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Journal of Bacteriology, February 2004, p. 750-757, Vol. 186, No. 3
0021-9193/04/$08.00+0 DOI: 10.1128/JB.186.3.750-757.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.
The Conserved Cys-X1-X2-Cys Motif Present in the TtcA Protein Is Required for the Thiolation of Cytidine in Position 32 of tRNA from Salmonella enterica serovar Typhimurium
Gunilla Jäger, Ramune Leipuviene, Michael G. Pollard, Qiang Qian,
and Glenn R. Björk*
Department of Molecular Biology, Umeå University, S-90187 Umeå, Sweden
Received 30 June 2003/
Accepted 22 October 2003
The modified nucleoside 2-thiocytidine (s2C) has so far been found in tRNA from organisms belonging to the phylogenetic domains Archaea and Bacteria. In the bacteria Escherichia coli and Salmonella enterica serovar Typhimurium, s2C is present in position 32 of only four tRNA species
,
,
, and
. An in-frame deletion of an S. enterica gene (designated ttcA, for "two-thio-cytidine") was constructed, and such a mutant has no detectable s2C in its tRNA. The TtcA protein family is characterized by the existence of both a PP-loop and a Cys-X1-X2-Cys motif in the central region of the protein but can be divided into two distinct groups based on the presence and location of additional Cys-X1-X2-Cys motifs in terminal regions of the sequence. Mutant analysis showed that both cysteines in this central conserved Cys-X1-X2-Cys motif are required for the formation of s2C. The
ttcA1 mutant grows at the same rate as the congenic wild-type strain, and no growth disadvantage caused by the lack of s2C was observed in a mixed-population experiment. Lack of s2C32 did not reduce the selection rate at the ribosomal aminoacyl-tRNA site (A-site) for
at any of its cognate CGN codons, whereas A-site selection at AGG by
was dependent on the presence of s2C32. The presence of s2C32 in peptidyl-
or in peptidyl-
interfered with decoding in the A-site. The presence of s2C32 in
decreased the rate of translation of the CGA codon but not that of the CGU codon.
* Corresponding author. Mailing address: Department of Molecular Biology, Umeå University, S-90187 Umeå, Sweden. Phone: 46 90 7856756. Fax: 46 90 772630. E-mail:
glenn.bjork{at}molbiol.umu.se.
Present address: Telecommunication System Inc., Annapolis, MD 21401.
Journal of Bacteriology, February 2004, p. 750-757, Vol. 186, No. 3
0021-9193/04/$08.00+0 DOI: 10.1128/JB.186.3.750-757.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.
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