Epitope Discovery via Nanocluster Presentation

描述(申请人提供)：这项建议的目的是开发一种多功能纳米颗粒平台，用于展示具有免疫学分析的金纳米簇上的表位，以创建针对免疫系统的纳米技术。基于纳米颗粒的疫苗的未来发展需要产生能够忠实地概括病原微生物保护性抗原构象的蛋白质结构。具有讽刺意味的是，我们逻辑设计具有保护性的重组或合成蛋白抗原的能力相当有限。由于天然蛋白质上的大多数表位可能是组装的，表位确定工作面临的最大挑战之一是识别构象组装的表位。我们假设，靶蛋白环结构在金纳米簇表面的呈现将呈现出统计上显着的潜在环构象的多样性，为发现致病毒素的未知表位的新方法提供基础。为了证明这种方法的可行性，我们正在创建一个模拟炭疽杆菌保护性抗原(PA)的表位标记纳米簇。在这项提案中，我们的目标是通过以下具体目标来完善和扩展我们在纳米团簇上展示表位的概念策略：1.在金纳米团簇表面设计、合成和组装线性和构象表位。2.确定哪个构象肽环结构忠实地代表天然蛋白质中自然产生的免疫原性结构。3.使用纳米簇上呈现的表位筛选小鼠和人类抗体库以及患者血清，致力于开发诊断微阵列。4.优化多功能纳米粒对小鼠的免疫应答。纳米技术中的具体挑战将是在纳米颗粒表面创建特定的亚结构，这些特定构象的免疫识别，多功能纳米颗粒的创建，以及多肽环结构与纳米颗粒大小和曲率的关系。生物学和医学的具体变化包括发现未映射的线性和构象表位，免疫系统对功能化纳米粒的反应，构象表位抗体库的筛选，以及最终向构象表位标记纳米粒作为潜在疫苗的发展。

DESCRIPTION (provided by applicant): The purpose of this proposal is to develop a multifunctional nanoparticle platform for epitope presentation on gold nanoclusters with immunological analysis for the creation of nanotechnologies that target the immune system. The future development of nanoparticle-based vaccines requires the generation of protein structures that faithfully recapitulate the conformation of protective antigens of pathogenic microorganisms. Ironically, our ability to logically design recombinant or synthetic protein antigens that are protective is quite limited. As most epitopes on native proteins are likely to be assembled, one of the most significant challenges facing epitope determination efforts is the identification of conformationally assembled epitopes. We hypothesize that the presentation of target protein loop structures on the surface of gold nanoclusters will present a statistically significant diversity of potential loop conformations to provide the foundation of a new method for discovering unknown epitopes of pathogenic toxins. To demonstrate the feasibility of this approach, we are creating an epitope-labeled nanocluster that mimics the protective antigen (PA) of Bacillus anthracis. In this proposal, our goal is to refine and extend our conceptual strategies for epitope presentation on nanoclusters via the following specific aims: 1. Design, synthesize, and assemble linear and conformational epitopes on the surface of gold nanoclusters. 2. Determine which of the conformational peptide loop constructs faithfully represent the naturally occurring immunogenic structures in native protein. 3. Screen mouse and human antibody libraries, and patient sera using epitopes presented on nanoclusters working towards the development of a diagnostic microarray. 4. Optimize the immunological response of multifunctional nanoparticles in mice. Specific challenges in nanotechnology will be the creation of specific substructures on a nanoparticle surface, immunorecognition of these specific conformations, creation of multifunctional nanoparticles, and correlation of peptide loop structure as a function of nanoparticle size and curvature. Specific changes in biology and medicine include the discovery of unmapped linear and conformational epitopes, the immune system response to functionalized nanoparticles, the screening of antibodies libraries for conformational epitopes, and ultimate development towards conformational epitope-labeled nanoparticles as potential vaccines.