Evolving the N-Terminal Domain of Pyrrolysyl-tRNA Synthetase for Improved Incorporation of Noncanonical Amino Acids. Sharma, V., Zeng, Y., Wang, W. W., Qiao, Y., Kurra, Y., & Liu, W. R. ChemBioChem, 19(1):26–30, 2018. _eprint: https://onlinelibrary.wiley.com/doi/pdf/10.1002/cbic.201700268Paper doi abstract bibtex By evolving the N-terminal domain of Methanosarcina mazei pyrrolysyl-tRNA synthetase (PylRS) that directly interacts with tRNAPyl, a mutant clone displaying improved amber-suppression efficiency for the genetic incorporation of Nϵ-(tert-butoxycarbonyl)-l-lysine threefold more than the wild type was identified. The identified mutations were R19H/H29R/T122S. Direct transfer of these mutations to two other PylRS mutants that were previously evolved for the genetic incorporation of Nϵ-acetyl-l-lysine and Nϵ-(4-azidobenzoxycarbonyl)-l-δ,ϵ-dehydrolysine also improved the incorporation efficiency of these two noncanonical amino acids. As the three identified mutations were found in the N-terminal domain of PylRS that was separated from its catalytic domain for charging tRNAPyl with a noncanonical amino acid, they could potentially be introduced to all other PylRS mutants to improve the incorporation efficiency of their corresponding noncanonical amino acids. Therefore, it represents a general strategy to optimize the pyrrolysine incorporation system-based noncanonical amino-acid mutagenesis.
@article{sharma_evolving_2018,
title = {Evolving the {N}-{Terminal} {Domain} of {Pyrrolysyl}-{tRNA} {Synthetase} for {Improved} {Incorporation} of {Noncanonical} {Amino} {Acids}},
volume = {19},
issn = {1439-7633},
url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/cbic.201700268},
doi = {10.1002/cbic.201700268},
abstract = {By evolving the N-terminal domain of Methanosarcina mazei pyrrolysyl-tRNA synthetase (PylRS) that directly interacts with tRNAPyl, a mutant clone displaying improved amber-suppression efficiency for the genetic incorporation of Nϵ-(tert-butoxycarbonyl)-l-lysine threefold more than the wild type was identified. The identified mutations were R19H/H29R/T122S. Direct transfer of these mutations to two other PylRS mutants that were previously evolved for the genetic incorporation of Nϵ-acetyl-l-lysine and Nϵ-(4-azidobenzoxycarbonyl)-l-δ,ϵ-dehydrolysine also improved the incorporation efficiency of these two noncanonical amino acids. As the three identified mutations were found in the N-terminal domain of PylRS that was separated from its catalytic domain for charging tRNAPyl with a noncanonical amino acid, they could potentially be introduced to all other PylRS mutants to improve the incorporation efficiency of their corresponding noncanonical amino acids. Therefore, it represents a general strategy to optimize the pyrrolysine incorporation system-based noncanonical amino-acid mutagenesis.},
language = {en},
number = {1},
urldate = {2021-11-24},
journal = {ChemBioChem},
author = {Sharma, Vangmayee and Zeng, Yu and Wang, W. Wesley and Qiao, Yuchen and Kurra, Yadagiri and Liu, Wenshe R.},
year = {2018},
note = {\_eprint: https://onlinelibrary.wiley.com/doi/pdf/10.1002/cbic.201700268},
keywords = {amber suppression, gene technology, mutagenesis, noncanonical amino acids, tRNA},
pages = {26--30},
}
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Direct transfer of these mutations to two other PylRS mutants that were previously evolved for the genetic incorporation of Nϵ-acetyl-l-lysine and Nϵ-(4-azidobenzoxycarbonyl)-l-δ,ϵ-dehydrolysine also improved the incorporation efficiency of these two noncanonical amino acids. As the three identified mutations were found in the N-terminal domain of PylRS that was separated from its catalytic domain for charging tRNAPyl with a noncanonical amino acid, they could potentially be introduced to all other PylRS mutants to improve the incorporation efficiency of their corresponding noncanonical amino acids. 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The identified mutations were R19H/H29R/T122S. Direct transfer of these mutations to two other PylRS mutants that were previously evolved for the genetic incorporation of Nϵ-acetyl-l-lysine and Nϵ-(4-azidobenzoxycarbonyl)-l-δ,ϵ-dehydrolysine also improved the incorporation efficiency of these two noncanonical amino acids. As the three identified mutations were found in the N-terminal domain of PylRS that was separated from its catalytic domain for charging tRNAPyl with a noncanonical amino acid, they could potentially be introduced to all other PylRS mutants to improve the incorporation efficiency of their corresponding noncanonical amino acids. Therefore, it represents a general strategy to optimize the pyrrolysine incorporation system-based noncanonical amino-acid mutagenesis.},\n\tlanguage = {en},\n\tnumber = {1},\n\turldate = {2021-11-24},\n\tjournal = {ChemBioChem},\n\tauthor = {Sharma, Vangmayee and Zeng, Yu and Wang, W. 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