C4â?²/H4â?² selective, non-uniformly sampled 4D HC(P)CH experiment for sequential assignments of 13C-labeled RNAs
Abstract
A through bond, C4â?²/H4â?² selective, â??out and stayâ?? type 4D HC(P)CH experiment is introduced which provides sequential connectivity via H4â?²(i)â??C4â?²(i)â??C4â?²(iâ??1)â??H4â?²(iâ??1) correlations. The 31P dimension (used in the conventional 3D HCP experiment) is replaced with evolution of better dispersed C4â?² dimension. The experiment fully utilizes 13C-labeling of RNA by inclusion of two C4â?² evolution periods. An additional evolution of H4â?² is included to further enhance peak resolution. Band selective 13C inversion pulses are used to achieve selectivity and prevent signal dephasing due to the of C4â?²â??C3â?² and C4â?²â??C5â?² 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 intra-nucleotide peaks are suppressed whereas inter-nucleotide peaks are enhanced to reduce the ambiguities. The performance of the experiment is verified on a fully 13C, 15N-labeled 34-nt hairpin RNA comprising typical structure elements.
Source: Journal of Biomolecular NMR