Document Detail


Oxidation of methyl and ethyl nitrosamines by cytochrome P450 2E1 and 2B1.
MedLine Citation:
PMID:  23186213     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Cytochrome P450 (P450) 2E1 is the major enzyme that oxidizes N-nitrosodimethylamine [N,N-dimethylnitrosamine (DMN)], a carcinogen and also a representative of some nitrosamines formed endogenously. Oxidation of DMN by rat or human P450 2E1 to HCHO showed a high apparent intrinsic kinetic deuterium isotope effect (KIE), ≥8. The KIE was not attenuated in noncompetitive intermolecular experiments with rat liver microsomes {(D)V = 12.5; (D)(V/K) = 10.9 [nomenclature of Northrop, D. B. (1982) Methods Enzymol. 87, 607-625]} but was with purified human P450 2E1 [(D)V = 3.3; (D)(V/K) = 3.7], indicating that C-H bond breaking is partially rate-limiting with human P450 2E1. With N-nitrosodiethylamine [N,N-diethylnitrosamine (DEN)], the intrinsic KIE was slightly lower and was not expressed [e.g., (D)(V/K) = 1.2] in noncompetitive intermolecular experiments. The same general pattern of KIEs was also seen in the (D)(V/K) results with DMN and DEN for the minor products resulting from the denitrosation reactions (CH(3)NH(2), CH(3)CH(2)NH(2), and NO(2)(-)). Experiments with deuterated N-nitroso-N-methyl-N-ethylamine demonstrated that the lower KIEs associated with ethyl versus methyl oxidation could be distinguished within a single molecule. P450 2E1 oxidized DMN and DEN to aldehydes and then to the carboxylic acids. No kinetic lags were observed in acid formation; pulse-chase experiments with carrier aldehydes showed only limited equilibration with P450 2E1-bound aldehydes, indicative of processive reactions, as reported for P450 2A6 [Chowdhury, G., et al. (2010) J. Biol. Chem. 285, 8031-8044]. These same features (no lag phase for HCO(2)H formation and a lack of equilibration in pulse-chase assays) were also seen with (rat) P450 2B1, which has a lower catalytic efficiency for DMN oxidation and a larger active site. Thus, the processivity of dialkyl nitrosamine oxidation appears to be shared by a number of P450s.
Authors:
Goutam Chowdhury; M Wade Calcutt; Leslie D Nagy; F Peter Guengerich
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural     Date:  2012-12-04
Journal Detail:
Title:  Biochemistry     Volume:  51     ISSN:  1520-4995     ISO Abbreviation:  Biochemistry     Publication Date:  2012 Dec 
Date Detail:
Created Date:  2012-12-18     Completed Date:  2013-02-19     Revised Date:  2014-01-09    
Medline Journal Info:
Nlm Unique ID:  0370623     Medline TA:  Biochemistry     Country:  United States    
Other Details:
Languages:  eng     Pagination:  9995-10007     Citation Subset:  IM    
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MeSH Terms
Descriptor/Qualifier:
Animals
Cytochrome P-450 CYP2B1 / metabolism*
Cytochrome P-450 CYP2E1 / metabolism*
Deuterium / metabolism
Diethylnitrosamine / metabolism*
Dimethylnitrosamine / metabolism*
Humans
Kinetics
Male
Microsomes, Liver / metabolism
Oxidation-Reduction
Rats
Rats, Sprague-Dawley
Grant Support
ID/Acronym/Agency:
F32 ES012123/ES/NIEHS NIH HHS; P30 ES000267/ES/NIEHS NIH HHS; P30 ES000267/ES/NIEHS NIH HHS; R37 CA0090426/CA/NCI NIH HHS; R37 CA090426/CA/NCI NIH HHS; T32 ES007028/ES/NIEHS NIH HHS; T32 ES007028/ES/NIEHS NIH HHS
Chemical
Reg. No./Substance:
3IQ78TTX1A/Diethylnitrosamine; AR09D82C7G/Deuterium; EC 1.14.14.1/Cytochrome P-450 CYP2B1; EC 1.14.14.1/Cytochrome P-450 CYP2E1; M43H21IO8R/Dimethylnitrosamine
Comments/Corrections

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