Escherichia coli MTC, a human NADPH P450 reductase competent mutagenicity tester strain for the expression of human cytochrome P450 isoforms 1A1, 1A2, 2A6, 3A4, or 3A5: catalytic activities and mutagenicity studies

Human Cytochrome P450 Oxidoreductase Deficiency Caused by the Y181D Mutation: Molecular Consequences and Rescue of Defect

“…Coexpression of CYPOR variants with CYP1A2 in the BMX100-derived MK_1A2_POR strain of Escherichia coli was achieved using a biplasmid system described previously (Kranendonk et al, 1999). Plasmid pLCM_POR contained human WT POR cDNA, based on National Center for Biotechnology Information sequence NM_000941, encoding the CYPOR consensus sequence NP_000932, lacking the first three amino acids.…”

Functional characterization of eight human cytochrome P450 1A2 gene variants by recombinant protein expression

“…28 The CYPOR content of membrane preparations was determined using the NADPH-cytochrome c reduction assay as described previously. 29 Western blot analysis Membrane proteins (10 mg) were separated by SDS-PAGE electrophoresis (10%) and electrotransferred onto nitrocellulose membranes Hybond-ECL from GE Healthcare (Uppsala, Sweden).…”

Recombinant human P450 forms 1A1, 1A2, and 1B1 catalyze the bioactivation of heterocyclic amine mutagens in Escherichia coli lacZ strains

“…It has also been suggested that high expression of P450 and P450 reductase may reduce bacterial cell viability [Kranendonk et al, 1999;Jansson et al, 2000]. TB medium was supplemented with ampicillin (50 g/mL), ␦-ALA (0.5 mM), IPTG (1 mM), and trace element mix [Josephy, 2000] (25 L per 100 mL).…”

“…Others Kranendonk et al, 1999] have routinely used cultures grown in TB for mutagenicity ex-periments. As noted earlier, only cultures grown in TB express P450 1A2 at very high levels.…”

“…In a previous study [Kranendonk et al, 1999] comparing several P450 forms, aflatoxin mutagenicity data sets for various P450-expressing strains were compared on the basis of calculated revertants per nmol mutagen per pmol expressed P450. Kushida et al [2000] used the same approach to compare strains expressing P450 2E1 and 2A6.…”