Currently three different methods have been taken to develop new mutagenicity tester strains containing human cytochrome P450s (CYPs). Each of these use a single expression vector. In this paper we describe a fourth approach, i.e., the coexpression of a CYP and its electron-transfer flavoprotein, NADPH CYP reductase (RED), encoded by two different expression vectors. The Escherichia coli mutagenicity tester strain BMX100 has been expanded to a strain, MTC which stably expresses human RED. This new tester strain permits the biplasmid coexpression of human CYP1A2 and RED (MTC1A2). This novel strain can be used for the determination of the mutagenicity of chemicals known to be procarcinogens and metabolized by CYP1A2. The mutagenicity tester strain MTC1A2 was compared with: (i) BMX100 using the post-mitochondrial rat liver fraction (S9); (ii) BMX100 with expressing CYP1A2 alone (iii) or with expressing CYP1A2 fused to rat RED or (iv) with expressing CYP1A2, bicistronically coexpressed with rat RED. The biplasmid RED/CYP coexpression system generated a strain with the highest methoxy- and ethoxy-resorufin dealkylase activities and highest mutagenic activities for the procarcinogens 2-aminoanthracene (2AA), aflatoxin B1 (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.
|Number of pages||14|
|Journal||Mutation Research - Genetic Toxicology and Environmental Mutagenesis|
|Publication status||Published - 19 Feb 1999|
- Chemical carcinogen
- E. coli
- Human cytochrome P450 1A2
- NADPH cytochrome P450 reductase