Cloning,Expression and Conversion Application of Glucose Isomerase from Alicyclobacillus sp.A4

doi:10.13560/j.cnki.biotech.bull.1985.2018-0408

Abstract

Abstract: Glucose isomerase is a key enzyme for the commercial preparation of high fructose corn syrup by isomerizing D-glucose to D-fructose in vitro. The glucose isomerase（A4GI）gene from the thermophilic Alicyclobacillus sp. A4 strain was cloned and successfully expressed in Escherichia coli BL21. His-tag protein-purified magnetic beads were used to purify the crude enzyme solution,and the detailed enzymatic properties and conversion rate of the purified recombinant A4GI were determined. The results showed that the optimal temperature and pH of A4GI were 65℃ and pH 7.5,respectively. The enzyme was stable over a broad pH range of 6.0-11.0. After incubation at 37℃ for 1 h,the enzyme retained over 99% of its initial activity at pH 6.0-11.0. Under optimal reaction conditions,the K_m and V_max values of the recombinant enzyme for D-glucose were 99.8 mmol/L and 3.75 µmol/min/mg. In the experiment of glucose conversion at different concentrations,the conversion rate of A4GI tended to increase along with the increased concentration of D-glucose,and the conversion rate reached the peak of 52.7% at the substrate concentration of 3 mol/L. Thus,high-efficiency conversion under high concentration of glucose by A4GI may be achieved,resulting in the reduced cost of post-concentration,and there is great potential for it to be applied in industry.

Key words: glucose isomerase, high fructose corn syrup, conversion rate, Alicyclobacillus sp. A4

DAI Chen-xia, CAO Hui-fang, LUO Hui-ying, YAO Bin, BAI Ying-guo, XU Bo. Cloning,Expression and Conversion Application of Glucose Isomerase from Alicyclobacillus sp.A4[J]. Biotechnology Bulletin, 2018, 34(11): 144-151.

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