Ionizing radiation produces a wide spectrum of DNA damages to the base and sugar moieties of DNA. The spectrum of damages induced by ionizing radiation in aqueous solution is affected by the dissolved gases, such as oxygen. Damages such as dihydrothymine, a-anomers, cyclopurines and pyrimidines are only formed under anoxic conditions. While the biological processing of a number of the oxic DNA damages are well understood, the repair and removal of most of the anoxic lesions remains unclear. It is well known that cell survival increased under hypoxia irradiation. The long range goal of this research is to elucidate mechanism of processing of DNA damage induced by ionizing radiation in hypoxic cells E. coli cells deficient in different repair enzymes will be studied with respect to their sensitivities to killing by ionizing radiation under anoxic and oxic conditions. This study should aid in the process of identifying novel enzymes that might be involved in the biological processing of these anoxic lesions. Duplex DNA containing unique anoxic lesions such as dihydrothymine, alpha-dA, 8,5'-cyclopurines and model DNA-protein crosslinks will be constructed and used as substrates for screening of enzymatic activities that might recognize these lesions. Novel enzymatic activities detected will be purified and characterized. The biological processing of these lesions will also be studied by the survival of plasmid DNA containing these lesions in E. coli hosts of different repair proficiency. The in vitro and in vivo processing of another important class of lesion, locally multiply damage sites, will also be studied in a similar manner. It is hoped that through a combination of enzymological and genetic studies, the mechanism of increased survival in hypoxic cells can be elucidated. The long range goal of this project is to develop methods that can enhance the radiation killing of hypoxic tumor cells and thus the therapeutic efficacy of radiotherapy.
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