A growing number of human diseases are correlated with point mutations in transfer RNA (tRNA) genes within the mitochondrial genome. The cause-and-effect basis for diseases associated with mutations in human mitochondrial (hs mt) tRNAs is an area of active investigation in medical and cell biology research, but the limited availability of information concerning the properties of these tRNAs has hampered the delineation of the molecular basis of these mitochondrial pathologies. ? ? Our goal is to characterize the unique tRNAs functioning in human mitochondria, and to elucidate how the functional and structural properties of hs mt tRNA mutants associated with disease are perturbed. The effects of pathogenic mutations in hs mt tRNAs will be investigated by: 1) monitoring structural properties using solution footprinting, in vitro selection, computational, and spectroscopic methods, 2) investigating functional properties, including the efficiency and fidelity of aminoacylation and ribosomal protein synthesis, using a variety of in vitro assays, and 3) probing the function of these molecules in vivo using human cell lines and a bacterial model system. This multipronged approach will allow the formulation of structure/function relationships for this uncharacterized class of molecules and will provide molecular-level information concerning the effects of disease-related tRNA mutations.
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