title: Human xanthine oxidase recombinant in E. coli: A whole cell catalyst for preparative drug metabolite synthesis.
abstract: Human xanthine oxidoreductase (XOR), which is responsible for the final steps of the purine metabolism pathway and involved in oxidative drug metabolism, was successfully expressed in Escherichia coli BL21(DE3) Gold. Recombinant human (rh) XOR yielded higher productivity with the gene sequence optimized for expression in E.coli than with the native gene sequence. Induction of XOR expression with lactose or IPTG resulted in complete loss of activity whereas shake flasks cultures using media rather poor in nutrients resulted in functional XOR expression in the stationary phase. LB medium was used for a 25L fermentation in fed-batch mode, which led to a 5 fold increase of the enzyme productivity when compared to cultivation in shake flasks. Quinazoline was used as a substrate on the semi-preparative scale using an optimized whole cell biotransformation protocol, yielding 73mg of the isolated product, 4-quinazolinone, from 104mg of starting material.
authors.affiliation: Edifício da Unidade Piloto do IBET, Estação Agronómica Nacional, Avenida da República, 2780-157 Oeiras, Portugal.
authors.forename: Márcia
authors.initials: M
authors.lastname: Ferreira Antunes
authors.affiliation: Novartis Pharma AG, 4002 Basel, Switzerland.
authors.forename: Fabian Kurt
authors.initials: FK
authors.lastname: Eggimann
authors.affiliation: Novartis Pharma AG, 4002 Basel, Switzerland. Electronic address: matthias.kittelmann@novartis.com.
authors.forename: Matthias
authors.initials: M
authors.lastname: Kittelmann
authors.affiliation: Novartis Pharma AG, 4002 Basel, Switzerland.
authors.forename: Stephan
authors.initials: S
authors.lastname: Lütz
authors.affiliation: F. Hoffmann-La Roche Ltd., 4070 Basel, Switzerland.
authors.forename: Steven P
authors.initials: SP
authors.lastname: Hanlon
authors.affiliation: F. Hoffmann-La Roche Ltd., 4070 Basel, Switzerland.
authors.forename: Beat
authors.initials: B
authors.lastname: Wirz
authors.affiliation: acib GmbH c/o Institute of Molecular Biotechnology, Graz University of Technology, Petersgasse 14, 8010 Graz, Austria.
authors.forename: Thorsten
authors.initials: T
authors.lastname: Bachler
authors.affiliation: acib GmbH c/o Institute of Molecular Biotechnology, Graz University of Technology, Petersgasse 14, 8010 Graz, Austria. Electronic address: margit.winkler@acib.at.
authors.forename: Margit
authors.initials: M
authors.lastname: Winkler
mesh_terms: D001709:Biotechnology; D004926:Escherichia coli / Q000235:genetics*; D005285:Fermentation; D006801:Humans; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D014969:Xanthine Oxidase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D014969:Xanthine Oxidase
keywords: Drug metabolites; E. coli; Purine; Whole cell biotransformation; Xanthine oxidase