Lysozyme aggregation and solution properties studied using PGSE NMR diffusion measurements

The solution behavior of lysozyme was studied as a function of protein concentration, NaCl concentration, pH, and temperature using pulsed-gradient spin-echo NMR diffusion measurements. The lysozyme solutions clearly exhibited nonideal behavior which was sensitive to both the salt concentration and pH. Lysozyme has an isoelectric point of pH 11, and it is often overlooked that at normal pH it has a net positive charge. Since lysozyme is a charged species, the changes in the diffusion coefficients were interpreted, considering the competing effects of salt-mediated changes in protein interactions (e.g., electrostatic repulsion) and aggregation. The behavior is in agreement with Derjaguin-Landau-Verwey-Overbeek (DLVO)-type modeling, accounting for the attractive and repulsive forces present. The diffusion data was compared with various self-association models, including corrections for the effects of self-obstruction. The diffusion coefficients of the higher oligomers were calculated, assuming that the monomers aggregated as hard spheres. Using an isodesmic association model, the equilibrium constant for the self-association of lysozyme at pH 4.6 and 298 K in the presence of 0.5 M NaCl was estimated to be 118 ± 12 M^-1.