NMR EVIDENCE OF THE STABILIZATION BY THE CARCINOGEN N-2-ACETYLAMINOFLUORENE OF A FRAMESHIFT MUTAGENESIS INTERMEDIATE

Two heteroduplexes (12)T(13)G(14)T(15)G(16)G(17)A(18)G(19)T(20)G(21)) containing a bulged guanine either unmodified or modified with the car cinogen N-2-acetylaminofluorene (AAF) have been studied by nuclear mag netic resonance (NMR) as models of slipped mutagenic intermediates (SM I). Conformational equilibria are observed in both the unmodified and the AAF-modified heteroduplexes. The major conformation of the unmodif ied duplex is one where the extra guanine is stacked in the helix and the major conformation of the AAF-modified heteroduplex is one where t he AAF is external to the helix. Unusual sugar proton chemical shifts of C5 - and G6-AAF indicate that the AAF ring is pointing out in the 5 ' direction. A strong increase in the modified heteroduplex melting te mperature (+ 15 degrees C) is observed. Moreover, in contrast to the u nmodified heteroduplex, which shows extensive melting in the vicinity of the bulged guanine, the base pairs around the bulge in the AAF-modi fied heteroduplex remain paired at temperatures up to 30 degrees C. Th is exceptional stability of the site around the bulged modified guanin e is suggested to be responsible for the high rate of -1 mutation indu ced by AAF at repetitive sequences.