High-level misincorporation of lysine for arginine at AGA codons in a fusion protein expressed in Escherichia coli

J Mol Biol. 1996 Oct 4;262(4):407-12. doi: 10.1006/jmbi.1996.0524.


The expression of eukaryotic genes in Escherichia coli is one of the most frequently used tools of modern science. The arginine codon AGA is a common codon in eukaryotic genes but is particularly rare in E. coli. We report here 36 to 42% misincorporation of lysine at three AGA codons in a well-expressed protein. This misincorporation yields a protein whose electrospray mass spectrum (ESMS) shows peaks at the expected mass (M), M-28, M-56 and M-84 with intensities representing 34.5(+/-0.7), 37.5(+/-1.1), 21.2(+/-1.7) and 6.6(+/-0.5) % of the total intensity, respectively. Replacement of either all three AGA codons or the two closest to the 3' end of the gene by the more common CGC arginine codon gave a protein with a single ESMS peak. Misincorporation could also be eliminated by the co-expression of the tRNA(UCL)Arg gene, argU. These studies demonstrate that misincorporation of amino acids at rare codons of recombinant proteins can be far higher than previously thought.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Arginine / chemistry*
  • Base Sequence
  • Codon*
  • Escherichia coli
  • Lysine / chemistry*
  • Mass Spectrometry
  • Molecular Sequence Data
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / genetics*


  • Codon
  • Recombinant Fusion Proteins
  • Arginine
  • Lysine