This project is directed towards the further elucidation of the participation of two highly substituted anticodon adjacent nucleosides in the structure and function of transfer RNA. N-(9-beta-D-ribofuranosyl) purin-6-ylcarbamoy1)threonine, t6A, is found at the 3'-end of the anticodon in tRNAs responding to adenosine-beginnning codons. High resolution nuclear magnetic resonance techniques (observing 1H, 13C and 31P nuclei) will be employed to test the hypothesis that t6A is a magnesium binding site. These procedures will also be used to investigate the involvement of the Y-nucleoside from torula yeast tRNAphe in the conformational flexibility of oligonucleotides isolated from the anticodon loop. Comparisons of the Y-containing oligonucleotides from torula with identical species from baker's yeast tRNAphe are planned to test the possiblity that the long hydrophobic side chain on the baker's yeast Y-base is involved in the apparent rigidity of these latter molecules.