title: Human FMO2-based microbial whole-cell catalysts for drug metabolite synthesis.
abstract: BACKGROUND
  Getting access to authentic human drug metabolites is an important issue during the drug discovery and development process. Employing recombinant microorganisms as whole-cell biocatalysts constitutes an elegant alternative to organic synthesis to produce these compounds. The present work aimed for the generation of an efficient whole-cell catalyst based on the flavin monooxygenase isoform 2 (FMO2), which is part of the human phase I metabolism.
  
  
  RESULTS
  We show for the first time the functional expression of human FMO2 in E. coli. Truncations of the C-terminal membrane anchor region did not result in soluble FMO2 protein, but had a significant effect on levels of recombinant protein. The FMO2 biocatalysts were employed for substrate screening purposes, revealing trifluoperazine and propranolol as FMO2 substrates. Biomass cultivation on the 100 L scale afforded active catalyst for biotransformations on preparative scale. The whole-cell conversion of trifluoperazine resulted in perfectly selective oxidation to 48 mg (46% yield) of the corresponding N (1)-oxide with a purity >98%.
  
  
  CONCLUSIONS
  The generated FMO2 whole-cell catalysts are not only useful as screening tool for human metabolites of drug molecules but more importantly also for their chemo- and regioselective preparation on the multi-milligram scale.
authors.affiliation: acib GmbH c/o Institute of Molecular Biotechnology, Graz University of Technology, NAWI Graz, Petersgasse 14, 8010, Graz, Austria. martina.geier@acib.at.
authors.forename: Martina
authors.initials: M
authors.lastname: Geier
authors.affiliation: acib GmbH c/o Institute of Molecular Biotechnology, Graz University of Technology, NAWI Graz, Petersgasse 14, 8010, Graz, Austria. thorsten.bachler@acib.at.
authors.forename: Thorsten
authors.initials: T
authors.lastname: Bachler
authors.affiliation: F. Hoffmann-La Roche Ltd., 4070, Basel, Switzerland. steven_paul.hanlon@roche.com.
authors.forename: Steven P
authors.initials: SP
authors.lastname: Hanlon
authors.affiliation: Novartis Pharma AG, 4002, Basel, Switzerland. fabian_kurt.eggimann@novartis.com.
authors.forename: Fabian K
authors.initials: FK
authors.lastname: Eggimann
authors.affiliation: Novartis Pharma AG, 4002, Basel, Switzerland. matthias.kittelmann@novartis.com.
authors.forename: Matthias
authors.initials: M
authors.lastname: Kittelmann
authors.affiliation: Institute of Organic Chemistry, Graz University of Technology, NAWI Graz, Stremayrgasse 9, 8010, Graz, Austria. hansjoerg.weber@tugraz.at.
authors.forename: Hansjörg
authors.initials: H
authors.lastname: Weber
authors.affiliation: Novartis Pharma AG, 4002, Basel, Switzerland. stephan.luetz@novartis.com.
authors.forename: Stephan
authors.initials: S
authors.lastname: Lütz
authors.affiliation: F. Hoffmann-La Roche Ltd., 4070, Basel, Switzerland. beat.wirz@roche.com.
authors.forename: Beat
authors.initials: B
authors.lastname: Wirz
authors.affiliation: acib GmbH c/o Institute of Molecular Biotechnology, Graz University of Technology, NAWI Graz, Petersgasse 14, 8010, Graz, Austria. margit.winkler@acib.at.
authors.forename: Margit
authors.initials: M
authors.lastname: Winkler
mesh_terms: D055162:Biocatalysis; D004138:Dinitrocresols / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D006899:Mixed Function Oxygenases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004364:Pharmaceutical Preparations / Q000378:metabolism*; D011433:Propranolol / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity; D014268:Trifluoperazine / Q000378:metabolism
chemical_list: D004138:Dinitrocresols; D004364:Pharmaceutical Preparations; D011994:Recombinant Proteins; C024132:4,6-dinitro-o-cresol; D014268:Trifluoperazine; D011433:Propranolol; D006899:Mixed Function Oxygenases