A convenient method for genetic incorporation of multiple noncanonical amino acids into one protein in Escherichia coli.

By overexpressing the C-terminal domain of the ribosomal protein L11 to decrease release factor 1-mediated termination of protein translation, enhanced amber suppression is achieved in E. coli. This enhanced amber suppression efficiency allows the genetic incorporation of three N(epsilon)-acetyl-l-lysines into one GFP(UV) protein in E. coli.