Binding of bufuralol, dextromethorphan and 3,4- methylenedioxymethylamphetamine to wild-type and F120A mutant cytochrome P450 2D6 studied by resonance Raman spectroscopy. Alois Bonifacio ‡ , Peter H.J.Keizers § , Jan N.M.Commandeur § , Nico P.E.Vermeulen § , Bruno Robert || , Cees Gooijer ‡ and Gert van der Zwan ‡* Laser Centre/Analytical Chemistry and Applied Spectroscopy, and LACDR/Molecular Toxicology - Department of Chemistry and Pharmaceutical Sciences, Vrije Universiteit Amsterdam, De Boelelaan 1083, 1081 HV Amsterdam, The Netherlands and Service de Biophysique des Fonctions Membranaires, DBJC/CEA & URA2096/CNRS, CEA- Saclay, 91191 Gif-sur-Yvette, Cedex, France Corresponding author: G. van der Zwan (zwan@few.vu.nl), Laser Centre/Analytical Chemistry and Applied Spectroscopy, Department of Chemistry and Pharmaceutical Sciences, Vrije Universiteit Amsterdam, De Boelelaan 1083, 1081 HV Amsterdam, The Netherlands, tel.0031 20 5987635, fax 0031 20 5987543 SHORT (PAGE HEADING) TITLE: Resonance Raman study of substrate-binding in P450 2D6 SYNOPSIS Cytochrome P450 2D6 (CYP2D6) is one of the most important drug-metabolizing enzymes in humans. Among others, it is involved in the metabolism of bufuralol, dextromethorphan and 3,4-methylenedioxymethylamphetamine (MDMA or ecstasy), which share similarity in having a basic N-atom and an adjacent aromatic moiety. Resonance Raman spectroscopy data, reported for the first time for CYP2D6, show that the CYP2D6 heme is found to be in a six-coordinated low-spin state in absence of substrates, and it is perturbed to different extents by the three compounds, despite their structural similarity. Dextromethorphan and MDMA induce in CYP2D6 a significant amount of five-coordinated high-spin heme species and reduce the polarity of its heme- pocket (as deduced by an upshift of the Fe-CO stretching frequency), whereas bufuralol ‡ Laser Centre/Analytical Chemistry and Applied Spectroscopy, Vrije Universiteit Amsterdam § LACDR/Molecular Toxicology, Vrije Universiteit Amsterdam || Service de Biophysique des Fonctions Membranaires, CEA/Saclay 1