Using in‐line probing to characterize pathogenic mutations in human mitochondrial tRNAs

Abstract

In‐line probing (Regulski, E.E. and Breaker, R.R., (2008) Methods Mol. Biol. 419, 53) provides a straightforward method for probing RNA structure. We have used this technique to characterize the human mitochondrial tRNAAla and four tRNA mutants associated with mitochondrial pathologies, including MERFF (myoclonic epilepsy with ragged red fibers), ophthalmoplegia, and mitochondrial myopathy. Our data suggests that two of these mutants, C5650T and A5628G, adopt abnormal secondary structures at room temperature, which may lead to reduced aminoacylation levels. This work was supported by Carleton College and the Howard Hughes Medical Institute.