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Title: Pyrrolysyl-tRNA Synthetase, an Aminoacyl-tRNA Synthetase for Genetic Code Expansion

Genetic code expansion (GCE) has become a central topic of synthetic biology. GCE relies on engineered aminoacyl-tRNA synthetases (aaRSs) and a cognate tRNA species to allow codon reassignment by co-translational insertion of non-canonical amino acids (ncAAs) into proteins. Introduction of such amino acids increases the chemical diversity of recombinant proteins endowing them with novel properties. Such proteins serve in sophisticated biochemical and biophysical studies both in vitro and in vivo, they may become unique biomaterials or therapeutic agents, and they afford metabolic dependence of genetically modified organisms for biocontainment purposes. In the Methanosarcinaceae the incorporation of the 22nd genetically encoded amino acid, pyrrolysine (Pyl), is facilitated by pyrrolysyl-tRNA synthetase (PylRS) and the cognate UAG-recognizing tRNAPyl. This unique aaRS•tRNA pair functions as an orthogonal translation system (OTS) in most model organisms. The facile directed evolution of the large PylRS active site to accommodate many ncAAs, and the enzyme’s anticodon-blind specific recognition of the cognate tRNAPyl make this system highly amenable for GCE purposes. The remarkable polyspecificity of PylRS has been exploited to incorporate >100 different ncAAs into proteins. Here we review the Pyl-OT system and selected GCE applications to examine the properties of an effective OTS.
Authors:
 [1] ;  [1] ;  [1] ;  [1]
  1. Yale Univ., New Haven, CT (United States)
Publication Date:
Grant/Contract Number:
FG02-98ER20311
Type:
Accepted Manuscript
Journal Name:
Croatica Chemica Acta
Additional Journal Information:
Journal Volume: 89; Journal Issue: 2; Journal ID: ISSN 0011-1643
Publisher:
The Croatian Chemical Society
Research Org:
Michigan State Univ., East Lansing, MI (United States)
Sponsoring Org:
USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22)
Country of Publication:
United States
Language:
English
Subject:
37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY
OSTI Identifier:
1434637

Crnkovic, Ana, Suzuki, Tateki, Soll, Dieter, and Reynolds, Noah M. Pyrrolysyl-tRNA Synthetase, an Aminoacyl-tRNA Synthetase for Genetic Code Expansion. United States: N. p., Web. doi:10.5562/cca2825.
Crnkovic, Ana, Suzuki, Tateki, Soll, Dieter, & Reynolds, Noah M. Pyrrolysyl-tRNA Synthetase, an Aminoacyl-tRNA Synthetase for Genetic Code Expansion. United States. doi:10.5562/cca2825.
Crnkovic, Ana, Suzuki, Tateki, Soll, Dieter, and Reynolds, Noah M. 2016. "Pyrrolysyl-tRNA Synthetase, an Aminoacyl-tRNA Synthetase for Genetic Code Expansion". United States. doi:10.5562/cca2825. https://www.osti.gov/servlets/purl/1434637.
@article{osti_1434637,
title = {Pyrrolysyl-tRNA Synthetase, an Aminoacyl-tRNA Synthetase for Genetic Code Expansion},
author = {Crnkovic, Ana and Suzuki, Tateki and Soll, Dieter and Reynolds, Noah M.},
abstractNote = {Genetic code expansion (GCE) has become a central topic of synthetic biology. GCE relies on engineered aminoacyl-tRNA synthetases (aaRSs) and a cognate tRNA species to allow codon reassignment by co-translational insertion of non-canonical amino acids (ncAAs) into proteins. Introduction of such amino acids increases the chemical diversity of recombinant proteins endowing them with novel properties. Such proteins serve in sophisticated biochemical and biophysical studies both in vitro and in vivo, they may become unique biomaterials or therapeutic agents, and they afford metabolic dependence of genetically modified organisms for biocontainment purposes. In the Methanosarcinaceae the incorporation of the 22nd genetically encoded amino acid, pyrrolysine (Pyl), is facilitated by pyrrolysyl-tRNA synthetase (PylRS) and the cognate UAG-recognizing tRNAPyl. This unique aaRS•tRNA pair functions as an orthogonal translation system (OTS) in most model organisms. The facile directed evolution of the large PylRS active site to accommodate many ncAAs, and the enzyme’s anticodon-blind specific recognition of the cognate tRNAPyl make this system highly amenable for GCE purposes. The remarkable polyspecificity of PylRS has been exploited to incorporate >100 different ncAAs into proteins. Here we review the Pyl-OT system and selected GCE applications to examine the properties of an effective OTS.},
doi = {10.5562/cca2825},
journal = {Croatica Chemica Acta},
number = 2,
volume = 89,
place = {United States},
year = {2016},
month = {6}
}