let-7 miRNAs control the timing of development of the nematode C. elegans. They bind two times in the 3’-UTR of lin-41 mRNAs and downregulate expression. We have determined by solution-state NMR the structure of a 34-nt RNA construct, which illustrates the interaction of let-7 miRNA to the first complementary binding site. Structure shows two non-canonical parts characteristic for the seed region: an asymmetric internal loop and a bulge. The internal loop demonstrates residues G23 and U25 bulged-out and the residue U24 looped-in. Residue A30 is not stacked-in but points into the minor groove.
In this review article we describe the preparation of the 33-mer RNA model construct mimicking interaction between let-7 miRNA and lin-41 mRNA. We explain NMR experiments used for assignment of 1H, 13C, 15N and 31P resonances and acquisition of NOE derived restraints and torsion angle restraints. Structural calculation showed that the 13C,15N-labeled RNA construct folds into a stable structure consisting of two stem regions separated by a defined asymmetric internal loop. Solution-state NMR makes important contribution toward deeper understanding of structural features of miRNA:mRNA complexes.