Wang Xiaowen, Chen Sibo, Fan Lin, Liu Hao, Chen Si, Ye Xianfeng, Li Zhoukun, Cui Zhongli, Huang Yan
Key Laboratory of Agricultural Environmental Microbiology, Ministry of Agriculture and Rural Affairs, College of Life Sciences, Nanjing Agricultural University, Nanjing, 210095, PR China.
Key Laboratory of Agricultural Environmental Microbiology, Ministry of Agriculture and Rural Affairs, College of Life Sciences, Nanjing Agricultural University, Nanjing, 210095, PR China.
Protein Expr Purif. 2022 Jun;194:106074. doi: 10.1016/j.pep.2022.106074. Epub 2022 Feb 23.
A new glycoside hydrolase family 2 (GH2) β-galactosidase encoding gene galM was cloned from Microvirga sp. strain MC18 and overexpressed in Escherichia coli. The recombinant β-galactosidase GalM showed optimal activity at pH 7.0 and 50 °C, with a stability at pH 6.0-9.0 and 20-40 °C, which are conditions suitable for the diary environment. The K and V values for o-nitrophenyl-β-d-galactopyranoside (oNPG) were 1.30 mmol/L and 15.974 μmol/(min·mg), respectively. GalM showed low product inhibition by galactose with a K of 73.18 mM and high tolerance for glucose that 86.5% activity retained in the presence of 500 mM glucose. It was also stable and active in 20% of methanol, ethanol and isopropanol. In addition, the enzyme activity of GalM was activated significantly over 0-2 mol/L NaCl (1.6-4.3 fold). These favorable properties make GalM a potential candidate for the industrial application.
从微小病毒属菌株MC18中克隆了一个新的糖苷水解酶家族2（GH2）β-半乳糖苷酶编码基因galM，并在大肠杆菌中进行了过表达。重组β-半乳糖苷酶GalM在pH 7.0和50℃时表现出最佳活性，在pH 6.0 - 9.0和20 - 40℃条件下具有稳定性，这些条件适合乳制品环境。对邻硝基苯基-β-D-吡喃半乳糖苷（oNPG）的K和V值分别为1.30 mmol/L和15.974 μmol/(min·mg)。GalM对半乳糖的产物抑制作用较低，K为73.18 mM，对葡萄糖具有高耐受性，在500 mM葡萄糖存在下仍保留86.5%的活性。它在20%的甲醇、乙醇和异丙醇中也稳定且有活性。此外，GalM的酶活性在0 - 2 mol/L NaCl范围内显著激活（1.6 - 4.3倍）。这些优良特性使GalM成为工业应用的潜在候选者。
