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Prototropic equilibria in DNA containing one-electron oxidized GC: intra-duplex vs. duplex to solvent deprotonation.

ABSTRACT: By use of ESR and UV-vis spectral studies, this work identifies the protonation states of one-electron oxidized G:C (viz. G?+:C, G(N1–H)?:C(+H+), G(N1–H)?:C, and G(N2-H)?:C) in a DNA oligomer d[TGCGCGCA]2. Benchmark ESR and UV-vis spectra from one electron oxidized 1-Me-dGuo are employed to analyze the spectral data obtained in one-electron oxidized d[TGCGCGCA]2 at various pHs. At pH ?7, the initial site of deprotonation of one-electron oxidized d[TGCGCGCA]2 to the surrounding solvent is found to be at N1 forming G(N1–H)?:C at 155 K. However, upon annealing to 175 K, the site of deprotonation to the solvent shifts to an equilibrium mixture of G(N1–H)?:C and G(N2–H)?:C. For the first time, the presence of G(N2–H)?:C in a ds DNA-oligomer is shown to be easily distinguished from the other prototropic forms, owing to its readily observable nitrogen hyperfine coupling (Azz(N2) = 16 G). In addition, for the oligomer in H2O, an additional 8 G N2–H proton HFCC is found. This ESR identification is supported by a UV-vis absorption at 630 nm which is characteristic for G(N2–H)? in model compounds and oligomers. We find that the extent of photo-conversion to the C1? sugar radical (C1??) in the one-electron oxidized d[TGCGCGCA]2 allows for a clear distinction among the various G:C protonation states which can not be easily distinguished by ESR or UV-vis spectroscopies with this order for the extent of photo-conversion: G?+:C > G(N1–H)?:C(+H+) ? G(N1–H)?:C. We propose that it is the G?+:C form that undergoes deprotonation at the sugar and this requires reprotonation of G within the lifetime of exited state

SUBMITTER: Adhikary A

PROVIDER: S-EPMC4677782 | biostudies-literature | 2010

REPOSITORIES: biostudies-literature

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Prototropic equilibria in DNA containing one-electron oxidized GC: intra-duplex vs. duplex to solvent deprotonation.

Adhikary Amitava A Kumar Anil A Munafo Shawn A SA Khanduri Deepti D Sevilla Michael D MD

Physical chemistry chemical physics : PCCP 20100101 20

By use of ESR and UV-vis spectral studies, this work identifies the protonation states of one-electron oxidized G:C (viz. G˙+:C, G(N1–H)˙:C(+H+), G(N1–H)˙:C, and G(N2-H)˙:C) in a DNA oligomer d[TGCGCGCA]2. Benchmark ESR and UV-vis spectra from one electron oxidized 1-Me-dGuo are employed to analyze the spectral data obtained in one-electron oxidized d[TGCGCGCA]2 at various pHs. At pH ≥7, the initial site of deprotonation of one-electron oxidized d[TGCGCGCA]2 to the surrounding solvent is found t ...[more]

PMID: 21491657

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Project description:Conformational flexibility in nucleic acids provides a basis for complex structures, binding, and signaling. One-base bulges directly neighboring single-base mismatches in nucleic acids can be present in a minimum of two distinct conformations, complicating the examination of the thermodynamics by calorimetry or UV-monitored melting techniques. To provide additional information about such structures, we demonstrate how electron paramagnetic resonance (EPR) active spin-labeled base analogues, base-specifically incorporated into the DNA, are monitors of the superposition of different bulge-mismatch conformations. EPR spectra provide information about the dynamic environments of the probe. This information is cast in terms of "dynamic signatures" that have an underlying basis in structural variations. By examining the changes in the equilibrium of the different states across a range of temperatures, the enthalpy and entropy of the interconversion among possible conformations can be determined. The DNA constructs with a single bulge neighboring a single-base mismatch ("bulge-mismatches") may be approximately modeled as an equilibrium between two possible conformations. This structural information provides insight into the local composition of the bulge-mismatch sequences. Experiments on the bulge-mismatches show that basepairing across the helix can be understood in terms of purine and pyrimidine interactions, rather than specific bases. Measurements of the enthalpy and entropy of formation for the bulge-mismatches by differential scanning calorimetry and UV-monitored melting confirm that the formation of bulge-mismatches is in fact more complicated than a simple two-state process, consistent with the base-specific spectral data that bulge-mismatches exist in multiple conformations in the premelting temperature region. We find that the calculations with the nearest-neighbor (NN) model for the two likely conformations do not correlate well with the populations of structures and thermodynamic parameters inferred from the base-specific EPR dynamics probe. We report that the base-specific spin probes are able to identify a bistable, temperature dependent, switching between conformations for a particular complex bulged construct.

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Dataset Information

Prototropic equilibria in DNA containing one-electron oxidized GC: intra-duplex vs. duplex to solvent deprotonation.

Publications

Prototropic equilibria in DNA containing one-electron oxidized GC: intra-duplex vs. duplex to solvent deprotonation.

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