Assignment of the low field proton nuclear magnetic resonance spectrum of yeast phenylalanine transfer RNA to specific base pairs

Abstract

High-resolution proton nuclear magnetic resonance spectra at 220 and 300 MHz have been used to investigate the base-pairing structure of fragments of yeast tRNA Phe, of chemically modified tRNA Phe and of intact tRNA Phe. To a very good approximation the positions of the fragment spectra are additive within 0·2 part per million, indicating that factors responsible for certain structural features in the intact molecule are already present in the smaller fragments (half molecules, hairpins and 3 4 molecules). A simple first-order ring-current shift theory taken in conjunction with the cloverleaf model for tRNA Phe (RajBhandary et al., 1967) has been used to predict the low-field (− 15 to −11 part per million) nuclear magnetic resonance spectra and make assignments of the resolved resonances to ring NH protons of specific base pairs. The general agreement between the predicted and observed spectra to within 0·2 part per million confirms in detail the cloverleaf model for the secondary structure of tRNA Phe in solution. It is also established that ring-current shifts are the principal factor responsible for the wide range of shifts observed in the low-field spectra. As a result it is evident that the resonances are very sensitive to small changes in the secondary structure and in some cases changes in the interbase distance as small as 0·2 Å could easily be detected. It is also clear from the analysis that certain of the resonances are sensitive to the tertiary structure of the molecule and specific examples are discussed. As with our previous study, we find no evidence for any strong Watson-Crick type base pairs beyond those predicted by the cloverleaf structure.