Investigating polymeric antibody assembly, structure, function and therapeutic potential

研究聚合抗体的组装、结构、功能和治疗潜力

• 批准号: 10493309
• 负责人: Beth M. Stadtmueller
• 金额: $ 53.3万
• 依托单位: UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-24 至 2026-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10493309
• 关键词: Antibodies; Antigens; Binding; Binding Sites; Biochemical; Biological Assay; Biological Models; Birds; Cells; Clostridium difficile; Complex; Cryoelectron Microscopy; Data; Electrons; Engineering; Experimental Models; Fab Immunoglobulins; Family suidae; Foundations; Heavy-Chain Immunoglobulins; Human; Immune response; Immunity; Immunoglobulin A; Immunoglobulin M; Immunoglobulins; Immunologics; Immunology; In Vitro; Investigation; Knowledge; Ligand Binding; Ligands; Mammals; Mediating; Methods; Modeling; Modification; Molecular; Molecular Conformation; Mucosal Immunity; Mucous Membrane; Mus; Mutation; Outcome; Pathogenicity; Play; Polymeric Immunoglobulin Receptors; Polymers; Positioning Attribute; Predictive Factor; Process; Publishing; Reporting; Reptiles; Research; Role; Salmonella enterica; Secretory Component; Secretory Immunoglobulin A; Side; Solvents; Spectrum Analysis; Structure; Structure-Activity Relationship; Surface Plasmon Resonance; Testing; Therapeutic; Vertebrates; Wing; Work; X-Ray Crystallography; base; biophysical techniques; commensal microbes; crosslink; dimer; experimental study; flexibility; functional outcomes; host-microbe interactions; improved; insight; microbial; monomer; pathogen; pathogenic bacteria; polymeric IgA; polymerization; porcine model; preservation; programs; receptor; receptor binding; teleost; teleost fish

This proposal aims to investigate the assembly mechanisms, structures, and functions of polymeric (p) immunoglobulins (Ig) that populate the mucosa. The pIgs are found in vertebrates and together form a structurally diverse group of antibodies. They comprise several Ig heavy chain classes, including mammalian IgA and IgM, which typically contain between two and five Ig monomers and one joining chain (JC); however, potential to assemble with the JC and/or to assemble into polymers of different size varies with vertebrate species and Ig heavy chain class. Following assembly, pIgs are transported to the mucosa by the polymeric Ig receptor (pIgR). In the mucosa, the pIgR ectodomain, called secretory component (SC), remains bound and the complex is referred to as a secretory (S) Ig. SIgA is the predominant mucosal antibody in mammals; it is typically found in dimeric (d) forms; however higher order polymers such as tetramers are functionally relevant. SIgA is associated with unique effector functions compared to monomeric, circulatory antibodies; it can coat, cross-link and agglutinate commensal and pathogenic antigens and also mediate interactions with receptors on host and microbial cells. Despite significance, the structural basis for pIg assembly and SIg functions remained poorly understood through decades of immunological research. In 2020 the cryo-electron microscopy structures of SIgM, SIgA and a dimeric (d) IgA precursor were published revealing unprecedented molecular insights into these crucial complexes and opening the door to new questions and structure-guided experiments. The structures of dIgA and dimeric forms of SIgA revealed two IgAs joined through the JC to form a pseudosymmetric, bent conformation that appears to restrict the positions of antigen-binding fragments (Fabs) and promote access to receptor-binding sites. The SC is asymmetrically bound to one side and is solvent accessible, suggesting it may promote yet uncharacterized interactions with host or microbial factors. These observations raise the questions of how structural differences among pIg are generated (e.g dimer versus tetramer and JC versus no JC), how the bent, asymmetric arrangement of components is induced and maintained, and how it contributes to function. The proposed research program will use structural and biophysical approaches to target these questions. Aim 1 will identify Ig heavy chain residues, structural motifs and/or conformational changes that promote pIg assembly and control pIg polymeric state, while also determining the structural basis for JC- independent pIg assembly and function. Aim 2 will characterize JC-specific mechanisms of pIg assembly and its structural contributions to the pseudosymmetric conformation of dIgA. Aim 3 will characterize the functional significance SC and its capacity to bind microbial ligands. These studies will deliver comprehensive mechanistic models for pIg assembly, generate new pIg structures and report new SIg structure-function relationships. This outcome will improve knowledge of mucosal immunity and provide a foundation for engineering pIg and SIg in order to explore their normal functions and therapeutic potential.

该建议旨在研究聚合物（P）的组装机制，结构和功能 填充粘膜的免疫球蛋白（IG）。猪在脊椎动物中发现，并在结构上一起形成一个 多种抗体组。它们包括几个IG重链类，包括哺乳动物IgA和IgM 通常包含两个至五个IG单体和一个连接链（JC）；但是，潜力 与JC组装和/或组装成不同大小的聚合物随脊椎动物和Ig的不同而变化 重链课。组装后，通过聚合物Ig受体（PIGR）将猪运送到粘膜。 在粘膜中，孢子域域称为分泌组件（SC），保持绑定，复合物为 称为分泌物Ig。 Siga是哺乳动物中主要的粘膜抗体。通常在 二聚体（d）形式；但是，高阶聚合物（例如四聚体）在功能上是相关的。 Siga是相关的 与单体循环抗体相比，具有独特的效应功能；它可以覆盖，交联和 凝集的共生和致病性抗原，并介导宿主和受体的相互作用 微生物细胞。尽管有重要意义，但猪组装和SIG功能的结构基础仍然很差 通过数十年的免疫学研究理解。 2020年，冷冻电子显微镜结构 Sigm，Siga和二聚体（D）IgA前体发表了前所未有的分子见解 这些关键的复合物，并为新问题和结构引导实验打开了大门。这 SIGA的Diga和Dimeric形式的结构揭示了两个IGA通过JC连接，形成了伪对称， 弯曲的构象似乎限制了抗原结合片段（FAB）的位置并促进通道 到受体结合位点。 SC不对称地绑定到一侧，可以溶剂访问，建议它 可能会促进与宿主或微生物因子的相互作用但未表征的相互作用。这些观察提出了 关于猪之间结构差异如何产生的问题（例如二聚体与四聚体，JC与NO JC），如何诱导和维护组件的弯曲，不对称排列，以及如何贡献 功能。拟议的研究计划将使用结构和生物物理方法来针对这些 问题。 AIM 1将确定IG重链残基，结构图案和/或构象变化 促进猪的组装和控制猪聚合物状态，同时还确定JC-的结构基础 独立的猪组件和功能。 AIM 2将表征猪组件的JC特异性机制和 它对DIGA的假构象的结构贡献。 AIM 3将表征功能 显着性SC及其结合微生物配体的能力。这些研究将提供全面的机理 猪装配的模型，生成新的猪结构并报告新的SIG结构功能关系。这 结果将提高粘膜免疫的知识，并为工程猪和SIG提供基础 为了探索其正常功能和治疗潜力。