Mechanism of protein salting in and salting out by divalent cation salts: balance between hydration and salt binding

Abstract

The preferential interactions of proteins with solvent components were studied in concentrated aqueous solutions of the sulfate, acetate and chloride salts of Mg2+, Ba2+, Ca2+, Mn2 + and Ni2+ [except for CaSO4, BaSO4, Mn-(OAc)2 and Ni(OAc)2], and results were compared with those of the Na+ salts. For all the salts, the preferential hydration increased in the order of Cl- < CH3-COO- < SO42- regardless of the cationic species used, in agreement with the anionic lyotropic series, and the same parameter exhibited a tendency to increase in the order of Mn2+, Ni2+ < Ca2+, Ba2+ < Mg2+ < Na+. The salting-out and stabilizing or salting-in and destabilizing effectiveness of the salts were interpreted in terms of the observed preferential interactions. The surface tension increment of salts, which is a major factor responsible for the preferential interactions of the Na+ salts, had no correlation with those of the divalent cation salts. The binding of divalent cations to the proteins overcomes the salt exclusion due to the surface tension increase, leading to a decrease in the preferential hydration. In conformity with this mechanism, the preferential interaction of MgCl2 was strongly pH dependent, because of the protein charge-dependent affinity of Mg2+ for prot eins, while NaCl showed no pH dependence of the preferential interaction. The proposed mechanism was supported by a strong correlation between the preferential interaction results and the interaction of these salts with the model peptide compound acetyltetraglycine ethyl ester, described by Robinson and Jencks.