Extrinsic protein stabilization by the naturally occurring osmolytes β-hydroxyectoine and betaine

Abstract

Thermodynamic aspects of protein stabilization by two widespread naturally occurring osmolytes, β-hydroxyectoine and betaine, were studied using differential scanning calorimetry (DSC) and bovine ribonuclease A (RNase A) as a model protein. The osmolyte β-hydroxyectoine purified from Marinococcus was found to be a very efficient stabilizer. At a concentration of 3 M it increased the melting temperature of RNase A (T _m ) by more than 12 K and gave rise to a stability increase of 10.6 kJ/mol at room temperature. The heat capacity difference between the folded and unfolded state (ΔC _p ) was found to be significantly increased. Betaine stabilized RNase A only at concentrations less than 3 M. Also, here ΔC _p was found to be increased. Calculation of the number of water molecules that additionally bind to unfolded RNase A resulted in surprisingly low numbers for both osmolytes. The significant stabilization of RNase A by β-hydroxyectoine makes this osmolyte an interesting stabilizer in biotechnological processes in which enzymes are applied in the presence of denaturants or at high temperature.