Structural dynamics of single Ebolavirus GP molecules.

单个埃博拉病毒 GP 分子的结构动力学。

• 批准号: 8954151
• 负责人: James B Munro
• 金额: $ 179.06万
• 依托单位: TUFTS UNIVERSITY BOSTON
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-30 至 2019-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8954151
• 关键词: Antigens; Antiviral Therapy; Biological Assay; Cell membrane; Cells; Development; Ebola virus; Engineering; Epidemic; Fc Receptor; Glycoproteins; Image; Individual; Kinetics; Life Cycle Stages; Light; Membrane; Molecular Conformation; Process; Role; Surface; Therapeutic Agents; Thermodynamics; Vaccine Design; Vaccines; Variant; Viral; Viral Proteins; Virion; Virus; base; design; insight; novel; public health relevance; receptor binding; single-molecule FRET; tool

 DESCRIPTION (provided by applicant): The persistence of the current epidemic is in part a result of the lack of any vaccine, and any approved post- exposure therapies. Progress in the development of such preventative and therapeutic agents has been slow due to a limited understanding of the Ebola viral life cycle. Like all enveloped viruses, entry of Ebolaviruses into target cells requires fusion of the viral membrane to a cell membrane. This process is catalyzed by the envelope glycoprotein (GP). GP is the only viral protein exposed on the surface of Ebola virions, making it an attractive target for vaccines and antiviral therapies. The significant energetic barrier to fusing viral and cell membranes is surmounted by transition of GP to successively lower energy conformations. But virtually no information presently exists on the structural dynamics of GP, the impact of receptor binding on GP conformation, or on the mechanism by which GP catalyzes the fusion of viral and cell membranes. Given the central role of envelope glycoprotein conformational changes in the mechanism of enveloped viral entry, an understanding of Ebolavirus entry requires a deep insight into the conformational landscape of GP. Here, we propose the application of an integrated framework involving smFRET imaging, and kinetic and thermodynamic analysis. We will apply this framework to probe the conformational dynamics of individual trimeric GP molecules on the surface of intact pseudovirions, interrogate their interaction with cellular receptors and antibodies, and elucidate the impact of changes in pH. This approach will bring new light to our understanding of Ebolavirus entry specifically, and enveloped virus entry in general. In addition, we will design GP variants that stably expose antigenic surfaces, and use our smFRET-based framework as a robust and high-throughput means of assaying the conformational landscape of these potential immunogens. We will thereby develop smFRET as a tool for structural vaccine design.

 描述（由适用提供）：当前流行病的持续性是由于缺乏任何疫苗和任何批准的暴露后疗法的结果。由于对埃博拉病毒生命周期的了解有限，这种预防剂和治疗剂的开发进展一直很慢。像所有包围病毒一样，将埃博拉病毒进入 靶细胞需要将病毒膜融合到细胞膜。该过程由包膜糖蛋白（GP）催化。 GP是暴露在埃博拉病毒表面上的唯一病毒蛋白，使其成为疫苗和抗病毒疗法的吸引力。通过GP过渡到成功降低能量构象的过渡，可以掩盖融合病毒和细胞膜的显着能屏障。但是几乎没有任何信息存在有关GP的结构动力学，受体结合对GP构象的影响或GP催化病毒和细胞机制融合的机制的影响。鉴于包膜糖蛋白构象变化在被包膜病毒进入机理中的核心作用，对埃博拉病毒进入的理解需要深入了解GP的构象景观。在这里，我们提出了涉及SMFRET成像以及动力学和热力学分析的集成框架的应用。我们将应用此框架来探测单个三聚体GP分子在完整伪造表面上的构象动力学，询问它们与细胞接收器和抗体的相互作用，并阐明pH变化的影响。这种方法将为我们对埃博拉病毒进入的理解带来新的启示，并总体上将病毒进入。此外，我们还将设计稳定暴露抗原表面的GP变体，并使用我们的基于SMFRET的框架作为一种强大而高通量的手段来主张这些潜在免疫原子的构象景观。因此，我们将开发SMFRET作为结构性疫苗设计的工具。