School of Biological Engineering, Henan University of Technology, Zhengzhou, 450001, People's Republic of China.
The National and Local Joint Engineering Research Center for Biomanufacturing of Chiral Chemicals, Zhejiang University of Technology, Hangzhou, 310014, People's Republic of China.
Appl Biochem Biotechnol. 2023 Jul;195(7):4399-4413. doi: 10.1007/s12010-023-04349-5. Epub 2023 Jan 25.
Fructose, which is produced by the isomerization of glucose isomerase, is a crucial precursor for the biosynthesis of rare sugars. In this study, thermophilic glucose isomerases (GI) from Caldicellulosiruptor acetigenus (CAGI), Thermoanaerobacter thermocopriae (TTGI), and Thermotoga petrophila (TPGI) were screened from GenBank database by a virtual probe and were successfully expressed in Escherichia coli BL21(DE3). The results of characterization demonstrated that the optimal pH for CAGI and TTGI were 8.0 and were maintained at 80% in a slightly acidic environment. The relative residual activities of CAGI and TTGI were found to be 40.6% and 52.6%, respectively, following an incubation period of 24 h at 90 ℃. Furthermore, CAGI and TTGI exhibited superior catalytic performance that their reaction equilibrium both reached only after an hour at 85 ℃ with 200 g/L glucose, and the highest conversion rates were 54.2% and 54.1%, respectively. This study identifies competitive enzyme candidates for fructose production in the industry with appreciable cost reduction.
果糖是由葡萄糖异构酶异构化产生的，是生物合成稀有糖的关键前体。在这项研究中，通过虚拟探针从 GenBank 数据库中筛选出了来源于嗜热纤维单胞菌（Caldicellulosiruptor acetigenus，CAGI）、热厌氧菌（Thermoanaerobacter thermocopriae，TTGI）和嗜热栖热菌（Thermotoga petrophila，TPGI）的耐热葡萄糖异构酶（GI），并在大肠杆菌 BL21（DE3）中成功表达。表征结果表明，CAGI 和 TTGI 的最适 pH 均为 8.0，在微酸性环境下可保持 80%的相对剩余活性。在 90℃孵育 24 h 后，CAGI 和 TTGI 的相对剩余活性分别为 40.6%和 52.6%。此外，CAGI 和 TTGI 表现出优异的催化性能，在 85℃下反应平衡仅需 1 小时，200 g/L 葡萄糖的最高转化率分别为 54.2%和 54.1%。本研究鉴定了工业中用于果糖生产的具有成本效益的竞争性酶候选物。
