Biotransformation of Isoeugenol into Vanillin Using Immobilized Recombinant Cells Containing Isoeugenol Monooxygenase Active Aggregates.

For efficiently enhancing the activity of isoeugenol monooxygenase, a whole cell overproducing active aggregate IEM720-18A was successfully fabricated via the fusion of amphiphilic short peptide 18A (EWLKAFYEKVLEKLKELF) and isoeugenol monooxygenase and then efficiently expressed in E. coli BL21 (DE3). Such resulting bacteria, E. coli BL21 (DE3) harboring pET30a-IEM720-18A was applied in the biotransformation of isoeugenol to vanillin with the optimization of cultivation conditions. Our results revealed that the vanillin concentration reached to the highest level (14.5 mmol/L) under the optimized reaction conditions including 1.5-g cells containing active aggregate of IEM720-18A, 10% (v/v) dimethyl sulfoxide (DMSO), 100 mmol/L isoeugenol, 50 mmol/L glycine-sodium hydroxide buffer (pH 10.5) in 10 mL reaction volume, and 200 rpm at 25 °C for 36 h. Moreover, the active aggregate IEM720-18A was immobilized with 100 mmol/L glutaraldehyde at 4 °C to improve the operational stability. The highest activity could be achieved when the reactions were carried out at 25 °C and the relative activity of the immobilized enzyme maintained over 60% after seven recycles. Our study provides a new approach to the biotransformation of isoeugenol into vanillin.