This paper reports the solution conformation of the covalent aminofluorene-C8-deoxyguanosine [AF]dG adduct positioned opposite a -2 deletion site in a DNA oligomer duplex. The combined NMR and molecular mechanics computational studies were undertaken on the [AF]dG adduct embedded in the d(C5-[AF]G6-A7-C8)·d(G17-G18) sequence context in a duplex containing 12 residues on the modified strand and 10 on the partner strand, with no bases opposite the [AF]dG6-dA7 segment. The exchangeable and nonexchangeable protons of the aminofluorene moiety and the nucleic acid were assigned following analysis of two-dimensional NMR data sets in H2O and D2O solution. The solution conformation of the [AF]dG·2del 12-mer duplex has been determined by incorporating intramolecular and intermolecular proton-proton distances defined by upper and lower bounds deduced from NOESY spectra as restraints in molecular mechanics computations in torsion angle space. The aminofluorene ring of [AF]dG6 is intercalated between intact Waston-Crick dC5·dG18 and dC8-dG17 base pairs with the deoxyguanosine base of [AF]dG6 in a syn alignment displaced into the major groove. The syn glycosidic torsion angle at [AF]dG6 is supported by both carbon and proton chemical shift data for the sugar resonances of the modified deoxyguanosine residue. The unpaired dA7 base is also looped out of the helix into the major groove with the purine rings of [AF]dG6 and dA7 stacking on each other in the groove. The long axis of the intercalated aminofluorene ring is parallel to the long axis of the flanking dG·dC base pairs. The intercalation site is wedge shaped with a pronounced propeller-twisting and buckling of the dC5·dG18 base pair. The deoxyguanosine base of [AF]dG6, which is positioned in the major groove, is inclined relative to the helix axis and stacks over the 5′-flanking dC5 residue in the solution structure. The intercalative base displacement structure of the [AF]dG·2del 12-mer duplex exhibits several unusually shifted proton resonances that can be readily accounted for by the ring current contributions of the deoxyguanosine purine and carcinogen fluorene aromatic rings of the [AF]dG6 adduct. We note similarities between the present conformation of [AF]dG positioned opposite a -2 deletion site with our earlier conformational studies of [AF]dG positioned opposite a -1 deletion site [Mao, B., Cosman, M., Hingerty, B. E., Broyde, S., & Patel, D. J. (1995) Biochemistry 34, 6226-6238]. For both conformations, the aminofluorene carcinogen inserts into the helix at the deletion site through base displacement of the modified deoxyguanosine in a syn alignment into the major groove and directed toward its 5′-neighbor in the sequence. These structures provide a molecular explanation of how transient strand slippage of the lesioncontaining segment can be accommodated by a double helix following translesion synthesis.
|Original language||English (US)|
|Number of pages||13|
|State||Published - 1995|
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