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Enhancement of cytochrome P-450 3A4 catalytic activities by cytochrome b(5) in bacterial membranes.
Drug Metab Dispos. 1999 Sep; 27(9):999-1004.DM

Abstract

Activities of testosterone, nifedipine, and midazolam oxidation by recombinant cytochrome P-450 (P-450) 3A4 coexpressed with human NADPH-P-450 reductase (NPR) in bacterial membranes (CYP3A4/NPR membranes) were determined in comparison with those of other recombinant systems and of human liver microsomes with high contents of CYP3A4. Growth conditions for Escherichia coli transformed with the bicistronic construct affected expression levels of CYP3A4 and NPR; an excess of NPR over P-450 in membrane preparations enhanced CYP3A4-dependent testosterone 6beta-hydroxylation activities of the CYP3A4/NPR membranes. Cytochrome b(5) (b(5)) and apolipoprotein b(5) further enhanced the testosterone 6beta-hydroxylation activities of CYP3A4/NPR membranes after addition to either bacterial membranes or purified enzymes. NPR was observed to enhance catalytic activity when added to the CYP3A4/NPR membranes, either in the form of bacterial membranes or as purified NPR (in combination with cholate and b(5)). Apparent maximal activities of testosterone 6beta-hydroxylation in CYP3A4/NPR membranes were obtained when the molar ratio of CYP3A4/NPR/b(5) was adjusted to 1:2:1 by mixing membranes containing each protein. Testosterone 6beta-hydroxylation, nifedipine oxidation, and midazolam 4- and 1'-hydroxylation activities in CYP3A4/NPR membranes plus b(5) systems were similar to those measured with microsomes of insect cells coexpressing CYP3A4 with NPR and/or of human liver microsomes, based on equivalent CYP3A4 contents. These results suggest that CYP3A4/NPR membrane systems containing b(5) are very useful models for prediction of the rates for liver microsomal CYP3A4-dependent drug oxidations.

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Authors+Show Affiliations

Yamazaki H
Faculty of Pharmaceutical Sciences, Kanazawa University, Kanazawa, Japan. yamazak@kenroku.kanazawa-u.ac.jp
Nakajima M
No affiliation info available
Nakamura M
No affiliation info available
Asahi S
No affiliation info available
Shimada N
No affiliation info available
Gillam EM
No affiliation info available
Guengerich FP
No affiliation info available
Shimada T
No affiliation info available
Yokoi T
No affiliation info available

MeSH

Anesthetics, IntravenousBacteriaCalcium Channel BlockersCytochrome P-450 CYP3ACytochrome P-450 Enzyme SystemCytochromes b5Escherichia coliHumansHydroxylationIn Vitro TechniquesKineticsMembranesMidazolamMixed Function OxygenasesNADPH-Ferrihemoprotein ReductaseNifedipineOxidation-ReductionRecombinant ProteinsTestosterone

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

10460798

Citation

Yamazaki, H, et al. "Enhancement of Cytochrome P-450 3A4 Catalytic Activities By Cytochrome B(5) in Bacterial Membranes." Drug Metabolism and Disposition: the Biological Fate of Chemicals, vol. 27, no. 9, 1999, pp. 999-1004.
Yamazaki H, Nakajima M, Nakamura M, et al. Enhancement of cytochrome P-450 3A4 catalytic activities by cytochrome b(5) in bacterial membranes. Drug Metab Dispos. 1999;27(9):999-1004.
Yamazaki, H., Nakajima, M., Nakamura, M., Asahi, S., Shimada, N., Gillam, E. M., Guengerich, F. P., Shimada, T., & Yokoi, T. (1999). Enhancement of cytochrome P-450 3A4 catalytic activities by cytochrome b(5) in bacterial membranes. Drug Metabolism and Disposition: the Biological Fate of Chemicals, 27(9), 999-1004.
Yamazaki H, et al. Enhancement of Cytochrome P-450 3A4 Catalytic Activities By Cytochrome B(5) in Bacterial Membranes. Drug Metab Dispos. 1999;27(9):999-1004. PubMed PMID: 10460798.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Enhancement of cytochrome P-450 3A4 catalytic activities by cytochrome b(5) in bacterial membranes. AU - Yamazaki,H, AU - Nakajima,M, AU - Nakamura,M, AU - Asahi,S, AU - Shimada,N, AU - Gillam,E M, AU - Guengerich,F P, AU - Shimada,T, AU - Yokoi,T, PY - 1999/8/26/pubmed PY - 1999/8/26/medline PY - 1999/8/26/entrez SP - 999 EP - 1004 JF - Drug metabolism and disposition: the biological fate of chemicals JO - Drug Metab Dispos VL - 27 IS - 9 N2 - Activities of testosterone, nifedipine, and midazolam oxidation by recombinant cytochrome P-450 (P-450) 3A4 coexpressed with human NADPH-P-450 reductase (NPR) in bacterial membranes (CYP3A4/NPR membranes) were determined in comparison with those of other recombinant systems and of human liver microsomes with high contents of CYP3A4. Growth conditions for Escherichia coli transformed with the bicistronic construct affected expression levels of CYP3A4 and NPR; an excess of NPR over P-450 in membrane preparations enhanced CYP3A4-dependent testosterone 6beta-hydroxylation activities of the CYP3A4/NPR membranes. Cytochrome b(5) (b(5)) and apolipoprotein b(5) further enhanced the testosterone 6beta-hydroxylation activities of CYP3A4/NPR membranes after addition to either bacterial membranes or purified enzymes. NPR was observed to enhance catalytic activity when added to the CYP3A4/NPR membranes, either in the form of bacterial membranes or as purified NPR (in combination with cholate and b(5)). Apparent maximal activities of testosterone 6beta-hydroxylation in CYP3A4/NPR membranes were obtained when the molar ratio of CYP3A4/NPR/b(5) was adjusted to 1:2:1 by mixing membranes containing each protein. Testosterone 6beta-hydroxylation, nifedipine oxidation, and midazolam 4- and 1'-hydroxylation activities in CYP3A4/NPR membranes plus b(5) systems were similar to those measured with microsomes of insect cells coexpressing CYP3A4 with NPR and/or of human liver microsomes, based on equivalent CYP3A4 contents. These results suggest that CYP3A4/NPR membrane systems containing b(5) are very useful models for prediction of the rates for liver microsomal CYP3A4-dependent drug oxidations. SN - 0090-9556 UR - https://www.unboundmedicine.com/medline/citation/10460798/Enhancement_of_cytochrome_P_450_3A4_catalytic_activities_by_cytochrome_b_5__in_bacterial_membranes_ L2 - http://dmd.aspetjournals.org/cgi/pmidlookup?view=long&pmid=10460798 DB - PRIME DP - Unbound Medicine ER -
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