The effect of aminoacylation and ternary complex formation with elongation factor Tu•GTP on the tertiary structure of yeast tRNAPhe was examined by 1H-NMR spectroscopy. Esterification of phenylalanine to tRNAPhe does not lead to changes with respect to the secondary and tertiary base pair interactions of tRNA. Complex formation of Phe-tRNAPhe with elongation factor Tu•GTP results in a broadening of all imino proton resonances of the tRNA. The chemical shifts of several NH proton resonances are slightly changed as compared to free tRNA, indicating a minor conformational rearrangement of Phe-tRNAPhe upon binding to elongation factor Tu•GTP. All NH proton resonances corresponding to the secondary and tertiary base pairs of tRNA, except those arising from the first three base pairs in the aminoacyl stem, are detectable in the Phe-tRNAPhe•elongation factor Tu•GTP ternary complex. Thus, although the interactions between elongation factor Tu and tRNA accelerate the rate of NH proton exchange in the aminoacyl stem-region, the Phe-tRNAPhe preserves its typical L-shaped tertiary structure in the complex. At high (> 10-4 M) ligand concentrations a complex between tRNAPhe and elongation factor Tu•GDP can be detected on the NMR time-scale. Formation of this complex is inhibited by the presence of any RNA not related to the tRNA structure. Using the known tertiary structures of yeast tRNAPhe and Thermus thermophilus elongation factor Tu in its active, GTP form, a model of the ternary complex was constructed.
Research Areas and Centers
- Academic Focus: Center for Infection and Inflammation Research (ZIEL)