Unspecific peroxygenases (UPOs) have attracted significant attention as biocatalysts due to their ability to function without expensive cofactors. In this study, the UPO from aspergillus brasiliensis (AbrUPO) is found to catalyze the aromatic hydroxylation of substituted benzenes, a feature that distinguishes AbrUPO from other reported wild-type UPOs which contain more phenylalanine residues in the active site and substrate access channel and primarily catalyze benzylic hydroxylation. A group of researchers from the Institute of Biochemistry, the Department of Biotechnology at Delft University of Technology and Faculty of Agriculture at Kindai University have constructed a number of variants with specific amino acid substitutions that mimic the active sites or substrate access channels of other UPOs to elucidate the differences in product distribution.
The publication recently appeared in ChemBioChem. Below you can read the abstract. The link to the whole publication can be found in the reference at the bottom.
“Heme-containing unspecific peroxygenases (UPOs) have attracted significant attention as biocatalysts for oxidation reactions due to their ability to function without expensive nicotinamide cofactors. In the recent study, the UPO from aspergillus brasiliensis (AbrUPO) is found to catalyze the aromatic hydroxylation of substituted benzenes, a feature that distinguishes AbrUPO from other reported wild-type UPOs. To elucidate the underlying factors in the active site and substrate access channel of AbrUPO—which contains fewer phenylalanine residues compared to other UPOs that primarily catalyze benzylic hydroxylation—twenty two AbrUPO variants with single, double, triple, or quadruple amino acid substitutions were constructed to mimic the active sites or substrate access channels of other UPOs. A number of mutated variants exhibited altered activity and selectivity, and several positions were identified that influence enzyme chemoselectivity. Among them, substitution of alanine at position 186 with bulkier residues such as phenylalanine or leucine lead to a shift in chemoselectivity toward alkyl chain hydroxylation of substituted benzenes. Molecular docking studies indicated that the A186F mutation restricts the flexibility and reorientation of ethylbenzene in the active site of AbrUPO, thereby preventing oxidation at the aromatic ring while promoting benzylic hydroxylation.”
Schmitz F, Hoffrogge M, Koschorreck K, Fukuta Y, Raffaele A, Tieves F, Hilberath T,
Hollmann F, Urlacher VB, 2025, Identification of Key Active-Site Positions Controlling the Chemoselectivity of Aspergillus Brasiliensis Unspecific Peroxygenase, ChemBioChem, https://doi.org/10.1002/cbic.202500181
© 2025 The Author(s). ChemBioChem published by Wiley-VCH GmbH.