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Design of S ‐Allylcysteine in Situ Production and Incorporation Based on a Novel Pyrrolysyl‐tRNA Synthetase Variant
Author(s) -
Exner Matthias P.,
Kuenzl Tilmann,
To Tuyet Mai T.,
Ouyang Zhaofei,
Schwagerus Sergej,
Hoesl Michael G.,
Hackenberger Christian P. R.,
Lensen Marga C.,
Panke Sven,
Budisa Nediljko
Publication year - 2017
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201600537
Subject(s) - bioorthogonal chemistry , transfer rna , amino acid , cysteine , translation (biology) , biochemistry , chemistry , alkene , chemical biology , protein biosynthesis , biosynthesis , aminoacyl trna synthetase , in situ , combinatorial chemistry , enzyme , click chemistry , rna , organic chemistry , messenger rna , gene , catalysis
The noncanonical amino acid S ‐allyl cysteine (Sac) is one of the major compounds of garlic extract and exhibits a range of biological activities. It is also a small bioorthogonal alkene tag capable of undergoing controlled chemical modifications, such as photoinduced thiol‐ene coupling or Pd‐mediated deprotection. Its small size guarantees minimal interference with protein structure and function. Here, we report a simple protocol efficiently to couple in‐situ semisynthetic biosynthesis of Sac and its incorporation into proteins in response to amber (UAG) stop codons. We exploited the exceptional malleability of pyrrolysyl‐tRNA synthetase (PylRS) and evolved an S ‐allylcysteinyl‐tRNA synthetase (SacRS) capable of specifically accepting the small, polar amino acid instead of its long and bulky aliphatic natural substrate. We succeeded in generating a novel and inexpensive strategy for the incorporation of a functionally versatile amino acid. This will help in the conversion of orthogonal translation from a standard technique in academic research to industrial biotechnology.