Informace o publikaci

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

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KRÄHENBÜHL Barbara LUKAVSKY Peter WIDER Gerhard

Rok publikování 2014
Druh Článek v odborném periodiku
Časopis / Zdroj Journal of Biomolecular NMR
Fakulta / Pracoviště MU

Středoevropský technologický institut

Citace
www http://link.springer.com/article/10.1007%2Fs10858-014-9841-3
Doi http://dx.doi.org/10.1007/s10858-014-9841-3
Obor Biochemie
Klíčová slova Nucleic acids; NMR; Projection spectroscopy; APSY; Automated assignment; FLYA; Novel sampling methods
Popis A procedure is presented for automated sequence-specific assignment of NMR resonances of uniformly [C-13, N-15]-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 C-13-H-1 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.

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