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Evidence for aminoacylation-induced conformational changes in human mitochondrial tRNAs

Enríquez, José A. and Attardi, Giuseppe (1996) Evidence for aminoacylation-induced conformational changes in human mitochondrial tRNAs. Proceedings of the National Academy of Sciences of the United States of America, 93 (16). pp. 8300-8305. ISSN 0027-8424. http://resolver.caltech.edu/CaltechAUTHORS:ENRpnas96

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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 tRNA(Lys) and tRNA(Leu(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.


Item Type:Article
Additional Information:Copyright © 1996 by the National Academy of Sciences. Contributed by Giuseppe Attardi, May 2, 1996. We thank Dr. Uttam RajBhandary for valuable comments on the manuscript, to Yuichi Michikawa for useful advice concerning the use of DGGE and to Patricio Fernandez-Silva for discussions. The expert technical assistance of A. Drew, B. Keeley and R. Zedan is gratefully acknowledged. This work was supported by National Institutes of Health Grant GM-11726 to G.A. and a Plan Formacion del Personal Investigador. Fellowship from the Spanish Ministry of Education to J.A.E. The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.
Subject Keywords:TRANSFER RNA-SYNTHETASE; PHENYLALANINE TRANSFER-RNA; INITIATOR TRANSFER-RNA; ESCHERICHIA-COLI; ACCEPTOR STEM; CRYSTAL-STRUCTURE; PROTEIN-SYNTHESIS; TERNARY COMPLEX; RIBOSOMAL-RNA; PHE-TRNA(PHE)
Record Number:CaltechAUTHORS:ENRpnas96
Persistent URL:http://resolver.caltech.edu/CaltechAUTHORS:ENRpnas96
Alternative URL:http://www.pnas.org/cgi/content/abstract/93/16/8300
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:1510
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:25 Jan 2006
Last Modified:26 Dec 2012 08:44

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