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The binding of purified Gly3-Phe-tRNA to Escherichia coli ribosomes was compared to that of crude Gly3-Phe-tRNA (containing bulk tRNA). Gly3-phe-tRNA from the purified preparation binds preferentially to the P site (peptidyl site) at both a high (30 mM) and a low (10 mM) magnesium acetate concentration, while peptidyl-tRNA from the crude preparation bound more to A site (aminoacyl site) than to the P site at 30 mM magnesium acetate under the same experimental conditions. The addition of tRNAPhe to the purified Gly3-Phe-tRNA preparation strongly inhibited the binding at 10 mM. At 30 mM Mg2+ the addition of the same amount of tRNAPhe caused the Gly3-Phe-tRNA to bind partially to the A site, but did not inhibit the total amount of Gly3-Phe-tRNA which binds to the ribosomes. The binding of purified Phe-tRNA in the presence of T factor and GTP is not inhibited by tetracycline. However, tetracycline strongly inhibited the enzymatic binding of Phe-tRNA when tRNAPhe was added. From the results presented in this paper it is proposed that in the presence of poly(U), Phe-tRNA or peptidyl-tRNAPhe can bind to A sites only of those ribosomes which have their P site occupied by tRNA or one of its derivatives (aminoacyl-tRNA or peptidyl-tRNA). [ABSTRACT FROM AUTHOR]

The non-enzymatic binding of aminoacyl-tRNA, N-blocked aminoacyl-tRNA, peptidyl-tRNA and N-blocked peptidyl-tRNA to E. coli ribosomes was measured at 0.01 M magnesium ion concentration in the presence of poly U. It was found that, the maximal amounts of phenylalanyl-tRNA and peptidyl-tRNA, that can be bound to the ribosomes are the same, but the amounts of N-blocked phenylalanyl-tRNA and N-blocked peptidyl-tRNA, that can maximally be bound to the ribosomes are much less. Phenytalanyl-tRNA and peptidyl-tRNA can be bound to ribosomes which are saturated in respect to N-blocked phenylalanyl-tRNA or N-blocked peptidyl-tRNA; however, the presence of the N-blocked substrates on the ribosomes partially inhibits the additional binding of phenylalanyl-tRNA or peptidyl-tRNA. Aminoacyl-tRNA added to ribosomes saturated with respect to peptidyl-tRNA, partially displaces the peptidyl-tRNA from its ribosomal binding site. Under the same conditions, peptidyl-tRNA exchanges with prebound aminoacyl-tRNA considerably less. [ABSTRACT FROM AUTHOR]