GTP hydrolysis by elongation factor Tu (EF-Tu) on the ribosome is induced by codon recognition. The mechanism by which a signal is transmitted from the site of codon-anticodon interaction in the decoding center of the 30S ribosomal subunit to the site of EF-Tu binding on the 50S subunit is not known. Here we examine the role of the tRNA in this process. We have used two RNA fragments, one which contains the anticodon and D hairpin domains (ACD oligomer) derived from tRNA(Phe) and the second which comprises the acceptor stem and T hairpin domains derived from tRNA(Ala) (AST oligomer) that aminoacylates with alanine and forms a ternary complex with EF-Tu·GTP. While the ACD oligomer and the ternary complex containing the Ala-AST oligomer interact with the 30S and 50S A site, respectively, no rapid GTP hydrolysis was observed when both were bound simultaneously. The presence of paromomycin, an aminoglycoside antibiotic that binds to the decoding site and stabilizes codon-anticodon interaction in unfavorable coding situations, did not increase the rate of GTP hydrolysis. These results suggest that codon recognition as such is not sufficient for GTPase activation and that an intact tRNA molecule is required for transmitting the signal created by codon recognition to EF-Tu.
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