Structural biology of antibody:antigen complexes

抗体的结构生物学：抗原复合物

• 批准号: 8377204
• 负责人: STEPHEN COPLAN HARRISON
• 金额: $ 25.45万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-08-01 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8377204
• 关键词: Affinity; Antibodies; Antigen-Antibody Complex; Antigens; B cell repertoire; Binding; Characteristics; Complex; Cryoelectron Microscopy; Devices; Dissection; Epitopes; Exposure to; Foundations; Glycoproteins; Goals; Immune response; Immunization; Immunodominant Epitopes; Infection; Influenza Hemagglutinin; Instruction; Knowledge; Lead; Link; Maps; Methods; Proteins; Resolution; Rotavirus; Structure; Technology; Testing; Vaccination; Vaccines; Viral Antigens; Virus; Work; base; design; influenzavirus; novel; particle; reconstruction; response; scaffold; structural biology; vaccine development

The overall goals of Project 3 are to understand the high-resolution structural correlates of immunodominance and to provide structural foundations for designing immunogens that can focus an immune response on particular, chosen epitopes. Our principal hypothesis is that structural contributions to immunodominance can be revealed by correlating structures of antibody-antigen complexes with knowledge of the repertoire from which the antibody was derived and reconstruction of the mutational sequence that led to the affinity matured antibody from its germline precursor. The high-throughput approaches described in Project 1 can give us the required information about repertoire and about affinity maturation of antibodies denved from the settings of vaccination vs. infection. We propose here to provide the corresponding three-dimensional information. We have the following four Aims. [1) What are the structural characteristics of cross-reactive antibodies bound with their HA epitopes? We will determine structures of selected representatives of the antibodies studied in projects 1 and 2, in complex with the relevant influenza virus HA. We will begin with standard crystallographic approaches, but proceed as rapidly as possible to higher-throughput cryoEM methods (Aim 2; see also Core C). (2) We will develop approaches for mounting "guest proteins" [with influenza HA as the principal target) onto icosahedral-virus scaffolds, to create a symmetric array suitable for structure determination by cryoEM. We will start with the rotavirus double-layer particle (DLP) as our scaffold, and the outer-layer glycoprotein, VP7, as the mounting device.; we will also test the other outer-layer protein, VP4, as a trimeric mounting protein. [3) What makes an epitope immunodominant? The opportunity afforded by the cryoEM approach to examine a systematic and relatively unbiased set of complexes will allow us to determine the structural correlates of immunodominance directly. We will determine structures of a selected HA in complex with antibodies representing the entire spectrum of a typical repertoire as revealed by the studies in Project 1 and mapped by Project 2. [4) Application to other viral antigens. We will apply to the mounting strategies developed in Aim 2 to other viral antigens for which a combination of repertoire analysis and structural dissection might lead to novel

项目 3 的总体目标是了解免疫优势的高分辨率结构相关性 并为设计免疫原提供结构基础，使免疫反应集中于 特别的、选择的表位。我们的主要假设是，结构对免疫优势的贡献可以 通过将抗体-抗原复合物的结构与来自的库的知识相关联来揭示 抗体的来源和导致亲和力成熟的突变序列的重建 来自其种系前体的抗体。项目 1 中描述的高通量方法可以为我们提供 所需的有关抗体库和有关从设置中衍生的抗体亲和力成熟的信息 疫苗接种与感染。我们这里建议提供相应的三维信息。我们 有以下四个目标。 [1) 结合的交叉反应抗体的结构特征是什么？ 他们的HA表位？我们将确定项目中研究的抗体的选定代表的结构 1和2，与相关流感病毒HA复合。我们将从标准晶体学开始 方法，但尽快转向更高通量的冷冻电镜方法（目标 2；另请参阅核心 C）。 (2) 我们将开发将“客体蛋白”（以流感 HA 为主要目标）安装到 二十面体病毒支架，创建适合冷冻电镜结构测定的对称阵列。我们将 从轮状病毒双层颗粒（DLP）作为我们的支架开始，外层糖蛋白 VP7 作为 安装装置。；我们还将测试另一种外层蛋白 VP4，作为三聚体安装蛋白。 [3) 什么 使表位具有免疫优势？冷冻电镜方法提供了检查 系统且相对公正的复合体集合将使我们能够确定以下结构的相关性： 直接免疫优势。我们将确定选定的 HA 与抗体复合物的结构 代表了项目 1 中的研究揭示的典型曲目的整个范围，并由 项目2. [4)在其他病毒抗原上的应用。我们将应用目标 2 中制定的安装策略来 其他病毒抗原，其库分析和结构解剖的结合可能会产生新的