The stimulatory role of human cytochrome b5 in the bioactivation activities of human CYP1A2, 2A6 and 2E1: a new cell expression system to study cytochrome P450-mediated biotransformation (a corrigendum report on Duarte et al. (2005) Mutagenesis 20, 93-100).

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Abstract

Cytochrome b(5) (b(5)) is increasingly recognized to be of importance for specific cytochrome P450 (CYP) activities. We developed human b(5)/CYP-competent mutagenicity tester bacteria to study the role of b(5) in the bioactivation activity of human CYP. These new tester bacteria were derived from the previously engineered human CYP-competent Escherichia coli K12 tester strain MTC, containing a bi-plasmid system for the co-expression of a specific CYP form (CYP1A2, 2A6 or 2E1) with human b(5), and human NADPH cytochrome P450 reductase (RED), resulting in the strain BTC-b(5)-1A2, BTC-b(5)-2A6 and BTC-b(5)-2E1, respectively. The relative content of b(5) with CYP and RED in these three BTC-b(5)-CYP strains demonstrated physiologically relevant co-expression levels and typical CYP-specific activities could be determined with their specific chemical probes. These strains were applied in mutagenicity assays along with their corresponding b(5)-void strains to determine the effect of b(5) on the CYP1A2-, CYP2A6- and CYP2E1-mediated bioactivation of several promutagens. For CYP1A2, of the 5 compounds tested [2-aminoanthracene (2AA), 1-aminopyrene, 6-aminochrysene, 2-amino-3-methylimidazo(4,5-f)quinoline and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)], only the mutagenicity of 2AA was slightly increased (similar to 1.5-fold) in the presence of b(5). The CYP2E1- and CYP2A6-dependent mutagenicity of N-nitrosodiethylamine increased similar to 3- and 23-fold, respectively when the bacteria contained b(5). The CYP2A6-mediated mutagenicity of NNK increased similar to 9-fold when co-expressed with b(5). The stimulatory effect of b(5) on the bioactivation of N-nitrosodi-n-propylamine was most striking. The mutagenicity of this procarcinogen was completely dependent on the co-expression of b(5) with CYP2A6 or CYP2E1. This demonstrates the prominent role of b(5) in the bioactivation of this carcinogen.
Original languageEnglish
Pages (from-to)75-81
Number of pages7
JournalMutagenesis
Volume22
Issue number1
DOIs
Publication statusPublished - 2007

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Cytochromes b5
Cytochrome P-450 CYP1A2
Mutagenesis
Biotransformation
Human Activities
Cytochrome P-450 Enzyme System
Cytochrome P-450 CYP2E1
NADPH-Ferrihemoprotein Reductase
Bacteria
2-amino-3-methylimidazo(4,5-f)quinoline
Propylamines
Escherichia coli K12
Diethylnitrosamine
corrigendum
Carcinogens
Escherichia coli
Assays
Plasmids

Cite this

@article{6d53e45654ca435492f4529db20f136b,
title = "The stimulatory role of human cytochrome b5 in the bioactivation activities of human CYP1A2, 2A6 and 2E1: a new cell expression system to study cytochrome P450-mediated biotransformation (a corrigendum report on Duarte et al. (2005) Mutagenesis 20, 93-100).",
abstract = "Cytochrome b(5) (b(5)) is increasingly recognized to be of importance for specific cytochrome P450 (CYP) activities. We developed human b(5)/CYP-competent mutagenicity tester bacteria to study the role of b(5) in the bioactivation activity of human CYP. These new tester bacteria were derived from the previously engineered human CYP-competent Escherichia coli K12 tester strain MTC, containing a bi-plasmid system for the co-expression of a specific CYP form (CYP1A2, 2A6 or 2E1) with human b(5), and human NADPH cytochrome P450 reductase (RED), resulting in the strain BTC-b(5)-1A2, BTC-b(5)-2A6 and BTC-b(5)-2E1, respectively. The relative content of b(5) with CYP and RED in these three BTC-b(5)-CYP strains demonstrated physiologically relevant co-expression levels and typical CYP-specific activities could be determined with their specific chemical probes. These strains were applied in mutagenicity assays along with their corresponding b(5)-void strains to determine the effect of b(5) on the CYP1A2-, CYP2A6- and CYP2E1-mediated bioactivation of several promutagens. For CYP1A2, of the 5 compounds tested [2-aminoanthracene (2AA), 1-aminopyrene, 6-aminochrysene, 2-amino-3-methylimidazo(4,5-f)quinoline and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)], only the mutagenicity of 2AA was slightly increased (similar to 1.5-fold) in the presence of b(5). The CYP2E1- and CYP2A6-dependent mutagenicity of N-nitrosodiethylamine increased similar to 3- and 23-fold, respectively when the bacteria contained b(5). The CYP2A6-mediated mutagenicity of NNK increased similar to 9-fold when co-expressed with b(5). The stimulatory effect of b(5) on the bioactivation of N-nitrosodi-n-propylamine was most striking. The mutagenicity of this procarcinogen was completely dependent on the co-expression of b(5) with CYP2A6 or CYP2E1. This demonstrates the prominent role of b(5) in the bioactivation of this carcinogen.",
keywords = "ESCHERICHIA-COLI MTC, POLYMORPHISMS, CHEMICAL CARCINOGENS, METABOLIZING ENZYMES, P450 REDUCTASE, HUMAN LIVER-MICROSOMES, MUTAGENICITY TESTER STRAIN, SALMONELLA-TYPHIMURIUM, B(5), CATALYTIC-ACTIVITIES",
author = "Duarte, {Maria Paula Amaro de Castilho} and Laires, {Ant{\'o}nio Jos{\'e} Cabrita Lucas} and Michel Kranendonk and J. Rueff",
year = "2007",
doi = "10.1093/mutage/gel054",
language = "English",
volume = "22",
pages = "75--81",
journal = "Mutagenesis",
issn = "0267-8357",
publisher = "OXFORD UNIV PRESS INC",
number = "1",

}

TY - JOUR

T1 - The stimulatory role of human cytochrome b5 in the bioactivation activities of human CYP1A2, 2A6 and 2E1: a new cell expression system to study cytochrome P450-mediated biotransformation (a corrigendum report on Duarte et al. (2005) Mutagenesis 20, 93-100).

AU - Duarte, Maria Paula Amaro de Castilho

AU - Laires, António José Cabrita Lucas

AU - Kranendonk, Michel

AU - Rueff, J.

PY - 2007

Y1 - 2007

N2 - Cytochrome b(5) (b(5)) is increasingly recognized to be of importance for specific cytochrome P450 (CYP) activities. We developed human b(5)/CYP-competent mutagenicity tester bacteria to study the role of b(5) in the bioactivation activity of human CYP. These new tester bacteria were derived from the previously engineered human CYP-competent Escherichia coli K12 tester strain MTC, containing a bi-plasmid system for the co-expression of a specific CYP form (CYP1A2, 2A6 or 2E1) with human b(5), and human NADPH cytochrome P450 reductase (RED), resulting in the strain BTC-b(5)-1A2, BTC-b(5)-2A6 and BTC-b(5)-2E1, respectively. The relative content of b(5) with CYP and RED in these three BTC-b(5)-CYP strains demonstrated physiologically relevant co-expression levels and typical CYP-specific activities could be determined with their specific chemical probes. These strains were applied in mutagenicity assays along with their corresponding b(5)-void strains to determine the effect of b(5) on the CYP1A2-, CYP2A6- and CYP2E1-mediated bioactivation of several promutagens. For CYP1A2, of the 5 compounds tested [2-aminoanthracene (2AA), 1-aminopyrene, 6-aminochrysene, 2-amino-3-methylimidazo(4,5-f)quinoline and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)], only the mutagenicity of 2AA was slightly increased (similar to 1.5-fold) in the presence of b(5). The CYP2E1- and CYP2A6-dependent mutagenicity of N-nitrosodiethylamine increased similar to 3- and 23-fold, respectively when the bacteria contained b(5). The CYP2A6-mediated mutagenicity of NNK increased similar to 9-fold when co-expressed with b(5). The stimulatory effect of b(5) on the bioactivation of N-nitrosodi-n-propylamine was most striking. The mutagenicity of this procarcinogen was completely dependent on the co-expression of b(5) with CYP2A6 or CYP2E1. This demonstrates the prominent role of b(5) in the bioactivation of this carcinogen.

AB - Cytochrome b(5) (b(5)) is increasingly recognized to be of importance for specific cytochrome P450 (CYP) activities. We developed human b(5)/CYP-competent mutagenicity tester bacteria to study the role of b(5) in the bioactivation activity of human CYP. These new tester bacteria were derived from the previously engineered human CYP-competent Escherichia coli K12 tester strain MTC, containing a bi-plasmid system for the co-expression of a specific CYP form (CYP1A2, 2A6 or 2E1) with human b(5), and human NADPH cytochrome P450 reductase (RED), resulting in the strain BTC-b(5)-1A2, BTC-b(5)-2A6 and BTC-b(5)-2E1, respectively. The relative content of b(5) with CYP and RED in these three BTC-b(5)-CYP strains demonstrated physiologically relevant co-expression levels and typical CYP-specific activities could be determined with their specific chemical probes. These strains were applied in mutagenicity assays along with their corresponding b(5)-void strains to determine the effect of b(5) on the CYP1A2-, CYP2A6- and CYP2E1-mediated bioactivation of several promutagens. For CYP1A2, of the 5 compounds tested [2-aminoanthracene (2AA), 1-aminopyrene, 6-aminochrysene, 2-amino-3-methylimidazo(4,5-f)quinoline and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)], only the mutagenicity of 2AA was slightly increased (similar to 1.5-fold) in the presence of b(5). The CYP2E1- and CYP2A6-dependent mutagenicity of N-nitrosodiethylamine increased similar to 3- and 23-fold, respectively when the bacteria contained b(5). The CYP2A6-mediated mutagenicity of NNK increased similar to 9-fold when co-expressed with b(5). The stimulatory effect of b(5) on the bioactivation of N-nitrosodi-n-propylamine was most striking. The mutagenicity of this procarcinogen was completely dependent on the co-expression of b(5) with CYP2A6 or CYP2E1. This demonstrates the prominent role of b(5) in the bioactivation of this carcinogen.

KW - ESCHERICHIA-COLI MTC

KW - POLYMORPHISMS

KW - CHEMICAL CARCINOGENS

KW - METABOLIZING ENZYMES

KW - P450 REDUCTASE

KW - HUMAN LIVER-MICROSOMES

KW - MUTAGENICITY TESTER STRAIN

KW - SALMONELLA-TYPHIMURIUM

KW - B(5)

KW - CATALYTIC-ACTIVITIES

U2 - 10.1093/mutage/gel054

DO - 10.1093/mutage/gel054

M3 - Article

VL - 22

SP - 75

EP - 81

JO - Mutagenesis

JF - Mutagenesis

SN - 0267-8357

IS - 1

ER -