THE T-ARM OF TRANSFER-RNA IS A SUBSTRATE FOR TRANSFER-RNA (M5U54)-METHYLTRANSFERASE

Fragments of Escherichia coli FUra-tRNA1Val as small as 15 nucleotides form covalent complexes with tRNA (m5U54)-methyltransferase (RUMT). The sequence essential for binding includes position 52 of the T-stem and the T-loop and extends toward the 3' acceptor end of FUra-tRNA. The in vitro synthesized 17mer T-arm of E. coli tRNA1Val, composed of the seven-base T-loop and 5-base-pair stem, is a good substrate for RUMT. The K(m) is decreased 5-fold and k(cat) is decreased 2-fold compared to the entire tRNA. The T-arm structure could be further reduced to an 11mer containing the loop and two base pairs and still retain activity; the K(m) was similar to that of the 17mer T-arm, whereas k(cat) was decreased and additional 20-fold. The data indicate that the primary specificity determinants for the RUMT-tRNA interaction are contained within the primary and secondary structure of the T-arm of tRNA.