AbstractBacterial inclusion bodies (IBs) have long been considered as inactive, unfolded waste material produced by heterologous overexpression of recombinant genes. In industrial applications, they are occasionally used as an alternative in cases where a protein cannot be expressed in soluble form and in high enough amounts. Then, however, refolding approaches are needed to transform inactive IBs into active soluble protein. While anecdotal reports about IBs themselves showing catalytic functionality/activity (CatIB) are found throughout literature, only recently, the use of protein engineering methods has facilitated the on-demand production of CatIBs. CatIB formation is induced usually by fusing short peptide tags or aggregation-inducing protein domains to a target protein. The resulting proteinaceous particles formed by heterologous expression of the respective genes can be regarded as a biologically produced bionanomaterial or, if enzymes are used as target protein, carrier-free enzyme immobilizates. In the present contribution, we review general concepts important for CatIB production, processing, and application.Key points• Catalytically active inclusion bodies (CatIBs) are promising bionanomaterials.• Potential applications in biocatalysis, synthetic chemistry, and biotechnology.• CatIB formation represents a generic approach for enzyme immobilization.• CatIB formation efficiency depends on construct design and expression conditions.