Solid-State NMR of Viral Fusion Peptides and Proteins

病毒融合肽和蛋白质的固态核磁共振

• 批准号: 7048792
• 负责人: David P Weliky
• 金额: $ 25.17万
• 依托单位: MICHIGAN STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-02-01 至 2011-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7048792
• 关键词: HIV envelope protein gp41; Orthomyxoviridae; acidity /alkalinity; cell fusion; conformation; crosslink; host organism interaction; human immunodeficiency virus; intermolecular interaction; membrane activity; membrane fusion; membrane model; microorganism hemagglutinin; nuclear magnetic resonance spectroscopy; peptide structure; protein protein interaction; radiotracer; structural biology; virus infection mechanism; virus protein

DESCRIPTION (provided by applicant): The long-term goal of the proposed research is to understand with atomic-resolution detail the basis for viral fusion protein-induced membrane fusion. Fusion of viral and target cell membranes is a key step in infection for many viruses important in disease and a detailed understanding of fusion mechanisms should aid development of anti-viral therapeutics whose mode of action is fusion inhibition. The proposed research focuses on the gp41 and the hemagglutinin HA2 fusion proteins of the human immunodeficiency virus (HIV) and the influenza virus (IFV), respectively, because of the significance of the diseases caused by these viruses and because these proteins serve as prototypes for other class I viral fusion proteins. There is particular emphasis on the "fusion peptide" (FP) domains of the fusion proteins, which represent approximately 20-residue apolar regions at the N-termini of the proteins. Numerous biochemical and biophysical studies have shown that the FP plays a key role in fusion and that chemically synthesized peptides with the FP sequence can serve as useful model systems to understand some aspects of viral/target cell fusion. The long-term objectives will be achieved with four specific aims. Two of the aims focus on solid-state nuclear magnetic resonance (SSNMR) structural analysis of chemically cross-linked dimeric and trimeric HIV FPs in membranes and additional liquid-state NMR studies of these peptides in detergent micelles. The oligomeric topology of these C-terminal cross-linked peptides reflects the expected topology of FPs in the HIV gp41 fusion protein. The third aim focuses on: (a) determination of the insertion orientation of FPs in membranes using samples in which the membranes are oriented between stacked glass plates; and (b) probing FP insertion depths using SSNMR distance measurements between 13C nuclei in the FP and 31P and 19F nuclei in the lipid. The fourth specific aim focuses on SSNMR structural and insertion depth measurements for large HIV and IFV fusion protein domains which contain FPs.

描述(由申请人提供)：拟议研究的长期目标是以原子分辨率详细了解病毒融合蛋白诱导的膜融合的基础。病毒和靶细胞膜的融合是许多疾病中重要病毒感染的关键步骤，对融合机制的详细了解将有助于以融合抑制为作用模式的抗病毒治疗的发展。建议的研究重点分别是人类免疫缺陷病毒(HIV)和流感病毒(IFV)的gp41和血凝素HA2融合蛋白，因为这些病毒引起的疾病的重要性，以及这些蛋白作为其他I类病毒融合蛋白的原型。特别强调融合蛋白的“融合肽”(FP)结构域，它代表了蛋白质N-末端的大约20个残基的非极区。大量的生化和生物物理研究表明，FP在融合中起着关键作用，化学合成的带有FP序列的多肽可以作为有用的模型系统来理解病毒/靶细胞融合的某些方面。长期目标将通过四个具体目标来实现。其中两个目标集中于膜上化学交联的二聚体和三聚体HIV FP的固态核磁共振(SS核磁共振)结构分析，以及洗涤剂胶束中这些多肽的额外液态核磁共振研究。这些C端交联肽的寡聚拓扑结构反映了HIV gp41融合蛋白中FPS的预期拓扑结构。第三个目标集中在：(A)使用膜取向在堆叠玻璃板之间的样品来确定FP在膜中的插入方向；以及(B)通过SSNMR测量FP中的13C核与脂质中的31P和19F核之间的距离来探测FP的插入深度。第四个具体目标集中在包含FP的大的HIV和IFV融合蛋白结构域的SSNMR结构和插入深度测量上。