融合蛋白CR2-GDH固定化及不对称还原制备(S)-4-氯-3-羟基丁酸乙酯

doi:10.16085/j.issn.1000-6613.2015.11.036

化工进展 ›› 2015, Vol. 34 ›› Issue (11): 4047-4053.DOI: 10.16085/j.issn.1000-6613.2015.11.036

融合蛋白CR2-GDH固定化及不对称还原制备(S)-4-氯-3-羟基丁酸乙酯

王爽, 穆晓清, 聂尧, 张荣珍, 徐岩

江南大学生物工程学院工业生物技术教育部重点实验室, 江苏无锡 214122

收稿日期:2015-03-06 修回日期:2015-04-08 出版日期:2015-11-05 发布日期:2015-11-05
通讯作者: 聂尧,教授,主要研究方向为生物催化。E-mailnieybird@hotmail.com。
作者简介:王爽(1989—),女,硕士研究生。

Immobilization of fusion enzyme CR2-GDH and bioreduction of ethyl (S)-4-chloro-3-hydroxybutyrate

WANG Shuang, MU Xiaoqing, NIE Yao, ZHANG Rongzhen, XU Yan

The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, Jiangsu, China

Received:2015-03-06 Revised:2015-04-08 Online:2015-11-05 Published:2015-11-05

摘要/Abstract

摘要： 比较介孔分子筛材料SBA-15、MCM-41、海藻酸钙、改性二氧化硅4种载体固定化融合蛋白CR2-GDH其酶固载量和酶活回收率,选择SBA-15为固定化载体。研究固定化条件对固定化融合酶量的影响以及固定化酶的稳定性,固定化酶在双相体系催化不对称还原反应。结果表明,在pH值为5.5、酶浓度为1.4mg/mL、反应1h条件下,固定化酶量为27.7mg/g。加入25mmol/L的Ca²⁺,固定化酶的酶活回收率由58.6%提高到78.1%。与游离酶相比,固定化酶的热稳定性显著提高,40℃条件下酶活回收率提高19.1%。固定化酶水相中反复使用7批次后,剩余活性仍超过30%,具有较好的操作稳定性。与游离酶相比,固定化酶更耐受烷烃类有机溶剂。在水/有机溶剂双相反应体系中,Ca²⁺/SBA-15固定化酶和游离酶催化相比,产物得率提高23.8%。

关键词: 融合蛋白, SBA-15, 固定化, 双相体系, 不对称还原

Abstract: Mesoporous molecular sieves SBA-15 was the best one among 4 kinds of carriers in terms of both carrier content and recovery of enzyme activity. The remaining carriers were MCM-41, modified silica, and calcium alginate. Immobilization of fusion enzyme CR2-GDH on SBA-15 was realized, and effect of immobilization conditions on immobilization content and activity, stabilization of immobilized enzyme, asymmetric reduction in biphasic system was investigated. The results showed that adsorption capacity was 27.7mg/g carrier under the condition of pH5.5, enzyme concentration 1.4mg/mL, reaction time of one hour. The recovery of immobilized enzyme activity was increased by near 20% adding metal ion Ca²⁺ of 25mmol/L. It was also found that the thermal and operational stabilities of the immobilized enzyme were higher than those of free enzyme. The recovery was increased by 19.1% at the temperature of 40℃. Immobilized enzyme was not more sensitive to organic solvents especially alkanes than free enzyme. In biphasic system, the production yield obtained by Ca²⁺/SBA-15 immobilized enzyme was increased by 23.8%.

Key words: fusion enzyme, SBA-15, immobilization, biphasic system, bioreduction

中图分类号:

Q814.2

王爽, 穆晓清, 聂尧, 张荣珍, 徐岩. 融合蛋白CR2-GDH固定化及不对称还原制备(S)-4-氯-3-羟基丁酸乙酯[J]. 化工进展, 2015, 34(11): 4047-4053.

WANG Shuang, MU Xiaoqing, NIE Yao, ZHANG Rongzhen, XU Yan. Immobilization of fusion enzyme CR2-GDH and bioreduction of ethyl (S)-4-chloro-3-hydroxybutyrate[J]. Chemical Industry and Engineering Progree, 2015, 34(11): 4047-4053.

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融合蛋白CR2-GDH固定化及不对称还原制备(S)-4-氯-3-羟基丁酸乙酯

Immobilization of fusion enzyme CR2-GDH and bioreduction of ethyl (S)-4-chloro-3-hydroxybutyrate

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