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2-amino-3-methylimidazo[4,5-f]quinoline (IQ) is a highly mutagenic heterocyclic amine formed in all cooked meats. IQ has been found to be a potent inducer of frameshift mutations in bacteria and carcinogenic in laboratory animals. Upon metabolic activation, IQ forms covalent adducts at the C8- and N2-positions of deoxyguanosine with a relative ratio of up to approximately 4:1. We have previously incorporated the major dGuo-C8-IQ adduct into oligonucleotides through the corresponding phosphoramidite reagent. We report here the sequence-specific synthesis of oligonucleotides containing the minor dGuo-N2-IQ adduct. Thermal melting analysis revealed that the dGuo-N2-IQ adduct significantly destabilizes duplex DNA.

2-Amino-3-methylimidazo[4,5-f]quinoline (IQ) is found in cooked meats and forms DNA adducts at the C8- and N2-positions of dGuo after appropriate activation. IQ is a potent inducer of frameshift mutations in bacteria and is carcinogenic in laboratory animals. We have incorporated both IQ-adducts into the G1- and G3-positions of the NarI recognition sequence (5'-G1G2CG3CC-3'), which is a hotspot for arylamine modification. The in vitro replication of the oligonucleotides was examined with Escherichia coli pol I Klenow fragment exo-, E. coli pol II exo-, and Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4), and the extension products were sequenced by tandem mass spectrometry. Replication of the C8-adduct at the G3-position resulted in two-base deletions with all three polymerases, whereas error-free bypass and extension was observed at the G1-position. The N2-adduct was bypassed and extended by all three polymerases when positioned at the G1-position, and the error-free product was observed. The N2-adduct at the G3-position was more blocking and was bypassed and extended only by Dpo4 to produce an error-free product. These results indicate that the replication of the IQ-adducts of dGuo is strongly influenced by the local sequence and the regioisomer of the adduct. These results also suggest a possible role for pol II and IV in the error-prone bypass of the C8-IQ-adduct leading to frameshift mutations in reiterated sequences, whereas noniterated sequences result in error-free bypass.

The site-specific synthesis of oligonucleotides containing the C8-deoxyguanosine adduct of the highly mutagenic heterocyclic amine 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) has been achieved, and the oligonucleotides were characterized by UV melting temperature analysis, circular dichroism, and UV absorption spectroscopy. Examination of these data indicated that the IQ-adduct is accommodated in dramatically different environments. This sequence-dependent conformational preference is likely to play a key role in the mutagenicity and repair of IQ-modified oligonucleotides.