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Literature summary extracted from

  • Serfling, R.; Lorenz, C.; Etzel, M.; Schicht, G.; Boettke, T.; Moerl, M.; Coin, I.
    Designer tRNAs for efficient incorporation of non-canonical amino acids by the pyrrolysine system in mammalian cells (2018), Nucleic Acids Res., 46, 1-10 .
    View publication on PubMedView publication on EuropePMC

Protein Variants

EC Number Protein Variants Comment Organism
6.1.1.26 Y349F the pyrrolysyl-tRNA synthetase/tRNAPyl pair is the most versatile and widespread system for the incorporation of non-canonical amino acids (ncAAs) into proteins in mammalian cells. Low yields of ncAA incorporation severely limit its applicability to relevant biological targets. Generation of two tRNAPyl variants that significantly boost the performance of the pyrrolysine system. Compared to the original tRNAPyl, the engineered tRNAs feature a canonical hinge between D- and T-loop, show higher intracellular concentrations and bear partially distinct post-transcriptional modifications. They allow efficient ncAA incorporation into a G-protein coupled receptor (GPCR) and simultaneous ncAA incorporation at two GPCR sites. Different combinations of recognition motifs for PylRS are introduced into Bos taurus mt-tRNASerCUA. Bicistronic plasmids are generated combining each tRNA with the PylRS from Methanosarcina barkeri, which bears the mutation Y349F for enhanced activity (PylRSF). In the presence of the PylRS substrate Lys(Boc), four tRNAs in the M series and five in the C series yield higher amber suppression compared to the tRNAPyl, whereas the other tRNAs show either lower efficiency or no amber suppression at all, overview Methanosarcina barkeri

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
6.1.1.26 Mg2+ required Methanosarcina barkeri

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
6.1.1.26 ATP + L-pyrrolysine + tRNAPyl Methanosarcina barkeri
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 AMP + diphosphate + L-pyrrolysyl-tRNAPyl
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
6.1.1.26 Methanosarcina barkeri Q6WRH6
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-

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
6.1.1.26 ATP + L-pyrrolysine + tRNAPyl
-
 Methanosarcina barkeri AMP + diphosphate + L-pyrrolysyl-tRNAPyl
-
 ?
6.1.1.26 additional information the archaeal tRNAPyl features a secondary structure that significantly diverges from that of canonical tRNAs Methanosarcina barkeri ?
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 ?

Synonyms

EC Number Synonyms Comment Organism
6.1.1.26 PylRS
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 Methanosarcina barkeri
6.1.1.26 pyrrolysyl-tRNA synthetase
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 Methanosarcina barkeri

Cofactor

EC Number Cofactor Comment Organism Structure
6.1.1.26 ATP
-
 Methanosarcina barkeri