The effect of iron to manganese substitution on microperoxidase 8 catalysed peroxidase and cytochrome P450 type of catalysis

This study describes the catalytic properties of manganese microperoxidase 8 [Mn(III)MP8] compared to iron microperoxidase 8 [Fe(III)MP8]. The mini-en zymes were tested for pH-dependent activity and operational stability in pe roxidase-type conversions, using 2-methoxyphenol and 3,3'-dimethoxybenzidin e, and in a cytochrome P450-like oxygen transfer reaction converting:anilin e to pam-aminophenol. For the peroxidase type of conversions the Fe to Mn r eplacement resulted in a less than 10-fold decrease in the activity at opti mal pH, whereas the aniline para-hydroxylation is reduced at least 30-fold. In addition it was observed that the peroxidase type of conversions are al l fully blocked by ascorbate and that aniline parahydroxylation by Fe(III)M P8 is increased by ascorbate whereas aniline para-hydroxylation by Mn(III)M P8 is inhibited by ascorbate. Altogether these results indicate that differ ent types of reactive metal oxygen intermediates are involved in the variou s conversions. Compound I/II, scavenged by ascorbate, may be the reactive s pecies responsible for the peroxidase reactions, the polymerization of anil ine and (part of) the oxygen transfer to aniline in the absence of ascorbat e. The para-hydroxylation of aniline by Fe(III)MP8, in the presence of asco rbate, must be mediated by another reactive iron-ore species which could be the electrophilic metal(III) hydroperoxide anion of microperoxidase 8 [M(I II)OOH MP8]. The lower oxidative potential of Mn, compared to Fe, may affec t the reactivity of both compound I/II and the metal(III) hydroperoxide ani on intermediate, explaining the differential effect of the Fe to Mn substit ution on the pH-dependent behavior, the rate of catalysis and the operation al stability of MP8.