Xie J, Zhao D, Zhao L, Pei J, Xiao W, Ding G, Wang Z, Xu J
College of Chemical Engineering, Nanjing Forestry University, Nanjing, China.
Jiangsu Key Laboratory of Biomass Based Green Fuels and Chemicals, Nanjing, China.
J Appl Microbiol. 2016 Mar;120(3):647-60. doi: 10.1111/jam.13040. Epub 2016 Feb 9.
(i) To investigate the enzymatic characterization of α-L-arabinofuranosidase from Thermotoga thermarum DSM5069. (ii) To evaluate the performance of its excellent properties on converting ginsenoside Rc to ginsenoside Rd.
The thermostable α-L-arabinofuranosidase (Tt-Afs) gene from T. thermarum DSM5069 was cloned and overexpressed. Recombinant Tt-Afs was purified, and its molecular weight was approx. 55 kDa. Its optimal activity was at pH 5·0 and 95°C. It has high selectivity for cleaving the outer arabinofuranosyl moieties at the C-20 carbon of ginsenoside Rc and its sugar-tolerance makes Tt-Afs a promising candidate for the production of ginsenoside Rd. In a reaction at 85°C and pH 5·0, 25 g l(-1) of ginsenoside Rc was transformed into 21·8 g l(-1) of Rd within 60 min, with a corresponding molar conversion of 99·4% and a high ginsenoside Rd productivity of 21800 mg l(-1) h(-1).
We have successfully cloned and overexpressed the novel α-l-arabinofuranosidase from T. thermarum DSM5069. The high ginsenoside Rd productivity and detailed characterization of recombinant Tt-Afs was provided.
The result shows a high productivity on the bioconversion from high concentration ginsenoside Rc to ginsenoside Rd, which also give rise to a potential commercial enzyme application.
（i）研究嗜热栖热菌DSM5069来源的α-L-阿拉伯呋喃糖苷酶的酶学特性。（ii）评估其将人参皂苷Rc转化为人参皂苷Rd的优异性能。
克隆并过表达了嗜热栖热菌DSM5069的热稳定α-L-阿拉伯呋喃糖苷酶（Tt-Afs）基因。对重组Tt-Afs进行了纯化，其分子量约为55 kDa。其最佳活性pH为5.0，温度为95°C。它对切割人参皂苷Rc C-20位碳原子上的外侧阿拉伯呋喃糖基部分具有高选择性，并且其耐糖性使Tt-Afs成为生产人参皂苷Rd的有前景的候选酶。在85°C和pH 5.0的反应中，25 g l⁻¹的人参皂苷Rc在60分钟内转化为21.8 g l⁻¹的Rd，相应的摩尔转化率为99.4%，人参皂苷Rd的生产率高达21800 mg l⁻¹ h⁻¹。
我们成功克隆并过表达了嗜热栖热菌DSM5069的新型α-L-阿拉伯呋喃糖苷酶。提供了重组Tt-Afs的高人皂苷Rd生产率和详细特性。
结果表明从高浓度人参皂苷Rc生物转化为人参皂苷Rd具有高生产率，这也为潜在的商业酶应用带来了可能。
