Nucleic Acids Research Advance Access originally published online on January 16, 2009
Nucleic Acids Research 2009 37(5):1616-1627; doi:10.1093/nar/gkp001
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Nucleic Acids Research, 2009, Vol. 37, No. 5 1616-1627
© 2009 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
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Unconventional decoding of the AUA codon as methionine by mitochondrial tRNAMet with the anticodon f5CAU as revealed with a mitochondrial in vitro translation system
1Department of Chemistry and Biotechnology, Graduate School of Engineering, University of Tokyo, Bunkyo-ku, Tokyo 113-8656, Japan, 2Department of Chemistry, University of North Carolina at Chapel Hill, NC 27599-3290, USA, 3Department of Medical Genome Science, Graduate School of Frontier Sciences, University of Tokyo, 5-1-5 Kashiwanoha, Kashiwa, Chiba 277-8562 and 4Department of Biomolecular Science, Faculty of Engineering, Gifu University, Gifu 501-1193, Japan
*To whom correspondence should be addressed. Tel: +81 45 503 9204; Fax: +81 45 503 9201; Email: chie{at}gsc.riken.jp
Correspondence may also be addressed to Kimitsuna Watanabe. Tel: +81 3 3599 8106; Fax: +81 3 5530 2064; Email: kim-watanabe{at}aist.go.jp
Received October 23, 2008. Revised December 27, 2008. Accepted December 30, 2008.
Mitochondrial (mt) tRNAMet has the unusual modified nucleotide 5-formylcytidine (f5C) in the first position of the anticodon. This tRNA must translate both AUG and AUA as methionine. By constructing an in vitro translation system from bovine liver mitochondria, we examined the decoding properties of the native mt tRNAMet carrying f5C in the anticodon compared to a transcript that lacks the modification. The native mt Met-tRNA could recognize both AUA and AUG codons as Met, but the corresponding synthetic tRNAMet lacking f5C (anticodon CAU), recognized only the AUG codon in both the codon-dependent ribosomal binding and in vitro translation assays. Furthermore, the Escherichia coli elongator tRNAMetm with the anticodon ac4CAU (ac4C = 4-acetylcytidine) and the bovine cytoplasmic initiator tRNAMet (anticodon CAU) translated only the AUG codon for Met on mt ribosome. The codon recognition patterns of these tRNAs were the same on E. coli ribosomes. These results demonstrate that the f5C modification in mt tRNAMet plays a crucial role in decoding the nonuniversal AUA codon as Met, and that the genetic code variation is compensated by a change in the tRNA anticodon, not by a change in the ribosome. Base pairing models of f5C-G and f5C-A based on the chemical properties of f5C are presented.
Present addresses: Chie Takemoto, Systems and Structural Biology Center, Yokohama Institute, RIKEN, Yokohama 230-0045, Japan
Kimitsuna Watanabe, Biomedicinal Information Research Center, National Institute of Advanced Industrial Science and Technology, 2-42 Aomi, Koto-ku, Tokyo 135-0064, Japan
Deceased.