Enzymatic alteration of the anticodon IAC of Torulopsis tRNAVal to IAU for isoleucine anticodon and aminoacylation of the variant

By enzymatic cleavage and ligation of tRNAVa1, its anticodon sequence IAC was altered to IAU, the anticodon of tRNAI1e. Valine acceptor activity of this variant tRNAVa1 (IAU) was reduced to the extent much lower than tyrosine acceptability of the previously prepared tRNATyr (GAA) (anticodon for tRNAPhe). Isoleucine acceptor activity was undetected, contrary to tRNATyr (GAA) which accepted phenylalanine weakly. Cleavage of tRNAVa1 (IAC) between IACA37 and C38 of its anticodon loop reduced the valine acceptor activity, suggesting some contribution of the conformation of the anticodon loop to the aminoacylation reaction.