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

Northwestern University, Evanston IL

Investigators

Abstract

NON-TECHNICAL SUMMARY This 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 SUMMARY This 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.