TY - JOUR
T1 - Escherichia coli MTC, a NADPH cytochrome P450 reductase competent mutagenicity tester strain for the expression of human cytochrome P450
T2 - Comparison of three types of expression systems
AU - Kranendonk, Michel
AU - Fisher, Charles W.
AU - Roda, Rita
AU - Carreira, Filipa
AU - Theisen, Patricia
AU - Laires, Antonio
AU - Rueff, J.
AU - Vermeulen, Nico P E
AU - Estabrook, Ronald W.
PY - 1999/2/19
Y1 - 1999/2/19
N2 - 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.
AB - 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.
KW - Biotransformation
KW - Chemical carcinogen
KW - CYP1A2
KW - E. coli
KW - Human cytochrome P450 1A2
KW - Mutagenicity
KW - NADPH cytochrome P450 reductase
UR - http://www.scopus.com/inward/record.url?scp=0033006924&partnerID=8YFLogxK
U2 - 10.1016/S1383-5718(98)00193-4
DO - 10.1016/S1383-5718(98)00193-4
M3 - Article
C2 - 10023085
AN - SCOPUS:0033006924
SN - 1383-5718
VL - 439
SP - 287
EP - 300
JO - Mutation Research - Genetic Toxicology and Environmental Mutagenesis
JF - Mutation Research - Genetic Toxicology and Environmental Mutagenesis
IS - 2
ER -