title: An aryl-alcohol oxidase of Pleurotus sapidus: heterologous expression, characterization, and application in a 2-enzyme system.
abstract: Aryl-alcohol oxidases (AAOs) are enzymes supporting the degradation of lignin by fungal derived class II peroxidases produced by white-rot fungi. AAOs are able to generate H2O2 as a by-product via oxidation of an aryl-alcohol into its correspondent aldehyde. In this study, an AAO was heterologously expressed in a basidiomycete host for the first time. The gene for an AAO of the white-rot fungus Pleurotus sapidus, a close relative to the oyster mushroom Pleurotus ostreatus, was cloned into an expression vector and put under control of the promotor of the glyceraldehyde-3-phosphate dehydrogenase gene 2 (gpdII) of the button mushroom Agaricus bisporus. The expression vector was transformed into the model basidiomycete Coprinopsis cinerea, and several positive transformants were obtained. The best producing transformants were grown in shake-flasks and in a stirred tank reactor reaching enzymatic activities of up to 125 U L(-1) using veratryl alcohol as a substrate. The purified AAO was biochemically characterized and compared to the previously described native and recombinant AAOs from other Pleurotus species. In addition, a two-enzyme system comprising a dye-decolorizing peroxidase (DyP) from Mycetinis scorodonius and the P. sapidus AAO was successfully employed to bleach the anthraquinone dye Reactive Blue 5.
authors.affiliation: Institute of Food Chemistry and Food Biotechnology, Justus Liebig University Giessen, Heinrich-Buff-Ring 17, 35392, Giessen, Germany.
authors.forename: Ilya
authors.initials: I
authors.lastname: Galperin
authors.affiliation: Institute of Food Chemistry and Food Biotechnology, Justus Liebig University Giessen, Heinrich-Buff-Ring 17, 35392, Giessen, Germany.
authors.forename: Aysha
authors.initials: A
authors.lastname: Javeed
authors.affiliation: Institute of Food Chemistry and Food Biotechnology, Justus Liebig University Giessen, Heinrich-Buff-Ring 17, 35392, Giessen, Germany.
authors.forename: Hanno
authors.initials: H
authors.lastname: Luig
authors.affiliation: Institute of Biochemistry, Justus Liebig University Giessen, Friedrichstrasse 24, 35392, Giessen, Germany.
authors.forename: Günter
authors.initials: G
authors.lastname: Lochnit
authors.affiliation: Institute of Food Chemistry and Food Biotechnology, Justus Liebig University Giessen, Heinrich-Buff-Ring 17, 35392, Giessen, Germany. martin.ruehl@lcb.chemie.uni-giessen.de.
authors.forename: Martin
authors.initials: M
authors.lastname: Rühl
mesh_terms: D000363:Agaricales / Q000235:genetics / Q000378:metabolism; D000429:Alcohol Oxidoreductases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D000880:Anthraquinones / Q000378:metabolism; D003001:Cloning, Molecular; D015870:Gene Expression; D009195:Peroxidase / Q000378:metabolism; D020076:Pleurotus / Q000201:enzymology* / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D000880:Anthraquinones; C544286:Reactive Blue 5; D011994:Recombinant Proteins; D000429:Alcohol Oxidoreductases; C077759:aryl-alcohol oxidase; D009195:Peroxidase
keywords: Aryl-alcohol oxidase; Coprinopsis cinerea; Dyp-type peroxidase; Enzyme kinetics; Heterologous expression; White-rot fungi