A sedimentation equilibrium study of the temperature-dependent association of bovine beta-casein

The temperature-dependent association of beta-casein was studied by the sedimentation equilibrium method. The weight-average molecular weight of the protein was determined in 0.2 M sodium phosphate buffer (pH 6.7) at 20, 15, 10 and 2 degrees C, and was found to be dependent markedly on both temperature and protein concentration. The data were well fitted by a monomer-n-mer association scheme, and the values of n, the second viral coefficient, and the free energy change for the association (association constant) were evaluated. The results show that temperature not only shifts the equilibrium but alters the polymer size: the values of n are 49, 22, and 12 at 20, 15, and 10 degrees C, respectively; the free energy change per monomer becomes more negative with increasing temperature, indicating that the attraction between the monomers in a polymer is stronger at higher temperature. This effect of temperature is in contrast to that of ionic strength which affects mainly the equilibrium. The enthalpy change, entropy change, and heat capacity change for the association were also estimated and were indicative of the formation of a considerable amount of hydrophobic bonds upon association.