ZFIN ID: ZDB-PUB-190528-23
Expanding the Zebrafish Genetic Code through Site-Specific Introduction of Azido-lysine, Bicyclononyne-lysine, and Diazirine-lysine
Syed, J., Palani, S., Clarke, S.T., Asad, Z., Bottrill, A.R., Jones, A.M.E., Sampath, K., Balasubramanian, M.K.
Date: 2019
Source: International Journal of Molecular Sciences   20(10): (Journal)
Registered Authors: Clarke, Scott, Sampath, Karuna
Keywords: click chemistry, crosslinking, genetic code expansion, protein engineering, proteomics, unnatural amino acid, zebrafish
MeSH Terms:
  • Animals
  • Codon, Terminator/genetics
  • Genetic Code
  • Glutathione Transferase/genetics
  • Green Fluorescent Proteins/genetics
  • Lysine/analogs & derivatives*
  • Lysine/genetics
  • Protein Engineering/methods*
  • Zebrafish/genetics*
  • Zebrafish Proteins/genetics
PubMed: 31130675 Full text @ Int. J. Mol. Sci.
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ABSTRACT
Site-specific incorporation of un-natural amino acids (UNAA) is a powerful approach to engineer and understand protein function. Site-specific incorporation of UNAAs is achieved through repurposing the amber codon (UAG) as a sense codon for the UNAA, using a tRNACUA that base pairs with an UAG codon in the mRNA and an orthogonal amino-acyl tRNA synthetase (aaRS) that charges the tRNACUA with the UNAA. Here, we report an expansion of the zebrafish genetic code to incorporate the UNAAs, azido-lysine (AzK), bicyclononyne-lysine (BCNK), and diazirine-lysine (AbK) into green fluorescent protein (GFP) and glutathione-s-transferase (GST). We also present proteomic evidence for UNAA incorporation into GFP. Our work sets the stage for the use of AzK, BCNK, and AbK introduction into proteins as a means to investigate and engineer their function in zebrafish.
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