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Laboratory evolved enzymes provide snapshots of the development of enantioconvergence in enzyme-catalyzed epoxide hydrolysis
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Biochemistry. (Widersten)
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology. (Kamerlin)
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Biochemistry. (Dobritzsch)
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC. (Widersten)
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2016 (English)In: ChemBioChem (Print), ISSN 1439-4227, E-ISSN 1439-7633, Vol. 17, no 18, 1693-1697 p.Article in journal (Refereed) Published
Abstract [en]

Engineered enzyme variants of potato epoxide hydrolase (StEH1) display varying degrees of enrichment of (2R)-3-phenylpropane-1,2-diol from racemic benzyloxirane. Curiously, the observed increase in the enantiomeric excess of the (R)-diol is not only due to changes in enantioselectivity for the preferred epoxide enantiomer, but also to changes in the regioselectivity of the epoxide ring opening of (S)-benzyloxirane. To probe the structural origin of these differences in substrate selectivities and catalytic regiopreferences, we have solved the crystal structures for the in-vitro evolved StEH1 variants. We have additionally used these structures as a starting point for docking the epoxide enantiomers into the respective active sites. Interestingly, despite the simplicity of our docking calculations, the apparent preferred binding modes obtained from the docking appears to rationalize the experimentally determined regioselectivities. These calculations could also identify an active site residue (F33) as a putatively important interaction partner, a role that could explain the high degree of conservation of this residue during evolution. Overall, our combined experimental, structural and computational studies of this system provide snapshots into the evolution of enantioconvergence in StEH1 catalyzed epoxide hydrolysis.

Place, publisher, year, edition, pages
2016. Vol. 17, no 18, 1693-1697 p.
Keyword [en]
enantioselectivity; epoxide hydrolysis; evolutionary snapshots; laboratory evolution; protein engineering
National Category
Biochemistry and Molecular Biology
Research subject
Biochemistry
Identifiers
URN: urn:nbn:se:uu:diva-298675DOI: 10.1002/cbic.201600330ISI: 000384425400004PubMedID: 27383542OAI: oai:DiVA.org:uu-298675DiVA: diva2:946852
Funder
Swedish Research CouncilEU, European Research Council, 306474Swedish National Infrastructure for Computing (SNIC), SNIC2015-16-12EU, FP7, Seventh Framework Programme, 283570
Available from: 2016-07-06 Created: 2016-07-06 Last updated: 2017-11-28Bibliographically approved
In thesis
1. Computational modelling of enzyme selectivity
Open this publication in new window or tab >>Computational modelling of enzyme selectivity
2017 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Enantioselective reactions are one of the ways to produce pure chiral compounds. Understanding the basis of this selectivity makes it possible to guide enzyme design towards more efficient catalysts. One approach to study enzymes involved in chiral chemistry is through the use of computational models that are able to simulate the chemical reaction taking place. The potato epoxide hydrolase is one enzyme that is known to be both highly enantioselective, while still being robust upon mutation of residues to change substrate scope. The enzyme was used to investigate the epoxide hydrolysis mechanism for a number of different substrates, using the EVB approach to the reaction both in solution and in several enzyme variants. In addition to this, work has been performed on new ways of performing simulations of divalent transition metals, as well as development of new simulation software.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2017. 104 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 1530
Keyword
enantiomer, epoxide hydrolase, chiral catalysis, empirical valence bond approach, method development
National Category
Biochemistry and Molecular Biology
Research subject
Biochemistry
Identifiers
urn:nbn:se:uu:diva-326108 (URN)978-91-513-0005-4 (ISBN)
Public defence
2017-09-13, A1:111 BMC, Husargatan 3, Uppsala, 09:00 (English)
Opponent
Supervisors
Available from: 2017-08-21 Created: 2017-07-02 Last updated: 2017-09-15

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