Improving the stability and catalyst lifetime of the halogenase RebH by directed evolution.
Publication Type:Journal Article
Source:Chembiochem, Volume 15, Issue 9, p.1286-9 (2014)
Keywords:Biocatalysis, Directed Molecular Evolution, Enzyme Stability, Models, Molecular, Molecular Structure, Oxidoreductases, Protein Engineering
<p>We previously reported that the halogenase RebH catalyzes selective halogenation of several heterocycles and carbocycles, but product yields were limited by enzyme instability. Here, we use directed evolution to engineer an RebH variant, 3-LR, with a Topt over 5 °C higher than that of wild-type, and 3-LSR, with a Tm 18 °C higher than that of wild-type. These enzymes provided significantly improved conversion (up to fourfold) for halogenation of tryptophan and several non-natural substrates. This initial evolution of RebH not only provides improved enzymes for immediate synthetic applications, but also establishes a robust protocol for further halogenase evolution.</p>