Distant Neighbor Base Sequence Context Effects in Human Nucleotide Excision Repair of a Benzo[a]pyrene-derived DNA Lesion

Abstract

The effects of non-nearest base sequences, beyond the nucleotides flanking a DNA lesion on either side, on nucleotide excision repair (NER) in extracts from human cells were investigated. We constructed two duplexes containing the same minor groove-aligned 10S (+)-trans-anti-B[a]P-N2-dG (G⁎) DNA adduct, derived from the environmental carcinogen benzo[a]pyrene (B[a]P): 5′-C-C-A-T-C-G⁎-C-T-A-C-C-3′ (CG⁎C-I), and 5′-C-A-C3-A4-C5-G⁎-C-A-C-A-C-3′ (CG⁎C-II). We used polyacrylamide gel electrophoresis to compare the extent of DNA bending, and molecular dynamics simulations to analyze the structural characteristics of these two DNA duplexes. The NER efficiencies are 1.6(±0.2)-fold greater in the case of the CG⁎C-II than the CG⁎C-I sequence context in 135-mer duplexes. Gel electrophoresis and self-ligation circularization experiments revealed that the CG⁎C-II duplex is more bent than the CG⁎C-I duplex, while molecular dynamics simulations showed that the unique -C3-A4-C5- segment in the CG⁎C-II duplex plays a key role. The presence of a minor groove-positioned guanine amino group, the Watson–Crick partner to C3, acts as a wedge; facilitated by a highly deformable local -C3-A4- base step, this amino group allows the B[a]P ring system to produce a more enlarged minor groove in CG⁎C-II than in CG⁎C-I, as well as a local untwisting and enlarged and flexible Roll only in the CG⁎C-II sequence. These structural properties fit well with our earlier findings that in the case of the family of minor groove 10S (+)-trans-anti-B[a]P-N2-dG lesions, flexible bends and enlarged minor groove widths constitute NER recognition signals, and extend our understanding of sequence context effects on NER to the neighbors that are distant to the lesion.