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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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Small molecule crystal structures of (a) phenoxazine (1) and (b) the nitroxide spin-labeled nucleobase ç.
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Figure 2: Small molecule crystal structures of (a) phenoxazine (1) and (b) the nitroxide spin-labeled nucleobase ç.

Mentions: Small molecule crystal structures were determined for the ç nucleobase as well as the phenoxazine analog 1 (Figure 2a and Supplementary Data). The phenoxazine nucleobase 1 adopts a planar conformation with almost no bend at the oxazine linkage between the cytosine and benzene rings (Figure 2a). In contrast, ç adopts a non-planar geometry with a bend of ∼20° at the oxazine linkage between the cytosine and benzene rings (Figure 2b and Supplementary Data). In order to rationalize the conformational differences of the phenoxazine moiety, DFT calculations have been performed on ç in the bent conformation as found in the small molecule crystal structure and on ç in the planar conformation as found in the small molecule crystal structure of 1. These calculations revealed that the bent form is only 1.31 kJ/mol higher in energy than the planar form and that all vibrational frequencies for both conformations are positive. Thus, both conformations are similar energy minima or might actually belong to the same energy minimum. This is supported by the finding that the frequencies for the bending motion differ by only 10 cm−1 (18 and 28 cm−1 for the planar and bent conformation, respectively). Taken together, we interpret these results as both conformations belonging to the same energy minimum with a low energy bending motion around the oxazine linkage. Because bending the phenoxazine moiety costs little energy, the surrounding environment such as crystal packing can drive it into either conformation. This raises the question as to whether ç is bent or planar when incorporated into an oligonucleotide structure. Thus, we proceeded to obtain a high-resolution structure of a DNA containing Ç to establish the preferred conformation of Ç within the context of a nucleic acid.


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)

Small molecule crystal structures of (a) phenoxazine (1) and (b) the nitroxide spin-labeled nucleobase ç.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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

Figure 2: Small molecule crystal structures of (a) phenoxazine (1) and (b) the nitroxide spin-labeled nucleobase ç.
Mentions: Small molecule crystal structures were determined for the ç nucleobase as well as the phenoxazine analog 1 (Figure 2a and Supplementary Data). The phenoxazine nucleobase 1 adopts a planar conformation with almost no bend at the oxazine linkage between the cytosine and benzene rings (Figure 2a). In contrast, ç adopts a non-planar geometry with a bend of ∼20° at the oxazine linkage between the cytosine and benzene rings (Figure 2b and Supplementary Data). In order to rationalize the conformational differences of the phenoxazine moiety, DFT calculations have been performed on ç in the bent conformation as found in the small molecule crystal structure and on ç in the planar conformation as found in the small molecule crystal structure of 1. These calculations revealed that the bent form is only 1.31 kJ/mol higher in energy than the planar form and that all vibrational frequencies for both conformations are positive. Thus, both conformations are similar energy minima or might actually belong to the same energy minimum. This is supported by the finding that the frequencies for the bending motion differ by only 10 cm−1 (18 and 28 cm−1 for the planar and bent conformation, respectively). Taken together, we interpret these results as both conformations belonging to the same energy minimum with a low energy bending motion around the oxazine linkage. Because bending the phenoxazine moiety costs little energy, the surrounding environment such as crystal packing can drive it into either conformation. This raises the question as to whether ç is bent or planar when incorporated into an oligonucleotide structure. Thus, we proceeded to obtain a high-resolution structure of a DNA containing Ç to establish the preferred conformation of Ç within the context of a nucleic acid.

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