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.
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References
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Acknowledgment
Our work is supported by research grant GM19559 from the National Institute of General Medical Sciences. DNA sequencing and primer synthesis were carried out by the Protein-DNA Core Facility at the University of Utah, which receives support from National Cancer Institute grant CA42014 to the Huntsman Cancer Institute.
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Ames, P., Parkinson, J.S. (2018). All-Codon Mutagenesis for Structure-Function Studies of Chemotaxis Signaling Proteins. In: Manson, M. (eds) Bacterial Chemosensing. Methods in Molecular Biology, vol 1729. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7577-8_8
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DOI: https://doi.org/10.1007/978-1-4939-7577-8_8
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Publisher Name: Humana Press, New York, NY
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Online ISBN: 978-1-4939-7577-8
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