Abstract:
: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 high-dimensional 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 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', 2', 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.
journal_name
J Biomol NMRjournal_title
Journal of biomolecular NMRauthors
Krähenbühl B,Lukavsky P,Wider Gdoi
10.1007/s10858-014-9841-3subject
Has Abstractpub_date
2014-08-01 00:00:00pages
231-40issue
4eissn
0925-2738issn
1573-5001journal_volume
59pub_type
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