Nucleic Acids Research Advance Access originally published online on September 18, 2009
Nucleic Acids Research 2009 37(20):6881-6895; doi:10.1093/nar/gkp697
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Nucleic Acids Research, 2009, Vol. 37, No. 20 6881-6895
© The Author(s) 2009. Published by Oxford University Press.
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.5/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
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Tertiary network in mammalian mitochondrial tRNAAsp revealed by solution probing and phylogeny
1Architecture et Réactivité de l'A;RN, Université de Strasbourg, CNRS, IBMC 15 rue René Descartes, 67084 Strasbourg, France and 2Department of Chemistry and Biotechnology, Graduate School of Engineering, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan
*To whom correspondence should be addressed. Tel: +33 3 88 41 70 59; Fax: +33 3 88 60 22 18; Email: C.Florentz{at}unistra.fr
Received May 25, 2009. Revised July 23, 2009. Accepted August 7, 2009.
Primary and secondary structures of mammalian mitochondrial (mt) tRNAs are divergent from canonical tRNA structures due to highly skewed nucleotide content and large size variability of D- and T-loops. The nonconservation of nucleotides involved in the expected network of tertiary interactions calls into question the rules governing a functional L-shaped three-dimensional (3D) structure. Here, we report the solution structure of human mt-tRNAAsp in its native post-transcriptionally modified form and as an in vitro transcript. Probing performed with nuclease S1, ribonuclease V1, dimethylsulfate, diethylpyrocarbonate and lead, revealed several secondary structures for the in vitro transcribed mt-tRNAAsp including predominantly the cloverleaf. On the contrary, the native tRNAAsp folds into a single cloverleaf structure, highlighting the contribution of the four newly identified post-transcriptional modifications to correct folding. Reactivities of nucleotides and phosphodiester bonds in the native tRNA favor existence of a full set of six classical tertiary interactions between the D-domain and the variable region, forming the core of the 3D structure. Reactivities of D- and T-loop nucleotides support an absence of interactions between these domains. According to multiple sequence alignments and search for conservation of Leontis–Westhof interactions, the tertiary network core building rules apply to all tRNAAsp from mammalian mitochondria.