Human FMO2-based microbial whole-cell catalysts for drug metabolite synthesis.

Fabian K Eggimann,Margit Winkler,Martina Geier,Matthias Kittelmann,Steven P Hanlon,Stephan Lütz,Hansjörg Weber,Thorsten Bachler,Beat Wirz

doi:10.1186/s12934-015-0262-0

Abstract

BackgroundGetting 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.ResultsWe 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 N1-oxide with a purity >98%.ConclusionsThe 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.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-015-0262-0) contains supplementary material, which is available to authorized users.

Highlights

Getting access to authentic human drug metabolites is an important issue during the drug discovery and development process
flavin monooxygenase isoform 2 (FMO2)*1 showed very little expression in E. coli BL21 compared to isoforms 3 and 5 [20]
Lidocaine, which is a diethyl amine, was not a substrate under the screening conditions used. Another compound, which we found to be metabolized by Human flavin monooxygenase 2 (hFMO2), was propranolol

Summary

Introduction

Getting access to authentic human drug metabolites is an important issue during the drug discovery and development process. 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. FMOs are involved in the phase 1 drug metabolism of compounds containing soft nucleophiles such as sulfur or nitrogen. The substrates typically described for human FMO2*1 are sulfur derived compounds such as thioureas [6, 7], thioetherorganophosphates [6], thiazetazone [8] and ethionamide [8, 9]. Rabbit FMO2 catalyzed the oxidation of prochlorperazine, desmethylperazine and trifluoperazine [10]. These are phenothiazine derived xenobiotics with antipsychotic effects

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