Characterization of a Lipid Hydroperoxide-Derived RNA Adduct in Rat Intestinal Epithelial Cells

Five major products (adducts A_1a, A_1b, A₂, A_3, and B) from the reaction of guanosine (Guo) with 4-oxo-2(E)-nonenal (ONE) were detected by liquid chromatography-mass spectrometry (LC-MS). Tandem MS (MS/MS) analysis of these compounds suggested that modifications to the nucleoside had been introduced. Adducts A_1a, A_1b, A₂, and A₃ were heptanone-ethano-2‘-Guo adducts that all decomposed to adduct B. Adducts A_1a and A_1b were isomeric hemi-ketal forms. Adducts A₂ and A₃ were diastereomers of the open chain ketone form. The structure of adduct B was shown by LC-MS/MS and NMR spectroscopy to be the heptanone-etheno-Guo (HεGuo) adduct, 3-(d-erythropentafuranosyl)imidazo-7-(heptane-2‘ ‘-one)-9-hydroxyl[1,2-α]purine. The overall reaction of Guo with ONE was very similar to its reaction with 2‘-deoxyguanosine. Reaction of ONE with yeast transfer RNA also resulted in the formation of HεGuo. Finally, HεGuo was detected and quantified in the RNA from rat intestinal epithelial cells that stably express cyclooxygenase-2. These data show that RNA is modified by the same bifunctional reactive electrophiles derived from lipid peroxidation that covalently modify DNA.