NDLI: Oxidation of Methoxyphenethylamines by Cytochrome P450 2D6 ANALYSIS OF RATE-LIMITING STEPS

Content Provider	Journal of Biological Chemistry (JBC)
Author	Guengerich, F. Peter Miller, Grover P. Hanna, Imad H. Sato, Hideaki Martin, Martha V.
Description	Microsomal cytochrome P450 (also termed heme-thiolate protein by the Enzyme Commission, EC 1.14.14.1 (1) (P450) Mass spectrometry. The conventions DV =Hkcat/Dkcatand D(V/K) =H(kcat/Km)/D(kcat/Km) of Northrop (2,3) are used in the designation of kinetic hydrogen isotope effects (MS)
Abstract	Cytochrome P450 (P450) 2D6 is involved in the oxidation of a large fraction (∼30%) of drugs used by humans and also catalyzes the O-demethylation of the model substrates 3- and 4-methoxyphenethylamine followed by subsequent ring hydroxylation to dopamine. Burst kinetics were not observed; rate-limiting step(s) must occur prior to product formation. Rates of reduction of ferric P450 2D6 were stimulated by 3- or 4-methoxyphenethylamine or the inhibitor quinidine; reduction is not the most rate-limiting step. The non-competitive intramolecular deuterium isotope effect, an estimate of the intrinsic isotope effect, for 4-methoxyphenethylamine O-demethylation was 9.6. Intermolecular non-competitive deuterium isotope effects of 3.1–3.8 were measured for k cat andk cat/K m for bothO-demethylation reactions, implicating at least partially rate-limiting C–H bond breaking. Simulation of steady-state kinetic data yielded a catalytic mechanism dominated by the rates of (i) Fe2+O protonation (plus O–O bond scission) and (ii) C–H bond breaking, consistent with the appearance of the spectral intermediates in the steady state, attributed to iron–oxygen complexes. However, all the rates of individual steps (or rates of combined steps) are considerably higher thank cat, and the contributions of several steps must be considered in understanding rates of the P450 2D6 reactions.
Related Links	http://www.jbc.org/content/277/37/33711.abstract
Ending Page	33719
Starting Page	33711
Page Count	9
File Format	HTM / HTML PDF
ISSN	00219258
Journal	Journal of Biological Chemistry (JBC)
Issue Number	37
Volume Number	277
DOI	10.1074/jbc.M205146200
e-ISSN	1083351X
Language	English
Publisher	American Society for Biochemistry and Molecular Biology
Publisher Date	2002-09-13
Access Restriction	Open
Subject Keyword	Tetrahydrofuran (THF) High performance liquid chromatography (HPLC) ENZYME CATALYSIS AND REGULATION
Alternative Title	Oxidation of Methoxyphenethylamines by Cytochrome P450 2D6
Content Type	Text
Resource Type	Article
Subject	Cell Biology Biochemistry Molecular Biology

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