STRUCTURAL ORIGINS OF BULKY OXIDATIVE DNA-ADDUCTS (TYPE-II I-COMPOUNDS) AS DEDUCED BY OXIDATION OF OLIGONUCLEOTIDES OF KNOWN SEQUENCE

Bulky DNA adducts, previously termed type II I-compounds, are detected by P-32-postlabeling following treatment of DNA with several Fenton-t ype oxygen radical-generating reagents, i.e., mixtures of Fe(II) or Ni (II) and H2O2. In an attempt to characterize the chemical nature and m echanism(s) of formation of these novel adducts, 16 single-stranded de oxyribooligonucleotides (20- and 21-mers) of known sequence were oxidi zed with Fe(II) or Ni(II) and H2O2, and the products were analyzed by P-32-postlabeling. Eight adducts were obtained reproducibly by oxidati on of DNA and test oligonucleotides in a sequence-dependent manner. On e major adduct (2) was formed only if the test oligonucleotide contain ed two adjacent adenine residues. Similarly, adducts 3 and 8 specifica lly originated in AC and CA sequences, respectively. Adduct 6 required a 5'-C-purine-3' sequence. On the other hand, GN sequences (where N i s any normal nucleotide) gave rise to adduct 1, another major product, and adduct 7. Similarly, adducts 4 and 5 were produced by the oxidati on of AN sequences. These observations are most readily explained if t he oxidation reactions caused intrastrand cross-links between adjacent nucleotides, leading to dimer formation. The observation that adducts 1, 4, 5, and 7 did not require a specific S'-nucleotide was consisten t with the notion that these nucleotides lacked a S'-base, suggesting the presence of a 5'-->3' purine-sugar cross-link in the oxidized prod ucts. The majority of the lesions came from AA and 5'-purine-N-3' sequ ences. The effects of Fe(II) and Ni(II) were qualitatively similar; ho wever, higher yields of products were observed with Fe(II) as the cata lyst. The definition of the chemical origins of these bulky DNA modifi cations, which represent a new type of DNA damage, is expected to cont ribute to a better understanding of the mechanism of metal carcinogene sis and to shed light upon the origins of certain endogenous DNA lesio ns. Recently, some of the major oxidative DNA adducts characterized he re were detected by P-32-postlabeling in the renal DNA of male rats tr eated with ferric nitrilotriacetate, a known potent prooxidative kidne y carcinogen in these animals [Randerath, E., Watson, W. P., Zhou, G. D., Chang, J., and Randerath, K. (1995) Mutat. Res. 341, 265-279].