Beschreibung:
<jats:title>Abstract</jats:title>
<jats:p>There is an increasing interest in the application of peroxygenases in biocatalysis, because of their ability to catalyse the oxyfunctionalisation reaction in a stereoselective fashion and with high catalytic efficiencies, while using hydrogen peroxide or organic peroxides as oxidant. However, enzymes belonging to this class exhibit a very low stability in the presence of peroxides. With the aim of bypassing this fast and irreversible inactivation, we study the use of a gradual supply of hydrogen peroxide to maintain its concentration at stoichiometric levels. In this contribution, we report a multienzymatic cascade for in situ generation of hydrogen peroxide. In the first step, in the presence of NAD<jats:sup>+</jats:sup> cofactor, formate dehydrogenase from <jats:italic>Candida boidinii</jats:italic> (FDH) catalysed the oxidation of formate yielding CO<jats:sub>2</jats:sub>. Reduced NADH was reoxidised by the reduction of the flavin mononucleotide cofactor bound to an old yellow enzyme homologue from <jats:italic>Bacillus subtilis</jats:italic> (YqjM), which subsequently reacts with molecular oxygen yielding hydrogen peroxide. Finally, this system was coupled to the hydroxylation of ethylbenzene reaction catalysed by an evolved peroxygenase from <jats:italic>Agrocybe aegerita</jats:italic> (r<jats:italic>Aae</jats:italic>UPO). Additionally, we studied the influence of different reaction parameters on the performance of the cascade with the aim of improving the turnover of the hydroxylation reaction.</jats:p>