The second virial coefficient as a predictor of protein aggregation propensity: A self-interaction chromatography study

The second osmotic virial coefficients (b(2)) of four proteins – lysozyme, recombinant human lactoferrin, concanavalin A and catalase were measured by self-interaction chromatography (SIC) in solutions of varying salt type, concentration and pH. Protein aggregate sizes based on the initial hydrodynamic radius of the protein solution species present were measured using dynamic light scattering, and the relationship between b(2) and protein aggregate size was studied. A linear correlation was established between b(2) values and protein aggregate hydrodynamic size for all proteins, and for almost all solution conditions. Aggregate sizes of <∼10 nm, indicative of non-aggregated protein systems, were consistently observed to have b(2) values >0. The observed b(2) trends as a function of solution conditions were very much protein dependent, with notable trends including the existence of attractive interactions (negative b(2) values) at low ionic strengths for catalase and concanavalin A, and the highly positive b(2) values observed for lactoferrin over a wide range of solution conditions, reflecting lactoferrin’s innately high stability. It is concluded that the quantification of protein–protein interactions using SIC based b(2) data is a potentially valuable screening tool for predicting protein aggregation propensity.