All-Enzyme Hydrogels for flow biocatalysis

Continuous flow biocatalysis is an emerging field of industrial biotechnology that uses enzymes immobilized in flow channels to produce value-added chemicals. Carrier-free enzyme immobilization techniques are an important development in the field of efficient and streamlined continuous synthetic processes using microreactors. With a recently established protein gelation strategy that utilizes a pair of genetically encoded reactive partners, SpyTag (ST) and SpyCatcher (SC), which spontaneously form a covalent isopeptide bond under physiological conditions, enzymes can be immobilized in the form of an All-Enzyme Hydrogel (AEH) without further chemical treatment. While this method was initially used only for the immobilization of homotetrameric proteins, the strategy can now be adapted to a wide range of biocatalysts, for example, by using recombinant ST- or SC-tagged variants of a highly active homodimeric phenolic acid decarboxylase, a homodimeric (S)-selective imine reductase or the monomeric ketoreductase Gre2p. These biocatalysts attracted much attention in recent years because of their ability to generate key structural motifs in active pharmaceutical ingredients.
... mehr
The functionality and stability of the novel biocatalytic AEH were demonstrated by continuous production of the corresponding products in a microfluidic flow reactor. Based on the imine reductase or the Gre2p enzyme in combination with the cofactor-regenerating glucose-1-dehydrogenase, a cofactor-regenerating hydrogel was used in a microfluidic reactor setup where it showed excellent conversions for over 40 hours. In contrast, the corresponding enzymes without a coupling system no longer showed stable turnovers after only 2 hours.