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

Patten, Chris; Ishizaki, Hiroyuki; Aoyama, Toru; Lee, Mao-Jung; Ning, Shu; Huang, Wen; Gonzalez, Frank J.; Yang, Chung S.

doi:10.1016/0003-9861(92)90258-x

Cited by 72 publications

(35 citation statements)

References 33 publications

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“…For most activities, the presence of b 5 increases V max of CYP3A4, but b 5 can also change K m , as observed for some CYP2E1-catalyzed reactions (28,29). Because CYP3A4 activity obeys sigmoidal rather than Michaelis-Menten kinetics, v vs. [S] data were fit to an allosteric sigmoidal model, to determine changes in V max and K 0.5 .…”

Section: Resultsmentioning

confidence: 65%

Two surfaces of cytochrome b5 with major and minor contributions to CYP3A4-catalyzed steroid and nifedipine oxygenation chemistries

Peng

Auchus

2014

Archives of Biochemistry and Biophysics

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Conserved human cytochrome b5 (b5) residues D58 and D65 are critical for interactions with CYP2E1 and CYP2C19, whereas E48 and E49 are essential for stimulating the 17,20-lyase activity of CYP17A1. Here, we show that b5 mutations E48G, E49G, D58G, and D65G have reduced capacity to stimulate CYP3A4-catalyzed progesterone and testosterone 6β-hydroxylation or nifedipine oxidation. The b5 double mutation D58G/D65G fails to stimulate these reactions, similar to CYP2E1 and CYP2C19, whereas mutation E48G/E49G retains 23–42% of wild-type stimulation. Neither mutation impairs the activity stimulation of wild-type b5, nor does mutation D58G/D65G impair the partial stimulation of mutations E48G or E48G/E49G. For assays reconstituted with a single phospholipid, phosphatidyl serine afforded the highest testosterone 6β-hydroxylase activity with wild-type b5 but the poorest activity with b5 mutation E48G/E49G, and the activity stimulation of mutation E48G/E49G was lost at [NaCl] > 50 mM. Cross-linking of CYP3A4 and b5 decreased in the order wild-type > E48G/E49G > D58G/D65G and varied with phospholipid. We conclude that two b5 acidic surfaces, primarily the domain including residues D58-D65, participate in the stimulation of CYP3A4 activities. Our data suggest that a minor population of CYP3A4 molecules remains sensitive to b5 mutation E48G/E49G, consistent with phospholipid-dependent conformational heterogeneity of CYP3A4.

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Section: Resultsmentioning

confidence: 65%

Two surfaces of cytochrome b5 with major and minor contributions to CYP3A4-catalyzed steroid and nifedipine oxygenation chemistries

Peng

Auchus

2014

Archives of Biochemistry and Biophysics

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“…Inhibition by human CYP2E1 is important because this enzyme occurs in many human tissues, including the esophagus (32,33), and because polymorphic forms of CYP2E1 may show different distributions in cases of cancer of the esophagus, nasopharynx, and oral cavity in China and Taiwan than in matched controls, suggesting that nitrosamine activation by this P450 is involved in the etiology of these cancers (34)(35)(36). Human CYP2E1 showed low K m values of 22 µM for DMN and 115 µM for MPN dealkylation (8,30). Inhibition by rat CYP2A3 was investigated because its K m of 8 µM for MPN depentylation (3) was much lower than that for other systems and because CYP2A3 resembles the P450 in REM that metabolizes MPN in that coumarin strongly inhibited MPN depentylation by both systems (3,8).…”

Section: Discussionmentioning

confidence: 98%

Inhibition by Allyl Sulfides and Phenethyl Isothiocyanate of Methyl-n-pentylnitrosamine Depentylation by Rat Esophageal Microsomes, Human and Rat CYP2E1, and Rat CYP2A3

Morris¹,

Chen²,

Zhou³

et al. 2004

Nutrition and Cancer

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Garlic and Cruciferae are associated with reduced risks of several human cancers, and some of their constituents are anticarcinogenic in animals. Here we studied inhibition of in vitro metabolism of the rat esophageal carcinogen methyl-n-pentylnitrosamine (MPN) by garlic-derived allyl sulfides and by Cruciferae-derived phenethyl isothiocyanate (PEITC) and sulforaphane. The test inhibitors were incubated with [3H]-MPN, NADPH-generating system and rat esophageal microsomes (REM) or a cytochrome P450 (CYP). [3H]-MPN activation by depentylation was assayed by HPLC with radiometric determination of [3H]-pentaldehyde 2,4-dinitrophenylhydrazone. IC50 for depentylation of 40 microM MPN by rat CYP2E1 was 5-12 microM for diallyl sulfide (DAS), diallyl disulfide (DADS), and PEITC and 10-20 microM for diallyl sulfone, allyl mercaptan, and diallyl trisulfide. Maximum inhibition required preincubation of rat CYP2E1 with DAS for 15 min and with DADS for 30 min. Using these preincubation times, Ki for MPN depentylation by REM, rat and human CYP2E1, and rat CYP2A3 was 0.6-1.6 microM for inhibition by DAS and 1.7-70 microM for inhibition by DADS. With PEITC, Ki for MPN depentylation by REM, rat CYP2E1, and rat CYP2A3 was 0.4-4.6 microM. These low Ki and IC50 values may help explain how garlic and Cruciferae inhibit carcinogenesis.

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“…Briefly after centrifugation the sup was taken and mixed with 2 M NaOH and the absorbance measured at 546 nm. 4-Nitrocatechol formation was quantitated by using an extinction coefficient of 10.28 mM À1 cm À1 [38].…”

Section: Estimation Of Cyp Activity From Liver Microsomesmentioning

confidence: 99%

Induction of necrosis in cadmium-induced hepatic oxidative stress and its prevention by the prophylactic properties of taurine

Sinha

Manna

Sil

2009

Journal of Trace Elements in Medicine and Biology

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

Cited by 72 publications

References 33 publications

Two surfaces of cytochrome b5 with major and minor contributions to CYP3A4-catalyzed steroid and nifedipine oxygenation chemistries

Two surfaces of cytochrome b5 with major and minor contributions to CYP3A4-catalyzed steroid and nifedipine oxygenation chemistries

Inhibition by Allyl Sulfides and Phenethyl Isothiocyanate of Methyl-n-pentylnitrosamine Depentylation by Rat Esophageal Microsomes, Human and Rat CYP2E1, and Rat CYP2A3

Induction of necrosis in cadmium-induced hepatic oxidative stress and its prevention by the prophylactic properties of taurine

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