Certain transition metals like nickel, chromium, cadmium, copper and cobalt are carcinogenic to humans and/or animals. Among others, their effects include transgenerational carcinogenesis in the progeny of fathers exposed to welding fumes and other metal dusts. However, mechanisms of the carcinogenic activity of these metals remain obscure. In recent years, we have formulated a novel hypothesis that one such mechanism would involve metal-mediated oxidative damage to DNA and nuclear proteins and found that the metals did induce promutagenic DNA base alterations typical for active oxygen attack. In 1996/97 we continued mechanistic in vitro studies of that hypothesis. To complete our investigations of Ni(II) interactions with histone H3, reported last year, we have confirmed the formation of Ni(II) complexes exclusively with the -CAIH- motif of this histone in a core histone tetramer, composed of two histone H3 molecules and two histone H4 molecules. Our further research was focused on chelation of Ni(II) and Cu(II) by the N-terminal motif of human protamine HP2, the major DNA- binding structural protein in the sperm nucleus. Such binding may damage the sperm DNA and thus become mechanistically involved in the transgenerational carcinogenesis. Using a synthetic pentadecapeptide modeling the N-terminus of HP2 (HP2-15), we characterized the Ni(II) and Cu(II) complexes with HP2-15, finding its Arg-Thr-His- motif to be the sole metal-binding site, and tested their catalytic potential toward oxidation of 2'-deoxyguanosine (dG) to promutagenic 8-oxo-dG in free dG and in DNA. The chelation of Ni(II) by HP2-15 enhanced the generation of 8-oxo-dG by hydrogen peroxide from both free and DNA-bound dG. In contrast, the complexation of Cu(II) inhibited the dG oxidation with hydrogen peroxide, as compared with unchelated Cu(II). Similar effects were observed for the oxidative single- and double strand breaking of DNA. Moreover, the complexed metals promoted oxidative degradation of the ligand itself, consisting of decreasing contents of its His (down to 50% of the control) and especially Tyr (down to 19% of the control) residues, and the appearance of aspartic acid (up to 22% vs the Gly residue). The above results support the oxidative hypothesis of metal- mediated toxicity and carcinogenesis. Also, our research provides an experimental basis for collaborative studies on oxidative damage by other carcinogenic insults (see projects Z01 BC 05301 and Z01 BC 05352).

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National Cancer Institute (NCI)
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National Cancer Institute Division of Basic Sciences
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