Effect of dextran on the thermodynamic stability and structure of ribonuclease A

Abstract

Pioneer works to understand protein-folding problem have been done in dilute solutions. However, the cellular environment is crowded with macromolecules where excluded volume affects protein stability and function. Here, we assess thermal denaturation curves of RNase A in the presence of various concentrations of dextran 70 at different pH values. Values of thermodynamic parameters such as Tm (midpoint of denaturation), ∆Hm (enthalpy change at Tm) and ∆Cp (constant pressure heat capacity change) were determined under a given solvent condition. Using values of Tm, ∆Hm and ∆Cp in Gibbs–Helmholtz equation, \(\Delta G_{\text{D}}^{ \circ }\) (Gibbs energy change at 25 °C) was determined. It has been observed that dextran 70 stabilizes RNase A in terms of Tm and \(\Delta G_{\text{D}}^{ \circ }\). The stabilization of the protein is more at lower pH where the effect of exclusion is more. We had also investigated the secondary and tertiary structure of RNase A in the presence of dextran 70 and found that dextran 70 induces secondary structure, which is more prominent at physiological pH. However, there is no effect of crowding agent on tertiary structure of the protein. Our study demonstrates that crowding enhances thermal stability as well as the secondary structure of the protein.