Identification of Ribose-Base Sequential NOEs According to Base Types in Uniformly ~(13)C-Labeled RNAs

摘要

Recent progress in molecular biology has facilitated the production of l3C/i5N-labeled RNAs (1-3), and the availability of such labeled molecules has permitted the development of novel NMR schemes for sequential resonance assignments based on through-bond coherence transfer along the phosphate-ribose backbone (4-10). Although these methods are contributing to the determination of detailed NMR solution structures for RNA molecules of up to about 30-35 nucleotides, because of limited spectral dispersion and broad linewidths of the phosphorus resonances, such methods may have limited utility in larger RNA molecules as a result of the long interpulse delays employed in such experiments. In these circumstances, it may be necessary to achieve sequential resonance assignments via NOE effects. A widely used approach uses the fact that in helical RNA structures, ribose HI' resonances give NOE cross peaks to their own H6/H8 resonance and to that of the next base in the sequence (11, 12). Unambiguous assignment of these sequential connectivities to the base type can facilitate sequence specific resonance assignments in helical regions of RNAs.