Abstract
Copper-radical oxidases (CROs) catalyze the two-electron oxidation of a large number of primary alcohols including carbohydrates, polyols and benzylic alcohols as well as aldehydes and α-hydroxy-carbonyl compounds while reducing molecular oxygen to hydrogen peroxide. Initially, CROs like galactose oxidase and glyoxal oxidase were identified only in fungal secretomes. Since the last decade, their representatives have also been identified in some bacteria. CROs are grouped in the AA5 family of "auxiliary activities" in the database of Carbohydrate-Active enzymes. Despite low overall sequence similarity and different substrate specificities, sequence alignments and the solved crystal structures revealed a conserved architecture of the active sites in all CROs, with a mononuclear copper ion coordinated to an axial tyrosine, two histidines, and a cross-linked cysteine-tyrosyl radical cofactor. This unique post-translationally modified protein cofactor has attracted much attention in the past, which resulted in a large number of reports that shed light on key steps of the catalytic cycle and physico-chemical properties of CROs. Thanks to their broad substrate spectrum accompanied by the only need for molecular oxygen for catalysis, CROs since recently experience a renaissance and have been applied in various biocatalytic processes. This review provides an overview of the structural features, catalytic mechanism and substrates of CROs, presents an update on the engineering of these enzymes to improve their expression in recombinant hosts and to enhance their activity, and describes their potential fields of biotechnological application.