Site-specifically located 8-amino-2 '-deoxyguanosine: thermodynamicstability and mutagenic properties in Escherichia coli

摘要

2-Nitropropane (2-NP), an important industrial solvent and a component of cigarette smoke, is mutagenic in bacteria and carcinogenic in rats, 8-Amino-2'-deoxyguanosine (8-amino-dG) is one of the types of DNA damage found in liver, the target organ in 2-NP-treated rats. To investigate the thermodynamic properties of 8-amino-dG opposite each of the four DNA bases, we have synthesized an 11mer, d(CCATCG*CTACC), in which G* represents the modified base, By annealing a complementary DNA strand to this modified 11mer, four sets of duplexes were generated each containing one of the four DNA bases opposite the lesion, Circular dichroism studies indicated that 8-amino-dG did not alter the global helical properties of natural right-handed B-DNA, The thermal stability of each duplex was examined by UV melting measurements and compared with its unmodified counterpart. For the unmodified 11mer, the relative stability of the complementary DNA bases opposite G was in the order C > T > G > A, as determined from their -DeltaG degrees values. The free energy change of each modified duplex was lower than its unmodified counterpart, except for the G*:G pair that exhibited a higher melting transition and a larger -DeltaG degrees than the G:G duplex. Nevertheless, the stability of the modified 11mer duplex also followed the order C > T > G > A when placed opposite 8-amino-dG. To explore if 8-amino-dG opposite another 8-amino-dG has any advantage in base pairing, a G*:G* duplex was evaluated, which showed that the stability of this duplex was similar to the G*:G duplex. Mutagenesis of 8-amino-dG in this sequence context was studied in Escherichia coli, which showed that the lesion is weakly mutagenic (mutation frequency similar to 10(-3)) but still can induce a variety of targeted and semi-targeted mutations.