Role of the conserved threonine 309 in mechanism of oxidation by cytochrome P450 2D6. Biochem Biophys Res Commun 2005 Dec 16;338(2):1065-74
Date
11/05/2005Pubmed ID
16269134DOI
10.1016/j.bbrc.2005.10.066Scopus ID
2-s2.0-27744459498 (requires institutional sign-in at Scopus site) 35 CitationsAbstract
Based on sequence alignments and homology modeling, threonine 309 in cytochrome P450 2D6 (CYP2D6) is proposed to be the conserved I-helix threonine, which is supposed to be involved in dioxygen activation by CYPs. The T309V mutant of CYP2D6 displayed a strong shift from O-dealkylation to N-dealkylation reactions in oxidation of dextromethorphan and 3,4-methylenedioxymethylamphetamine. This may be explained by an elevated ratio of hydroperoxo-iron to oxenoid-iron of the oxygenating species. In consistence, using cumene hydroperoxide, which directly forms the oxenoid-iron, the T309V mutant again selectively catalyzed the O-dealkylation reactions. The changed ratio of oxygenating species can also explain the decreased activity and changed regioselectivity that were observed in 7-methoxy-4-(aminomethyl)-coumarin and bufuralol oxidation, respectively, by the T309V mutant. Interestingly, the T309V mutant always showed a significantly increased, up to 75-fold, higher activity compared to that of the wild-type when using cumene hydroperoxide. These results indicate that T309 in CYP2D6 is involved in maintaining the balance of multiple oxygenating species and thus influences substrate and regioselectivity.
Author List
Keizers PH, Schraven LH, de Graaf C, Hidestrand M, Ingelman-Sundberg M, van Dijk BR, Vermeulen NP, Commandeur JNMESH terms used to index this publication - Major topics in bold
Amino Acid SubstitutionCytochrome P-450 CYP2D6
Enzyme Activation
Escherichia coli
Mutagenesis, Site-Directed
Oxidation-Reduction
Structure-Activity Relationship
Substrate Specificity
Threonine
