Structural analysis of HCV E1E2 glycoproteins

HCV E1E2 糖蛋白的结构分析

• 批准号: 10409764
• 负责人: Richard J. Kuhn
• 金额: $ 30.81万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-06-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10409764
• 关键词: Animals; Antibodies; Antigens; Architecture; B-Lymphocytes; Binding; Bioinformatics; Biology; Biophysics; CD81 gene; Cavia; Complex; Computer Models; Cryo-electron tomography; Cryoelectron Microscopy; Crystallography; Data; Electron Microscopy Facility; Engineering; Epitopes; Fc Receptor; Flavivirus; Genotype; Glycoproteins; Goals; Hepatitis C; Hepatitis C Vaccine; Hepatitis C virus; Human; Immunity; Immunology procedure; Laboratories; Lead; Length; Light; Mediating; Membrane Glycoproteins; Methods; Modification; Molecular Conformation; Molecular Virology; Mutation; Nature; Outcome; Polysaccharides; Preparation; Production; Protein Engineering; Proteins; Protocols documentation; Recombinant Proteins; Recombinants; Research; Resolution; Role; Structure; Surface; Synchrotrons; T-Lymphocyte; Testing; Time; Transmembrane Domain; Vaccine Design; Vaccines; Viral; Viral Pathogenesis; Virion; Virus; Virus Receptors; Work; X-Ray Crystallography; base; design; experience; glycoprotein structure; glycosylation; human pathogen; in vivo; insight; lead candidate; molecular modeling; nanodisk; neutralizing antibody; nonhuman primate; novel; particle; plasmid DNA; programs; receptor; receptor binding; response; structural biology; tool

ABSTRACT – PROJECT 4 Despite extensive research efforts to understand the biology of hepatitis C virus (HCV), it remains a major human pathogen without a vaccine. In addition, there is a remarkable absence of information regarding the structure of the virion and organization of its component proteins. We intend to use our extensive structural biology experience with the related flaviviruses, to focus on determining the structure of the viral glycoproteins, the virus particle, and complexes of potent neutralizing antibodies bound to the surface glycoproteins. This project will rely heavily on and interact extensively with Project 1 to interrogate the structure and provide guidance into the features of the glycoproteins that are important for eliciting broad and potent neutralizing antibodies. Iterative analysis and re-engineering of the E1E2 glycoproteins will be conducted to guide the antibody component of a dual B and T cell HCV vaccine, the focus of this U19 proposal. This specific project will use three aims to determine the structure of the E1E2 heterodimer, the virus particle, and neutralizing antibody Fabs bound to the glycoproteins. Based on biophysical and immunological assays, computational modeling and structure analyses, the E1E2 glycoproteins will be engineered to present epitopes that elicit strong neutralizing and broadly reactive antibodies and minimize the presentation of decoy non-productive epitopes. Furthermore, the organization of the glycoproteins and the structure of the virus particle will be determined. The tools of molecular virology, biophysics, computational modeling, x-ray crystallography and cryo-electron microscopy will be used to produce atomic-level details of the epitope landscape of HCV. The gained structural information will be utilized to design novel antigens in Project 1, determine the structures of designed immunogens experimentally in Project 4 and tested in vivo in Projects 1, 2 and 3. We will guide Project 1 at various times during the project to allow them to utilize the optimal E1E2 immunogen for studies in guinea pigs and, ultimately, a lead candidate for an optimized E1E2 immunogen for nonhuman primate studies. This information will be invaluable in deciphering the nature of neutralizing antibodies and their mechanisms of action against HCV and will shed light on the assembly and entry of hepatitis C virions.

摘要 – 项目 4 尽管为了解丙型肝炎病毒 (HCV) 的生物学特性进行了广泛的研究工作，但它仍然是人类的主要病毒。 此外，还明显缺乏有关其结构的信息。 我们打算利用我们广泛的结构生物学来研究病毒体及其组成蛋白的组织。 相关黄病毒的经验，重点确定病毒糖蛋白的结构，病毒 该项目将依赖于颗粒和与表面糖蛋白结合的有效中和抗体复合物。 重点关注项目 1 并主要与项目 1 互动，以询问其结构并为项目 1 提供指导 糖蛋白的特征对于引发广泛且有效的中和抗体非常重要。 将进行E1E2糖蛋白的分析和重新设计，以指导抗体成分 B 细胞和 T 细胞 HCV 疫苗是该 U19 提案的重点，该具体项目将使用三个目标。 确定E1E2异二聚体、病毒颗粒和与病毒颗粒结合的中和抗体Fab的结构 基于生物物理和免疫学测定、计算模型和结构分析， E1E2 糖蛋白将被设计为呈现能引发强中和和广泛反应的表位 抗体并最大限度地减少诱饵非生产性表位的呈现。 糖蛋白和病毒颗粒的结构将被确定分子病毒学的工具， 生物物理学、计算模型、X射线晶体学和冷冻电子显微镜将用于生产 HCV 表位景观的原子级细节将用于设计。 项目1中的新抗原，项目4中通过实验确定设计的免疫原的结构，以及 在项目1、2和3中进行了体内测试。我们将在项目期间的不同时间指导项目1，让他们能够 利用最佳 E1E2 免疫原在豚鼠中进行研究，并最终成为优化的主要候选者 用于非人类灵长类动物研究的 E1E2 免疫原对于破译 E1E2 的本质非常宝贵。 中和抗体及其针对 HCV 的作用机制，并将揭示其组装和 丙型肝炎病毒粒子的进入。