Bending and circularization of site-specific and stereoisomeric carcinogen-DNA adducts

Rong Xu, Bing Mao, Shantu Amin, Nicholas Geacintov

Research output: Contribution to journalArticle

Abstract

The potent tumorigen and mutagen (+)-7(R),8(S)-dihydroxy-9(S),10(R)- epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene ((+)-anti-BPDE) is a metabolite of benzo[a]pyrene that binds predominantly to the exocyclic amino group of guanine residues in DNA in vivo and in vitro. While the (-)-7S, 8R, 9R, 10S enantiomer, (-)-anti-BPDE, also reacts with DNA to form similar covalent N2- deoxyguanosyl adducts, this diol epoxide is nontumorigenic and its mutagenic activities are different from those of (+)-anti-BPDE. In this work, T4 ligase-induced cyclization methods have been employed to demonstrate that the (+)-anti-[BP]-N2-dG lesions (G*) cause significantly greater amounts of bending and circularization of the one-base overhang undecamer duplex 5'- d(CACAT[G*]TACAC)·d(TGTACATGTGG) than the stereoisomeric oligonucleotide duplex with G* = (-)-anti-[BP]-N2-dG. In the case of the (+)-anti-BPDE- modified oligonucleotides, the ratio of circular to linear DNA multimers reaches values of 8-9 for circle contour sizes of 99-121 base pairs, while for the (-)-anti-[BP]-N2-dG-modified DNA this ratio reaches a maximum value of only ~1 at 154-176 base pairs. Assuming a planar circle DNA model, the inferred bending angles for 90-92% of the observed circular ligation products range from 30 to 51°per (+)-trans-anti-[BP]-N2-dG lesion and from 20 to 40°per (-)-trans-anti-[BP]-N2-dG lesion. In the case of unmodified DNA, the probability of circular product formation is at least 1 order of magnitude less efficient than in the BPDE-modified sequences and about 90% of the circular products exhibit bending angles in the range of 14-19°. In the most abundant circular products observed experimentally, the bending angles are 40°and 26 ± 2°per (+)-anti-[BP]- or (-)-anti-[BP]-modified 11-mer; these values correspond to a net contribution of 21-26°and 5-19% respectively, to the observed overall bending per lesion. The coexistence of circular DNA molecules of different sizes and, therefore, different average bending angles per lesion, suggest that the lesions induce both torsional flexibility and flexible bends, which permit efficient cyclization, especially in the case of (+)-trans-[BP]-N2-dG adducts. The NMR characteristics of (+)-trans-[BP]-N2-dG lesion in the 11-mer duplex 5'- d(CACAT[G*]TACAC)·d(GTGTACATGTG) indicate that all base pairs are intact, except at the underlined base pairs. This suggests a distortion in the normal conformation of the duplex on the 5'-side of the modified guanosine residue, which may be due to bending enhanced base pair opening and bending induced by the bulky carcinogen residue. The implications of base sequence-dependent flexibilities and conformational mobilities of anti[BP]-N2-dG lesions on DNA replication and mutation are discussed.

Original languageEnglish (US)
Pages (from-to)769-778
Number of pages10
JournalBiochemistry
Volume37
Issue number2
DOIs
StatePublished - Jan 13 1998

Fingerprint

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide
DNA Adducts
Base Pairing
Carcinogens
DNA
Circular DNA
Cyclization
Oligonucleotides
Guanosine
Benzo(a)pyrene
Epoxy Compounds
Mutagens
Guanine
Ligases
DNA Replication
Enantiomers
Ligation
Metabolites
Conformations
Mutation

ASJC Scopus subject areas

  • Biochemistry

Cite this

Bending and circularization of site-specific and stereoisomeric carcinogen-DNA adducts. / Xu, Rong; Mao, Bing; Amin, Shantu; Geacintov, Nicholas.

In: Biochemistry, Vol. 37, No. 2, 13.01.1998, p. 769-778.

Research output: Contribution to journalArticle

Xu, Rong ; Mao, Bing ; Amin, Shantu ; Geacintov, Nicholas. / Bending and circularization of site-specific and stereoisomeric carcinogen-DNA adducts. In: Biochemistry. 1998 ; Vol. 37, No. 2. pp. 769-778.
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T1 - Bending and circularization of site-specific and stereoisomeric carcinogen-DNA adducts

AU - Xu, Rong

AU - Mao, Bing

AU - Amin, Shantu

AU - Geacintov, Nicholas

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N2 - The potent tumorigen and mutagen (+)-7(R),8(S)-dihydroxy-9(S),10(R)- epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene ((+)-anti-BPDE) is a metabolite of benzo[a]pyrene that binds predominantly to the exocyclic amino group of guanine residues in DNA in vivo and in vitro. While the (-)-7S, 8R, 9R, 10S enantiomer, (-)-anti-BPDE, also reacts with DNA to form similar covalent N2- deoxyguanosyl adducts, this diol epoxide is nontumorigenic and its mutagenic activities are different from those of (+)-anti-BPDE. In this work, T4 ligase-induced cyclization methods have been employed to demonstrate that the (+)-anti-[BP]-N2-dG lesions (G*) cause significantly greater amounts of bending and circularization of the one-base overhang undecamer duplex 5'- d(CACAT[G*]TACAC)·d(TGTACATGTGG) than the stereoisomeric oligonucleotide duplex with G* = (-)-anti-[BP]-N2-dG. In the case of the (+)-anti-BPDE- modified oligonucleotides, the ratio of circular to linear DNA multimers reaches values of 8-9 for circle contour sizes of 99-121 base pairs, while for the (-)-anti-[BP]-N2-dG-modified DNA this ratio reaches a maximum value of only ~1 at 154-176 base pairs. Assuming a planar circle DNA model, the inferred bending angles for 90-92% of the observed circular ligation products range from 30 to 51°per (+)-trans-anti-[BP]-N2-dG lesion and from 20 to 40°per (-)-trans-anti-[BP]-N2-dG lesion. In the case of unmodified DNA, the probability of circular product formation is at least 1 order of magnitude less efficient than in the BPDE-modified sequences and about 90% of the circular products exhibit bending angles in the range of 14-19°. In the most abundant circular products observed experimentally, the bending angles are 40°and 26 ± 2°per (+)-anti-[BP]- or (-)-anti-[BP]-modified 11-mer; these values correspond to a net contribution of 21-26°and 5-19% respectively, to the observed overall bending per lesion. The coexistence of circular DNA molecules of different sizes and, therefore, different average bending angles per lesion, suggest that the lesions induce both torsional flexibility and flexible bends, which permit efficient cyclization, especially in the case of (+)-trans-[BP]-N2-dG adducts. The NMR characteristics of (+)-trans-[BP]-N2-dG lesion in the 11-mer duplex 5'- d(CACAT[G*]TACAC)·d(GTGTACATGTG) indicate that all base pairs are intact, except at the underlined base pairs. This suggests a distortion in the normal conformation of the duplex on the 5'-side of the modified guanosine residue, which may be due to bending enhanced base pair opening and bending induced by the bulky carcinogen residue. The implications of base sequence-dependent flexibilities and conformational mobilities of anti[BP]-N2-dG lesions on DNA replication and mutation are discussed.

AB - The potent tumorigen and mutagen (+)-7(R),8(S)-dihydroxy-9(S),10(R)- epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene ((+)-anti-BPDE) is a metabolite of benzo[a]pyrene that binds predominantly to the exocyclic amino group of guanine residues in DNA in vivo and in vitro. While the (-)-7S, 8R, 9R, 10S enantiomer, (-)-anti-BPDE, also reacts with DNA to form similar covalent N2- deoxyguanosyl adducts, this diol epoxide is nontumorigenic and its mutagenic activities are different from those of (+)-anti-BPDE. In this work, T4 ligase-induced cyclization methods have been employed to demonstrate that the (+)-anti-[BP]-N2-dG lesions (G*) cause significantly greater amounts of bending and circularization of the one-base overhang undecamer duplex 5'- d(CACAT[G*]TACAC)·d(TGTACATGTGG) than the stereoisomeric oligonucleotide duplex with G* = (-)-anti-[BP]-N2-dG. In the case of the (+)-anti-BPDE- modified oligonucleotides, the ratio of circular to linear DNA multimers reaches values of 8-9 for circle contour sizes of 99-121 base pairs, while for the (-)-anti-[BP]-N2-dG-modified DNA this ratio reaches a maximum value of only ~1 at 154-176 base pairs. Assuming a planar circle DNA model, the inferred bending angles for 90-92% of the observed circular ligation products range from 30 to 51°per (+)-trans-anti-[BP]-N2-dG lesion and from 20 to 40°per (-)-trans-anti-[BP]-N2-dG lesion. In the case of unmodified DNA, the probability of circular product formation is at least 1 order of magnitude less efficient than in the BPDE-modified sequences and about 90% of the circular products exhibit bending angles in the range of 14-19°. In the most abundant circular products observed experimentally, the bending angles are 40°and 26 ± 2°per (+)-anti-[BP]- or (-)-anti-[BP]-modified 11-mer; these values correspond to a net contribution of 21-26°and 5-19% respectively, to the observed overall bending per lesion. The coexistence of circular DNA molecules of different sizes and, therefore, different average bending angles per lesion, suggest that the lesions induce both torsional flexibility and flexible bends, which permit efficient cyclization, especially in the case of (+)-trans-[BP]-N2-dG adducts. The NMR characteristics of (+)-trans-[BP]-N2-dG lesion in the 11-mer duplex 5'- d(CACAT[G*]TACAC)·d(GTGTACATGTG) indicate that all base pairs are intact, except at the underlined base pairs. This suggests a distortion in the normal conformation of the duplex on the 5'-side of the modified guanosine residue, which may be due to bending enhanced base pair opening and bending induced by the bulky carcinogen residue. The implications of base sequence-dependent flexibilities and conformational mobilities of anti[BP]-N2-dG lesions on DNA replication and mutation are discussed.

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