(640c) Ion Exchange Equilibrium and Kinetics of Associating Proteins– Effects of Urea and Salt Concentrations

In this work, we probe the self-association and ion exchange behavior of Apolipoprotein A-I Milano using two anion exchange resins, Q-Sepharose-HP and Macro-Prep-HQ, and urea as a modifier of protein-protein interactions and protein structure. Macroscopic measurements, such as equilibrium isotherms, batch uptake, and column experiments are used to determine equilibrium binding capacities and mass transfer rates under different processing conditions, while refractive index-based microscopy is used to provide insight into the underlying mass transfer mechanisms. In addition, protein cross-linking and dynamic light scattering experiments are used to characterize the protein association and diffusivity in solution. High equilibrium binding capacities are found in the absence of urea for both resins. However, these high binding capacities in the absence of urea are accompanied by slow mass transfer under binding conditions due to the self-association of the protein. Conversely, rapid mass transfer, i.e. faster than the free solution diffusivity of the protein, is observed at high urea concentrations where self-association is minimal, but at the expense of greatly reduced binding capacities. Under non-binding conditions, relatively small changes in the protein mass transfer rate are observed as a function of urea, with higher urea concentrations leading to slower rates, in opposition to the protein's behavior under binding conditions.