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Crystal structure of a DNA containing the planar, phenoxazine-derived bi-functional spectroscopic probe C.

Edwards TE, Cekan P, Reginsson GW, Shelke SA, Ferré-D'Amaré AR, Schiemann O, Sigurdsson ST - Nucleic Acids Res. (2011)

Bottom Line: To understand the effect of Ç on nucleic acid structure, we undertook a detailed crystallographic analysis.These results indicate a small degree of flexibility around the oxazine linkage, which may be a consequence of the antiaromaticity of a 16-π electron ring system.This structural analysis shows that the Ç forms a planar, structurally non-perturbing base pair with G indicating it can be used with high confidence in EPR- or fluorescence-based structural and dynamics studies.

View Article: PubMed Central - PubMed

Affiliation: Emerald BioStructures, Bainbridge Island, WA 98110, USA.

ABSTRACT
Previously, we developed the deoxycytosine analog Ç (C-spin) as a bi-functional spectroscopic probe for the study of nucleic acid structure and dynamics using electron paramagnetic resonance (EPR) and fluorescence spectroscopy. To understand the effect of Ç on nucleic acid structure, we undertook a detailed crystallographic analysis. A 1.7 Å resolution crystal structure of Ç within a decamer duplex A-form DNA confirmed that Ç forms a non-perturbing base pair with deoxyguanosine, as designed. In the context of double-stranded DNA Ç adopted a planar conformation. In contrast, a crystal structure of the free spin-labeled base ç displayed a ∼ 20° bend at the oxazine linkage. Density function theory calculations revealed that the bent and planar conformations are close in energy and exhibit the same frequency for bending. These results indicate a small degree of flexibility around the oxazine linkage, which may be a consequence of the antiaromaticity of a 16-π electron ring system. Within DNA, the amplitude of the bending motion is restricted, presumably due to base-stacking interactions. This structural analysis shows that the Ç forms a planar, structurally non-perturbing base pair with G indicating it can be used with high confidence in EPR- or fluorescence-based structural and dynamics studies.

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(a) Structures of phenoxazine-derived nitroxide spin labels Ç and ç, and the unmodified phenoxazine derivative 1. The spin labels are shown base-paired with guanine (G), with hydrogen bonds indicated by dashed lines (b) Sequence and secondary structure of the duplex DNA used to obtain a high-resolution crystal structure of a Ç-containing DNA helix. dR = 2′-deoxyribose. 2′OMeU = 2′-O-methyluridine.
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Figure 1: (a) Structures of phenoxazine-derived nitroxide spin labels Ç and ç, and the unmodified phenoxazine derivative 1. The spin labels are shown base-paired with guanine (G), with hydrogen bonds indicated by dashed lines (b) Sequence and secondary structure of the duplex DNA used to obtain a high-resolution crystal structure of a Ç-containing DNA helix. dR = 2′-deoxyribose. 2′OMeU = 2′-O-methyluridine.

Mentions: Here we report a detailed crystallographic characterization of Ç. This analysis includes small molecule crystal structures of the ç nucleobase, which has been used for non-covalent spin labeling of nucleic acids containing an abasic site (26), and its phenoxazine analog (1) (Figure 1) as well as a 1.7 Å resolution crystal structure of a decamer DNA duplex containing Ç. The high-resolution nucleic acid structure demonstrates that within the context of the nucleic acid, the nitroxide spin label Ç adopts a planar conformation while forming a standard three hydrogen bond base pair with deoxyguanosine. These results validate the interpretation of distance and orientation measurements between two Ç reporters described previously and provide the basis for further structural and dynamics studies on oligonucleotides with unknown folds.Figure 1.


Crystal structure of a DNA containing the planar, phenoxazine-derived bi-functional spectroscopic probe C.

Edwards TE, Cekan P, Reginsson GW, Shelke SA, Ferré-D'Amaré AR, Schiemann O, Sigurdsson ST - Nucleic Acids Res. (2011)

(a) Structures of phenoxazine-derived nitroxide spin labels Ç and ç, and the unmodified phenoxazine derivative 1. The spin labels are shown base-paired with guanine (G), with hydrogen bonds indicated by dashed lines (b) Sequence and secondary structure of the duplex DNA used to obtain a high-resolution crystal structure of a Ç-containing DNA helix. dR = 2′-deoxyribose. 2′OMeU = 2′-O-methyluridine.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3105401&req=5

Figure 1: (a) Structures of phenoxazine-derived nitroxide spin labels Ç and ç, and the unmodified phenoxazine derivative 1. The spin labels are shown base-paired with guanine (G), with hydrogen bonds indicated by dashed lines (b) Sequence and secondary structure of the duplex DNA used to obtain a high-resolution crystal structure of a Ç-containing DNA helix. dR = 2′-deoxyribose. 2′OMeU = 2′-O-methyluridine.
Mentions: Here we report a detailed crystallographic characterization of Ç. This analysis includes small molecule crystal structures of the ç nucleobase, which has been used for non-covalent spin labeling of nucleic acids containing an abasic site (26), and its phenoxazine analog (1) (Figure 1) as well as a 1.7 Å resolution crystal structure of a decamer DNA duplex containing Ç. The high-resolution nucleic acid structure demonstrates that within the context of the nucleic acid, the nitroxide spin label Ç adopts a planar conformation while forming a standard three hydrogen bond base pair with deoxyguanosine. These results validate the interpretation of distance and orientation measurements between two Ç reporters described previously and provide the basis for further structural and dynamics studies on oligonucleotides with unknown folds.Figure 1.

Bottom Line: To understand the effect of Ç on nucleic acid structure, we undertook a detailed crystallographic analysis.These results indicate a small degree of flexibility around the oxazine linkage, which may be a consequence of the antiaromaticity of a 16-π electron ring system.This structural analysis shows that the Ç forms a planar, structurally non-perturbing base pair with G indicating it can be used with high confidence in EPR- or fluorescence-based structural and dynamics studies.

View Article: PubMed Central - PubMed

Affiliation: Emerald BioStructures, Bainbridge Island, WA 98110, USA.

ABSTRACT
Previously, we developed the deoxycytosine analog Ç (C-spin) as a bi-functional spectroscopic probe for the study of nucleic acid structure and dynamics using electron paramagnetic resonance (EPR) and fluorescence spectroscopy. To understand the effect of Ç on nucleic acid structure, we undertook a detailed crystallographic analysis. A 1.7 Å resolution crystal structure of Ç within a decamer duplex A-form DNA confirmed that Ç forms a non-perturbing base pair with deoxyguanosine, as designed. In the context of double-stranded DNA Ç adopted a planar conformation. In contrast, a crystal structure of the free spin-labeled base ç displayed a ∼ 20° bend at the oxazine linkage. Density function theory calculations revealed that the bent and planar conformations are close in energy and exhibit the same frequency for bending. These results indicate a small degree of flexibility around the oxazine linkage, which may be a consequence of the antiaromaticity of a 16-π electron ring system. Within DNA, the amplitude of the bending motion is restricted, presumably due to base-stacking interactions. This structural analysis shows that the Ç forms a planar, structurally non-perturbing base pair with G indicating it can be used with high confidence in EPR- or fluorescence-based structural and dynamics studies.

Show MeSH
Related in: MedlinePlus