Moutai Jiuqu is a famous aromatic raw material of Maotai flavor liquor in China. It is brewed at high temperature and contains many kinds of bacteria, molds, and yeasts. There are many useful glycoside hydrolases in these microfloras, from which efficient glycoside hydrolases can be screened for biotransformation of natural saponins. In this study, an α-L-arabinofuranosidase gene (CaAraf51, 1524 bp, 507 amino acid, 55.07 kDa, and pI = 4.8) was cloned from Cellulosimicrobium aquatile Lyp51, which was isolated from the Maotai Jiuqu. The CaAraf51 was heterogeneously expressed in E. coli BL21 (DE3) and purified by N-terminal His-tag with the Ni2+-affinity column chromatography. The results show that purified CaAraf51 has a 6.8-fold purification factor and specific activity of 15 U/mg. Under optimal conditions (pH 5.0, temperature 40 °C), kinetic parameters Km of CaAraf51 for pNPαAraf and Rc were 1.1 and 0.57 mM, the Vmax were 25 and 6.25 μmol/min/mg, respectively. 90% of 0.87 mg Rc substrate can be transformed by 9.6 U purified CaAraf51 in 1 mL reaction system under suitable conditions (30 °C, pH 7.5 phosphate buffer, 1 h). In addition, we also tested the effects of metal ions and chemical agents on the activity of CaAraf51. According to systematically studied its function and enzymatic properties, CaAraf51 has excellent value and potential of biotransformation Rc into Rd.

中文翻译：

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人参皂苷水解 α-L-阿拉伯呋喃糖苷酶 CaAraf51 的克隆和表征，来自 Cellulosimicrobium aquatile Lyp51

茅台酒是我国著名的茅台风味白酒的芳香原料。它是在高温下酿造的，含有多种细菌、霉菌和酵母菌。这些微生物群中有许多有用的糖苷水解酶，从中可以筛选出有效的糖苷水解酶用于天然皂苷的生物转化。在本研究中，从茅台酒曲中分离的纤维素水生菌 Lyp51 中克隆了 α-L-阿拉伯呋喃糖苷酶基因（CaAraf51，1524 bp，507 个氨基酸，55.07 kDa，pI = 4.8）。CaAraf51 在大肠杆菌 BL21 (DE3) 中异质表达，并通过 Ni2+ 亲和柱色谱法通过 N 端 His 标签纯化。结果表明，纯化的 CaAraf51 具有 6.8 倍的纯化因子和 15 U/mg 的比活。在最佳条件下（pH 5.0，温度 40 °C），CaAraf51 对 pNPαAraf 和 Rc 的动力学参数 Km 分别为 1.1 和 0.57 mM，Vmax 分别为 25 和 6.25 μmol/min/mg。在合适的条件下（30°C，pH 7.5 磷酸盐缓冲液，1 小时），90% 的 0.87 mg Rc 底物可以被 9.6 U 纯化的 CaAraf51 在 1 mL 反应系统中转化。此外，我们还测试了金属离子和化学试剂对 CaAraf51 活性的影响。根据系统研究其功能和酶学性质，CaAraf51具有将Rc生物转化为Rd的优异价值和潜力。我们还测试了金属离子和化学试剂对 CaAraf51 活性的影响。根据系统研究其功能和酶学性质，CaAraf51具有将Rc生物转化为Rd的优异价值和潜力。我们还测试了金属离子和化学试剂对 CaAraf51 活性的影响。根据系统研究其功能和酶学性质，CaAraf51具有将Rc生物转化为Rd的优异价值和潜力。