Protein-Protein Interactions in Dilute to Concentrated Solutions: alpha-Chymotrypsinogen in Acidic Conditions

Protein-protein interactions were investigated for alpha-chymotrypsinogen by static and dynamic light scattering (SLS and DLS, respectively), as well as small-angle neutron scattering (SANS), as a function of protein and salt concentration at acidic conditions. Net protein-protein protein concentration with a local Taylor series approach, which does not require one to assume the underlying form or nature of intermolecular interactions. In addition, G(22) and S(q) were further analyzed by traditional methods involving fits to effective interaction potentials. Although the fitted model parameters were not always physically realistic, the numerical values for G(22) and the static structure factor S(q) from SLS and SANS data. G(22) was obtained by regressing the Rayleigh ratio versus protein concentration with a local Taylor series approach, which does not require one to assume the underlying form or nature of intermolecular interactions. In addition, G(22) and S(q) were further analyzed by traditional methods involving fits to effective interaction potentials. Although the fitted model parameters were not always physically realistic, the numerical values for G(22) and S(q -> 0) were in good agreement from SLS and SANS as a function of protein concentration. In the dilute regime, fitted G(22) values agreed with those obtained via the osmotic second virial coefficient B-22 and showed that electrostatic interactions are the dominant contribution for colloidal interactions in alpha-chymotrypsinogen solutions. However, as protein concentration increases, the strength of protein protein interactions decreases, with a more pronounced decrease at low salt concentrations. The results are consistent with an effective "crowding" or excluded volume contribution to G(22) due to the long-ranged electrostatic repulsions that are prominent even at the moderate range of protein concentrations used here (<40 g/L). These apparent crowding effects were confirmed and quantified by assessing the hydrodynamic factor H(q -> 0), which is obtained by combining measurements of the collective diffusion coefficient from DLS data with measurements of S(q -> 0). H(q -> 0) was significantly less than that for a corresponding hard-sphere system and showed that hydrodynamic nonidealities can lead to qualitatively incorrect conclusions regarding B-22, G(22), and static protein-protein interactions if one uses only DLS to assess protein interactions.