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Enzymatic Biotransformation of Ginsenoside Rb2 into Rd by Recombinant α-L-Arabinopyranosidase from Blastococcus saxobsidens.
Journal of Microbiology and Biotechnology ( IF 2.5 ) Pub Date : 2020-1-3 , DOI: 10.4014/jmb.1910.10065
Ju-Hyeon Kim 1, 2 , Jung-Mi Oh 3 , Sungkun Chun 3 , Hye Yoon Park 4 , Wan-Taek Im 1, 2, 5
Affiliation  

In this study, we used a novel α-L-arabinopyranosidase (AbpBs) obtained from ginsenoside-converting Blastococcus saxobsidens that was cloned and expressed in Escherichia coli BL21 (DE3), and then applied it in the biotransformation of ginsenoside Rb2 into Rd. The gene, termed AbpBs, consisting of 2,406 nucleotides (801 amino acid residues), and with a predicted translated protein molecular mass of 86.4 kDa, was cloned into a pGEX4T-1 vector. A BLAST search using the AbpBs amino acid sequence revealed significant homology with a family 2 glycoside hydrolase (GH2). The over-expressed recombinant AbpBs in Escherichia coli BL21 (DE3) catalyzed the hydrolysis of the arabinopyranose moiety attached to the C-20 position of ginsenoside Rb2 under optimal conditions (pH 7.0 and 40°;C). Kinetic parameters for α-Larabinopyranosidase showed apparent Km and Vmax values of 0.078 ± 0.0002 micrometer and 1.4 ± 0.1 μmol/min/mg of protein against p-nitrophenyl-α-L-arabinopyranoside. Using a purified AbpBs (1 μg/ml), 0.1% of ginsenoside Rb2 was completely converted to ginsenoside Rd within 1 h. The recombinant AbpBs could be useful for high-yield, rapid, and low-cost preparation of ginsenoside Rd from Rb2.

中文翻译:

通过来自 Blastococcus saxobsidens 的重组 α-L-Arabinopyranosidase 将人参皂苷 Rb2 酶促生物转化为 Rd。

在这项研究中,我们使用从人参皂苷转化Blastococcus saxobsidens中获得的一种新型 α-L-阿拉伯吡喃糖苷酶 (AbpBs),该酶在大肠杆菌BL21 (DE3)中克隆和表达,然后将其应用于人参皂苷 Rb 2向 Rd 的生物转化。该基因称为AbpBs,由 2,406 个核苷酸(801 个氨基酸残基)组成,预测的翻译蛋白分子量为 86.4 kDa,被克隆到 pGEX4T-1 载体中。使用 AbpBs 氨基酸序列的 BLAST 搜索揭示了与家族 2 糖苷水解酶 (GH2) 的显着同源性。大肠杆菌中过表达的重组 AbpBs在最佳条件(pH 7.0 和 40°;C)下,BL21 (DE3) 催化连接到人参皂苷 Rb2 C-20 位的阿拉伯吡喃糖部分的水解。α-Larabinopyranosidase 的动力学参数显示表观 K m和 V最大值为 0.078 ± 0.0002 微米和 1.4 ± 0.1 μmol/min/mg 蛋白质对抗硝基苯基-α-L-arabinopyranoside。使用纯化的 AbpBs (1 μg/ml),0.1% 的人参皂苷 Rb 2在 1 小时内完全转化为人参皂苷 Rd。重组AbpBs可用于从Rb 2高产、快速、低成本地制备人参皂苷Rd 。
更新日期:2020-08-21
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