Ni2+螯合的琼脂糖凝胶载体用于重组酶SUMO-Hep I-His的固定化

Immobilization of an Sumo-Heparinase I-His Fusion Enzyme on Ni2+ Chelating Agarose Gel Carrier

DOI：10.3969/j.issn.1673-1689.2018.08.007

中文关键词: 肝素酶I GLK-Gel Ni 固定化 酶活

英文关键词: Hep I,GLK-Gel Ni,immobilization,enzyme activity

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中文摘要:

以Ni2+螯合的琼脂糖凝胶（GLK-Gel Ni） 作为固定化载体，对融合肝素酶Sumo-Hep I-His的固定化进行了研究，实现了融合酶的一步纯化和固定化。通过对固定化条件的优化，得到较优的固定化体系：载体与粗酶质量比为30∶1，固定化pH为7.0，吸附时间为6 h，该条件下获得的固定化酶酶活10.07±0.35 IU/mL载体。酶学性质研究结果表明：固定化酶在30 ℃的热稳定性相对于游离酶显著提高，固定化酶半衰期是游离酶的8倍，最适反应温度提高了3 ℃，反应pH的耐受性也有了明显的提高。此外，与游离酶相比，固定化酶的储藏稳定性和可重复利用性良好，在4 ℃下放置60 d能保持76%的初始活性；固定化酶重复使用8次后，酶活仍剩余75.8%；而且GLK-Gel Ni载体本身具有良好的重复利用性，重复固定5次Sumo-Hep I-His后，载体对酶的活性吸附仍能达到88.9%。整体而言，固定化Sumo-Hep I-His显现了较好的工业应用潜能。

英文摘要:

In this study，immobilization of Sumo-Hep I-His was investigated using Ni2+ chelating agarose gel as carrier. Under optimal conditions，i.e.，weight ratio of barrier to rude enzyme 30∶1，pH 7.0 and immobilized time 6 h，Sumo-Hep I-His was successfully immobilized with an apparent enzymatic activity of 10.07±0.35 IU/mL carrier. Compared with free enzyme，immobilized Sumo-Hep I-His showed great improvement in thermo stability （the half-live was 8 times of free Hep I at 30 ℃ as well as pH stability. The storage and operation stability of the immobilized enzyme was also significantly improved，as 76% relative enzymatic activity was remained after storing for 60 days at 4 ℃ and 75.8% relative enzymatic activity was remained after 8 reaction cycles. GLK-Gel Ni gel also showed good recycling stability that 88.9% adsorptive capacity was remained after 5 cycles immobilization. Conclusively，immobilized Hep I showed great potential in industrial application.

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