Life without tRNAIle-lysidine synthetase: translation of the isoleucine codon AUA in Bacillus subtilis lacking the canonical tRNA2Ile |
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| Authors: | Caroline K?hrer Debabrata Mandal Kirk W Gaston Henri Grosjean Patrick A Limbach Uttam L RajBhandary |
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| Institution: | 1.Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA, 2.Department of Chemistry, Rieveschl Laboratories for Mass Spectrometry, University of Cincinnati, Cincinnati, OH 45221, USA and 3.Centre de Génétique Moléculaire, CNRS, Gif-sur-Yvette, F-91198, France |
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| Abstract: | Translation of the isoleucine codon AUA in most prokaryotes requires a modified C (lysidine or agmatidine) at the wobble position of tRNA2Ile to base pair specifically with the A of the AUA codon but not with the G of AUG. Recently, a Bacillus subtilis strain was isolated in which the essential gene encoding tRNAIle-lysidine synthetase was deleted for the first time. In such a strain, C34 at the wobble position of tRNA2Ile is expected to remain unmodified and cells depend on a mutant suppressor tRNA derived from tRNA1Ile, in which G34 has been changed to U34. An important question, therefore, is how U34 base pairs with A without also base pairing with G. Here, we show (i) that unlike U34 at the wobble position of all B. subtilis tRNAs of known sequence, U34 in the mutant tRNA is not modified, and (ii) that the mutant tRNA binds strongly to the AUA codon on B. subtilis ribosomes but only weakly to AUG. These in vitro data explain why the suppressor strain displays only a low level of misreading AUG codons in vivo and, as shown here, grows at a rate comparable to that of the wild-type strain. |
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