Contribution of dihydrouridine in folding of the D-arm in tRNAElectronic supplementary information (ESI) available: Complementary NMR and structural data. See DOI: 10.1039/c5ob00164a

Posttranscriptional modifications of transfer RNAs (tRNAs) are proven to be critical for all core aspects of tRNA function. While the majority of tRNA modifications were discovered in the 1970s, their contribution in tRNA folding, stability, and decoding often remains elusive. In this work an NMR study was performed to obtain more insight in the role of the dihydrouridine (D) modification in the D-arm of tRNA i Met from S. pombe . While the unmodified oligonucleotide adopted several undefined conformations that interconvert in solution, the presence of a D nucleoside triggered folding into a hairpin with a stable stem and flexible loop region. Apparently the D modification is required in the studied sequence to fold into a stable hairpin. Therefore we conclude that D contributes to the correct folding and stability of D-arm in tRNA. In contrast to what is generally assumed for nucleic acids, the sharp 'imino' signal for the D nucleobase at 10 ppm in 90% H 2 O is not indicative for the presence of a stable hydrogen bond. The strong increase in p K a upon loss of the aromatic character in the modified nucleobase slows down the exchange of its 'imino' proton significantly, allowing its observation even in an isolated D nucleoside in 90% H 2 O in acidic to neutral conditions. NMR studies of the D-arm of tRNA i Met revealed the crucial role of dihydrouridine nucleoside in folding of the oligo.