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Translational quality control is monitored at several steps, including substrate selection by aminoacyl-tRNA synthetases (aaRSs), and discrimination of aminoacyl-tRNAs by elongation factor Tu (EF-Tu) and the ribosome. Phenylalanyl-tRNA synthetase (PheRS) misactivates Tyr but is able to correct the mistake using a proofreading activity named editing. Previously we found that overproduction of editing-defective PheRS resulted in Tyr incorporation at Phe-encoded positions in vivo , although the misreading efficiency could not be estimated. This raised the question as to whether or not EF-Tu and the ribosome provide further proofreading mechanisms to prevent mistranslation of Phe codons by Tyr. Here we show that, after evading editing by PheRS, Tyr-tRNAPhe is recognized by EF-Tu as efficiently as the cognate Phe-tRNAPhe. Kinetic decoding studies using full-length Tyr-tRNAPhe and Phe-tRNAPhe, as well as a poly(U)-directed polyTyr/polyPhe synthesis assay, indicate that the ribosome lacks discrimination between Tyr-tRNAPhe and Phe-tRNAPhe. Taken together, these data suggest that PheRS editing is the major proofreading step that prevents infiltration of Tyr into Phe codons during translation.


This article was originally published in RNA, volume 13, in 2007.


RNA Society



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