Bleomycin versus OH-radical-induced malonaldehydic-product formation in DNA

Purpose: To compare the actions of bleomycin and ionizing radiation on DNA regarding the formation of malonaldehyde-like products. Materials and methods: Calf thymus DNA was treated with iron/bleomycin or g amma-radiation at pH 7. Products Mere analysed by HPLC. The thiobarbituric- acid reactivity of the samples was determined directly or after HPLC by pos t-column derivatization. ESI mass spectra were taken on-line following HPLC . Results: Malonaldehyde and malonaldehyde-like products as detected by the s ensitive 2-thiobarbituric acid (TBA) assay are formed in gamma-irradiated D NA and thymidine solutions as well as upon treatment of DNA with bleomycin/ iron. In gamma-irradiated DNA solutions in the presence of oxygen, no base propenals were detected, and the major TEA-active product was malonaldehyde . In the gamma-radiolysis of thymidine, thymine propenal was formed only in traces (not more than 0.07 per cent of the OH-radical yield). Malonaldehyd e was practically absent after treatment with bleomycin; three other TEA-ac tive products were seen by HPLC which have been identified as the cytosine thymine and adenine propenals. Guanine propenal was not detected under our conditions. Conclusions: The absence of these base propenals upon gamma-radiolysis impl ies that although the initiating step of OH-radical and bleomycin action [i .e. H-abstraction at C(4')] may be the same, the bleomycin-iron complex mus t participate in subsequent steps en route to the base propenals. It is pro posed that the bleomycin pathway may involve the interaction of the C(4')-p eroxyl radical with the 'spent' bleomycin-iron complex by ligand exchange, under formation of a bleomycin-iron-peroxyl-radical complex, Blm(Fe4+, (OOR )-O-.), which then decomposes by heterolysis into the alkoxy cation precurs or +OR of the base propenal and reconstitution of the bleomycin-iron comple x Blm(Fe, O)(3+), i.e. gives rise to base propenal formation without the in volvement of a C(4')-hydroperoxide.