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)·] in DNA and nitrogen dioxide (·NO₂) 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 SO₄·^- 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 CO₃·^- and ·NO₂ radicals by one-electron oxidation of the bicarbonate and nitrite anions. In turn, the CO₃·^- radicals site-selectively generate G(-H)· radicals in DNA that combine with ·NO₂ to form 8-nitro-dG lesions in the oligonucleotide. The nitrated oligonucleotides were purified by reversed-phase HPLC techniques and are stable at 4 °C for at least 4 days, but depurinate at ambient temperatures of 23 °C at pH 7 with a half-life of ∼20 h. The nature of the reaction and decomposition products were studied by a combination of ESI and MALDITOF mass spectrometric techniques.