Abstract
Damaged DNA precursors are formed by reactive oxygen species induced by ionizing radiation. After damaged DNA precursors are incorporated into DNA, they might be removed by DNA repair enzymes. In this study, we examined whether a nucleotide excision repair enzyme, UvrABC, could suppress mutations induced by oxidized nucleotides in vivo .
We introduced oxidized DNA precursors, 8-hydroxy-dGTP and 2-hydroxy-dATP, into uvrA and uvrB Escherichia coli strains, and analyzed mutations in the chromosomal rpoB gene, as a mutagenesis target. Oxidized DNA precursors induced mutations in the wild type, but not in the uvrA and uvrB strains. Next, mutT, mutT/uvrA, and mutT/uvrB E. coli strains were treated with hydrogen peroxide (H2O2), and the rpoB mutant frequencies were calculated. Frequency of the H2O2-induced mutations was increased in the all strains tested, and the increase was two- to three-fold lower in the mutT/uvrA and mutT/uvrB strains than in the mutT strain. Thus, it is suggested that the UvrABC nuclease is involved in fixation, but not suppression, of the mutations induced by oxidized nucleotides. We hypothesized that incision of the complementary strand, not that of the strand containing oxidized base damage, by UvrABC might fix the mutation. Although duplex oligonucleotides containing 8-hydroxyguanine and 2-hydroxyadenine were treated with the purified Thermus thermophilus HB8 UvrABC proteins, the expected activity was not observed. Thus, the UvrABC proteins would fix the mutation by another mechanism.
