THE CHEMISTRY OF SINGLE-STRANDED 4'-DNA RADICALS - INFLUENCE OF THE RADICAL PRECURSOR ON ANAEROBIC AND AEROBIC STRAND CLEAVAGE

Background: Deoxyribosylnucleotide radicals with a radical center at t he 4'-position are important intermediates in radical-induced DNA stra nd cleavage. In the presence of O-2, these DNA radicals yield cleavage products that are partly oxidized. In the past, the postulated peroxi de intermediates could not be detected directly because they were unst able under the conditions of either radical generation, the work-up pr ocedure, or the analytical techniques used. We set out to generate and analyze these crucial intermediates in radical-induced DNA strand cle avage under mild conditions. Results: Photolysis experiments with modi fied single-stranded oligonucleotides generated 4'-DNA radicals that w ere trapped by O-2. Using MALDT-MS, DNA peroxides could be detected di rectly. Depending upon the precursor, these peroxides are formed eithe r before or after the cleavage of the single-stranded DNA radical. Rea ctions in the presence of O-18(2) and/or (H2O)-O-18 as well as subsequ ent transformations to the oxidized cleavage products confirmed the st ructure of the DNA peroxides. Conclusions: Our technique of selective DNA radical generation under mild conditions makes it possible to dete ct labile reaction products of single-stranded DNA radicals and to gai n further insight into their cleavage reactions. In cases where a radi cal pair is formed, the shielding effect protects the DNA radical from external attack so that cleavage of the single strand competes succes sfully with trapping by O-2. This shielding effect might be of general importance if the DNA radicals are generated by reagents that bind to the DNA.