Purification and characterization of pyranose oxidase from the white rot fungus Trametes multicolor.
We purified an intracellular pyranose oxidase from mycelial extracts of the white rot fungus Trametes multicolor by using ammonium sulfate fractionation, hydrophobic interaction, ion-exchange chromatography, and gel filtration. The native enzyme has a molecular mass of 270 kDa as determined by equilibrium ultracentrifugation and is composed of four identical 68-kDa subunits as determined by matrix-assisted laser desorption ionization mass spectrometry. Each subunit contains one covalently bound flavin adenine dinucleotide as its prosthetic group. The enzyme oxidizes several aldopyranoses specifically at position C-2, and its preferred electron donor substrates are D-glucose, D-xylose, and L-sorbose. During this oxidation reaction electrons are transferred to oxygen, yielding hydrogen peroxide. In addition, the enzyme catalyzes the two-electron reduction of 1,4-benzoquinone, several substituted benzoquinones, and 2,6-dichloroindophenol, as well as the one-electron reduction of the ABTS [2,2'-azinobis(3-ethylbenzthiazolinesulfonic acid)] cation radical. As judged by the catalytic efficiencies (k(cat)/K(m)), some of these quinone electron acceptors are much better substrates for pyranose oxidase than oxygen. The optimum pH of the pyranose oxidase-catalyzed reaction depends strongly on the electron acceptor employed and varies from 4 to 8. It has been proposed that the main metabolic function of pyranose oxidase is as a constituent of the ligninolytic system of white rot fungi that provides peroxidases with H(2)O(2). An additional function could be reduction of quinones, key intermediates that are formed during mineralization of lignin.
Leitner
C
C
Division of Biochemical Engineering, Institute of Food Technology, University of Agricultural Sciences Vienna, A-1190 Vienna, Austria.
Volc
J
J
Haltrich
D
D
D000595:Amino Acid Sequence; D001487:Basidiomycota / Q000201:enzymology* / Q000254:growth & development; D002237:Carbohydrate Dehydrogenases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D013696:Temperature
D002237:Carbohydrate Dehydrogenases; C022489:pyranose oxidase; D005947:Glucose