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Charging tRNA with pyrrolysine

United States Patent Application

View the Complete Application at the US Patent & Trademark Office
Provided herein is a method for preparing a cell that, when exposed to the non-canonical amino acid, pyrrolysine or a derivative thereof and transformed or transfected with a polynucleotide comprising an in-frame UAG or TAG codon incorporates the pyrrolysine residue or derivative thereof into the protein or polynucleotide into the protein or polypeptide encoded by the polynucleotide. The method comprises introducing a first polynucleotide comprising a sequence that encodes a protein having pyrrolysyl-tRNA synthetase activity and a second polynucleotide comprising a sequence that encodes a pyrrolysine specific tRNA into the cell and maintining the cell under conditions that permit expression of the first and second polynucleotide. Also provided herein are modified cells produced in accordance with the present method and methods of making peptides, polypeptides, and proteins which utilize such modified cells. Also provide are kits comprising pyrrolysine or a pyrrolysine or both. The kits also provide the first polynucleotide and the second polynucelotide or cells produced in accordance with the present methods or both.
Chan, Michael K. (Hilliard, OH), Krzycki, Joseph A. (Columbus, OH)
11/ 204,295
August 15, 2005
STATEMENT REGARDING FEDERALLY FUNDED RESEARCH [0002] This work was supported at least in part by grants from the National Science Foundation (MCB-9808914); the Department of Energy (DE-FG02-92ER20042); and the National Institutes of Health (GM 061796). The Federal Government may have certain rights in this invention.