Investigating the Architecture of HIV Envelope Glycoprotein

研究 HIV 包膜糖蛋白的结构

• 批准号: 8262992
• 负责人: Miklos Guttman
• 金额: $ 4.92万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-02-01 至 2014-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8262992
• 关键词: Address; Antibodies; Antigens; Architecture; Binding; Binding Sites; CD4 Antigens; Carbohydrates; Cataloging; Catalogs; Cells; Comparative Study; Complementarity Determining Regions; Complex; Coupled; Data; Deuterium; Development; Enzyme-Linked Immunosorbent Assay; Epitopes; Glycoproteins; Goals; HIV; HIV Envelope Protein gp120; Hydrogen; Imagery; Immune; Immune response; Immunoprecipitation; Kinetics; Link; Maps; Masks; Mass Spectrum Analysis; Measures; Mediating; Membrane; Membrane Proteins; Methods; Modeling; Molecular; Monitor; Monoclonal Antibodies; Nature; Polysaccharides; Property; Proteins; Resolution; Roentgen Rays; Role; Shapes; Site; Solvents; Spatial Distribution; Structure; System; Techniques; Testing; Uncertainty; Vaccine Design; Vaccines; Viral; base; design; env Glycoproteins; flexibility; glycosylation; gp160; improved; interest; neutralizing antibody; next generation; receptor; receptor binding; reconstruction; vaccine development; virus envelope

DESCRIPTION (provided by applicant): The human immunodeficiency virus (HIV) employs an array of properties that enable it to evade antibody neutralization. Rational vaccine design has been limited due to the insufficient structural understanding of the HIV envelope glycoprotein (Env), the primary target for neutralizing antibodies. Advances in structural determination have been hindered due to the presence of flexible hypervariable loops and the high carbohydrate content of this protein and the arrangement of the full Env complex remains controversial. The nature of the glycan shield, thought to mask conserved sites, is also of key interest as it mediates immune escape. Furthermore the binding of host cell CD4 receptor is known to induce massive conformational changes in Env, but a detailed examination of these changes has yet to be addressed. This proposal aims to combine two complementary approaches, small-angle X-ray scattering (SAXS) with ab initio shape reconstruction and hydrogen-deuterium exchange (HXMS) with mass spectrometry analysis to address several questions regarding the architecture of trimeric Env. The extensive catalog of constructs available from the Hu lab enables comparative studies of Env ranging from monomeric subunits to trimeric complexes, in the context of the same isolate. Identification of the trimeric interface, conformational changes induced upon receptor binding, and epitope occlusion by variable loops and glycans can be addressed with these techniques. The interaction of several well-characterized monoclonal antibodies directed against the CD4 binding site will also be compared to address the question of how conformational changes induced by antibody binding differ from those resulting from CD4 binding, and whether there are potentially correlates with neutralizing potency. Understanding these properties of Env will assist current efforts to identify targets for development of broadly neutralizing antibodies, necessary for an effective vaccine. PUBLIC HEALTH RELEVANCE: The poor understanding of the primary surface protein of the Human Immunodeficiency Virus (HIV) has been a major hindrance to vaccine development. A thorough understanding of the organization of this protein is critical for designing next generation immunogens to induce an effective immune response. Using two emerging techniques we will address uncertainties regarding the architecture of this surface protein, identify how critical antigenic regions are protected, and identify ways to improve vaccine design.

描述（由申请人提供）：人类免疫缺陷病毒（HIV）利用一系列特性使其能够逃避抗体中和。由于对中和抗体的主要靶标 HIV 包膜糖蛋白 (Env) 的结构了解不足，合理的疫苗设计受到限制。由于柔性高变环的存在和该蛋白质的高碳水化合物含量，结构确定的进展受到阻碍，并且完整的 Env 复合物的排列仍然存在争议。聚糖屏蔽的性质被认为可以掩盖保守位点，但由于它介导免疫逃逸，因此也具有重要意义。此外，已知宿主细胞 CD4 受体的结合会诱导 Env 发生巨大的构象变化，但尚未对这些变化进行详细检查。 该提案旨在结合两种互补的方法，即小角度 X 射线散射 (SAXS) 与从头形状重建和氢氘交换 (HXMS) 与质谱分析，以解决有关三聚 Env 结构的几个问题。 Hu 实验室提供的广泛构建体目录使得能够在同一分离物的背景下对从单体亚基到三聚体复合物的 Env 进行比较研究。通过这些技术可以解决三聚体界面的识别、受体结合引起的构象变化以及可变环和聚糖导致的表位封闭。还将比较几种针对 CD4 结合位点的已充分表征的单克隆抗体的相互作用，以解决抗体结合引起的构象变化与 CD4 结合引起的构象变化有何不同，以及是否与中和效力潜在相关的问题。了解 Env 的这些特性将有助于当前确定开发有效疫苗所必需的广泛中和抗体的目标。 公共卫生相关性：对人类免疫缺陷病毒 (HIV) 主要表面蛋白的了解不足一直是疫苗开发的主要障碍。彻底了解这种蛋白质的组织对于设计下一代免疫原以诱导有效的免疫反应至关重要。使用两种新兴技术，我们将解决有关这种表面蛋白结构的不确定性，确定如何保护关键抗原区域，并确定改进疫苗设计的方法。