C 4 0 / H 4 0 selective , non - uniformly sampled 4 D HC ( P ) CH experiment for sequential assignments of 13 C - labeled RNAs

A through bond, C40/H40 selective, ‘‘out and stay’’ type 4D HC(P)CH experiment is introduced which provides sequential connectivity via H4(i)–C4 0 (i)–C4 0 (i-1)– H4(i-1) correlations. The P dimension (used in the conventional 3D HCP experiment) is replaced with evolution of better dispersed C40 dimension. The experiment fully utilizes C-labeling of RNA by inclusion of two C40 evolution periods. An additional evolution of H40 is included to further enhance peak resolution. Band selective C inversion pulses are used to achieve selectivity and prevent signal dephasing due to the of C40–C30 and C40– C50 homonuclear couplings. For reasonable resolution, non-uniform sampling is employed in all indirect dimensions. To reduce sensitivity losses, multiple quantum coherences are preserved during shared-time evolution and coherence transfer delays. In the experiment the intranucleotide peaks are suppressed whereas inter-nucleotide peaks are enhanced to reduce the ambiguities. The performance of the experiment is verified on a fully C, Nlabeled 34-nt hairpin RNA comprising typical structure elements.