The anticodon and discriminator base are important for aminoacylation of Escherichia coli tRNA(Asn)

Abstract

A gel shift assay that distinguishes the aminoacylated form from the deacylated form of tRNAs was used to study the requirements for aminoacylation of Escherichia coli tRNA(Asn) in vivo. tRNA(Asn) derivatives containing single base changes in their anticodons or discriminator bases were constructed, and the extent of in vivo aminoacylation was determined directly. Substitution of U35 with C35 or U36 with C36 abolished aminoacylation of the tRNA. Substitution of G34 with C34 converted tRNA(Asn) into a lysine acceptor. Thus, each of the anticodon nucleotides are important for aminoacylation of tRNA(Asn). Substitution of discriminator base G73 with A73 affected the extent of aminoacylation in vivo indicating that the discriminator base also contributes to aminoacylation of tRNA(Asn).