THERMOSTABILITY OF RIBONUCLEASE-A IN ORGANIC-SOLVENTS - A CALORIMETRIC AND SPECTROSCOPIC STUDY

The enhancement of ribonuclease A (RNAase) thermostability in water-im miscible organic solvents with respect to water has been studied by di fferential scanning calorimetry (DSC) and IR spectroscopy. RNAase has been adsorbed onto an inert support (Celite) in order to avoid aggrega tion effects. The dependence on hydration of the thermodynamic paramet ers of unfolding, such as the enthalpy change, Delta H, and the transi tion temperature, Tm, have been studied in the absence of any solvent and in the presence of selected water-immiscible organic solvents. In both cases, RNAase shows a progressive enhancement of the thermal stab ility as the water content is lowered. In highly hydrophobic solvents, RNAase stability does not change significantly with respect to the no -solvent state at the same hydration level. More hydrophilic solvents enhance stability by further dehydrating the protein molecule. A corre lation has been found between the protein stability parameters (Delta H and Tm) and the dielectric constant of the bulk solvent, suggesting that electrostatic interactions may play a relevant role in protein st ability in aqueous solution. A poorer correlation was found with solve nt hydrophobicity indexes, such as log P. The presence of organic solv ent perturbs the IR spectra of the denatured RNAase more significantly than the corresponding spectra of the thermally native protein.