Conformational Heterogeneity of Arylamine-Modified DNA:  ¹⁹F NMR Evidence

Abstract

F NMR spectroscopy was used to investigate the conformational heterogeneity of two arylamine-modified DNA duplexes, d[CTTCTTG*ACCTC]‚d [GAGGTCAAGAAG], in which G* is either N-(deoxyguanosin-8-yl)-4′-fluoro-4-aminobiphenyl (dG-C8-FABP) (I) or N-(deoxyguanosin-8-yl)-7-fluoro-2-aminofluorene (dG-C8-FAF) (II). The 19 F NMR spectrum of I showed a single peak, while that of II revealed two prominent signals with a 55:45 ratio, in good agreement with previous 1 H NMR results (Cho et al. Biochemistry 1992, 31, 9587-9602;1994, 33, 1373-1384. Slow interconversion between the two conformations of II was established by temperature-dependent two-dimensional 19 F NMR chemical exchange spectra. On the basis of magnetic anisotropy effects and isotopic solvent-induced shifts, the 19 F signals at -117.31 and -118.09 ppm in the 19 F NMR spectrum of II were assigned to a relatively undisturbed "B-type" conformer and a highly perturbed "stacked" conformer, respectively. Analysis of the temperature dependent (5-40 °C) line shapes by computer simulation yielded an interconversion barrier (∆G q ) of 14.0 kcal/mol with a chemical exchange time of 2 ms at 30 °C. This new 19 F approach should be very useful in investigating the sequence-dependent conformational heterogeneity of arylamine-modified DNA.

Examination of the Long-range Effects of Aminofluorene-induced Conformational Heterogeneity and Its Relevance to the Mechanism of Translesional DNA Synthesis

“…The 19 F NMR method takes advantage of the sensitivity of the fluorine nucleus to the tertiary structure of DNA, thereby requiring fluorine-tagged FAF as a model probe (Fig. 1a) (12). Induced circular dichroism in the 290-360 nm FAF-absorbing range (ICD 290-360nm ) serves as a sensitive conformational marker for probing AF-induced B-S-W heterogeneities (19,40).…”

“…In contrast, the N-deacetylated AF-adduct possesses flexiblility around the glycosidyl bond (χ), enabling it to adopt multiple conformational motifs depending upon the location of the aminofulorene moiety, including a major-groove binding "B-type" (B) conformation and a minor-groove binding "wedged" (W) conformation in addition to the S conformation ( Fig. 1b) (12)(13)(14)(15)(16)(17)(18). In general, fully base-paired DNA duplexes that have incorporated AF are present in an S/B equilibrium in which the tendency for the molecules to favor the S or the B conformation is dependent upon the flanking-sequence context (17)(18)(19).…”

“…The rationale for the H/D isotope effect is that the 19 F resonance of the exposed FAF residue in a B-or W-conformer should be more susceptible to solvent-induced shielding (usually > 0.2 ppm) than the buried FAF in an Sconformer (usually < 0.1 ppm) when the deuterium content is increased from 10 to 100% (12,42). While the ICD 290-360nm of a B-conformer is mostly negative, W-and S-conformers are characterized by positive ICD 290-360nm values with much greater intensity for the former (40).…”

Fluorescence Probing of Aminofluorene-Induced Conformational Heterogeneity in DNA Duplexes

“…We demonstrated previously the utility of the FAF fluorine probe for studying the structurefunction relationship of AF adducts (7,12). A H/F swap at the C7 position of the aminofluorene moiety generally had little effect on the spectroscopic and repair characteristics in different sequence contexts.…”

“…However, they are not well-suited for studying multiple conformations in dynamic equilibriums, as is the case for AF. As a remedy, we introduced dynamic 19 F nuclear magnetic resonance ( 19 F NMR) spectroscopy coupled with the fluorine probe FAF ( Figure 1A) to study AF-induced conformational heterogeneities in various DNA sequence contexts (7,11,12). Theoretical calculations showed that both the stacking and the electrostatic interactions between the carcinogen and the neighboring base pairs are equally important to the observed sequence effect (12).…”