Experimental methods for scanning unnatural amino acid mutagenesis.

Site-specific incorporation of unnatural amino acids into proteins in vivo relies on the genetic reassignment of nonsense or quadruplet codons. Here, we describe a general procedure for the random introduction of these codons into open reading frames resulting in protein libraries that are scanned with unnatural amino acid residues. These libraries can enable large-scale mutagenesis experiments aimed at understanding and improving protein function.