Beyond the natural proteome: nondegenerate saturation mutagenesis - methodologies and advantages

M.M. Ferreira Amaral, L. Frigotto, A.V. Hine*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review


Beyond the natural proteome, high-throughput mutagenesis offers the protein engineer an opportunity to “tweak” the wild-type activity of a protein to create a recombinant protein with required attributes. Of the various approaches available, saturation mutagenesis is one of the core techniques employed by protein engineers and in recent times, nondegenerate saturation mutagenesis is emerging as the approach of choice. This review compares the current methodologies available for conducting nondegenerate saturation mutagenesis with traditional, degenerate saturation and briefly outlines the options available for screening the resulting libraries, to discover a novel protein with the required activity and/or specificity.
Original languageEnglish
Pages (from-to)111-133
JournalMethods in Enzymology
Publication statusPublished - 3 Jan 2017

Bibliographical note

Proteomics in biology, part A ISBN: 978-0-1280-9742-7.
© 2016, Elsevier. Licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International


  • gene shuffling
  • molecular evolution
  • directed evolution
  • synthetic biology
  • saturation mutagenesis
  • non-degenerate
  • diversity
  • protein engineering
  • antibody engineering


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