ArticleProtein engineering, design & selection : PEDS2012
A method for multi-codon scanning mutagenesis of proteins based on asymmetric transposons.
Article in Protein engineering, design & selection : PEDS, 2012. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 7 papers.
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Who cites it
7 citing papers in PubMed.
- Unravelling molecular dynamics in living cells: Fluorescent protein biosensors for cell biology.Journal of microscopy · 2025Review
- Accessing unexplored regions of sequence space in directed enzyme evolution via insertion/deletion mutagenesis.Nature communications · 2020Article
- Structural plasticity of green fluorescent protein to amino acid deletions and fluorescence rescue by folding-enhancing mutations.BMC biochemistry · 2015Article
- Functional genomics in the study of yeast cell polarity: moving in the right direction.Philosophical transactions of the Royal Society of London. Series B, Biological sciences · 2013Review
- Rational protein sequence diversification by multi-codon scanning mutagenesis.Methods in molecular biology (Clifton, N.J.) · 2013Article
- Random mutagenesis MAPPIT analysis identifies binding sites for Vif and Gag in both cytidine deaminase domains of Apobec3G.PloS one · 2012Article
- PFunkel: efficient, expansive, user-defined mutagenesis.PloS one · 2012Article
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2 authors.
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Abstract
Random mutagenesis followed by selection or screening is a commonly used strategy to improve protein function. Despite many available methods for random mutagenesis, nearly all generate mutations at the nucleotide level. An ideal mutagenesis method would allow for the generation of 'codon mutations' to change protein sequence with defined or mixed amino acids of choice. Herein we report a method that allows for mutations of one, two or three consecutive codons. Key to this method is the development of a Mu transposon variant with asymmetric terminal sequences. As a demonstration of the method, we performed multi-codon scanning on the gene encoding superfolder GFP (sfGFP). Characterization of 50 randomly chosen clones from each library showed that more than 40% of the mutants in these three libraries contained seamless, in-frame mutations with low site preference. By screening only 500 colonies from each library, we successfully identified several spectra-shift mutations, including a S205D variant that was found to bear a single excitation peak in the UV region.
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