Cross-Linked Enzyme Aggregates as Industrial Biocatalysts

The immobilization of enzymes as cross-linked enzyme aggregates (CLEAs) and their applications in industrially relevant biotransformations are reviewed. The preparation of CLEAs involves precipitation from aqueous buffer followed by cross-linking with a bifunctional reagent, usually a dialdehyde such as glutaraldehyde or dextran polyaldehyde. The technique is exquisitely simple and broadly applicable. CLEAs have several benefits in the context of industrial biocatalysis. They have a significantly enhanced shelf life and operational stability, are easy to recover and reuse, and are completely stable towards leaching in aqueous media. They circumvent the use of, often expensive, carriers and have high volumetric and catalyst productivities as a result of the absence of noncatalytic ballast. The applications of CLEAs for a wide variety of enzymes, including various types of hydrolases, oxidoreductases, and lyases in industrially relevant biotransformations, are discussed. Co-aggregation and cross-linking of two or more enzymes affords combi-CLEAs that are ideally suited for catalyzing one-pot, enzymatic cascade processes, for example, the successful application of a triple-decker combi-CLEA comprising a hydroxynitrile lyase, a nitrilase, and an amidase in the one-pot conversion of benzaldehyde to S-mandelic acid in >99% ee at 96% conversion. Finally, reactor concepts are discussed in the context of industrial applications.