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2′-OH vs 3′-OH Specificity in tRNA Aminoacylation

Sidney M. Hecht


Although the aminoacylation of tRNA and participation of the amino-acylated species in the partial reactions of protein biosynthesis have been studied in detail for a number of years, until recently little data were available concerning the (2′ or 3′) positional isomer of tRNA that participated in each of these reactions. The source of ambiguity has been the aminoacyl-tRNA itself, which is believed to exist in solution as a rapidly equilibrating mixture of 2′- and 3′-O-aminoacylated molecules. Thus, whereas the activation of a given tRNA has been thought to involve attachment of the amino acid to a single hydroxyl group at the 3′ terminal of the tRNA, early efforts at trapping the initially formed isomer prior to equilibration were unsuccessful (Feldmann and Zachau 1964; Wolfenden et al. 1964; McLaughlin and Ingram 1965). Equilibration of the formed aminoacyl-tRNAs (Fig. 1) has also precluded the use of single isomers of the aminoacylated species to probe the partial reactions subsequent to aminoacylation.

In 1973, two laboratories described the preparation of tRNAs terminating in isomeric, modified adenine nucleotides (Sprinzl et al. 1973; Hecht et al. 1973), and such positionally defined, isomeric tRNAs were shown to be of utility for studying certain of the individual transformations in the overall process of peptide bond formation (Sprinzl and Cramer 1973; Fraser and Rich 1973). Attention has now focused on tRNAs terminating in 2′- and 3′-deoxyadenosine, which are good analogs of unmodified tRNA to a reasonable level of refinement and have been useful in identifying the positional isomers of...

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