CONFORMATIONAL RIGIDITY OF SPECIFIC PYRIMIDINE RESIDUES IN TRANSFER-RNA ARISES FROM POSTTRANSCRIPTIONAL MODIFICATIONS THAT ENHANCE STERIC INTERACTION BETWEEN THE BASE AND THE 2'-HYDROXYL GROUP

In order to elucidate roles of the 2'-O-methylation of pyrimidine nucleotide residues of tRNAs, conformations of 2'-O-methyluridylyl(3' --> 5')uridine (UmpU), 2'-O-methyluridine 3'-monophosphate (Ump), and 2'-O-methyluridine (Um) in (H2O)-H-2 solution were analyzed by one- and two-dimensional proton NMR spectroscopy and compared with those of related nucleotides and nucleoside. As for UpU and UmpU, the 2'-O-methylation was found to stabilize the C3'-endo form of the 3'-nucleotidyl unit (Up-/Ump-moiety). This stabilization of the C3'-endo form is primarily due to an intraresidue effect, since the conformation of the 5'-nucleotidyl unit (-pU moiety) was only slightly affected by the 2'-O-methylation of the 3'-nucleotide unit. In fact even for Up and Ump, the 2'-O-methylation significantly stabilizes the C3'-endo form by 0.8 kcal.mol-1. By contrast, for nucleosides (U and Um), the C3'-endo form is slightly stabilized by 0.1 kcal.mol-1. Accordingly, the stabilization of the C3'-endo form by the 2'-O-methylation is primarily due to the steric repulsion among the 2-carbonyl group, the 2'-O-methyl group and the 3'-phosphate group in the C2'-endo form. For some tRNA species, 2-thiolation of pyrimidine residues is found in positions where the 2'-O-methylation is found for other tRNA species. A common feature is now found between the 2'-O-methylated and 2-thiolated pyrimidine nucleotides; the predominance of the C3'-endo form is due to an enhancement of the intraresidue steric repulsion between the groups in positions 2 and 2' (and 3') in the C2'-endo form. Such conformational rigidity of the modified pyrimidine nucleotide residues contributes to the correct codon recognition and thermostability of tRNA molecules.