Collaborative Research: DMREF: GOALI: High-Affinity Supramolecular Peptide Materials for Selective Capture and Recovery of Proteins

该项目是一个综合的实验计算方法，旨在开发一类基于肽的超分子材料，作为非色谱蛋白质纯化的高亲和力沉淀剂。从生物资源中分离和纯化治疗性蛋白质是生物制剂工业生产效率和成本效益的一个重要限制。尽管亲和层析的介质成本高，负载能力有限，但它仍然是大规模工业蛋白质纯化中最广泛使用的捕获方法。由于哺乳动物细胞培养的进步和持续的工艺发展，上游滴度的快速增长进一步挑战了下游制造的效率。亲和沉淀可以潜在地克服与柱大小和配体固定化相关的色谱限制。根据材料基因组计划（MGI）的目标，该项目将设计、合成和开发能够特异性结合、选择性捕获和有效分离蛋白质的自组装肽材料。此外，该项目将为各级学生提供新的教育和推广机会，让他们参与STEM研究，并体验学术界和工业界不同的实验室环境。技术概述：该项目旨在解决下游蛋白质生产中基于肽的亲和沉淀剂发展的关键基本挑战。使用亲和沉淀剂进行蛋白质纯化基本上有三个步骤。这些是：(1)选择性捕获感兴趣的蛋白质；(2)结合诱导上清相分离；(3)蛋白质的回收。该项目包括三个具体目标，每个目标都涵盖了成功开发用于非层析蛋白纯化的肽基超分子免疫纤维的关键步骤。第一个重点是设计和合成用于选择性捕获目标单克隆抗体（mab）的免疫纤维。第二个目的是了解、确定和优化mAb结合触发宏观相分离的条件。第三个目标集中在蛋白质恢复过程和系统适用性和可扩展性的评估。理论和多尺度模型将为免疫纤维的超分子设计和共组装策略提供指导原则，以优化配体的呈现以最大限度地捕获蛋白质，并有助于阐明免疫纤维组装和解离的热力学和动力学方面，以及结合触发的相分离机制。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

NON-TECHNICAL SUMMARYThis project is an integrated experimental-computational approach that aims to develop a class of peptide-based supramolecular materials as high-affinity precipitants for non-chromatographic protein purification. The separation and purification of therapeutic proteins from their biological resources are a significant limitation for industrial manufacturing of biologics in terms of their efficiency and cost-effectiveness. Despite the high media cost and limited loading capacity, affinity chromatography remains the most widely used capture method for large-scale industrial protein purification. The rapid growth of upstream titers, due to advancements in mammalian cell culture and continuous process development, has further challenged the efficiency of downstream manufacturing. Affinity precipitation can potentially overcome the chromatography limitations associated with column size and ligand immobilization. In line with the goals of the Materials Genome Initiative (MGI), this project will design, synthesize, and develop self-assembling peptide materials that can specifically bind, selectively capture, and effectively separate proteins from their bio-based resources. In addition, this project will foster new educational and outreach opportunities for students at all levels to participate in STEM research and to experience different laboratory settings that range across academia and industry.TECHNICAL SUMMARYThis project aims to address the key fundamental challenges in the development of peptide-based affinity precipitants for downstream protein manufacturing. There are essentially three steps in the use of affinity precipitants for protein purification. These are the: (1) selective capture of proteins of interest; (2) binding-induced phase separation from supernatant; and (3) recovery of proteins. The project includes three specific aims, each covering a key step toward the successful development of peptide-based supramolecular immunofibers for non-chromatographic protein purification. The first thrust focuses on the design and synthesis of immunofibers for selective capture of target monoclonal antibodies (mAbs). The second is intended to understand, determine, and optimize the conditions for mAb binding-triggered macroscopic phase separation. The third aim centers on the protein recovery processes and the assessment of system applicability and scalability. Theory and multiscale models will be developed to provide guiding principles for the supramolecular design of immunofibers and the co-assembly strategies to optimize the ligand presentation for maximal protein capture, and to help elucidate the thermodynamic and kinetic aspects of immunofiber assembly and dissociation, as well as the binding-triggered phase separation mechanisms.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.