Fusion of a Coiled-Coil Domain Facilitates the High-Level Production of Catalytically Active Enzyme Inclusion Bodies

The increasing number of biocatalytic reactions implemented in chemical synthesis routes raises the urgent need for large amounts of enzymes. Hence, new generic methods are required for their simple and cost-efficient production. Here, we describe a generally applicable method based on the production of catalytically active inclusion bodies (CatIBs). CatIBs represent a promising new form of biologically produced, carrier-free, biodegradable enzyme immobilizate. CatIBs are produced in Escherichia coli by expression of a gene fusion consisting of a coiled-coil domain and a target enzyme. Employing this strategy, the lipase A of Bacillus subtilis (BsLA), the hydroxynitrile lyase of Arabidopsis thaliana (AtHNL), and the 2-succinyl-5-enolpyruvyl-6-hydroxy-3-cyclohexene-1-carboxylate synthase MenD of E. coli (EcMenD) were successfully produced as CatIBs and used in aqueous and micro-aqueous organic solvent based reaction systems, showing excellent stability and recyclability.