Multi-enzymatic assemblies offer the opportunity of bringing in proximity several enzymes that are enabled to work together for the catalysis of multi-step reactions. Especially, the development of robust nanobiocatalytic systems comprising of several enzymes has gained considerable attention over the last few years for the catalysis of complex reactions and the production of high added-value products. In the present chapter, we describe the methodology for the development of a bi-enzymatic nanobiocatalyst consisting of the enzymes β-glucosidase from Thermotoga maritima and lipase A from Candida antarctica (CalA) co-immobilized on chitosan-coated magnetic nanoparticles. This nanobiocatalyst can be efficiently applied for the biotransformation of oleuropein to hydroxytyrosol, a reaction of increased biotechnological interest. Several techniques, as well as methodologies that are required for the characterization of the structure and the activity of such systems are also comprehensively described.
Keywords: Biotransformation; Chitosan-coated; Hydroxytyrosol; Lipase; Magnetic nanoparticles; Multi-enzymatic assemblies; Nanobiocatalyst; Oleuropein; β-glucosidase.
© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.