Protein Separations Using Affinity Ultrafiltration with Small Charged Ligands

Abstract

Proposal Title: Protein Adsorption and Transport in Chromatographic Bioprocessing

Proposal Number: CTS-0345528

Principal Investigator: Andrew Zydney

Institution: Pennsylvania State University

The objective of this project is to examine a novel approach for protein purification using small, charged, affinity ligands to achieve high-resolution separations by affinity ultrafiltration. Affinity ultrafiltration exploits the high selectivity that can be obtained by molecular recognition, but the product and ligand are allowed to interact in free solution, thereby eliminating the mass transfer limitations and concerns about ligand accessibility that often hinder applications of packed column affinity chromatography. The binding interaction causes a change in electrical charge of the protein-ligand complex, and this is used to obtain high product retention by exploiting electrostatic interactions during ultrafiltration. Initial experimental studies will be performed using Cibacron Blue as the charged affinity ligand and human serum albumin as the model protein. Preliminary theoretical calculations indicate that the change in protein charge will provide high selectivity for membrane separations between albumin and protein impurities that are unable to bind Cibacron Blue. These results will demonstrate the feasibility of this technique, while providing a quantitative framework for design and optimization of affinity ultrafiltration processes using small charged affinity ligands. In terms of the broader impacts, this project will provide graduate and undergraduate students with hands-on experience dealing with membrane processes and protein separations. The results of this work may aid in the development of new technologies that provide high-resolution protein purification at a price, scale, and quality appropriate for high-value products.