Dataset Information

1,N2-Etheno-2'-deoxyguanosine adopts the syn conformation about the glycosyl bond when mismatched with deoxyadenosine.

ABSTRACT: The oligodeoxynucleotide 5'-CGCATXGAATCC-3'·5'-GGATTCAATGCG-3' containing 1,N(2)-etheno-2'-deoxyguanosine (1,N(2)-?dG) opposite deoxyadenosine (named the 1,N(2)-?dG·dA duplex) models the mismatched adenine product associated with error-prone bypass of 1,N(2)-?dG by the Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4) and by Escherichia coli polymerases pol I exo(-) and pol II exo(-). At pH 5.2, the T(m) of this duplex was increased by 3 °C as compared to the duplex in which the 1,N(2)-?dG lesion is opposite dC, and it was increased by 2 °C compared to the duplex in which guanine is opposite dA (the dG·dA duplex). A strong NOE between the 1,N(2)-?dG imidazole proton and the anomeric proton of the attached deoxyribose, accompanied by strong NOEs to the minor groove A(20) H2 proton and the mismatched A(19) H2 proton from the complementary strand, establish that 1,N(2)-?dG rotated about the glycosyl bond from the anti to the syn conformation. The etheno moiety was placed into the major groove. This resulted in NOEs between the etheno protons and T(5) CH(3). A strong NOE between A(20) H2 and A(19) H2 protons established that A(19), opposite to 1,N(2)-?dG, adopted the anti conformation and was directed toward the helix. The downfield shifts of the A(19) amino protons suggested protonation of dA. Thus, the protonated 1,N(2)-?dG·dA base pair was stabilized by hydrogen bonds between 1,N(2)-?dG N1 and A(19) N1H(+) and between 1,N(2)-?dG O(9) and A(19)N(6)H. The broad imino proton resonances for the 5'- and 3'-flanking bases suggested that both neighboring base pairs were perturbed. The increased stability of the 1,N(2)-?dG·dA base pair, compared to that of the 1,N(2)-?dG·dC base pair, correlated with the mismatch adenine product observed during the bypass of 1,N(2)-?dG by the Dpo4 polymerase, suggesting that stabilization of this mismatch may be significant with regard to the biological processing of 1,N(2)-?dG.

SUBMITTER: Shanmugam G

PROVIDER: S-EPMC3138413 | biostudies-literature | 2011 Jul

REPOSITORIES: biostudies-literature

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1,N2-Etheno-2'-deoxyguanosine adopts the syn conformation about the glycosyl bond when mismatched with deoxyadenosine.

Shanmugam Ganesh G Kozekov Ivan D ID Guengerich F Peter FP Rizzo Carmelo J CJ Stone Michael P MP

Chemical research in toxicology 20110616 7

The oligodeoxynucleotide 5'-CGCATXGAATCC-3'·5'-GGATTCAATGCG-3' containing 1,N(2)-etheno-2'-deoxyguanosine (1,N(2)-εdG) opposite deoxyadenosine (named the 1,N(2)-εdG·dA duplex) models the mismatched adenine product associated with error-prone bypass of 1,N(2)-εdG by the Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4) and by Escherichia coli polymerases pol I exo(-) and pol II exo(-). At pH 5.2, the T(m) of this duplex was increased by 3 °C as compared to the duplex in which the 1,N(2)-εdG les ...[more]

PMID: 21675798

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Project description:Oligonucleotides were synthesized containing the 7-(2-oxoheptyl)-etheno-dGuo adduct, which is derived from the reaction of dGuo and the lipid peroxidation product 4-oxo-2-nonenal. The in vitro replication of 7-(2-oxoheptyl)-etheno-dGuo by the model Y-family polymerase Sulfolobus solfataricus P2 DNA Polymerase IV (Dpo4) was examined in two sequences. The extension products were sequenced using an improved LC-ESI-MS/MS protocol developed in our laboratories, and the results were compared to that of the 1,N(2)-etheno-dGuo adduct in the same sequence contexts. Both etheno adducts were highly miscoding when situated in 5'-TXG-3' local sequence contexts with <4% of the extension products being derived from error-free bypass. The major extension products resulted from the misinsertion of Ade opposite the adduct and a one-base deletion. The major extension products from replication of the etheno lesions in a 5'-CXG-3' local sequence context were the result of misinsertion of Ade, a one-base deletion, and error-free bypass. Other minor extension products were also identified. The 7-(2-oxoheptyl)-etheno-dGuo lesion resulted in a larger frequency of misinsertion of Ade, whereas the 1,N(2)-etheno-dGuo gave more of the one-base deletion product. Conformational studies of duplex DNA containing the 7-(2-oxoheptyl)-etheno-dGuo in a 5'-TXG-3' sequence context by NMR indicated the presence of a pH-dependent conformational transition, likely involving the glycosyl bond at the adducted guanosine; the pK(a) for this transition was lower than that observed for the 1,N(2)-epsilon-dGuo lesion. However, the 7-(2-oxoheptyl)-etheno-dGuo lesion, the complementary Cyt, and both flanking base pairs remained disordered at all pH values, which is attributed to the presence of the hydrophobic heptyl group of the 7-(2-oxoheptyl)-etheno-dGuo lesion. The altered pK(a) value and the structural disorder at the 7-(2-oxoheptyl)-etheno-dGuo lesion site, as compared to the same sequence containing the 1,N(2)-etheno-dGuo, may contribute to higher frequency of misinsertion of Ade.

Project description:ConspectusCarbohydrates (glycans, saccharides, and sugars) are essential molecules in all domains of life. Research on glycoscience spans from chemistry to biomedicine, including material science and biotechnology. Access to pure and well-defined complex glycans using synthetic methods depends on the success of the employed glycosylation reaction. In most cases, the mechanism of the glycosylation reaction is believed to involve the oxocarbenium ion. Understanding the structure, conformation, reactivity, and interactions of this glycosyl cation is essential to predict the outcome of the reaction. In this Account, building on our contributions on this topic, we discuss the theoretical and experimental approaches that have been employed to decipher the key features of glycosyl cations, from their structures to their interactions and reactivity.We also highlight that, from a chemical perspective, the glycosylation reaction can be described as a continuum, from unimolecular SN1 with naked oxocarbenium cations as intermediates to bimolecular SN2-type mechanisms, which involve the key role of counterions and donors. All these factors should be considered and are discussed herein. The importance of dissociative mechanisms (involving contact ion pairs, solvent-separated ion pairs, solvent-equilibrated ion pairs) with bimolecular features in most reactions is also highlighted.The role of theoretical calculations to predict the conformation, dynamics, and reactivity of the oxocarbenium ion is also discussed, highlighting the advances in this field that now allow access to the conformational preferences of a variety of oxocarbenium ions and their reactivities under SN1-like conditions.Specifically, the ground-breaking use of superacids to generate these cations is emphasized, since it has permitted characterization of the structure and conformation of a variety of glycosyl oxocarbenium ions in superacid solution by NMR spectroscopy.We also pay special attention to the reactivity of these glycosyl ions, which depends on the conditions, including the counterions, the possible intra- or intermolecular participation of functional groups that may stabilize the cation and the chemical nature of the acceptor, either weak or strong nucleophile. We discuss recent investigations from different experimental perspectives, which identified the involved ionic intermediates, estimating their lifetimes and reactivities and studying their interactions with other molecules. In this context, we also emphasize the relationship between the chemical methods that can be employed to modulate the sensitivity of glycosyl cations and the way in which glycosyl modifying enzymes (glycosyl hydrolases and transferases) build and cleave glycosidic linkages in nature. This comparison provides inspiration on the use of molecules that regulate the stability and reactivity of glycosyl cations.

Dataset Information

1,N2-Etheno-2'-deoxyguanosine adopts the syn conformation about the glycosyl bond when mismatched with deoxyadenosine.

Publications

1,N2-Etheno-2'-deoxyguanosine adopts the syn conformation about the glycosyl bond when mismatched with deoxyadenosine.

Similar Datasets

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