Pyranose 2-oxidase from Phanerochaete chrysosporium

Abstract

Enzymes with glucose 2-oxidase activity, tentatively assigned as pyranose 2-oxidase have so far been isolated from mycelia cultures of two basidiomycetes, Polyporus obtusus and Trametes versicolor . The enzyme is also present in extracts of a number of other higher fungi and appears to play an important metabolic role. This chapter discusses assay method and purification procedure of the enzyme. Glucose 2-oxidase activity is measured using a modification of the chromogenic assay for peroxidase. The chapter also discusses the properties of the isolated glucose 2-oxidase. The purified enzyme has a specific activity of 15.5 U/mg proteins. It is stable at 4° in 30 mM sodium phosphate buffer, pH 6.5 and has a broad pH optimum with maximum activity at pH 7.5. The oxidized form of the enzyme exhibits absorption maxima at 360 and 457 nm. Reduction of the enzyme by the substrate glucose under nitrogen atmosphere, or by sodium dithionite, results in elimination of the absorption maximum at 457 nm. These spectral properties are characteristic of a flavoprotein.