Formation of 8-oxo-7,8-dihydroguanine-radicals in gamma-irradiated DNA by multiple one-electron oxidations

Electron spin resonance (ESR) studies of radicals formed by radiation-induced multiple one-electron oxidations of guanine moieties in DNA are reported in this work. Annealing of gamma-irradiated DNA from 77 to 235 K results in the hydration of one electron oxidized guanine (G^•+) to form the 8-hydroxy-7,8-dihydroguanin-7-yl-radical (•GOH) having one β-proton coupling of 17–28 G and an anisotropic nitrogen coupling, A_‖, of ~20 G, A = 0 with g_‖ = 2.0026 and g = 2.0037. Further annealing to 258 K results in the formation of a sharp singlet at g = 2.0048 with line-width of 5.3 G that is identified as the 8-oxo-7,8-dihydroguanine one-electron-oxidized radical (8-oxo-G^•+). This species is formed via two one-electron oxidations of •GOH. These two one-electron oxidation steps leading to the formation of 8-oxo-G^•+ from •GOH in DNA, are in accordance with the expected ease of oxidation of •GOH and 8-oxo-G. The incorporation of oxygen from water in G^•+ leading to •GOH and to 8-oxo-G^•+ is verified by ESR studies employing ¹⁷O isotopically enriched water, which provide unambiguous evidence for the formation of both radicals. ESR analysis of irradiated-DNA in the presence of the electron scavenger, Tl³⁺, demonstrates that the cationic pathway leads to the formation of the 8-oxo-G^•+. In irradiated DNA–Tl³⁺ samples, Tl³⁺ captures electrons. Tl²⁺ thus produced is a strong oxidant (2.2 V), which is metastable at 77 K and is observed to increase the formation of G^•+ and subsequently of 8-oxo-G^•+ upon annealing. We find that in the absence of the electron scavenger the yield of 8-oxo-G^•+ is substantially reduced as a result of electron recombinations with G^•+ and possible reaction with •GOH.