All-Codon Mutagenesis for Structure-Function Studies of Chemotaxis Signaling Proteins

Abstract

The technique of all-codon mutagenesis can generate mutants that represent all possible amino acid replacements at any particular residue in a protein. It is thus a powerful tool to probe structure-function relationships in proteins of interest. In this chapter, we describe how we used all-codon mutagenesis to obtain mutants of the Escherichia coli serine receptor Tsr with amino acid replacements at residue F373, a functionally important site in this protein. We provide general protocols for mutagenesis of a target codon in a plasmid-borne gene and for the selection and screening of the resultant mutants. These techniques should be adaptable for the study of a variety of bacterial proteins.