A Simple Method for Beta-glucosidase Immobilization and Its Application in Soybean Isoflavone Glycosides Hydrolysis
A Simple Method for Beta-glucosidase Immobilization and Its Application in Soybean Isoflavone Glycosides Hydrolysis
复制标题
β-葡萄糖苷酶的简易固定化方法及其在大豆异黄酮苷水解中的应用
DOI：
10.1007/s12257-017-0434-3
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发表时间：
2018-02-01
影响因子：
3.2
通讯作者：
Hong, Jiong
中科院分区：
文献类型：
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作者：
Hu, Shenglin;Wang, Dongmei;Hong, Jiong
本研究构建了一种简单、廉价、快速的β-葡萄糖苷酶固定化体系，并对该固定化体系进行了初步的研究。利用毕赤酵母KM 71 H重组表达了橙色嗜热菌IFO 9748的β-葡萄糖苷酶基因，并通过融合纤维素结合模块3将其固定在再生无定形纤维素（RAC）上。将纤维素酶和粗酶液在室温下混合15 min，可将96.04%的β-葡萄糖苷酶固定化在RAC上。固定化后5AC使β-葡萄糖苷酶的最适酶活温度提高。固定化酶在60 ℃热失活的半衰期（tA 1/2）提高了8倍以上。在40 ℃循环30次后，大豆苷和染料木苷的水解率分别为96.9%和98.9%。在0.2mL/min的流量下，水解30 h后，染料木苷和大豆苷的水解率仍分别为95.6%和90.2%。该固定化酶系统简单、高效，纤维素价格低廉，具有广阔的工业应用前景。
In this study, a simple, inexpensive and fast beta-glucosidase immobilization system was constructed and evaluated in isoflavone glycosides hydrolysis. A beta-glucosidase gene from Thermoascus aurantiacus IFO9748 was recombinantly expressed in Pichia pastoris KM71H and immobilized on regenerated amorphous cellulose (RAC) by fused cellulose binding module 3. Through simple mixing cellulose and crude enzyme for 15 min under room temperature, 96.04% beta-glucosidase was immobilized onto RAC. The optimum temperature for beta-glucosidase activity was increased by 5AC after immobilization. The half-life (tA 1/2) of heat inactivation of immobilized enzyme at 60oC was improved over 8 folds. After 30 rounds recycled at 40oC, 96.9% daidzin and 98.9% genistin could still be hydrolyzed. A continuous hydrolysis system was also constructed, and at the flow rate of 0.2 mL/min after 30 h hydrolysis, 95.6% genistin and 90.2% daidzin can still be hydrolyzed. Combined the simple and high efficient enzyme immobilization procedure and inexpensive cellulose, this scalable and practical system may have broad prospects for industrial utilization.