Quantitative Characterization of Protein-Protein and Protein-Polymer Interaction via Nonideal Tracer Sedimentation Equilibrium

Abstract

The interaction between three dilute ‘tracer’ proteins (bovine serum albumin, superoxide dismutase, and ovomucoid) at a concentration of 2 mg/ml and three ‘crowder’ macromolecules (Ficoll 70, Dextran 70 and ovomucoid) at concentrations up to 100 mg/ml was characterized by analysis of dependence of the equilibrium gradients of both tracer and crowder upon the concentration of crowder. The equilibrium gradients of all three crowder species were found to be independent of temperature over the range 5 - 37 degree. The equilibrium gradients of BSA and ovomucoid in crowder were likewise found to be independent of temperature over this range, indicating that interaction between these tracers and all three crowders is predominantly repulsive and essentially entirely entropic in nature. However, composition dependence of the equilibrium gradient of SOD was found to be significantly dependent upon temperature in a manner indicating the presence of significant temperature-dependent (i.e., enthalpic) attractive interaction between SOD and all three crowder species. The experimental data are analyzed thermodynamically and in the context of effective hard particle theory [1] generalized to allow for the presence of attractive enthalpic interaction between solute species [2].[1] Fodeke A.A. and Minton A.P. (2010) J. Phys. Chem B. 114: 10876 - 10880[2] Jiao M et. al. (2010) Biophys. J. 99: 914-923.