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Wild-type CYP102A1 as a biocatalyst: turnover of drugs usually metabolised by human liver enzymes

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Abstract

This work provides functional data showing that the bacterial CYP102A1 recognises compounds metabolised by human CYP3A4, CYP2E1 and CYP1A2 and is able to catalyse different reactions. Wild-type cytochrome CYP102A1 from Bacillus megaterium is a catalytically self-sufficient enzyme, containing an NADPH-dependent reductase and a P450 haem domain fused in a single polypeptidie chain. An NADPH-dependent method (Tsotsou et al. in Biosens. Bioelectron. 17:119–131, 2002) together with spectroscopic assays were applied to investigate the catalytic activity of CYP102A1 towards 19 xenobiotics, including 17 commercial drugs. These molecules were chosen to represent typical substrates of the five main families of drug-metabolising human cytochromes P450. Liquid chromatography–mass spectrometry analysis showed that CYP102A1 catalyses the hydroxylation of chlorzoxazone, aniline and p-nitrophenol, as well as the N-dealkylation of propranolol and the dehydrogenation of nifedipine. These drugs are typical substrates of human CYP2E1 and CYP3A4. The K M values calculated for these compounds were in the millimolar range: 1.21 ± 0.07 mM for chlorzoxazone, 2.52 ± 0.08 mM for aniline, 0.81 ± 0.04 mM for propranolol. The values of v max for chlorzoxazone and propranolol were 46.0 ± 9.0 and 7.6 ± 3.4 nmol min−1 nmol−1, respectively. These values are higher then those measured for the human enzymes. The v max value for aniline was 9.4 ± 1.3 nmol min−1 nmol−1, comparable to that calculated for human cytochromes P450. The functional data were found to be in line with the sequence alignments, showing that the identity percentage of CYP102A1 with CYP3A4 and CYP2E1 is higher than that found for CYP1A2, CYP2C9 and CYP2D6 families.

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Acknowledgements

G.D.N. and G.G. gratefully acknowledge the ISI foundation for a Lagrange postdoctoral fellowship and Programma di Ricareca di Interesse Nazionale (PRIN) for financial support. Thanks are due to Graham Taylor for help with MS experiments.

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Authors and Affiliations

  1. Department of Human and Animal Biology, University of Turin, via Accademia Albertina 13, 10123, Turin, Italy

    Giovanna Di Nardo, Paola Panicco, Carlo Sassone, Carlo Giunta & Gianfranco Gilardi

  2. Division of Molecular Biosciences, Imperial College London, London, UK

    Andrea Fantuzzi, Anastasia Sideri & Gianfranco Gilardi

  3. NanoBioDesign Ltd, London, UK

    Andrea Fantuzzi, Paola Panicco, Carlo Sassone & Gianfranco Gilardi

Authors
  1. Giovanna Di Nardo
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  2. Andrea Fantuzzi
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  3. Anastasia Sideri
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  4. Paola Panicco
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  5. Carlo Sassone
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  6. Carlo Giunta
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  7. Gianfranco Gilardi
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Correspondence to Gianfranco Gilardi.

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Di Nardo, G., Fantuzzi, A., Sideri, A. et al. Wild-type CYP102A1 as a biocatalyst: turnover of drugs usually metabolised by human liver enzymes. J Biol Inorg Chem 12, 313–323 (2007). https://doi.org/10.1007/s00775-006-0188-4

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  • DOI: https://doi.org/10.1007/s00775-006-0188-4

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