Extracellular Oxidase from the Neonothopanus nambi Fungus as a Promising Enzyme for Analytical Applications

The extracellular enzyme with oxidase function was extracted from the Neonothopanus nambi luminescent fungus by using mild processing of mycelium with β-glucosidase and then isolated by gel-filtration chromatography. The extracted enzyme is found to be a FAD-containing protein, catalyzing phenol co-oxidation with 4-aminoantipyrine without addition of H₂O₂, which distinguishes it from peroxidases. This fact allowed us to assume that this enzyme may be a mixed-function oxidase. According to gel-filtration chromatography and SDS-PAGE, the oxidase has molecular weight of 60 kDa. The enzyme exhibits maximum activity at 55-70 °C and pH 5.0. Kinetic parameters K_m and V_max of the oxidase for phenol were 0.21 mM and 0.40 µM min^-1. We suggest that the extracted enzyme can be useful to develop a simplified biosensor for colorimetric detection of phenol in aqueous media, which does not require using hydrogen peroxide.