Plasmid-mediated expression of the UmuDC mutagenesis proteins in an Escherichia coli strain engineered for human cytochrome P450 1A2-catalyzed activation of aromatic amines

The mutagenic actions of many chemicals depend on the activities of bacteri al "mutagenesis proteins", which allow replicative bypass of DNA lesions. G enes encoding these proteins occur on bacterial chromosomes and plasmids, o ften in the form of an operon (such as umuDC or mucAB) encoding two protein s. Many bacterial strains used in mutagenicity testing carry mutagenesis pr otein genes borne on plasmids, such as pKM101. Our objective was to introdu ce mutagenesis protein function into Escherichia coli strain DJ4309. This s train expresses recombinant human cytochrome P450 1A2 and NADPH-P450 reduct ase and carries out the metabolic conversion of aromatic and heterocyclic a mines into DNA-reactive mutagens. We discovered that many mutagenesis-prote in plasmids severely inhibit the response of strain DJ4309 to 2-amino-3,4-d imethylimid-azo[4,5-f]quinoline (MeIQ), a typical heterocyclic amine mutage n. Among many plasmids examined, one, pGY8294, a pSC101 derivative carrying the umuDC operon, did not inhibit MeIQ mutagenesis. Strain DJ4309 pGY8294 expresses active mutagenesis proteins, as shown by its response to mutagens such as 1-nitropyrene and 4-nitroquinoline 1-oxide (4-NQO), and is as sens itive as the parent strain DJ4309 to P450-dependent mutagens, such as MeIQ and 1-aminopyrene. (C) 1999 Elsevier Science B.V. All rights reserved.