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Title: Pyranose 2-oxidase from Phanerochaete chrysosporium-Expression in E. coli and biochemical characterization

Source: Journal of Biotechnology 142(2009)97-106; 2009

Author(s)Pisanelli, Ines; Kujawa, Magdalena; Spadiut, Oliver; Kittl, Roman; Halada, Petr; Volc, Jindrich; Mozuch, Michael D.; Kersten, Philip; Haltrich, dietmar, Peterbauer, Clemens

Publication Year: 2009  View PDF »

Category: Journal Articles
Associated Research Project(s):   FPL-4712-3A

Abstract: The presented work reports the isolation and heterologous expression of the p2ox gene encoding the flavoprotein pyranose 2-oxidase (P2Ox) from the basidiomycete Phanerochaete chrysosporium. The p2ox cDNAwas insertedinto the bacterial expression vector pET21a(+) and successfully expressed in Escherichia coli.We obtained active, fully flavinylated recombinant P2Ox in yields of approximately 270 mg/lmedium. The recombinant enzyme was provided with an N-terminal T7-tag and a C-terminal His6-tag to facilitate simple one-step purification.We obtained an apparently homogenous enzyme preparation with a specific activity of 16.5 U/mg. Recombinant P2Ox from P. chrysosporium was characterized in some detail with respect to its physical and catalytic properties, both for electron donor (sugar substrates) and - for the first time - alternative electron acceptors (1,4-benzoquinone, substituted quinones, 2,6-dichloroindophenol and ferricenium ion). As judged from the catalytic efficiencies kcat/Km, some of these alternative electron acceptors are better substrates than oxygen, which might have implications for the proposed in vivo function of pyranose 2-oxidase.

Keywords: Pyranose 2-oxidase, Phanerochaete chrysosporium, lignocellulose degradation, heterologous expression, lignocellulose, biodegradation, enzymes, Basidiomycetes, molecular genetics, fungi, genetics, wood-decaying fung, industrial applications, gene expression, biotechnology, pyranose, Escherichia coli, Bacteria, flavoproteins, recombinant DNA, sugars, microbial metabolism, chemical kinetics, oxygen

Publication Review Process: Informally Refereed (Peer-Reviewed)

File size: 727 kb(s)

Date posted: 02/09/2010

This publication is also viewable on Treesearch:  view
RITS Product ID: 31951
Current FPL Scientist associated with this product
Kersten, Philip J.
Research Microbiologist

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