The Effect of Ethylene Glycol on the Thermal Denaturation of Ribonuclease A and Chymotrypsinogen A as Measured by Differential Scanning Calorimetry

Abstract

Abstract The thermal denaturation of ribonuclease A(RNase A) and chymotrypsinogen A in aqueous solutions of ethylene glycol (EG) was investigated by differential scanning calorimetry at several pH values in the acidic region. The calorimetric enthalpies of denaturation, ΔHd, of these proteins increased monotonously with increasing the EG concentration under all the conditions studied. These proteins, however, differed from one another in the variation of the denaturation temperature, Td, with the addition of EG. The Td of chemotrypsinogen decreased slightly with an increase in EG concentration under all the conditions, producing a greater effect at high pH. The Td of RNase A, on the other hand, was almost independent of the EG content at pH 3.8, while at pH 2 and 3, the Td increased slightly with increasing the EG concentration and decreased slightly at pH 5. That is, EG stabilized or unstabilized these proteins against thermal denaturation, in a way which seemed to be dependent on temperature rather than the pH. The standard thermodynamic parameters for denaturation, ΔG°, ΔH°, and ΔS°, which were calculated using Td and ΔH° and assuming a constant heat capacity change, suggested that the slight increase or decrease in ΔG° induced by EG is attributed to whether ΔH° increases as much as ΔS° does or not. These results may be explained on the basis of the effect of EG on the solvation around hydrophobic groups of the protein. The EG effect may depends on the structural properties of proteins such as hydrophobicity and polarity, as well as on temperature.