Thermodynamic and mutagenesis studies of 8-amino-2'-deoxyguanosine, a lesion generated by 2-nitropropane
Date of Completion
2-nitropropane, an important industrial solvent and a component of cigarette smoke, is mutagenic in bacteria and carcinogenic in rats. 8-amino-2 ′-deoxyguanosine is one of the types of DNA damage found in liver, the target organ of 2-NP treated rats. It has been shown to be mutagenic producing mainly G → T transversions when studied in mammalian cells. ^ To investigate the thermodynamic properties of 8-amino-2′ -deoxyguanosine, we synthesized an 11-mer, d(CCATCG*CTACC), in which G* represents the modified base. By annealing a complementary DNA strand to this modified 11-mer, four sets of duplexes were generated, each containing one of the four DNA bases opposite the lesion. The thermal stability of each duplex was examined by UV melting measurements and compared to its unmodified counterpart. ^ The relative stability of the different DNA bases opposite 8-aminoguanine was in the same order as that observed for the normal guanine base (C > T > G > A). The 8-amino-2′-deoxyguanosine lesion was shown to reduce duplex thermal stability relative to the corresponding lesion-free duplex, except in the case of G*:G duplex, which was found to be slightly more stable than the G:G duplex. This suggests that the amination of C8 position of guanine destabilizes the helix but does not cause severe perturbations and that the G*:G pair may have some additional favorable interactions relative to the G:G mismatched pair. In addition, a G*:G* duplex was also evaluated and the results from that study indicated that even though G*:G* is more stable than the G:G pair, the difference is small and it does not appear to stabilize the duplex by some novel ordered structure. ^ Using recombinant DNA techniques, 8-amino-dG was incorporated into an M13 bacteriophage at a preselected site. The lesion containing single stranded DNA was transfected into Escherichia coli and the cytotoxicity and mutagenicity were evaluated in two different sequences (the above mentioned 11-mer and a 6-mer). In both cases, the genome containing the lesion showed almost 100% survival, indicating that the lesion is not cytotoxic. Also, the lesion was found to be weakly mutagenic in the 11-mer and non-mutagenic in the 6-mer sequence. ^
Venkatarangan, Lata Mahadevan, "Thermodynamic and mutagenesis studies of 8-amino-2'-deoxyguanosine, a lesion generated by 2-nitropropane" (2001). Doctoral Dissertations. AAI9999699.