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Formation of aminyl radicals on electron attachment to AZT: abstraction from the sugar phosphate backbone versus one-electron oxidation of guanine.

ABSTRACT: Employing electron spin resonance (ESR) spectroscopy, we have characterized the radicals formed in 3'-azido-3'-deoxythymidine (3'-AZT) and in its 5'-analog 5'-azido-5'-deoxythymidine (5'-AZT) after electron attachment in gamma-irradiated aqueous (H(2)O or D(2)O) glassy (7.5 M LiCl) systems. ESR spectral studies and theoretical calculations show that the predominant site of electron capture in 3'-AZT and in 5'-AZT is at the azide group and not at the thymine moiety. The azide group in AZT is therefore more electron affinic than the most electron affinic DNA base, thymine. Electron attachment to 3'-AZT and 5'-AZT results in an unstable azide anion radical intermediate (RN(3)*(-)) that is too short-lived to be observed in our work even at 77 K. At 77 K, we observe the neutral aminyl radical (RNH*) after loss of N(2) from RN(3)*(-) followed by protonation of nitrene anion radical (RN*(-)) to give RNH*. The expected RN*(-) intermediate is not observed as protonation from water is complete at 77 K even under highly basic conditions. Formation of RND* in D(2)O solutions confirms water as the source of the NH proton in the RNH*. Our assignments to these radicals are aided by DFT calculations for hyperfine coupling constants that closely match the experimental values. On annealing to higher temperatures (ca. 160-170 K), RNH* undergoes bimolecular hydrogen abstraction reactions from the thymine methyl group and the sugar moiety resulting in the formation of the thymine allyl radical (UCH(2)*) and two sugar radicals, C3'* and C5'*. RNH* also results in one-electron oxidation of the guanine base in 3'-AZG. This work provides a potential mechanism for the reported radiosensitization effects of AZT.

SUBMITTER: Adhikary A

PROVIDER: S-EPMC2912441 | biostudies-literature | 2010 Jul

REPOSITORIES: biostudies-literature

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Formation of aminyl radicals on electron attachment to AZT: abstraction from the sugar phosphate backbone versus one-electron oxidation of guanine.

Adhikary Amitava A Khanduri Deepti D Pottiboyina Venkata V Rice Cory T CT Sevilla Michael D MD

The journal of physical chemistry. B 20100701 28

Employing electron spin resonance (ESR) spectroscopy, we have characterized the radicals formed in 3'-azido-3'-deoxythymidine (3'-AZT) and in its 5'-analog 5'-azido-5'-deoxythymidine (5'-AZT) after electron attachment in gamma-irradiated aqueous (H(2)O or D(2)O) glassy (7.5 M LiCl) systems. ESR spectral studies and theoretical calculations show that the predominant site of electron capture in 3'-AZT and in 5'-AZT is at the azide group and not at the thymine moiety. The azide group in AZT is ther ...[more]

PMID: 20575557

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Project description:DNA sequence context has emerged as a critical determinant of the location and quantity of nucleobase damage caused by many oxidizing agents. However, the complexity of nucleobase and 2-deoxyribose damage caused by strong oxidants such as ionizing radiation and the Fenton chemistry of Fe2+-EDTA/H2O2 poses a challenge to defining the location of nucleobase damage and the effects of sequence context on damage chemistry in DNA. To address this problem, we developed a gel-based method that allows quantification of nucleobase damage in oxidized DNA by exploiting Escherichia coli exonuclease III to remove fragments containing direct strand breaks and abasic sites. The rigor of the method was verified in studies of guanine oxidation by photooxidized riboflavin and nitrosoperoxycarbonate, for which different effects of sequence context have been demonstrated by other approaches (Margolin, Y., Cloutier, J. F., Shafirovich, V., Geacintov, N. E., and Dedon, P. C. (2006) Nat. Chem. Biol. 2, 365-366). Using duplex oligodeoxynucleotides containing all possible three-nucleotide sequence contexts for guanine, the method was used to assess the role of DNA sequence context in hydroxyl radical-induced guanine oxidation associated with gamma-radiation and Fe2+-EDTA/H2O2. The results revealed both differences and similarities for G oxidation by hydroxyl radicals and by one-electron oxidation by riboflavin-mediated photooxidation, which is consistent with the predominance of oxidation pathways for hydroxyl radicals other than one-electron oxidation to form guanine radical cations. Although the relative quantities of G oxidation produced by hydroxyl radicals were more weakly correlated with sequence-specific ionization potential than G oxidation produced by riboflavin, damage produced by both hydroxyl radical generators and riboflavin within two- and three-base runs of G showed biases in location that are consistent with a role for electron transfer in defining the location of the damage products. Furthermore, both gamma-radiation and Fe2+-EDTA/H2O2 showed relatively modest effects of sequence context on the proportions of different damage products sensitive to E. coli formamidopyrimidine DNA glycosylase and hot piperidine, although GT-containing sequence contexts displayed subtle biases in damage chemistry (formamidopyrimidine DNA glycosylase/piperidine ratio). Overall, the results are consistent with the known chemistry of guanine oxidation by hydroxyl radical and demonstrate that charge migration plays a relatively minor role in determining the location and chemistry of hydroxyl radical-mediated oxidative damage to guanine in DNA.

Dataset Information

Formation of aminyl radicals on electron attachment to AZT: abstraction from the sugar phosphate backbone versus one-electron oxidation of guanine.

Publications

Formation of aminyl radicals on electron attachment to AZT: abstraction from the sugar phosphate backbone versus one-electron oxidation of guanine.

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