Conformational differences of the C8-deoxyguanosine adduct of 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) within the NarI recognition sequence

Abstract

2-amino-3-methylimidazo[4,5-f]quinoline (IQ) is a highly mutagenic heterocyclic amine found in cooked meats. The major DNA adduct of IQ is at the C8-position of dGuo. We have previously reported the incorporation of the C8-IQ adduct into oligonucleotides, namely, the G1-position of codon 12 of the N-ras oncogene sequence (G1G2T) and the G3-position of the NarI recognition sequence (G1G2CG3CC) (Elmquist et al. (2004) J. Am. Chem. Soc. 126, 11189-11201). Ultraviolet spectroscopy and circular dichroism studies indicated that the conformation of the adduct in the two oligonucleotides was different, and they were assigned as groove-bound and base-displaced intercalated, respectively. The conformation of the latter was subsequently confirmed through NMR and restrained molecular dynamics studies (Wang et al. (2006) J. Am. Chem. Soc. 128, 10085-10095). We report here the incorporation of the C8-IQ adduct into the G1- and G2-positions of the NarI sequence. A complete analysis of the UV, CD, and NMR chemical shift data for the IQ protons are consistent with the IQ adduct adopting a minor groove-bound conformation at the G1- and G2-positions of the NarI sequence. To further correlate the spectroscopic data with the adduct conformation, the C8-aminofluorene (AF) adduct of dGuo was also incorporated into the NarI sequence; previous NMR studies demonstrated that the AF-modified oligonucleotides were in a sequence-dependent conformational exchange between major groove-bound and base-displaced intercalated conformations. The spectroscopic data for the IQ- and AF-modified oligonucleotides are compared. The sequence-dependent conformational preferences are likely to play a key role in the repair and mutagenicity of C8-arylamine adducts.

Figure 1

C8-dGuo adducts of IQ, AF, and AAF.

Figure 2

(A–C) CD spectra of the unmodified duplex (—), the C8-IQ-modified single strand (---), and the C8-IQ-modified duplex (---) oligonucleotides at positions G₁ (2a, panel A), G₂ (3a, panel B), and G₃ (4a, panel C) of the NarI restriction sequence. (D–F) UV spectra of the C8-modified single strand (---) and double strand (—) oligonucleotides at positions G₁ (2a, panel D), G₂ (3a, panel E), and G₃ (4a, panel F) of the NarI restriction sequence.

Figure 3

(A–C) CD spectra of the C8-AF-modified single strand (---) and duplex (---) oligonucleotides at positions G₁ (2b, panel A), G₂ (3b, panel B), and G₃ (4b, panel C) of the NarI restriction sequence. (D–F) UV spectra of the C8-AF-modified single strand (–––) and double strand (—) oligonucleotides at positions G₁ (2b, panel D), G₂ (3b, panel E), and G₃ (4b, panel F) of the NarI restriction sequence.

Figure 4

Expanded plots of the COSY (A) and NOESY (B) spectra showing the IQ aromatic region of duplex 4a. The NOESY spectrum was recorded at 15 °C with the mixing time at 90 ms or 25 °C with a mixing time of 200 ms. 1 and 2 are the NOESY cross-peaks between the IQ N-CH₃ group and the 3′-flanking Cyt H5 and 3′-flanking Gua H8 of the complementary strand, respectively, and are consistent with a base-displaced intercalated conformation.