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.
Key wordsScanning mutagenesis tRNA Mu transposon Codon Unnatural amino acids
The authors thank the National Institutes of Health (GM084396) for financial support. We are also grateful to Prof. Stefan Lutz (Emory University) for the plasmid pInSALect and Prof. Peter G. Schultz for the plasmid pSUP.
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