Immobilization of Clostridium cellulolyticum D-psicose 3-epimerase on artificial oil bodies.

Abstract

The rare sugar D-psicose possesses several fundamental biological functions. D-Psicose 3-epimerase from Clostridium cellulolyticum (CC-DPEase) has considerable potential for use in D-psicose production. In this study, CC-DPEase was fused to the N terminus of oleosin, a unique structural protein of seed oil bodies and was overexpressed in Escherichia coli as a CC-DPEase-oleosin fusion protein. After reconstitution into artificial oil bodies (AOBs), refolding, purification, and immobilization of the active CC-DPEase were simultaneously accomplished. Immobilization of CC-DPEase on AOB increased the optimal temperature but decreased the optimal pH of the enzyme activity. Furthermore, the AOB-immobilized CC-DPEase had a thermal stability and a bioconversion rate similar to those of the free-form enzyme and retained >50% of its initial activity after five cycles of enzyme use. Thus, AOB-immobilized CC-DPEase has potential application in the production of d-psicose at a lower cost than the free-form enzyme.