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

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

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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Ampicillin Resistance / genetics
  • Caulobacter crescentus / genetics*
  • Caulobacter crescentus / metabolism*
  • Codon, Terminator*
  • Escherichia coli / genetics*
  • Escherichia coli / metabolism*
  • Gene Expression
  • Gene Expression Regulation, Bacterial
  • Histidine-tRNA Ligase / metabolism*
  • RNA, Transfer, His / genetics*
  • RNA, Transfer, His / metabolism

Substances

  • Codon, Terminator
  • RNA, Transfer, His
  • Histidine-tRNA Ligase