HEW lysozyme salting by high-concentration NaCl solutions followed by titration calorimetry

Concentration dependence of NaCl salting of 0-1.5 mM lysozyme solution in 0.1 M sodium acetate buffer, pH 4.25, was investigated for NaCl concentration varying up to 0.9 M. Calorimetric experiments demonstrated that depending on the salt concentration the estimated number of the binding sites on the lysozyme surface varied in the range of 5 up to 13, and the increase of salt concentration caused the decrease of the number of accessible sites. The small, but significant, local maximum centered at 0.63 M NaCl concentration indicated the specific salting-out of the lysozyme accompanied by binding of ∼2-3 chloride anions. Generalized McMillan and Mayer's approach reduced to the third-order virial coefficients demonstrates the domination of lysozyme aggregation upon salt addition (a21-hxxy) and salt organization on the lysozyme surface (a12-hxyy) processes.