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Updated: Jun 25, 2026

Residue-specific Incorporation of Noncanonical Amino Acids into Model Proteins Using an Escherichia coli Cell-free Transcription-translation System
Published on: August 1, 2016
Aminoacyl-tRNA synthesis by pre-translational amino acid modification
Liang Feng1, Kelly Sheppard, Suk Namgoong
1Department of Molecular Biophysics, Yale University, New Haven, Connecticut 06520-8114, USA.
Abstract:
Aminoacyl-tRNAs (aa-tRNAs) are essential substrates for ribosomal translation, and are generally synthesized by aminoacyl-tRNA synthetases (aaRSs). It was expected earlier that every organism would contain a complete set of twenty aaRSs, one for each canonical amino acid. However, analysis of the many known genome sequences and biochemical studies revealed that most organisms lack asparaginyl- and glutaminyl-tRNA synthetases, and thus are unable to attach asparagine and glutamine directly onto their corresponding tRNA. Instead, a pretranslational amino acid modification is required to convert Asp-tRNA(Asn) and Glu-tRNA(Gln) to the correctly charged Asn-tRNA(Asn) and Gln-tRNA(Gln), respectively. This transamidation pathway of amide aa-tRNA synthesis is common in most bacteria and archaea. Unexpected results from biochemical, genetic and genomic studies showed that a large variety of different bacteria rely on tRNA-dependent transamidation for the formation of the amino acid asparagine. Pretranslational modifications are not restricted to asparagine and glutamine but are also found in the biosynthesis of some other aa-tRNAs, such as the initiator tRNA fmet-tRNA(Met)(i) and Sec-tRNA(Sec) specifying selenocysteine, the 21(st) cotranslationally inserted amino acid. tRNA-dependent amino acid modification is also involved in the generation of aminolevulinic acid, the first precursor for porphyrin biosynthesis in many organisms.
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