Aromatic Halogenation by Using Bifunctional Flavin Reductase-Halogenase Fusion Enzymes
Mary C Andorfer1, Ketaki D Belsare1, Anna M Girlich1
1Department of Chemistry, University of Chicago, 5735 South Ellis Avenue, SCL 302, Chicago, IL, 60637, USA.
Chembiochem : a European Journal of Chemical Biology
|September 8, 2017
Summary
Engineered flavin-dependent halogenase-flavin reductase (FDH-FR) fusion proteins enhance biocatalysis. These fused proteins improve product yields in vivo compared to separate enzymes, advancing synthetic biology applications.
Area of Science:
- Biocatalysis and Synthetic Biology
- Enzyme Engineering
- Biotechnology
Background:
- Flavin-dependent halogenases (FDHs) are valuable biocatalysts due to their selectivity and broad substrate scope.
- FDHs can be engineered for altered substrate specificity and site selectivity.
- FDHs are utilized in vivo for synthesizing chlorinated aromatics, but require a reduced flavin cofactor typically from a flavin reductase (FR).
Purpose of the Study:
- To develop and characterize functional, fused flavin-dependent halogenase-flavin reductase (FDH-FR) proteins.
- To evaluate the performance of these fusion proteins in in vivo biocatalysis.
- To improve product titers in engineered organisms through enzyme fusion.
Main Methods:
- Construction of synthetic FDH-FR fusion proteins with various FDH and FR components and linkers.
- Expression and characterization of fusion proteins in Escherichia coli.
- Comparison of product titers achieved using fusion proteins versus individual FDH and FR components.
Main Results:
- Demonstrated functional activity of the synthesized FDH-FR fusion proteins.
- Showed that FDH-FR fusion proteins can increase product titers compared to separate FDH and FR enzymes.
- Validated the utility of fused FDH-FR proteins for in vivo biocatalysis.
Conclusions:
- FDH-FR fusion proteins represent an effective strategy to enhance biocatalytic efficiency.
- Enzyme fusion simplifies cofactor regeneration and improves overall pathway performance.
- This approach offers a promising tool for the development of novel biocatalytic systems and the synthesis of valuable chlorinated compounds.
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