Characteristic of immobilized cephalosporin C acylase and its application in one-step enzymatic conversion of cephalosporin C to 7-aminocephalosporanic acid |
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Authors: | Xiangwei Zhu Hui Luo Yanhong Chang Houbo Su Qiang Li Huimin Yu Zhongyao Shen |
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Institution: | (1) Department of Biological Science and Technology, University of Science and Technology Beijing, 100083 Beijing, China;(2) Department of Environmental Engineering, University of Science and Technology Beijing, 100083 Beijing, China;(3) Department of Chemical Engineering, Institute of Biochemical Engineering, Tsinghua University, 100084 Beijing, China; |
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Abstract: | Cephalosporin C (CPC) acylase is an enzyme which hydrolyzes CPC to 7-aminocephalosporanic acid (7-ACA) directly, and therefore
has great potential in industrial application. In this study, the CPC acylase from a recombinant Escherichia coli was purified to high purity by immobilized metal affinity chromatography, and the CPC acylase was covalently attached to
three kinds of epoxy supports, BB-2, ES-V-1 and LX-1000EP. The immobilized CPC acylase with LX-1000EP as the support shows
the highest activity (81 U g−1) suggesting its potential in industrial 7-ACA production. The activity of immobilized enzyme was found to be optimal at pH
between 8.5 and 9.5 and to increase with temperature elevation until 55 °C. Immobilized CPC acylase showed good stability
at pH between 8.0 and 9.5 and at temperature up to 40 °C. To avoid product degradation, the production of 7-ACA utilizing
immobilized enzyme was carried out at 25 °C, pH 8.5 in a designed reactor. Under optimal reaction conditions, a very high
7-ACA yield of 96.7% was obtained within 60 min. In the results of repeated batch production of 7-ACA, 50% activity of the
initial cycle was maintained after being recycled 24 times and the average conversion rate of CPC reached 98%. |
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