Interactions between proteins in aqueous solutions are influenced by the addition of ions, steaming from the disolved low molecular weight salts. The phenomenon depends on the salt-type and is known as the Hofmeister effect. Understanding the origins of these effect is important, for example, when trying to avoid protein aggregation in drug formulations [1]. To study interactions between the bovine serum albumin (BSA) and various low molecular weight salts isothermal titration calorimetry technique was used. Experiments were performed with BSA in four different buffer solutions (acetic, Tris, HEPS, and MOPS) and at three temperatures (5, 15, and 35 °C). BSA molecules in acetic buffer (pH = 4.0) adopt a net positive charge, whereas in Tris, HEPES, and MOPS buffers (pH = 7.2 - 8.0) the protein molecules are negatively charged as a whole [2]. For this reason, the influence of anions — Cl-, Br-, I-, NO3-, and SCN- — acting as counterions to the BSA could be studied in acetic buffer. On the other hand, the investigation of the influence of cations — Li+, Na+, K+, Rb+, and Cs+ — is possible in Tris, HEPS, and MOPS buffer solutions of BSA. Preliminary results indicate a relatively weak dependence of the protein-salt mixing enthalpy on the chemical nature of the cation (exception being the lithium ion). In contrast, the salt-specific effect of anions is much more pronounced. The exothermic effect decreases in the following order: SCN- &gt; I- &gt; NO3- &gt; Br- &gt; Cl- (sodium salts were used). The series corroborates with the inverse Hofmeister series for anions. We proposed a thermodynamic model of BSA-counterion binding, and used the measured enthalpy changes at different temperatures to obtain the thermodynamics fingerprint of the protein-salt interaction (standard Gibbs free energy, entropy, and heat capacity changes). Stability of BSA solutions in mixture with low molecular weight salts was further investigated also in the light of solubility. Polyethylenglycol (PEG) was added to solutions in order to lower the solubility of BSA at ambient conditions. Liquid-liquid phase separation was observed, where one phase was protein reach and the other protein poor. The systematic investigation of the salt identity and the nature of the buffer on these phenomena is being undertaken.

Stability of bovine serum albumin solutions in presence of salts: A calorimetric and solubility study

253rd National Meeting (2017)

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