is a crucial first step in any NMR study of biomolecules and will address open questions regarding the secondary of structure of various domains of the protein. Research with respect to Aim 2 will characterize site-specific dynamics of the Gag polyprotein assembled into VLPs; quantifying timescales of dynamic processes in the protein will lend insight into the roles specific residues and regions of the protein play with respect to function. Characterization of atomic-level interactions of RNA with Gag assembled into VLPs, which are key given conformational changes that the polyprotein undergoes during viral maturation, will be accomplished in Aim 3. This research project addresses a critical gap in the understanding of HIV viral maturation and replication: the structural and dynamic features of Gag at the molecular level and the details of its interactions with the RNA viral genome. The insight gained from this research may be potentially important in the development of new therapeutics to treat AIDS.