13C-direct detected NMR experiments for the sequential J-based resonance assignment of RNA oligonucleotides

Abstract

We present here a set of 13C-direct detected NMR experiments to facilitate the resonance assignment of RNA oligonucleotides. Three experiments have been developed: (1) the (H)CC-TOCSY-experiment utilizing a virtual decoupling scheme to assign the intraresidual ribose 13C-spins, (2) the (H)CPC-experiment that correlates each phosphorus with the 4C©nuclei of adjacent nucleotides via J(C,P) couplings and (3) the (H)CPC-CCH-TOCSYexperiment that correlates the phosphorus nuclei with the respective 1C©, 1H© ribose signals. The experiments were applied to two RNA hairpin structures. The current set of 13C-direct detected experiments allows direct and unambiguous assignment of the majority of the hetero nuclei and the identification of the individual ribose moieties following their sequential assignment. Thus, 13C-direct detected NMR methods constitute useful complements to the conventional 1H-detected approach for the resonance assignment of oligonucleotides that is often hindered by the limited chemical shift dispersion. The developed methods can also be applied to large deuterated RNAs. © The Author(s) 2010.