Immobilised Alcohol Dehydrogenase and Ene‐Reductase Work in Concert to Reduce Cyclohex‐2‐enone in Bulk Organic Solvent

• Published in: ChemCatChem Vol. 16; no. 21
• Main Authors: Pintor, Antía, Volkov, Alexey, Lavandera, Iván, Gotor‐Fernández, Vicente
• Format: Journal Article
• Language: English
• Published: Weinheim Wiley Subscription Services, Inc 11-11-2024
• Subjects: Alcohol dehydrogenase; Alkene; Bioreduction; Dehydrogenases; Ene-reductase; Enzyme activity; Enzyme immobilisation; Enzymes; Isooctane; Moisture content; Nicotinamide; Organic synthesis; Reductases; Solvents; Toluene; Zymomonas mobilis
• ISSN: 1867-3880; 1867-3899
• DOI: 10.1002/cctc.202400416

Enone reduction by ene‐reductase (ERED) has not been reported yet in bulk organic media probably due to expected challenges with the cofactor recycling using a second enzyme and the exchange of the NAD(P)‐cofactor between the two enzymes. Herein, the combination of an ERED from Zymomonas mobilis (NCR‐ERED) and Thermoanaerobacter ethanolicus alcohol dehydrogenase (TeSADH) has been found as an efficient biocatalytic redox system for the reduction of cyclohex‐2‐enone into cyclohexanol in bulk organic solvents. Both enzymes were successfully immobilised on three EziG supports, finding Coral as the most promising carrier for both of them. The highest enzymatic activities were found in hydrophobic solvents such as toluene and isooctane. It was observed that the control of water content and post‐immobilisation treatments enabled retention of enzyme activity. Both enzymes were co‐immobilised on the same bead together with NADP+, leading to the production of a self‐sufficient catalytic system, although the external supplementation of the nicotinamide cofactor was necessary for an efficient enzyme reuse. In this case, no ERED activity loss was detected after two additional cycles, while the ADH experienced less than 20 % activity loss upon reuse.
The immobilisation of ene‐reductase from Zymomonas mobilis (NCR‐ERED) and alcohol dehydrogenase from Thermoanaerobacter ethanolicus (TeSADH) onto EziG supports enabled the development of a self‐sufficient catalytic system for cyclohex‐2‐enone bioreduction in bulk organic solvents. Controlling water content in enzyme formulation together with post‐immobilisation treatments were key parameters, whereas, external addition of the nicotinamide cofactor was necessary for efficient of co‐immobilised enzymes.