Halide assays are important for the study of enzymatic dehalogenation, a topic of great industrial and scientific importance. Here we describe the development of a very sensitive halide assay that can detect less than a picomole of bromide ions, making it very useful for quantifying enzymatic dehalogenation products. Halides are oxidised under mild conditions using the vanadium-dependent chloroperoxidase from Curvularia inaequalis, forming hypohalous acids that are detected using aminophenyl fluorescein. The assay is up to three orders of magnitude more sensitive than currently available alternatives, with detection limits of 20 nM for bromide and 1 μM for chloride and iodide. We demonstrate that the assay can be used to determine specific activities of dehalogenases and validate this by comparison to a well-established GC-MS method. This new assay will facilitate the identification and characterisation of novel dehalogenases and may also be of interest to those studying other halide-producing enzymes.

Biocatalysis and Enzyme Engineering Laboratory Department of Biochemistry School of Life Sciences University of Hyderabad Gachibowli 500046 India

Department of Biotechnology and Enzyme Catalysis Institute of Biochemistry Greifswald University Greifswald 17487 Germany

Department of Biotechnology Delft University of Technology Delft 2629 HZ (The Netherlands

Institute of Applied Synthetic Chemistry TU Wien Vienna 1060 Austria

International Clinical Research Center St Anne's University Hospital Brno Brno 656 91 Czech Republic

Loschmidt Laboratories Department of Experimental Biology and RECETOX Faculty of Science Masaryk University Brno 625 00 Czech Republic

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Engineering Dehalogenase Enzymes Using Variational Autoencoder-Generated Latent Spaces and Microfluidics