Migration of Electrons and Holes in DNA Exposed to Low LET Irradiation: A Study of Free Radical Trapping in DNA and DNA Model Systems.A thesis byMichael T. Milano |
Abstract |
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Free radicals are intermediate precursors to stable damage in DNA exposed to ionizing radiation. Electron and hole migration and trapping are the underlying events determining the extent and distribution of radical trapping in DNA. This research elucidates the influence of variables, such as DNA hydration, conformation and packing, on the fate of electrons and holes in irradiated DNA. Electron paramagnetic resonance spectroscopy was used to examine the free radicals generated in DNA model systems irradiated at cryogenic temperatures (4 K).
In crystalline d(CGATCG)-anthracycline, only anthracycline radicals were observed, suggesting that most or all electrons and holes migrate over a two base pair range. One of the radicals was formed on the anthracycline's amino sugar group, located outside the base stack. The presence of this radical implies a mechanism of hole transfer from the DNA hydration waters to the helix backbone. The hydration dependence of free radical yields (FRYs) observed in Na DNA films and lyophilized powders argues that DNA solvation waters stabilize DNA radicals, most likely via proton transfer reactions. The difference in FRYs between DNA films and powders suggests that DNA packing is also critical. In tightly packed (powder) DNA, radicals are more likely to be scavenged onto separate DNA molecules, enabling dielectric shielding of radicals and consequent enhancement of radical trapping. This thesis is consistent with the hydration dependent decrease in Na DNA FRYs associated with increased interhelical spacing. In Li DNA, packing variability caused FRY variability of greater than �50-70%. The high yield and relative thermal stability of radicals generated in crystalline d(CCAACGTTGG) further support the importance of packing and proton transfer in radical trapping. Clearly, DNA behaves more as an insulator than a conductor. In contrast, the radicals in d(CGATCG)-anthracycline were thermally unstable, suggesting a mechanism of thermally induced anthracycline radical combination. An attempt was made to characterize the radicals formed in crystalline d(CCAACGTTGG). The analyses were compromised by relatively low spectral resolution. The spectra were consistent with electron trapping primarily at cytosine/thymine and hole trapping at guanine. Upon thermal annealing, the thymine hydrogen adduct is formed; spectral features are also consistent with a formamidopyrimidine radical, which arises from oxidized guanine. |
