Orthogonal Protein Translation Using Pyrrolysyl-tRNA Synthetases for Single- and Multiple-Noncanonical Amino Acid Mutagenesis

Adv Biochem Eng Biotechnol. 2018:162:1-19. doi: 10.1007/10_2016_37.

Abstract

To date, the two systems most extensively used for noncanonical amino acid (ncAA) incorporation via orthogonal translation are based on the Methanococcus jannaschii TyrRS/tRNA CUATyr and the Methanosarcina barkeri/Methanosarcina mazei PylRS/tRNA CUAPyl pairs. Here, we summarize the development and usage of the pyrrolysine-based system for orthogonal translation, a process that allows for the recombinant production of site-specifically labeled proteins and peptides. Via stop codon suppression in Escherichia coli and mammalian cells, genetically encoded biomolecules can be equipped with a great diversity of chemical functionalities including click chemistry handles, post-translational modifications, and photocaged sidechains.

Keywords: Expanded genetic code; Noncanonical amino acid; Orthogonal translation; Pyrrolysyl-tRNA synthetase; Stop codon suppression.

Publication types

  • Review

MeSH terms

  • Amino Acyl-tRNA Synthetases* / genetics
  • Amino Acyl-tRNA Synthetases* / metabolism
  • Bacterial Proteins* / genetics
  • Bacterial Proteins* / metabolism
  • Codon, Terminator* / genetics
  • Codon, Terminator* / metabolism
  • Escherichia coli* / genetics
  • Escherichia coli* / metabolism
  • Methanococcus* / genetics
  • Methanococcus* / metabolism
  • Methanosarcina / genetics
  • Methanosarcina / metabolism
  • Mutagenesis
  • Protein Biosynthesis / genetics*
  • Suppression, Genetic / genetics*

Substances

  • Bacterial Proteins
  • Codon, Terminator
  • Amino Acyl-tRNA Synthetases