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Structural insights into xenobiotic and inhibitor binding to human aldehyde oxidase

Nature Chemical Biology volume 11, pages 779–783 (2015)Cite this article

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Abstract

Aldehyde oxidase (AOX) is a xanthine oxidase (XO)-related enzyme with emerging importance due to its role in the metabolism of drugs and xenobiotics. We report the first crystal structures of human AOX1, substrate free (2.6-Å resolution) and in complex with the substrate phthalazine and the inhibitor thioridazine (2.7-Å resolution). Analysis of the protein active site combined with steady-state kinetic studies highlight the unique features, including binding and substrate orientation at the active site, that characterize human AOX1 as an important drug-metabolizing enzyme. Structural analysis of the complex with the noncompetitive inhibitor thioridazine revealed a new, unexpected and fully occupied inhibitor-binding site that is structurally conserved among mammalian AOXs and XO. The new structural insights into the catalytic and inhibition mechanisms of human AOX that we now report will be of great value for the rational analysis of clinical drug interactions involving inhibition of AOX1 and for the prediction and design of AOX-stable putative drugs.

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Figure 1: Surface representation of the hAOX1 crystal structure homodimer.
Figure 2: Close up of the hAOX1-free (green), hAOX1–Pht–Thi (blue) and mAOX3 (gray) active sites and their surrounding residues.
Figure 3: Binding site of the noncompetitive inhibitor thioridazine and localization of the two gates that determine substrate (phthalazine) access.

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Acknowledgements

This work was financially supported by the Fundação para a Ciência e Tecnologia (FCT-MEC) through projects UID/Multi/04378/2013, EXCL/QEQ-COM/0394/2012, PTDC/BIA-PRO/118377/2010 (M.J.R., T.S.-S., C.C.), SFRH/BPD/84581/2012 (C.C.) and DAAD-441.00 (M.J.R., T.S.-S., S.L.) and by Deutsche Forschungsgemeinschaft Grant Le1171/8-1 (S.L.). We thank the I02 staff of the Diamond Light Source (DLS, Didcot, United Kingdom) and X06DA-PXIII staff from the Swiss Light Source (SLS, Paul Scherrer Institut, Villigen, Switzerland) for assistance during data collection. We also thank the staff from beamlines ID14-1, ID29-1 and ID23-1 from the European Synchrotron Radiation Facility (ESRF, Grenoble, France). The research leading to these results has received funding from the European Community's Seventh Framework Programme (FP7/2007-2013) under BioStruct-X (grant agreement no. 283570). We thank R. Hille (University of California, Riverside) for providing bXO for the inhibition experiments and A. Palma (Universidade Nova de Lisboa, Portugal) for critical reading of the manuscript.

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Authors and Affiliations

  1. Departamento de Química, Research Unit on Applied Molecular Biosciences–Rede de Química e Tecnologia (UCIBIO-REQUIMTE), Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, Caparica, Portugal

    Catarina Coelho, Teresa Santos-Silva & Maria João Romão

  2. Department of Molecular Enzymology, Institute of Biochemistry and Biology, University of Potsdam, Potsdam, Germany

    Alessandro Foti, Tobias Hartmann & Silke Leimkühler

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  1. Catarina Coelho
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Contributions

C.C., T.S.-S. and M.J.R. conceived and designed the crystallization experiments and performed 3D structure determination. A.F., T.H. and S.L. purified the protein. A.F. and S.L. conducted kinetic experiments. C.C., A.F., T.S.-S., S.L. and M.J.R. performed data analysis. C.C., T.S.-S., S.L. and M.J.R. wrote the manuscript.

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Correspondence to Maria João Romão.

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Supplementary Results, Supplementary Tables 1 and 2 and Supplementary Figures 1–5. (PDF 1884 kb)

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Coelho, C., Foti, A., Hartmann, T. et al. Structural insights into xenobiotic and inhibitor binding to human aldehyde oxidase. Nat Chem Biol 11, 779–783 (2015). https://doi.org/10.1038/nchembio.1895

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