Pathogenesis-related mutations in the T-loops of human mitochondrial tRNAs affect 3′ end processing and tRNA structure

Numerous mutations in the mitochondrial genome are associated with maternally transmitted diseases and syndromes that affect muscle and other high energy-demand tissues. The mitochondrial genome encodes 13 polypeptides, 2 rRNAs and 22 interspersed tRNAs via long bidirectional polycistronic primary transcripts, requiring precise excision of the tRNAs. Despite making up only ~10% of the mitochondrial genome, tRNA genes harbor most of the pathogenesis-related mutations. tRNase Z endonucleolytically removes the pre-tRNA 3' trailer. The flexible arm of tRNase Z recognizes and binds the elbow (including the T-loop) of pre-tRNA. Pathogenesis-related T-loop mutations in mitochondrial tRNAs could thus affect tRNA structure, reduce tRNase Z binding and 3' processing, and consequently slow mitochondrial protein synthesis. Here we inspect the effects of pathogenesis-related mutations in the T-loops of mitochondrial tRNAs on pre-tRNA structure and tRNase Z processing. Increases in K(M) arising from 59A > G substitutions in mitochondrial tRNA(Gly) and tRNA(Ile) accompany changes in T-loop structure, suggesting impaired substrate binding to enzyme.