Wobble modification defect in tRNA disturbs codon–anticodon interaction in a mitochondrial disease

We previously showed that in mitochondrial tRNALys with an A8344G mutation responsible for myoclonus epilepsy associated with ragged‐red fibers (MERRF), a subgroup of mitochondrial encephalomyopathic diseases, the normally modified wobble base (a 2‐thiouridine derivative) remains unmodified. Since wobble base modifications are essential for translational efficiency and accuracy, we used mitochondrial components to estimate the translational activity in vitro of purified tRNALys carrying the mutation and found no mistranslation of non‐cognate codons by the mutant tRNA, but almost complete loss of translational activity for cognate codons. This defective translation was not explained by a decline in aminoacylation or lowered affinity toward elongation factor Tu. However, when direct interaction of the codon with the mutant tRNALys defective anticodon was examined by ribosomal binding analysis, the wild‐type but not the mutant tRNALys bound to an mRNA–ribosome complex. We therefore concluded that the anticodon base modification defect, which is forced by the pathogenic point mutation, disturbs codon–anticodon pairing in the mutant tRNALys, leading to a severe reduction in mitochondrial translation that eventually could result in the onset of MERRF.