Immobilization of Sucrose Isomerase from Erwinia sp. with Graphene Oxide and Its Application in Synthesizing Isomaltulose

Abstract

Sucrose isomerase (SIase) is a key enzyme used for the production of isomaltulose from sucrose. In this study, an SIase gene from Erwinia sp. Ejp617 (ErSIase) was heterologously expressed in Escherichia coli BL21(DE3), and the recombinant ErSIase was served as biocatalyst combined with the graphene oxide (GO) as carrier for ErSIase immobilization. The Fourier transform infrared spectroscopy, transmission electron microscope, and confocal laser microscopy analyses showed that ErSIase was successfully immobilized on the surface of GO to form ErSIase-GO. The loading capacity of ErSIase on GO reached up to 460 mg/g with a specific activity of 727.04 U/mg protein when the optimal immobilization time of 12 h and the ErSIase/GO ratio of 7.4:4 (w/w) were applied. A high conversion rate of 95.3% was reached from sucrose to isomaltulose using ErSIase-GO as biocatalyst with 600 g/L sucrose as substrate, after 180 min at 40 °C and pH 6.0. Moreover, stabilities of the immobilized ErSIase-GO in the aspects of thermal, pH, and storage were improved, and its activity after 10 batches still remained around 80% under the optimal conditions. The Km value of ErSIase-GO was 29.32 mM, and the kcat/Km was increased to 27.34 s−1 mM−1 when 0.1% (w/v) detergent NP40 was added. These results indicated that the ErSIase was well immobilized onto GO, and the ErSIase-GO is a promising biocatalyst with high operational stability and catalytic activity for industrial production of isomaltulose.