Limits...
Conformational and thermodynamic properties modulate the nucleotide excision repair of 2-aminofluorene and 2-acetylaminofluorene dG adducts in the NarI sequence.

Jain V, Hilton B, Patnaik S, Zou Y, Chiarelli MP, Cho BP - Nucleic Acids Res. (2012)

Bottom Line: Our (19)F-NMR/ICD results showed that FAAF at G(1) and G(3) prefer syn S- and W-conformers, whereas anti B-conformer was predominant for G(2).The melting and thermodynamic data indicate that the S- and W-conformers produce greater DNA distortion and thermodynamic destabilization.The present results provide valuable conformational insight into the sequence-dependent UvrABC incisions of the bulky aminofluorene DNA adducts.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedical and Pharmaceutical Sciences, University of Rhode Island, Kingston, RI 02881, USA.

ABSTRACT
Nucleotide excision repair (NER) is a major repair pathway that recognizes and corrects various lesions in cellular DNA. We hypothesize that damage recognition is an initial step in NER that senses conformational anomalies in the DNA caused by lesions. We prepared three DNA duplexes containing the carcinogen adduct N-(2'-deoxyguanosin-8-yl)-7-fluoro-2-acetylaminofluorene (FAAF) at G(1), G(2) or G(3) of NarI sequence (5'-CCG(1)G(2)CG(3)CC-3'). Our (19)F-NMR/ICD results showed that FAAF at G(1) and G(3) prefer syn S- and W-conformers, whereas anti B-conformer was predominant for G(2). We found that the repair of FAAF occurs in a conformation-specific manner, i.e. the highly S/W-conformeric G(3) and -G(1) duplexes incised more efficiently than the B-type G(2) duplex (G(3)∼G(1)> G(2)). The melting and thermodynamic data indicate that the S- and W-conformers produce greater DNA distortion and thermodynamic destabilization. The N-deacetylated N-(2'-deoxyguanosin-8-yl)-7-fluoro-2-aminofluorene (FAF) adducts in the same NarI sequence are repaired 2- to 3-fold less than FAAF: however, the incision efficiency was in order of G(2)∼G(1)> G(3), a reverse trend of the FAAF case. We have envisioned the so-called N-acetyl factor as it could raise conformational barriers of FAAF versus FAF. The present results provide valuable conformational insight into the sequence-dependent UvrABC incisions of the bulky aminofluorene DNA adducts.

Show MeSH

Related in: MedlinePlus

19F-NMR spectra of (a) FAAF-modified NarI 16-mer, (b) FAAF-modified non-NarI 12-mer and (c) FAF-modified NarI 12-mer duplexes at 5°C. *unknown conformers; #impurity.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3351159&req=5

gkr1307-F4: 19F-NMR spectra of (a) FAAF-modified NarI 16-mer, (b) FAAF-modified non-NarI 12-mer and (c) FAF-modified NarI 12-mer duplexes at 5°C. *unknown conformers; #impurity.

Mentions: Figure 4a shows the 19F-NMR spectra of FAAF–NarI 16-mer G1-, G2- and G3-duplexes measured at 5°C, in which 19F signals are in slow chemical exchange. These NarI-FAAF duplexes exhibited three to five 19F signals, each representing a particular conformation. The percent population ratios shown were calculated on the basis of line simulations as shown in Supplementary Figure S9. Assignments of the different 19F signals of each duplex were necessary to carry out meaningful structure-activity-relationship studies. The signal assignments in Figure 4a were made initially on the basis of chemical exchange, ring current effect and chemical shift pattern recognition as have been done for a number of FAAF- and FAF-adducts in various sequence contexts (30,35). It has been demonstrated that AF and AAF adducts adopt the S/B- and S/B/W-conformational equilibrium, respectively (Figure 1c and d) and their 19F chemical shifts are independent of overall sequence and its length, but strongly rely on the nature of the bases flanking the lesion (15,30). The major 19F signals in Figure 4a correlate well with the S/B/W-profiles reported previously for FAAF adducts (15), i.e. B-, S- and W-conformers going from downfield to upfield, i.e. −115.0 to −115.5, −115.5 to −117.0 and −117.0 to −118.0 ppm, respectively. Additional signals were observed in the −114.0∼−115.0 ppm range for the NarI–G2 and –G3 duplexes (Figure 4a, see Supplementary Table S2 for exact chemical shifts). Consistent with this observation, their proton spectra displayed a mixture of broad imino signals arising not only from those involved in Watson–Crick hydrogen bonds (12–14 ppm), but also from the lesion site and its vicinity (11–12 ppm) (Supplementary Figure S10).Figure 4.


Conformational and thermodynamic properties modulate the nucleotide excision repair of 2-aminofluorene and 2-acetylaminofluorene dG adducts in the NarI sequence.

Jain V, Hilton B, Patnaik S, Zou Y, Chiarelli MP, Cho BP - Nucleic Acids Res. (2012)

19F-NMR spectra of (a) FAAF-modified NarI 16-mer, (b) FAAF-modified non-NarI 12-mer and (c) FAF-modified NarI 12-mer duplexes at 5°C. *unknown conformers; #impurity.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3351159&req=5

gkr1307-F4: 19F-NMR spectra of (a) FAAF-modified NarI 16-mer, (b) FAAF-modified non-NarI 12-mer and (c) FAF-modified NarI 12-mer duplexes at 5°C. *unknown conformers; #impurity.
Mentions: Figure 4a shows the 19F-NMR spectra of FAAF–NarI 16-mer G1-, G2- and G3-duplexes measured at 5°C, in which 19F signals are in slow chemical exchange. These NarI-FAAF duplexes exhibited three to five 19F signals, each representing a particular conformation. The percent population ratios shown were calculated on the basis of line simulations as shown in Supplementary Figure S9. Assignments of the different 19F signals of each duplex were necessary to carry out meaningful structure-activity-relationship studies. The signal assignments in Figure 4a were made initially on the basis of chemical exchange, ring current effect and chemical shift pattern recognition as have been done for a number of FAAF- and FAF-adducts in various sequence contexts (30,35). It has been demonstrated that AF and AAF adducts adopt the S/B- and S/B/W-conformational equilibrium, respectively (Figure 1c and d) and their 19F chemical shifts are independent of overall sequence and its length, but strongly rely on the nature of the bases flanking the lesion (15,30). The major 19F signals in Figure 4a correlate well with the S/B/W-profiles reported previously for FAAF adducts (15), i.e. B-, S- and W-conformers going from downfield to upfield, i.e. −115.0 to −115.5, −115.5 to −117.0 and −117.0 to −118.0 ppm, respectively. Additional signals were observed in the −114.0∼−115.0 ppm range for the NarI–G2 and –G3 duplexes (Figure 4a, see Supplementary Table S2 for exact chemical shifts). Consistent with this observation, their proton spectra displayed a mixture of broad imino signals arising not only from those involved in Watson–Crick hydrogen bonds (12–14 ppm), but also from the lesion site and its vicinity (11–12 ppm) (Supplementary Figure S10).Figure 4.

Bottom Line: Our (19)F-NMR/ICD results showed that FAAF at G(1) and G(3) prefer syn S- and W-conformers, whereas anti B-conformer was predominant for G(2).The melting and thermodynamic data indicate that the S- and W-conformers produce greater DNA distortion and thermodynamic destabilization.The present results provide valuable conformational insight into the sequence-dependent UvrABC incisions of the bulky aminofluorene DNA adducts.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedical and Pharmaceutical Sciences, University of Rhode Island, Kingston, RI 02881, USA.

ABSTRACT
Nucleotide excision repair (NER) is a major repair pathway that recognizes and corrects various lesions in cellular DNA. We hypothesize that damage recognition is an initial step in NER that senses conformational anomalies in the DNA caused by lesions. We prepared three DNA duplexes containing the carcinogen adduct N-(2'-deoxyguanosin-8-yl)-7-fluoro-2-acetylaminofluorene (FAAF) at G(1), G(2) or G(3) of NarI sequence (5'-CCG(1)G(2)CG(3)CC-3'). Our (19)F-NMR/ICD results showed that FAAF at G(1) and G(3) prefer syn S- and W-conformers, whereas anti B-conformer was predominant for G(2). We found that the repair of FAAF occurs in a conformation-specific manner, i.e. the highly S/W-conformeric G(3) and -G(1) duplexes incised more efficiently than the B-type G(2) duplex (G(3)∼G(1)> G(2)). The melting and thermodynamic data indicate that the S- and W-conformers produce greater DNA distortion and thermodynamic destabilization. The N-deacetylated N-(2'-deoxyguanosin-8-yl)-7-fluoro-2-aminofluorene (FAF) adducts in the same NarI sequence are repaired 2- to 3-fold less than FAAF: however, the incision efficiency was in order of G(2)∼G(1)> G(3), a reverse trend of the FAAF case. We have envisioned the so-called N-acetyl factor as it could raise conformational barriers of FAAF versus FAF. The present results provide valuable conformational insight into the sequence-dependent UvrABC incisions of the bulky aminofluorene DNA adducts.

Show MeSH
Related in: MedlinePlus