Differential excision from DNA of the C-8 and N2 guanosine adducts of N-acetyl-2-aminofluorene by single strand-specific endonucleases

Abstract

Purified duck reticulocyte DNA was reacted in vitro with [9-14C]-N-acetoxy-N-acetyl-2-aminofluorene. Hydrolysis of the [14C]-N-acetyl-2-aminofluorene-modified DNA followed by Sephadex LH-20 column chromatography showed that 85% of the DNA-bound [14C]-N-acetyl-2-aminofluorene was N-(deoxyguanosin-8-yl)-N-acetyl-2-aminofluorene and 15% was 3-(deoxyguanosin-N2-yl)-N-acetyl-2-aminofluorene. When this modified DNA was incubated with the single strand-specific nuclease, S1, and the undigested fraction of the DNA was analyzed, there was preferential loss of the quanosine C-8 adduct from the DNA. Moreover, analysis of the nucleosides released by exposure of N-acetyl-2-aminofluorene-modified DNA to a single strand-specific nuclease from Neurospora crassa showed only the guanosine C-8 adduct in the supernatant fraction. These results suggest that, whereas the N-(deoxyguanosin-8-yl)-N-acetyl-2-aminofluorene adduct in DNA causes major conformational changes in the double-stranded helix and localized regions of denaturation, the 3-(deoxyguanosin-N2-yl)-N-acetyl-2-aminofluorene adduct does not cause major distortions of the native DNA structure.