Recognition of the anticodon loop of trna^l₁^le by isoleucyl-trna synthetase from escherichia coli

For Escherichia coli tRNA₁ ^llehaving anticodon G34-A35-U36, two vari- ants with substitution and/or insertion in the anticodon loop were prepared by in vitro recombinant RNA method. A variant with replacement of the N4(9-β-D-ribofuranosy1-purine-6-yl)carbamoyl)threonine (t⁶A) residue at position 37 with an unmodified adenosine exhibited a drastic reduction in isoleucine-accepting activity. This shows that t⁶A37 plays a crucial role in the recognition by isoleucyl-tRNA synthetase (IleRS). Into this A37 variant, unmodified A was further inserted at position 36 so that a sequence of GAAUA was created. This insertion did not show further reduction in isoleucine-accepting activity, indicating that IleRS recognizes the three anficodon residues, which have already been found to be identity elements, individually but not as a whole.