The overall goal of the proposed research is to probe the DNA mutagenicity and repair of three lesions that are produced in nucleic acids via a variety of oxidative stress mechanisms. These lesions all lack the presence of a nucleobase, and result from formal oxidation of the carbohydrate moiety. The presence of these oxidized abasic lesions in DNA in vivo is associated with cancer and aging. Despite their importance, the effects of these abasic lesions on the function of DNA at the molecular level are not well understood. Using the fundamental knowledge gained from these and previous related studies, we are designing enzyme inhibitors of DNA repair processes. First generation molecules that inhibit the lyase step of base excision repair (BER) enzymes are presented in this proposal. Our general experimental approach involves the unambiguous synthesis (and characterization) of oligonucleotides containing individual oxidized abasic lesions site specifically incorporated. The effects of the lesions on duplex stability and enzyme activity are examined using these chemically synthesized substrates.
Specific aims i nclude: 1. Investigation of the in vitro and in vivo mutagenicity of the lesions. 2. Investigation of the repair of these lesions by BER enzymes in vitro and in vivo. 3. Examination of the lethality of these lesions in E. coli and in yeast. 4. Design of suicide inhibitors of BER. Increased understanding of the effects of oxidized abasic lesions on nucleic acid structure and function will be useful for understanding the association between nucleic acid damage and aging, as well as the etiology of diseases such as cancer. Application of these studies, such as the design of enzyme inhibitors may provide new therapeutic agents.
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