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PLoS One. 2013 Dec 30;8(12):e83630. doi: 10.1371/journal.pone.0083630. eCollection 2013.

Suppression of amber codons in Caulobacter crescentus by the orthogonal Escherichia coli histidyl-tRNA synthetase/tRNAHis pair.

Author information

1
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut, United States of America.
2
Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, Connecticut, United States of America.
3
Howard Hughes Medical Institute, Yale University, New Haven, Connecticut, United States of America.
4
Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, Connecticut, United States of America ; Howard Hughes Medical Institute, Yale University, New Haven, Connecticut, United States of America ; Department of Microbial Pathogenesis, Yale School of Medicine, New Haven, Connecticut, United States of America.
5
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut, United States of America ; Department of Chemistry, Yale University, New Haven, Connecticut, United States of America.

Abstract

While translational read-through of stop codons by suppressor tRNAs is common in many bacteria, archaea and eukaryotes, this phenomenon has not yet been observed in the α-proteobacterium Caulobacter crescentus. Based on a previous report that C. crescentus and Escherichia coli tRNA(His) have distinctive identity elements, we constructed E. coli tRNA(His) CUA, a UAG suppressor tRNA for C. crescentus. By examining the expression of three UAG codon- containing reporter genes (encoding a β-lactamase, the fluorescent mCherry protein, or the C. crescentus xylonate dehydratase), we demonstrated that the E. coli histidyl-tRNA synthetase/tRNA(His) CUA pair enables in vivo UAG suppression in C. crescentus. E. coli histidyl-tRNA synthetase (HisRS) or tRNA(His) CUA alone did not achieve suppression; this indicates that the E. coli HisRS/tRNA(His) CUA pair is orthogonal in C. crescentus. These results illustrate that UAG suppression can be achieved in C. crescentus with an orthogonal aminoacyl-tRNA synthetase/suppressor tRNA pair.

PMID:
24386240
PMCID:
PMC3875453
DOI:
10.1371/journal.pone.0083630
[Indexed for MEDLINE]
Free PMC Article
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