纳米花固定化葡萄糖异构酶的制备及性能分析

Preparation and Performance Analysis of Nanoflower-immobilized Glucose Isomerase

摘要:

该研究采用共沉淀法制备了葡萄糖异构酶（Glucose Isomerase，GI）纳米花，对固定化条件进行了优化，同时对纳米花固定化酶的形态特征以及酶学性质进行了探究。结果表明，40 μL酶液中加入9 mL、pH值7.4的PBS缓冲液后与30 μL CuSO4混合，在35 ℃条件下静置反应18 h，制得的纳米花固定化葡萄糖异构酶（Glucose Isomerase @ Nano flowers，GI@NFs）的酶活回收率高达183.06%。SEM表征结果显示GI@NFs有完整的纳米花结构，傅里叶红外光谱显示GI@NFs具有酶和PO43-的特征吸收，X-射线衍射结果进一步证明其载体为Cu3(PO4)2。酶学性质研究发现，GI@NFs的最适反应温度为60 ℃，比自由酶的提高了10 ℃；最适反应pH值为8，比游离酶的最适pH更高；GI@NFs的温度稳定性和pH稳定性均比自由酶的明显提高；固定化酶被循环使用8次，其酶活力仍保持最初活力的60.32%。实验结果表明，纳米花结构提高了葡萄糖异构酶的酶活，表现出较好的循环性能和稳定性，具有一定的应用价值。

Abstract:

In this paper, glucose isomerase (GI) nanoflowers were prepared using the co-precipitation method, and the immobilization conditions were optimized. The morphological characteristics and enzymatic properties of nanoflower-immobilized enzymes were also explored. The results showed that by adding 9 mL of PBS buffer at pH value 7.4 to 40 μL of enzyme solution and mixing with 30 μL CuSO4, and then allowing the reaction to proceed at 35 ℃ for 18 h, Glucose Isomerase @ Nanoflowers (GI@NFs) were obtained, with a recovery rate of enzyme activity at 183.06%. Scanning electron microscopy characterization showed that GI@NFs had a complete nanoflower structure, while Fourier transform infrared spectrometry showed that GI@NFs had characteristic absorption of enzymes and PO43-, and X-ray diffraction results further proved that its carrier was Cu3(PO4)2. In the study of enzymatic properties, the optimum reaction temperature for GI@NFs was 60 ℃, which was 10 ℃ higher than that for the free enzyme; the optimal reaction pH was 8, which was higher than that for the free enzyme. The temperature stability and pH stability of GI@NFs were significantly higher than those for the free enzyme. The immobilized enzymes were reused eight times, and the remaining enzyme activity was 60.32% of the original activity. The results showed that the nanoflower structure improved the activity of glucose isomerase, exhibited better reusability and stability and had certain potential for application.