HYDROGEN PEROXIDE-DEPENDENT 4-T-BUTYLPHENOL HYDROXYLATION BY TYROSINASE - A NEW CATALYTIC ACTIVITY

The aim of this work was to study the hydroxylation by tyrosinase of 4 -t-butylphenol to 4-t-butylcatechol, in the presence of hydrogen perox ide. This hydroxylation reaction does not take place without the addit ion of hydrogen peroxide. Some properties of this new hydroxylating ac tivity have been analysed. The kinetic parameters of mushroom tyrosina se for hydrogen peroxide (K-m = 4.9 mM, V-m = 48.1 mu M/min) and 4-t-b utylphenol (K-m = 16 mu M/min, V-m = 6.7 mu M/min) were evaluated. A l ag period appeared, which was similar to the characteristic lag of mon ophenolase activity at the expense of molecular oxygen. The length of the lag phase decreased with increasing hydrogen peroxide concentratio ns but was longer with higher 4-t-butylphenol concentrations. The pH o ptimum for this hydroxylating activity was close to 5.5, The lag also varied with pH, reaching its highest value at pH 4.8. The lag was shor tened by the addition of increasing amounts of 4-t-butylcatechol, and was abolished at 24.5 mu M of 4-t-butylcatechol. 4-t-Butylphenol was o xidized by mushroom tyrosinase in the presence of 24.5 mu M 4-t-butylc atechol and in the absence of hydrogen peroxide although the enzymatic activity tailed off. The presence of hydrogen peroxide is necessary t o maintain a constant steady-state rate of 4-t-butylphenol oxidation b y tyrosinase.