title: Mechanism-based inactivation of human CYP2E1 by diethyldithocarbamate.
abstract: Although the ability of disulfiram to inactivate CYP2E1 has been known for more than 20 years, the mechanism has not yet been elucidated. A metabolite of disulfiram, diethyldithocarbamate (DDC), is converted by CYP2E1 to a reactive intermediate that subsequently inactivates the protein, leading to mechanism-based inactivation. Mass spectral analysis of the inactivated human 2E1 protein demonstrates that the inactivation is due to the formation of an adduct of the reactive metabolite of DDC with the apoprotein. These data, along with mass spectral analysis of a reactive intermediate trapped with GSH, indicate the involvement of a reactive intermediate with a molecular mass of 116 Da. Our results suggest that this binding involves formation of a disulfide bond with one of the eight cysteines in CYP2E1. The inactivation of wild-type CYP2E1 as well as two of its polymorphic mutants, CYP2E1*2 and CYP2E1*4, was also investigated. For wild-type CYP2E1, the K(I) was 12.2 μM and the k(inact) was 0.02 min(-1). The K(I) values for the two polymorphic mutants were 227.6 and 12.4 μM for CYP2E1.2 and CYP2E1.4, and the k(inact) values were 0.0061 and 0.0187 min(-1), respectively. These data indicate that DDC is a much less efficient inactivator of CYP2E1.2 than it is of either the wild-type or the CYP2E1.4 variant.
authors.affiliation: Department of Pharmacology, University of Michigan, Ann Arbor, MI 48109-5632, USA.
authors.forename: Matthew
authors.initials: M
authors.lastname: Pratt-Hyatt
authors.forename: Hsia-lien
authors.initials: HL
authors.lastname: Lin
authors.forename: Paul F
authors.initials: PF
authors.lastname: Hollenberg
mesh_terms: D002851:Chromatography, High Pressure Liquid; D019392:Cytochrome P-450 CYP2E1 / Q000235:genetics; D065691:Cytochrome P-450 CYP2E1 Inhibitors; D004221:Disulfiram / Q000494:pharmacology; D004050:Ditiocarb / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D007700:Kinetics; D009154:Mutation; D053719:Tandem Mass Spectrometry
chemical_list: D065691:Cytochrome P-450 CYP2E1 Inhibitors; D004050:Ditiocarb; D019392:Cytochrome P-450 CYP2E1; D004221:Disulfiram
references.citation: Chem Res Toxicol. 1998 Oct;11(10):1154-61
references.pmid: 9778311
references.citation: Pharmacogenetics. 1998 Dec;8(6):543-52
references.pmid: 9918138
references.citation: J Biol Chem. 1986 Dec 15;261(35):16689-97
references.pmid: 3782137
references.citation: J Stud Alcohol. 1977 Jan;38(1):96-113
references.pmid: 319300
references.citation: Lancet. 1948 Dec 25;2(6539):1001-4
references.pmid: 18103475
references.citation: Physiol Rev. 1997 Apr;77(2):517-44
references.pmid: 9114822
references.citation: Biochem J. 1992 Feb 1;281 ( Pt 3):689-95
references.pmid: 1536649
references.citation: Cancer Epidemiol Biomarkers Prev. 2009 Feb;18(2):433-43
references.pmid: 19190172
references.citation: J Biol Chem. 1988 May 25;263(15):7170-5
references.pmid: 3366772
references.citation: J Biol Chem. 2008 Nov 28;283(48):33698-707
references.pmid: 18818195
references.citation: Cancer Res. 1989 May 15;49(10):2689-92
references.pmid: 2713853
references.citation: Arch Biochem Biophys. 2001 Nov 1;395(1):57-68
references.pmid: 11673866
references.citation: Drug Metab Dispos. 1999 Jun;27(6):717-23
references.pmid: 10348802
references.citation: Xenobiotica. 2003 Jun;33(6):575-86
references.pmid: 12851035
references.citation: J Pharmacol Exp Ther. 2009 Apr;329(1):26-37
references.pmid: 19168709
references.citation: Mol Pharmacol. 1997 Mar;51(3):370-6
references.pmid: 9058590
references.citation: Chem Res Toxicol. 2002 Dec;15(12):1561-71
references.pmid: 12482238
references.citation: Mol Pharmacol. 2009 Nov;76(5):1011-8
references.pmid: 19720728
references.citation: Chem Biol Interact. 2001 Jan 30;130-132(1-3):93-102
references.pmid: 11306034
references.citation: J Pharmacol Exp Ther. 1966 Apr;152(1):56-61
references.pmid: 5937402
references.citation: Biochem Pharmacol. 1996 Jul 26;52(2):197-204
references.pmid: 8694843
references.citation: Chem Res Toxicol. 1994 Jul-Aug;7(4):526-33
references.pmid: 7981417
references.citation: Biochim Biophys Acta. 1992 Dec 29;1171(2):141-6
references.pmid: 1282830
references.citation: Mol Interv. 2009 Aug;9(4):175-87
references.pmid: 19720750
references.citation: J Pharmacol Exp Ther. 1977 Sep;202(3):724-31
references.pmid: 197231
references.citation: Chem Res Toxicol. 1991 Mar-Apr;4(2):168-79
references.pmid: 1664256
references.citation: FASEB J. 1992 Jan 6;6(2):724-30
references.pmid: 1537462
references.citation: J Pharmacol Exp Ther. 2005 Apr;313(1):154-64
references.pmid: 15608076
references.citation: J Biol Chem. 1991 Apr 25;266(12):7321-4
references.pmid: 2019566
references.citation: Pharmacol Rev. 1966 Mar;18(1):77-82
references.pmid: 5904190