Catalytic properties of the human cytochrome P450 2E1 produced by cDNA expression in mammalian cells. 1992

C J Patten, and H Ishizaki, and T Aoyama, and M Lee, and S M Ning, and W Huang, and F J Gonzalez, and C S Yang
Laboratory for Cancer Research, College of Pharmacy, Rutgers University, Piscataway, New Jersey 08855.

A full-length cDNA encoding human cytochrome P450 2E1 was expressed in mammalian cell lines using the vaccinia virus expression system. Immunoblot analysis showed that the expressed protein reacted with a polyclonal antibody against rat 2E1 and comigrated with P450 2E1 from human liver microsomes. P450 2E1 expressed in Hep G2 cells, a human cell line which contains both cytochrome b5 and NADPH:P450 oxidoreductase, was able to metabolize several known P450 2E1 substrates: N-nitrosodimethylamine (NDMA), N-nitrosomethylbenzylamine (NMBzA), p-nitrophenol, phenol, and acetaminophen. Apparent Km and Vmax values for NDMA demethylation were 22 microM and 173 pmol/min/mg microsomal protein, respectively. P450 2E1 expressed in TK-143 cells, which do not contain b5, displayed Km and Vmax values of 31 microM and 34 pmol/min/mg microsomal protein, respectively. Incorporation of purified rat liver b5 into TK-143 microsomes increased the Vmax 2.2-fold and decreased the Km to 22 microM. Addition of b5 to Hep G2 microsomes resulted in a 1.6-fold increase in Vmax, but showed no effect on the Km. P450 2E1 expressed in Hep G2 cells was shown to metabolize NMBzA with a Km of 47 microM and Vmax of 213 pmol/min/mg microsomal protein. Addition of b5 lowered the Km to 27 microM, but had no effect on Vmax. These results demonstrate conclusively that P450 2E1 is responsible for the low Km forms of NDMA demethylase and NMBzA debenzylase observed in liver microsomes and that these activities are affected by cytochrome b5.

UI MeSH Term Description Entries
D007700 Kinetics The rate dynamics in chemical or physical systems.
D008861 Microsomes Artifactual vesicles formed from the endoplasmic reticulum when cells are disrupted. They are isolated by differential centrifugation and are composed of three structural features: rough vesicles, smooth vesicles, and ribosomes. Numerous enzyme activities are associated with the microsomal fraction. (Glick, Glossary of Biochemistry and Molecular Biology, 1990; from Rieger et al., Glossary of Genetics: Classical and Molecular, 5th ed) Microsome
D008862 Microsomes, Liver Closed vesicles of fragmented endoplasmic reticulum created when liver cells or tissue are disrupted by homogenization. They may be smooth or rough. Liver Microsomes,Liver Microsome,Microsome, Liver
D009603 Nitroso Compounds Organic compounds containing the nitroso (-N Compounds, Nitroso
D010089 Oxidoreductases, N-Demethylating N-Demethylase,N-Demethylases,Oxidoreductases, N Demethylating,Demethylating Oxidoreductases, N,N Demethylase,N Demethylases,N Demethylating Oxidoreductases,N-Demethylating Oxidoreductases
D010636 Phenols Benzene derivatives that include one or more hydroxyl groups attached to the ring structure.
D011994 Recombinant Proteins Proteins prepared by recombinant DNA technology. Biosynthetic Protein,Biosynthetic Proteins,DNA Recombinant Proteins,Recombinant Protein,Proteins, Biosynthetic,Proteins, Recombinant DNA,DNA Proteins, Recombinant,Protein, Biosynthetic,Protein, Recombinant,Proteins, DNA Recombinant,Proteins, Recombinant,Recombinant DNA Proteins,Recombinant Proteins, DNA
D002273 Carcinogens Substances that increase the risk of NEOPLASMS in humans or animals. Both genotoxic chemicals, which affect DNA directly, and nongenotoxic chemicals, which induce neoplasms by other mechanism, are included. Carcinogen,Oncogen,Oncogens,Tumor Initiator,Tumor Initiators,Tumor Promoter,Tumor Promoters,Initiator, Tumor,Initiators, Tumor,Promoter, Tumor,Promoters, Tumor
D002460 Cell Line Established cell cultures that have the potential to propagate indefinitely. Cell Lines,Line, Cell,Lines, Cell
D003577 Cytochrome P-450 Enzyme System A superfamily of hundreds of closely related HEMEPROTEINS found throughout the phylogenetic spectrum, from animals, plants, fungi, to bacteria. They include numerous complex monooxygenases (MIXED FUNCTION OXYGENASES). In animals, these P-450 enzymes serve two major functions: (1) biosynthesis of steroids, fatty acids, and bile acids; (2) metabolism of endogenous and a wide variety of exogenous substrates, such as toxins and drugs (BIOTRANSFORMATION). They are classified, according to their sequence similarities rather than functions, into CYP gene families (>40% homology) and subfamilies (>59% homology). For example, enzymes from the CYP1, CYP2, and CYP3 gene families are responsible for most drug metabolism. Cytochrome P-450,Cytochrome P-450 Enzyme,Cytochrome P-450-Dependent Monooxygenase,P-450 Enzyme,P450 Enzyme,CYP450 Family,CYP450 Superfamily,Cytochrome P-450 Enzymes,Cytochrome P-450 Families,Cytochrome P-450 Monooxygenase,Cytochrome P-450 Oxygenase,Cytochrome P-450 Superfamily,Cytochrome P450,Cytochrome P450 Superfamily,Cytochrome p450 Families,P-450 Enzymes,P450 Enzymes,Cytochrome P 450,Cytochrome P 450 Dependent Monooxygenase,Cytochrome P 450 Enzyme,Cytochrome P 450 Enzyme System,Cytochrome P 450 Enzymes,Cytochrome P 450 Families,Cytochrome P 450 Monooxygenase,Cytochrome P 450 Oxygenase,Cytochrome P 450 Superfamily,Enzyme, Cytochrome P-450,Enzyme, P-450,Enzyme, P450,Enzymes, Cytochrome P-450,Enzymes, P-450,Enzymes, P450,Monooxygenase, Cytochrome P-450,Monooxygenase, Cytochrome P-450-Dependent,P 450 Enzyme,P 450 Enzymes,P-450 Enzyme, Cytochrome,P-450 Enzymes, Cytochrome,Superfamily, CYP450,Superfamily, Cytochrome P-450,Superfamily, Cytochrome P450

Related Publications

C J Patten, and H Ishizaki, and T Aoyama, and M Lee, and S M Ning, and W Huang, and F J Gonzalez, and C S Yang
July 1994, Archives of biochemistry and biophysics,
C J Patten, and H Ishizaki, and T Aoyama, and M Lee, and S M Ning, and W Huang, and F J Gonzalez, and C S Yang
June 1993, European journal of biochemistry,
C J Patten, and H Ishizaki, and T Aoyama, and M Lee, and S M Ning, and W Huang, and F J Gonzalez, and C S Yang
March 1995, Archives of biochemistry and biophysics,
C J Patten, and H Ishizaki, and T Aoyama, and M Lee, and S M Ning, and W Huang, and F J Gonzalez, and C S Yang
April 1998, Molecular pharmacology,
C J Patten, and H Ishizaki, and T Aoyama, and M Lee, and S M Ning, and W Huang, and F J Gonzalez, and C S Yang
December 2001, Hunan yi ke da xue xue bao = Hunan yike daxue xuebao = Bulletin of Hunan Medical University,
C J Patten, and H Ishizaki, and T Aoyama, and M Lee, and S M Ning, and W Huang, and F J Gonzalez, and C S Yang
July 1995, European journal of pharmacology,
C J Patten, and H Ishizaki, and T Aoyama, and M Lee, and S M Ning, and W Huang, and F J Gonzalez, and C S Yang
October 1995, Biochemical pharmacology,
C J Patten, and H Ishizaki, and T Aoyama, and M Lee, and S M Ning, and W Huang, and F J Gonzalez, and C S Yang
March 1997, Biochemical pharmacology,
C J Patten, and H Ishizaki, and T Aoyama, and M Lee, and S M Ning, and W Huang, and F J Gonzalez, and C S Yang
July 2005, Life sciences,
C J Patten, and H Ishizaki, and T Aoyama, and M Lee, and S M Ning, and W Huang, and F J Gonzalez, and C S Yang
December 2003, World journal of gastroenterology,
Copied contents to your clipboard!