Single-Plasmid-Based System for Efficient Noncanonical Amino Acid Mutagenesis in Cultured Mammalian Cells.
We describe a new expression system for efficient non-canonical amino acid mutagenesis in cultured mammalian cells by using the pyrrolysine tRNA synthetase/tRNACUA (Pyl) pair. A significant improvement in the incorporation of non-canonical amino acids into proteins was obtained by combining all the required genetic components into a single and compact vector that can be efficiently delivered to different mammalian cell lines by conventional transfection reagents.
Department of Chemistry, The National Institute for Biotechnology in the Negev, Ben-Gurion University of the Negev, Beer-Sheva, 84105, Israel.
Sarit
S
Cohen
Department of Chemistry, The National Institute for Biotechnology in the Negev, Ben-Gurion University of the Negev, Beer-Sheva, 84105, Israel. arbely@bgu.ac.il.|Department of Life-Sciences, Ben-Gurion University of the Negev, Beer-Sheva, 84105, Israel. arbely@bgu.ac.il.
Eyal
E
Arbely
D000107:Acetylation; D000596:Amino Acids / Q000378:metabolism*; D000604:Amino Acyl-tRNA Synthetases / Q000378:metabolism; D000818:Animals; D005815:Genetic Code; D045325:HCT116 Cells; D057809:HEK293 Cells; D006801:Humans; D008239:Lysine / Q000031:analogs & derivatives / Q000378:metabolism; D051379:Mice; D016296:Mutagenesis; D041681:NIH 3T3 Cells; D010957:Plasmids / Q000235:genetics / Q000378:metabolism*; D015202:Protein Engineering; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism
D000596:Amino Acids; D016159:Tumor Suppressor Protein p53; D000604:Amino Acyl-tRNA Synthetases; C456839:pyrrolysine; D008239:Lysine
amber suppression; genetic code expansion; protein engineering; protein expression; pyrrolysine