Synergetic effect of polyols with tetrabutylammonium bromide and urea on the thermal stability of lysozyme

Abstract

The thermal denaturation of lysozyme was studied in 2.0 molal aqueous solutions of polyols at pH 2.50 and varying concentration of glycerol at pH 2.50 and 6.00 using differential scanning calorimetry (DSC). The transition temperature, heat capacity, enthalpy, entropy and free energy of stabilization have been determined by a least square fit of the excess heat capacity data to the two-state model. Polyols are found to stabilize lysozyme and the stabilization increases with an increase in the number of hydroxyl groups. The stabilization increases with an increasing concentration of glycerol. The stabilization has been explained in terms of preferential hydration or due to the strengthening of the water structure which in turn intensify the hydrophobic interactions of the protein. The calorimetric studies have been done on the thermal denaturation of lysozyme in the presence of 0.5 m tetrabutylammonium bromide (Bu₄NBr) + 2.0 m polyols at pH 2.50 and 0.5 m Bu₄NBr or urea + varying concentration of glycerol (0.0-10.0m) at pH 2.50 and 6.00. The effect is found to be nearly additive in the case of Bu₄NBr and polyols. The results on the combined systems of Bu₄NBr and varying concentration of glycerol shows that effect is additive at low pH but not at high pH. This has been explained in terms of predominance of the enthalpic contribution of glycerol in comparison to the entropic contribution of Bu₄NBr. The comparison of the combined effect of 0.5 m urea + glycerol and 0.5 m Bu₄NBr + glycerol indicates that Bu₄NBr is a stronger destabilizer than urea.