Rapid structural characterization of most promising novel mAbs and vaccines

最有前途的新型单克隆抗体和疫苗的快速结构表征

• 批准号: 9057431
• 负责人: Andrew Barrett Ward
• 金额: $ 58.47万
• 依托单位: VANDERBILT UNIVERSITY MEDICAL CENTER
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9057431
• 关键词: Antibodies; Antigens; Binding Sites; Complex; Cryoelectron Microscopy; Crystallography; Development; Electron Microscopy; Engineering; Experimental Designs; Generations; Goals; Hemagglutinin; Housing; Influenza; Joints; Methods; Modeling; Negative Staining; Proteins; Research; Research Project Grants; Resolution; Structure; Therapeutic; Therapeutic antibodies; Vaccine Antigen; Vaccine Research; Vaccines; Validation; X-Ray Crystallography; anti-influenza; antibody engineering; base; combat; design; influenzavirus; neutralizing antibody; novel; programs; protein complex; receptor binding; scaffold; screening; structural genomics; therapeutic vaccine; tool

The Structure Core will support all three projects in this U19. The goal of our U19 research program is to use structure-based rational design to combat influenza virus from both the therapeutic antibody and vaccine perspective. These endeavors, targeting the receptor-binding site (RBS) on hemagglutinin (HA), require many rounds of design and experimental validation executed in an iterative manner. Thus, the focus of this Core is three-fold: (i) Naturally occurring antibodies from Project 1 will be studied in complex with natural HA molecules. (ii) to solve structures of HA in complex with engineered antibodies from Project 2 for therapeutic applications, and (iii) to solve structures of HA engineered scaffolds in complex with known broadly neutralizing antibodies (bnAbs) in Project 3 for vaccine applications. We will use both electron microscopy (EM) and x-ray crystallography to accomplish these aims and generate sufficient structural coverage to enable development of anti-influenza solutions. Specifically, we will use electron microscopy (EM) to solve structures of HA in complex with engineered antibodies. EM is uniquely suited to rapidly generate structural information of protein complexes in a high-throughput manner. The information from EM can directly inform further structural and design efforts. We will employ both negative stain and cryo-EM methods to evaluate antibody-HA complexes at moderate and high resolution, respectively. We will also use x-ray crystallography to solve structures of HA and engineered HA scaffolds with antibodies. X-ray crystallography is routinely used to reveal atomic-level details of proteins and their complexes. Using the crystal screening capacity of The Joint Center for Structural Genomics housed at Scripps (Dr. Wilson is the PI), we can screen large numbers of antibodies, antigens, and conjugate complexes. We will use this pipeline to solve atomic-resolution structures of antibody-HA and antibody-scaffold complexes.

结构核心将支持本U19中的所有三个项目。我们U19的目标 研究计划是使用基于结构的理性设计来打击流感病毒 从治疗抗体和疫苗的角度来看。这些努力，目标 血凝素（HA）上的受体结合位点（RB）需要多轮设计 以及以迭代方式执行的实验验证。因此，重点 核心是三倍：（i）将研究项目1的自然发生抗体 与天然HA分子的复合物。 （ii）解决HA的结构与 项目2的工程抗体用于治疗应用，以及（iii）解决 HA工程脚手架的结构中，已知广泛中和 疫苗应用项目3中的抗体（BNAB）。我们将使用两个电子 显微镜（EM）和X射线晶体学以实现这些目标并产生 足够的结构覆盖范围，可以开发抗激素解决方案。 具体而言，我们将使用电子显微镜（EM）在复合物中求解HA的结构 使用工程抗体。 EM非常适合快速生成结构 蛋白质复合物的信息以高通量方式。来自EM的信息 可以直接告知进一步的结构和设计工作。我们将使用否定 污渍和冷冻EM方法评估中等和高的抗体-HA复合物 分别解决。我们还将使用X射线晶体学来求解HA的结构 和具有抗体的工程HA脚手架。 X射线晶体学通常用于 揭示蛋白质及其复合物的原子水平细节。使用晶体筛选 在Scripps所容纳的结构基因组合联合中心的能力（Wilson博士是 PI），我们可以筛选大量抗体，抗原和偶联络合物。 我们将使用此管道来解决抗体-HA和 抗体 - 损伤复合物。