title: Evidence for aminoacylation-induced conformational changes in human mitochondrial tRNAs.
abstract: Analysis by acid polyacrylamide/urea gel electrophoresis of 14 individual mitochondrial tRNAs (mt-tRNAs) from human cells has revealed a variable decrease in mobility of the aminoacylated relative to the nonacylated form, with the degree of separation of the two forms not being correlated with the mass, polar character, or charge of the amino acid. Separation of the charged and uncharged species has been found to be independent of tRNA denaturation, being observed also in the absence of urea. In another approach, electrophoresis through a perpendicular denaturing gradient gel of several individual mt-tRNAs has shown a progressive unfolding of the tRNA with increasing denaturant concentration, which is consistent with an initial disruption of tertiary interactions, followed by the sequential melting of the four stems of the cloverleaf structure. A detailed analysis of the unfolding process of charged and uncharged tRNALys and tRNALeu(UUR) has revealed that the separation of the two forms of these tRNAs persisted throughout the almost entire range of denaturant concentrations used and was lost upon denaturation of the last helical domain(s), which most likely included the amino acid acceptor stem. These observations strongly suggest that the electrophoretic retardation of the charged species reflects an aminoacylation-induced conformational change of the 3'-end of these mt-tRNAs, with possible significant implications in connection with the known role of the acceptor end in tRNA interactions with the ribosomal peptidyl transferase center and the elongation factor Tu.
authors.affiliation: Division of Biology, California Institute of Technology, Pasadena 91125, USA.
authors.forename: J A
authors.initials: JA
authors.lastname: Enríquez
authors.forename: G
authors.initials: G
authors.lastname: Attardi
mesh_terms: D001483:Base Sequence; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008928:Mitochondria / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D015345:Oligonucleotide Probes / Q000737:chemistry; D014176:Protein Biosynthesis; D012313:RNA / Q000648:ultrastructure*; D000077278:RNA, Mitochondrial; D012346:RNA, Transfer, Amino Acyl / Q000648:ultrastructure*; D046249:Transfer RNA Aminoacylation
chemical_list: D015345:Oligonucleotide Probes; D000077278:RNA, Mitochondrial; D012346:RNA, Transfer, Amino Acyl; D012313:RNA