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A Computational Library Design Protocol for Rapid Improvement of Protein Stability: FRESCO

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Protein Engineering

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1685))

Abstract

The ability to stabilize enzymes and other proteins has wide-ranging applications. Most protocols for enhancing enzyme stability require multiple rounds of high-throughput screening of mutant libraries and provide only modest improvements of stability. Here, we describe a computational library design protocol that can increase enzyme stability by 20–35 °C with little experimental screening, typically fewer than 200 variants. This protocol, termed FRESCO, scans the entire protein structure to identify stabilizing disulfide bonds and point mutations, explores their effect by molecular dynamics simulations, and provides mutant libraries with variants that have a good chance (>10%) to exhibit enhanced stability. After experimental verification, the most effective mutations are combined to produce highly robust enzymes.

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Acknowledgments

This research was supported by the European Union Seventh framework project Kyrobio (KBBE-2011-5, 289646), by the European Union Horizon 2020 program (project LEIT-BIO-2014-1, 635734) by NWO (Netherlands Organization for Scientific Research) through an ECHO grant, and by the Dutch Ministry of Economic Affairs through BE-Basic (www.be-basic.org).

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Authors and Affiliations

  1. Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 4, 9747, Groningen, The Netherlands

    Hein J. Wijma, Maximilian J. L. J. Fürst & Dick B. Janssen

Authors
  1. Hein J. Wijma
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  2. Maximilian J. L. J. Fürst
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  3. Dick B. Janssen
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Corresponding authors

Correspondence to Hein J. Wijma or Dick B. Janssen .

Editor information

Editors and Affiliations

  1. Department of Biotechnology and Enzyme Catalysis Institute of Biochemistry, Greifswald University, Greifswald, Germany

    Uwe T. Bornscheuer

  2. Protein Biochemistry Institute of Biochemistry, Greifswald University, Greifswald, Germany

    Matthias Höhne

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Wijma, H.J., Fürst, M.J.L.J., Janssen, D.B. (2018). A Computational Library Design Protocol for Rapid Improvement of Protein Stability: FRESCO. In: Bornscheuer, U., Höhne, M. (eds) Protein Engineering. Methods in Molecular Biology, vol 1685. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7366-8_5

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  • DOI: https://doi.org/10.1007/978-1-4939-7366-8_5

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7364-4

  • Online ISBN: 978-1-4939-7366-8

  • eBook Packages: Springer Protocols

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