Published online 28 June 2006
© 2006 The Author(s)
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Methods Online |
Short, synthetic and selectively 13C-labeled RNA sequences for the NMR structure determination of protein–RNA complexes
Institut des Science et Ingénierie Chimiques, Ecole Polytechnique Fédérale de Lausanne EPFL-BCH, 1015 Lausanne, Switzerland 1 Institute for Molecular Biology and Biophysics, Biology Department, Swiss Federal Institute of Technology Zürich ETH-Hönggerberg, CH-8093 Zürich, Switzerland
*To whom correspondence should be addressed. Tel: 0041 21 6939380; Fax: 0041 21 6939380; Email: Stefan.Pitsch{at}epfl.ch
*Correspondence may also be addressed to Frédéric H.-T. Allain. Tel: 0041 44 6333940; Fax: 0041 44 6331294; Email: allain@mol.biol.ethz.ch
Received April 3, 2006. Revised May 3, 2006. Accepted May 29, 2006.
We report an optimized synthesis of all canonical 2'-O-TOM protected ribonucleoside phosphoramidites and solid supports containing [13C5]-labeled ribose moieties, their sequence-specific introduction into very short RNA sequences and their use for the structure determination of two protein–RNA complexes. These specifically labeled sequences facilitate RNA resonance assignments and are essential to assign a high number of sugar–sugar and intermolecular NOEs, which ultimately improve the precision and accuracy of the resulting structures. This labeling strategy is particularly useful for the study of protein–RNA complexes with single-stranded RNA in solution, which is rapidly an increasingly relevant research area in biology.