Abstract
New base-type-edited transverse-relaxation optimized CT-HCN(C) experiments are presented that yield intra-base and sugar-to-base correlations for 13C−15N labeled RNA. High spectral resolution in the 13C and 15N dimensions is achieved by constant time (CT) frequency editing. A spectral editing filter applied during the CT 15N labeling period separates the correlation peaks arising from G/U and A/C nucleotide bases. This provides the increased spectral resolution required to unambiguously connect the 1H and 13C resonances in sugar and base moieties of RNA nucleotides. In addition, the experiment allows base type identification for each residue, and therefore presents an attractive spectroscopic alternative to nucleotide-specific isotope labeling. Application to a 33-nucleotide RNA aptamer demonstrates the performance of the new pulse scheme.
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Acknowledgements
We thank Ad Bax for the sample of a complex formed by two RNA stem loops, and Marella Canny for sample preparation of the 33-nucleotide aptamer. This work was supported by the Commissariat à l’Energie Atomique, the Centre National de la Recherche Scientifique, and NIH (AI33098). H.V.M. acknowledges the receipt of a fellowship from the C.E.A. J.B. acknowledges support from the HFSP Organization.
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Melckebeke, H.V., Pardi, A., Boisbouvier, J. et al. Resolution-Enhanced Base-Type-Edited HCN Experiment for RNA. J Biomol NMR 32, 263–271 (2005). https://doi.org/10.1007/s10858-005-8872-1
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DOI: https://doi.org/10.1007/s10858-005-8872-1