A Fungal Ascorbate Oxidase with Unexpected Laccase Activity

Ascorbate oxidases are an enzyme group that has not been explored to a large extent. So far, mainly ascorbate oxidases from plants and only a few from fungi have been described. Although ascorbate oxidases belong to the well-studied enzyme family of multi-copper oxidases, their function is still unclear. In this study, _AO1, an enzyme from the fungus , was characterized. Sequence analyses and copper content determination demonstrated _AO1 to belong to the multi-copper oxidase family. Biochemical characterization and 3D-modeling revealed a similarity to ascorbate oxidases, but also to laccases. _AO1 had a 10-fold higher affinity to ascorbic acid ( = 0.16 ± 0.03 mM) than to ABTS ( = 1.89 ± 0.12 mM). Furthermore, the best fitting 3D-model was based on the ascorbate oxidase from var. . The laccase-like activity of _AO1 on ABTS ( = 11.56 ± 0.15 µM/min/mg) was, however, not negligible. On the other hand, other typical laccase substrates, such as syringaldezine and guaiacol, were not oxidized by _AO1. According to the biochemical and structural characterization, _AO1 was classified as ascorbate oxidase with unusual, laccase-like activity.