PuSH - Publication Server of Helmholtz Zentrum München: Site-specific isotope-labeling of inosine phosphoramidites and NMR analysis of an inosine-containing RNA duplex.

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Dallmann, A. ; Beribisky, A.V. ; Gnerlich, F.* ; Rübbelke, M. ; Schiesser, S.* ; Carell, T.* ; Sattler, M.

Site-specific isotope-labeling of inosine phosphoramidites and NMR analysis of an inosine-containing RNA duplex.

Chem. Eur. J. 22, 15350-15359 (2016)

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Structural features and internal dynamics of inosine-containing RNAs are poorly understood. NMR studies of such RNAs require 13C,15N-labeling, which cannot be achieved using in vitro transcription as inosine and guanosine are not distinguished by RNA polymerase. Herein, we report the synthesis of an inosine phosphoramidite with selective 13C8 and 15N7-isotope incorporation in the base and uniform 13C-labeling of the ribose. Chemical synthesis of an RNA duplex containing four consecutive IU base pairs with this optimized isotope-labeling scheme greatly simplifies NMR spectra and resolves signal overlap. The absence of detectable NMR signals of imino protons and unusual inter-residue NOE correlations in this RNA indicate deviations from standard A-form geometry, consistent with reduced stability of this duplex seen in UV melting studies compared to its nonedited RNA counterparts. These studies indicate that the introduction of IU base pairs distorts and destabilizes RNA helices significantly compared to the also noncanonical GU base-pairs. Our optimized isotope-labeling scheme enables high-resolution NMR studies of inosine-edited RNAs.

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Publication type Article: Journal article

Document type Scientific Article

Corresponding Author

Keywords Biosynthesis ; Isotopic Labeling ; Nmr Spectroscopy ; Rna Structures ; Structural Biology; Double-stranded-rna; Relaxation Dispersion; Protein P100; Base-pairs; Tudor-sn; Spectroscopy; Dynamics; Cleavage; Ribonucleotides; Complex

ISSN (print) / ISBN 0947-6539

e-ISSN 1521-3765

Journal Chemistry - A European Journal

Quellenangaben Volume: 22, Issue: 43, Pages: 15350-15359

Publisher Wiley

Publishing Place Weinheim

Non-patent literature Publications

Reviewing status Peer reviewed

Institute(s) Institute of Structural Biology (STB)