Inactivation and aggregation of R-specific 1-(4-hydroxyphenyl)-ethanol dehydrogenase from Aromatoleum aromaticum

R-specific 1-(4-hydroxyphenyl)-ethanol dehydrogenase (R-HPED) is a promising biotool for stereoselective synthesis of chiral aromatic alcohols. This work focused on the evaluation of its stability under storage and in-process conditions in the pH range from 5.5 to 8.5. The relationship between the dynamics of aggregation and activity loss under various pH conditions and in the presence of glucose, serving as a stabilizer, was analysed using spectrophotometric techniques and dynamic light scattering. pH 8.5 was indicated as a representative environment in which the enzyme, despite relatively low activity, shows high stability and the highest total product yield. Based on a series of inactivation experiments, the mechanism of thermal inactivation at pH 8.5 was modelled. The irreversible first-order mechanism of R-HPED inactivation in the temperature range of 47.5–60 °C was verified by isothermal and multi-temperature evaluation of data, confirming that in the alkaline pH 8.5, R-HPED aggregation is the secondary process occurring at already inactivated protein molecules. The rate constants were from 0.029 min−1 to 0.380 min−1 for a buffer solution but they decreased to 0.011 min−1 and 0.161 min−1, respectively, when 1.5 M glucose was added as a stabilizer. The activation energy was however about 200 kJ mol−1 in both cases. •R-specific 1-(4-hydroxyphenyl)-ethanol dehydrogenase stability was investigated.•Dynamics of aggregation and activity loss was analysed by different techniques.•The mechanism of thermal inactivation at pH 8.5 was modelled.•The irreversible first-order mechanism of inactivation was verified at 47.5–60 °C.