Volume 252, 2024

High-throughput selection of (new) enzymes: phage display-mediated isolation of alkyl halide hydrolases from a library of active-site mutated epoxide hydrolases

Abstract

Epoxide hydrolase StEH1, from potato, is similar in overall structural fold and catalytic mechanism to haloalkane dehalogenase DhlA from Xanthobacter autotrophicus. StEH1 displays low (promiscuous) hydrolytic activity with (2-chloro)- and (2-bromo)ethanebenzene producing 2-phenylethanol. To investigate possibilities to amplify these very low dehalogenase activities, StEH1 was subjected to targeted randomized mutagenesis at five active-site amino acid residues and the resulting protein library was challenged for reactivity towards a bait chloride substrate. Enzymes catalyzing the first half-reaction of a hydrolytic cycle were isolated following monovalent phage display of the mutated proteins. Several StEH1 derived enzymes were identified with enhanced dehalogenase activities.

Graphical abstract: High-throughput selection of (new) enzymes: phage display-mediated isolation of alkyl halide hydrolases from a library of active-site mutated epoxide hydrolases

  • This article is part of the themed collection: Biocatalysis

Associated articles

Supplementary files

Article information

Article type
Paper
Submitted
02 jan 2024
Accepted
17 jan 2024
First published
03 jun 2024
This article is Open Access
Creative Commons BY-NC license

Faraday Discuss., 2024,252, 115-126

High-throughput selection of (new) enzymes: phage display-mediated isolation of alkyl halide hydrolases from a library of active-site mutated epoxide hydrolases

M. Blazic, C. Gautier, T. Norberg and M. Widersten, Faraday Discuss., 2024, 252, 115 DOI: 10.1039/D4FD00001C

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