The effects of pH and electrolyte concentration on protein-protein interactions in lysozyme and chymotrypsinogen solutions were investigated by static light scattering (SLS) and small angle neutron scattering (SANS). Very good agreement between the values of the virial coefficients measured by SLS and SANS was obtained without use of adjustable parameters. At low electrolyte concentration the virial coefficients depend strongly on pH, and change from positive to negative as the pH increases. All coefficients at high salt concentration are slightly negative and depend weakly on pH. For lysozyme, the coefficients always decrease with increasing electrolyte concentration. However, for chymotrypsinogen there is a crossover point around pH 5.2 above which the virial coefficients decrease with increasing ionic strength, indicating the presence of attractive electrostatic interactions. The data are in agreement with DLVO-type modeling, accounting for the repulsive and attractive electrostatic, van der Waals and excluded volume interactions of equivalent colloid spheres. This model is, however, unable to resolve the complex short-ranged orientational interactions. The results of protein precipitation and crystallization experiments are in qualitative correlation with the patterns of the virial coefficients and demonstrate that interaction mapping could help outline new crystallization regions.
The onset of protein precipitation or crystallization and the morphology of any separated protein phase will be predominantly determined by the mechanism of molecular approach, reorientation and incorporation, which is governed by the strength and range of the colloidal interactions. Further theoretical and experimental study of these interactions is the key to better understanding of protein crystallization. A pioneering study of George and Wilson (Acta Cryst. D50, 361) demonstrated the relation between the protein osmotic second virial coefficient and protein crystallization behavior. The most versatile tool for investigating the bulk colloidal properties of protein solutions are the scattering methods. In the present study, we describe and compare systematic interaction and crystallization data obtained with lysozyme and a -chymotrypsinogen.
OVERVIEW OF THE LIGHT AND NEUTRON SCATTERING METHODS USED
SANS probes both the interaction and molecular structure data
g(r) is the radial distribution function. In the limit of q -> 0
SLS from proteins probes the interactions only
In the practical static light scattering experiment
where K is a constant calculated from the optical properties of the system
The above methods were used to characterize the interactions in protein solutions at varying pH and electrolyte concentration.
© Copyright O. D. Velev et al. 1999
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