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Update on Pure Translation Display with Unnatural Amino Acid Incorporation

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Ribosome Display and Related Technologies

Part of the book series: Methods in Molecular Biology ((MIMB,volume 805))

Abstract

The identification of peptide and protein ligands by directed evolution in vitro has been of enormous utility in molecular biology and biotechnology. However, the translation step in almost all polypeptide selection methods is performed in vivo or in crude extracts, restricting applications. These restrictions include a limited library size due to transformation efficiency, unwanted competing reactions in translation, and an inability to incorporate multiple unnatural amino acids (AAs) with high fidelity and efficiency. These restrictions can be addressed by “pure translation display” where the translation step is performed in a purified system. To date, all pure translation display selections have coupled genotype to phenotype in a ribosome display format, though other formats also should be practical. Here, we detail the original, proof-of-principle, pure-translation-display method because this version should be the most suitable for encoding multiple unnatural AAs per peptide product toward the goal of “peptidomimetic evolution.” Challenges and progress toward this ultimate goal are discussed and are mainly associated with improving the efficiency of ribosomal polymerization of multiple unnatural AAs.

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Acknowledgments

We thank Tarjani Thaker for help with Fig. 2, and Craig Goodwin, Seth Villarreal, and the editors for comments on the manuscript. This work was supported by the National Institutes of Health and the American Cancer Society.

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Authors and Affiliations

  1. Department of Pharmacology, Vanderbilt Institute of Chemical Biology, Vanderbilt University Medical Center, Nashville, TN, USA

    R. Edward Watts

  2. Department of Cell and Molecular Biology, Uppsala University, Uppsala, Sweden

    Anthony C. Forster

Authors
  1. R. Edward Watts
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  2. Anthony C. Forster
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Corresponding author

Correspondence to Anthony C. Forster .

Editor information

Editors and Affiliations

  1. , Antibody Discovery & Protein Engineering, MedImmune Limited, Granta Park, Cambridge, CB21 6GH, United Kingdom

    Julie A. Douthwaite

  2. , Antibody Discovery & Protein Engineering, MedImmune Limited, Granta Park, Cambridge, CB21 6GH, United Kingdom

    Ronald H. Jackson

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Watts, R.E., Forster, A.C. (2012). Update on Pure Translation Display with Unnatural Amino Acid Incorporation. In: Douthwaite, J., Jackson, R. (eds) Ribosome Display and Related Technologies. Methods in Molecular Biology, vol 805. Springer, New York, NY. https://doi.org/10.1007/978-1-61779-379-0_20

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  • DOI: https://doi.org/10.1007/978-1-61779-379-0_20

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  • Publisher Name: Springer, New York, NY

  • Print ISBN: 978-1-61779-378-3

  • Online ISBN: 978-1-61779-379-0

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