KINETICS OF THERMAL INACTIVATION OF HORSERADISH-PEROXIDASE - STABILIZING EFFECT OF METHOXYPOLY(ETHYLENE GLYCOL)

The horseradish peroxidase has been modified by reaction of methoxypol y(ethylene glycol) (molecular mass 5000 Da) with external lysine resid ues, The thermal stability and the sensitivity to pH of the native and the modified horseradish peroxidase have been analysed over a tempera ture range of 40-80 degrees C and from pH 4.0-12.0, The effect of the addition of free methoxypoly(ethylene glycol) has also been investigat ed. The enzyme inactivation was modelled according to a single-step in activation scheme leading to a final steady-state residual activity, B oth the covalent modification by methoxypoly(ethylene glycol) and the addition of the free polymer ire the medium improved the stability of the enzyme towards pH, Similarly, the activation energy values calcula ted from Arrhenius plots showed an increased thermostability of the pe roxidase, In all instances, stabilization of horseradish peroxidase ap peared to occur both by suppressing initial unfolding and by lessening the extent of inactivation of the final enzyme form, The enhanced sta bility of the methoxypoly(ethylene glycol)-modified horseradish peroxi dase compared to that of the native enzyme was interpreted as due main ly to a decrease in the positive net charge of protein.