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Installing hydrolytic activity into a completely de novo protein framework

Research output: Contribution to journalArticle

Original languageEnglish
Pages (from-to)837-844
Number of pages8
JournalNature Chemistry
Volume8
Issue number9
Early online date4 Jul 2016
DOIs
DateAccepted/In press - 23 May 2016
DateE-pub ahead of print - 4 Jul 2016
DatePublished (current) - 1 Sep 2016

Abstract

Designing enzyme-like catalysts tests our understanding of sequence-to-structure/function relationships in proteins. Here, we install hydrolytic activity predictably into a completely de novo and thermo-stable α-helical barrel, which comprises 7 helices arranged around an accessible channel. We show that the lumen of the barrel accepts 21 mutations to functional polar residues. The resulting variant, which has cysteine-histidine-glutamic acid triads on each helix, hydrolyses p-nitrophenyl acetate with catalytic efficiencies matching the most-efficient redesigned hydrolases based on natural protein scaffolds. This is the first report of a functional catalytic triad engineered into a de novo protein framework. The flexibility of our system also allows the facile incorporation of unnatural side chains to improve activity and probe the catalytic mechanism. Such predictable and robust construction of truly de novo biocatalysts holds promise for applications in chemical and biochemical synthesis.

    Research areas

  • Biocatalysis, Hydrolases, Protein design

    Structured keywords

  • Bristol BioDesign Institute
  • BrisSynBio

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    Rights statement: This is the author accepted manuscript (AAM). The final published version (version of record) is available online via Nature at http://www.nature.com/nchem/journal/vaop/ncurrent/full/nchem.2555.html. Please refer to any applicable terms of use of the publisher.

    Accepted author manuscript, 3 MB, PDF document

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