| Summary: | 碩士 === 逢甲大學 === 化學工程學所 === 100 === This study was aimed to immobilize the cellulose-hydrolyzing enzymes (cellulases) using plant oil bodies. First, the celA gene (encoding endocellulase), the celK gene (encoding exocellulase), the gls gene (encoding ??glucosidase) were individually fused with the ole gene, whose product is the membrane protein of plant seed oil bodies. These fused genes were then overexpressed in Escherichia coli. The fusion proteins in the insoluble form were recovered for assembly of oil bodies, which in turn facilitates the refolding and immobilization of enzymes. The optimal conditions for enzyme immobilization were investigated by the Box-Behnken design while the optimal reaction conditions were located with the aid of the central composite design. The result revealed that immobilized enzymes exhibited higher thermal stability than free enzymes. Immobilized enzymes had a half-life of 15 h when operation at 66 oC and were recycled for 5 times with 50% of the original activity remained. Overall, our results illustrate the feasibility of immobilizing the cellulose-hydrolyzing enzymes on oil bodies. This proposed immobilization method is featured with easy operation, high efficiency, and cost effectiveness. In particular, immobilization of cellulases on oil bodies is a mimic of the naturally occurring cellulosome.
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