Enhanced catalytic efficiency and thermostability of glucose isomerase from Thermoanaerobacter ethanolicus via site-directed mutagenesis
Enhanced catalytic efficiency and thermostability of glucose isomerase from Thermoanaerobacter ethanolicus via site-directed mutagenesis
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DOI：
10.1016/j.enzmictec.2021.109931
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发表时间：
2021-10-09
影响因子：
3.4
通讯作者：
Zheng, Yu-Guo
中科院分区：
文献类型：
--
作者：
Jin, Li-Qun;Jin, Yi-Ting;Zheng, Yu-Guo
葡萄糖异构酶（GI）是高果糖玉米糖浆（HFCS）制备过程中的关键酶。本研究采用定点突变的方法，从嗜热厌氧杆菌CCSD 1菌株中获得了一株葡萄糖异构酶（TEGI-M）突变株TEGI-M-L38 M/V137 L（TEGI-M2）。TEGI-M2在85 ℃和pH 6.5下与二价阳离子Co2+和Mg 2+显示出最佳活性。基于同源模建和分子对接研究了TEGI-M和TEGI-M2的结构差异，以阐明酶性质改善的机制。与原酶相比，TEGI-M2的酶活提高了2.0倍，Km值从234.2 mM降低到85.9 mM。最后，尝试将突变体TEGI-M2应用于HFCS一步合成，D-果糖产量为67.3%，比原酶提高了14.3%。这对一步法制备D-果糖的工业化生产具有重要意义。
Glucose isomerase (GI) is a key enzyme in the preparation of high fructose corn syrup (HFCS). In this study, a mutant TEGI-M-L38 M/V137 L (TEGI-M2) of glucose isomerase (TEGI-M) originated from Thermoanaerobacter ethanalicus CCSD1 was obtained by site-directed mutagenesis. The TEGI-M2 showed an optimal activity at 85 degrees C and pH 6.5 with the divalent cations Co2+ and Mg2+. The structural differences between TEGI-M and TEGI-M2 were investigated based on the homology modeling and molecular docking, to elucidate the mechanism of improvement in the enzymatic properties. Compared with the original enzyme, the TEGI-M2 showed a 2.0-fold increased enzyme activity and a decreased K-m from 234.2 mM to 85.9 mM. Finally, the application of mutant TEGI-M2 in HFCS one-step biosynthesis was attempted, resulting in a D-fructose yield of 67.3 %, which was 14.3 % higher than that of TEGI-M. This improved catalytic performance of TEGI-M2 was of great importance for the industrial preparation of D-fructose in one-step process.