Role of cytochrome b5 in the modulation of the enzymatic activities of cytochrome P450 17α-hydroxylase/17,20-lyase (P450 17A1)

Bhatt, Megh Raj; Khatri, Yogan; Rodgers, Raymond J.; Martin, Lisandra L.

doi:10.1016/j.jsbmb.2016.02.033

Cited by 44 publications

(32 citation statements)

References 252 publications

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“…Recently, it was reported that CYP17A1 yields different by‐products in the presence of iodosobenzene and of NADPH‐P450 reductase to form compound I . The effect of cytochrome b5 on the catalytic activity and product formation of CYPs has been widely studied, and shown to induce the activity of specific CYPs . In another experiment, MycG, a multifunctional P450 monooxygenase derived from a bacterial source ( Micromonospora griseorubida ) showed an altered type of catalysis in conjunction with an alternative surrogate redox partner, thus highlighting the roles played by variant redox partners in protein–protein interactions in the catalytic activity of CYP enzymes .…”

Section: Discussionmentioning

confidence: 99%

Effects of Alternative Redox Partners and Oxidizing Agents on CYP154C8 Catalytic Activity and Product Distribution

Dangi

Park

2018

ChemBioChem

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CYP154C8 catalyzes the hydroxylation of diverse steroids, as has previously been demonstrated, by using an NADH-dependent system including putidaredoxin and putidaredoxin reductase as redox partner proteins carrying electrons from NADH. In other reactions, CYP154C8 reconstituted with spinach ferredoxin and NADPH-dependent ferredoxin reductase displayed catalytic activity different from that of the NADH-dependent system. The NADPH-dependent system showed multistep oxidation of progesterone and other substrates including androstenedione, testosterone, and nandrolone. (Diacetoxyiodo)benzene was employed to generate compound I (FeO ), actively supporting the redox reactions catalyzed by CYP154C8. In addition to 16α-hydroxylation, progesterone and 11-oxoprogesterone also underwent hydroxylation at the 6β-position in reactions supported by (diacetoxyiodo)benzene. CYP154C8 was active in the presence of high concentrations (>10 mm) of H O , with optimum conversion surprisingly being achieved at ≈75 mm H O . More importantly, H O tolerance by CYP154C8 was evident in the very low heme oxidation rate constant (K) even at high concentrations of H O . Our results demonstrate that alternative redox partners and oxidizing agents influence the catalytic efficiency and product distribution of a cytochrome P450 enzyme. More importantly, these choices affected the type and selectivity of reaction catalyzed by the P450 enzyme.

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Section: Discussionmentioning

confidence: 99%

Effects of Alternative Redox Partners and Oxidizing Agents on CYP154C8 Catalytic Activity and Product Distribution

Dangi

Park

2018

ChemBioChem

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“…The addition of b5 was to support CYP activities and serve as an obligate component of the reaction [19,20]. The reaction was allowed to preincubate for 5 min at 37°C in a Fisher shaking water bath.…”

Section: Methodsmentioning

confidence: 99%

Effects of CYP2C19 Variants on Fluoxetine Metabolism in vitro

Fang

He²,

Han³

et al. 2017

Pharmacology

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Aims: CYP2C19 is an important member of the cytochrome P450 enzyme superfamily. We recently identified 31 CYP2C19 alleles in the Han Chinese population. The aim of this study was to assess the catalytic activities of these allelic isoforms and their effects on the metabolism of fluoxetine in vitro. Methods: The wild-type and 30 CYP2C19 variants were expressed in insect cells and each variant was characterized using fluoxetine as the substrate. Reactions were performed at 37°C with 20-1,000 µmol/L substrate for 30 min. By using ultra-high performance liquid chromatography-mass spectrometry to detect the products, the kinetic parameters K_m, V_max, and intrinsic clearance (V_max/K_m) of norfluoxetine were determined. Results: Among the CYP2C19 variants tested, T130M showed similar intrinsic clearance (V_max/K_m) values with CYP2C19*1, while the intrinsic clearance values of other variants were significantly decreased (from 9.56 to 77.77%). In addition, CYP2C19*3 and *35FS could not be detected because they have no detectable enzyme activity. Conclusion: In China, the assessment of CYP2C19 variants in vitro offers valuable information relevant to the personalized medicine for CYP2C19-metabolized drug.

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“…The incubation mixture consisted of (final assay concentration) 5–10 pmol of P450 from insect microsomes (5 pmol for CYP2C19*1 or 10 pmol for other CYP2C19 mutants), 5–10 pmol of purified cytochrome b5 (5 pmol for CYP2C19*1 or 10 pmol for other CYP2C19 mutants, CYP2C19/b5 = 1:1), and 176.9 μL Tris‐HCl buffer (pH 7.4) and 3.09 μL of MTD were added to the mixture. The addition of b5 was to support CYP activities and serve as an obligate component of the reaction . The concentrations of MTD used in the incubation mixture for kinetic analysis were 50, 100, 200, 500, 1000, 1500, and 2000 μM.…”

Section: Methodsmentioning

confidence: 99%

Effects of CYP2C19 variants on methadone metabolism in vitro

Lan

Yuan

et al. 2016

Drug Testing and Analysis

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CYP2C19 is an important member of the cytochrome P450 (CYP450) enzyme super family and is responsible for clearing approximately 10% of commonly used clinical drugs that undergo phase I metabolism. Genetic polymorphisms of CYP2C19 significantly influence the efficacy and safety of some drugs, which might cause undesirable adverse effects or cure failure at standard dosages. The aim of this study was to clarify the catalytic activities of 31 CYP2C19 alleles on the oxidative in vitro metabolism of methadone. Insect microsomes expressing the CYP2C19 alleles were incubated with 50-2000 μM methadone for 30 min at 37 °C and terminated by cooling to -80 °C immediately. Methadone and its metabolite EDDP were analyzed by an ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) system. Of the 31 tested CYP2C19 allelies variants, CYP2C19*1 is the wild-type. Compared with CYP2C19*1, two CYP2C19 variants (CYP2C19*3 and *35FS) had no detectable enzyme activity, one variant L16F exhibited slightly increased intrinsic clearance values, and one variant N277K showed no significant difference. In addition, 26 variants exhibited significantly decreased values (from 1.48% to 80.40%). These findings suggest that more attention should be paid in clinical administration of methadone to individuals carrying these CYP2C19 alleles. Copyright © 2016 John Wiley & Sons, Ltd.

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Role of cytochrome b5 in the modulation of the enzymatic activities of cytochrome P450 17α-hydroxylase/17,20-lyase (P450 17A1)

Cited by 44 publications

References 252 publications

Effects of Alternative Redox Partners and Oxidizing Agents on CYP154C8 Catalytic Activity and Product Distribution

Effects of Alternative Redox Partners and Oxidizing Agents on CYP154C8 Catalytic Activity and Product Distribution

Effects of CYP2C19 Variants on Fluoxetine Metabolism in vitro

Effects of CYP2C19 variants on methadone metabolism in vitro

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