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

We previously showed that in mitochondrial tRNA^Lys 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 tRNA^Lys 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 tRNA^Lys defective anticodon was examined by ribosomal binding analysis, the wild-type but not the mutant tRNA^Lys bound to an mRNAribosome 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 tRNA^Lys, leading to a severe reduction in mitochondrial translation that eventually could result in the onset of MERRF.