Photochemically Catalyzed Generation of Site-Specific 8-Nitroguanine Adducts in DNA by the Reaction of Long-Lived Neutral Guanine Radicals with Nitrogen Dioxide

A novel photochemical approach is described for synthesizing site-specific 8-nitro-2'-deoxyguanosine (8-nitro-dG) adducts DNA. The method is based on the bimolecular reaction of a neutral, deprotonated guanine radical [G(-H) super( times )] in DNA and nitrogen dioxide ( super( times )NO sub(2)) radicals. This approach is illustrated using the single-stranded oligodeoxyribonucleotide 5'-d(CCATCGCTACC) dissolved in an aqueous solution of nitrite and bicarbonate anions at pH 7.5. The photochemical synthesis was triggered by the selective photodissociation of persulfate anions to yield SO super( times -) radical anions by either 308 nm XeCl excimer laser pulses or by a continuous irradiation with 290-340 nm light from a 1000 W Xe lamp. The sulfate radicals formed generate the CO super( times -) and super( times )NO sub(2) radicals by one-electron oxidation of the bicarbonate and nitrite anions. In turn, the CO super( times -) radicals site-selectively generate G(-H) super( times ) radicals in DNA that combine with super( times )NO sub(2) to form 8-nitro-dG lesions in the oligonucleotide. The nitrated oligonucleotides were purified by reversed-phase HPLC techniques and are stable at 4 degree C for at least 4 days, but depurinate at ambient temperatures of 23 degree C at pH 7 with a half-life of similar to 20 h. The nature of the reaction and decomposition products were studied by a combination of ESI and MALDI-TOF mass spectrometric techniques.