Identity determinants ofE.colitryptophan tRNA

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Abstract
The first base pair of the acceptor stem A1-U72 and the discriminator base G73, as well as the anticodon nucleotides, characterize the tryptophan tRNA in E. coli. To determine the contribution of these nucleotides to the tryptophan acceptor activity, various transcripts of E. coli tryptophan tRNA mutants were constructed. Substitutions of the discriminator base G73, which is conserved within prokaryotic tryptophan tRNAs, impaired aminoacylation with tryptophan. Substitutions of other purine-pyrimidine pairs for A1-U72 revealed that only U72 weakly contributed to recognition by tryptophanyl-tRNA synthetase. The E. coli aspartic acid tRNA transcript introducing the tryptophan anticodon CCA showed almost the same tryptophan charging activity as the tryptophan tRNA transcript possessing a G1-C72 base pair. Only a low activity was detected in the mutant tryptophan tRNA transcript possessing a set of G1-C72 and A73, which is observed in eukaryotic tryptophan tRNAs. These results indicate that the anticodon and G73 are major identity determinants of tryptophan tRNA in E. coli, whereas the A1-U72 base pair is only a weak recognition element.