Statistical optimization of one-step immobilization process for recombinant endoglucanase from Clostridium thermocellum |
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| Authors: | Chung-Jen Chiang Po Ting Chen Chien Yu Yeh Yun-Peng Chao |
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| Affiliation: | 1. Department of Medical Laboratory Science and Biotechnology, China Medical University, No. 91, Hsueh-Shih Road, Taichung 40402, Taiwan;2. Department of Biotechnology, Southern Taiwan University of Science and Technology, No. 1, Nantai Street, Tainan 71005, Taiwan;3. Department of Chemical Engineering, Feng Chia University, 100 Wen-Hwa Road, Taichung 40724, Taiwan;4. Department of Health and Nutrition Biotechnology, Asia University, Taichung 41354, Taiwan;5. Department of Medical Research, China Medical University Hospital, Taichung 40447, Taiwan |
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| Abstract: | ![]()
Endoglucanase CelA from Clostridium thermocellum (CtCelA) is a thermophilic endo-β-1,4-glucanase and has a low solubility when expressed in Escherichia coli. To make industrial application of CtCeA more appealing, artificial oil bodies (AOBs) was implemented for one-step renaturation and immobilization of recombinant CtCelA. CtCelA was first fused with oleosin (Ole-CtCelA), a structural protein of plant seed oils. Ole-CtCelA was overexpressed in E. coli, and its insoluble form was recovered and mixed with plant oils to assemble AOBs. Moreover, the Box–Behnken design and the central composite design were employed to optimize the condition for assembly of AOBs and the enzymatic reaction condition, respectively. Consequently, the approach led to the resumption of active CtCelA on AOBs. CtCelA-bound AOBs exhibited an optimum activity at 69 °C and pH 6.3 while the immobilized protein remained stable for several hours at 70 °C and after 5 repeated uses. Overall, it indicates a promise of this novel approach for direct processing and immobilization of recombinant CtCelA. |
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| Keywords: | Endoglucanase Artificial oil body Enzyme immobilization Response surface methodology |
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