Pyrrolysyl-tRNA synthetase: An ordinary enzyme but an outstanding genetic code expansion tool

• Published in: Biochimica et biophysica acta 2014-06, Vol.1844 (6), p.1059-1070
• Main Authors: Wan, Wei, Tharp, Jeffery M., Liu, Wenshe R.
• Format: Article
• Language: English
• Subjects: Amber codon; Amino Acyl-tRNA Synthetases - genetics; Amino Acyl-tRNA Synthetases - metabolism; Codon, Terminator; Genetic Code; Genetic code expansion; Genetic Engineering - methods; Hydroxy acids; Lysine - analogs & derivatives; Lysine - metabolism; Methane - metabolism; Methanosarcina - genetics; Methanosarcina - metabolism; Methylamines - metabolism; Models, Molecular; Non-canonical amino acids; Protein Biosynthesis; Pyrrolysine; RNA, Transfer - metabolism; Substrate Specificity
• ISSN: 1570-9639; 0006-3002; 1878-1454
• DOI: 10.1016/j.bbapap.2014.03.002

The genetic incorporation of the 22nd proteinogenic amino acid, pyrrolysine (Pyl) at amber codon is achieved by the action of pyrrolysyl-tRNA synthetase (PylRS) together with its cognate tRNAPyl. Unlike most aminoacyl-tRNA synthetases, PylRS displays high substrate side chain promiscuity, low selectivity toward its substrate α-amine, and low selectivity toward the anticodon of tRNAPyl. These unique but ordinary features of PylRS as an aminoacyl-tRNA synthetase allow the Pyl incorporation machinery to be easily engineered for the genetic incorporation of more than 100 non-canonical amino acids (NCAAs) or α-hydroxy acids into proteins at amber codon and the reassignment of other codons such as ochre UAA, opal UGA, and four-base AGGA codons to code NCAAs. •PylRS has high substrate side chain promiscuity.•PylRS has high substrate α-substituent promiscuity.•PylRS doesn't specifically recognize its tRNA anticodon.•Using the Pyl system, 100 non-canonical amino acids or α-hydroxy acids are encoded.•Using the Pyl system, amber, opal, ochre, and AGGA codons are reassigned.