Escherichia coli MTC, a NADPH cytochrome P450 reductase competent mutagenicity tester strain for the expression of human cytochrome P450: Comparison of three types of expression systems

Citation
M. Kranendonk et al., Escherichia coli MTC, a NADPH cytochrome P450 reductase competent mutagenicity tester strain for the expression of human cytochrome P450: Comparison of three types of expression systems, MUT RES-GTE, 439(2), 1999, pp. 287-300
Citations number
32
Categorie Soggetti
Molecular Biology & Genetics
Journal title
MUTATION RESEARCH-GENETIC TOXICOLOGY AND ENVIRONMENTAL MUTAGENESIS
ISSN journal
13835718 → ACNP
Volume
439
Issue
2
Year of publication
1999
Pages
287 - 300
Database
ISI
SICI code
1383-5718(19990219)439:2<287:ECMANC>2.0.ZU;2-#
Abstract
Currently three different methods have been taken to develop new mutagenici ty tester strains containing human cytochrome P450s (CYPs), Each of these u se a single expression vector. In this paper we describe a fourth approach, i.e., the coexpression of a CYP and its electron-transfer flavoprotein, NA DPH CYP reductase (RED), encoded by two different expression vectors. The E scherichia coli mutagenicity tester strain BMX100 has been expanded to a st rain, MTC which stably expresses human RED. This new tester strain permits the biplasmid coexpression of human CYP1A2 and RED (MTC1A2), This novel str ain can be used for the determination of the mutagenicity of chemicals know n to be procarcinogens and metabolized by CYP1A2, The mutagenicity tester s train MTC1A2 was compared with: (i) BMX100 using the post-mitochondrial rat liver fraction (S9); (ii) BMX100 with expressing CYP1A2 alone (iii) or wit h expressing CYP1A2 fused to rat RED or (iv) with expressing CYP1A2, bicist ronically coexpressed with rat RED, The biplasmid RED/CYP coexpression syst em generated a strain with the highest methoxy- and ethoxy-resorufin dealky lase activities and the highest mutagenic activities for the procarcinogens 2-aminoanthracene (2AA), aflatoxin BI (AFB1) and 2-amino-3-methylimidazo(4 ,5-f)quinoline (IQ). Furthermore, the metabolism of 2AA and IQ was detected more efficiently using the MTC1A2 strain than with the BMX100 strain plus the standard rodent liver S9 metabolic system. (C) 1999 Elsevier Science B. V. All rights reserved.