Calorimetric studies of interactions between low molecular weight salts and bovine serum albumin in water at p H values below and above the isoionic point.

Isothermal titration calorimetry was used to determine the temperature and salt concentration dependence of the enthalpy of mixing, Δmix H , of bovine serum albumin (BSA) in aqueous buffer solutions with several low molecular weight salts. Three buffers were used: acetate ( p H = 4.0), MOPS (7.2), and borate (9.2). Since the isoionic point of BSA is at p I ≈ 4.7, the net charge of BSA in acetate buffer was positive (≈ +20), while in the other two buffer solutions it was negative (≈ -15 in MOPS and ≈ -25 in borate). The majority of the recorded heat effects were exothermic, while only at p H = 9.2 a weak endothermic effect upon mixing BSA with LiCl, NaCl, and KCl was observed. For all buffer solutions the absolute values of Δmix H of sodium salts followed the order: NaCl < NaBr < NaNO3 < NaI < NaSCN, which is the reverse Hofmeister series for anions. The magnitude of the effects was the largest in acetate buffer and decreased with an increasing p H value of the solution. While the effect of varying the anion of the added salts was strongly pronounced at all p H values, the effect of the cation (LiCl, NaCl, KCl, RbCl and CsCl salts) was weak. The most interesting feature of the results obtained for p H > p I was the fact that Δmix H were considerably more sensitive to the anion (co-ion to the net BSA charge) than to the cation species. This indicated that anions interacted quite strongly with the BSA even at p H values where the net charge of the protein was negative. We showed that Δmix H at high addition of salts correlated well with the enthalpy of hydration of the corresponding salt anion. This finding suggested, consistently with some previous studies, that a part of the exothermic contribution to Δmix H originated from the hydration changes upon the protein-salt interaction. Theoretical analysis, based on the primitive model of highly asymmetric electrolyte solutions solved within the mean spherical approximation, was used to estimate Coulomb effects upon mixing.