Strategy for automated NMR resonance assignment of RNA: application to 48-nucleotide K10

Barbara Krähenbühl, Peter Lukavsky, Gerhard Wider
<span title="2014-06-05">2014</span> <i title="Springer Nature"> <a target="_blank" rel="noopener" href="https://fatcat.wiki/container/cimmcbrwkbf4xblwfxyaslsraa" style="color: black;">Journal of Biomolecular NMR</a> </i> &nbsp;
A procedure is presented for automated sequence-specific assignment of NMR resonances of uniformly [ 13 C, 15 N]-labeled RNA. The method is based on a suite of four through-bond and two through-space highdimensional automated projection spectroscopy (APSY) experiments. The approach is exemplified with a 0.3 mM sample of an RNA stem-loop with 48 nucleotides, K10, which is responsible for dynein-mediated localization of Drosophila fs(1)K10 mRNA transcripts. The automated analysis of the APSY data
more &raquo; ... led to highly accurate and precise 3-to 4-dimensional peak lists. They provided a reliable basis for the subsequent sequence-specific resonance assignment with the algorithm FLYA and resulted in the fully automated resonance assignment of more than 80 % of the resonances of the 13 C-1 H moieties at the 1 0 , 2 0 , 5, 6, and 8 positions in the nucleotides. The procedure was robust with respect to numerous impurity peaks, low concentration of this for NMR comparably large RNA, and structural features such as a loop, single-nucleotide bulges and a non-Watson-Crick wobble base pairs. Currently, there is no precise chemical shift statistics (as used by FLYA) for RNA regions which deviate from the regular A-form helical structure. Reliable and precise peak lists are thus required for automated sequence-specific assignment, as provided by APSY.
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