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Protein synthesis requires both high speed and accuracy to ensure a healthy cellular environment. Estimates of errors during protein synthesis in Saccharomyces cerevisiae have varied from 10-3 to 10-4 errors per codon. Here, we show that errors made by ${\rm{tRNA}}^{\rm Glu}_{\rm UUC}$ in yeast can vary 100-fold, from 10-6 to 10-4 errors per codon. The most frequent errors require a G•U mismatch at the second position for the near cognate codon GGA (Gly). We also show, contrary to our previous results, that yeast tRNAs can make errors involving mismatches at the wobble position but with low efficiency. We have also assessed the effect on misreading frequency of post-transcriptional modifications of tRNAs, which are known to regulate cognate codon decoding in yeast. We tested the roles of mcm5s2U34 and t6A37 and show that their effects depend on details of the codon anticodon interaction including the position of the modification with respect to the base mismatch and the nature of that mismatch. Both mcm5 and s2 modification of wobble uridine strongly stabilizes G2•U35 mismatches when ${\rm{tRNA}}^{\rm Glu}_{\rm UUC}$ misreads the GGA Gly codon but has weaker effects on other mismatches. By contrast, t6A37 destabilizes U1•U36 mismatches when ${\rm{tRNA}}^{\rm Lys}_{\rm UUU}$ misreads UAA or UAG but stabilizes mismatches at the second and wobble positions.

Codon-specific effects of tRNA anticodon loop modifications on translational misreading errors in the yeastSaccharomyces cerevisiae