Department of Bioscience and Biotechnology, Konkuk University, Seoul 143-701, Republic of Korea.
Appl Microbiol Biotechnol. 2013 Jul;97(14):6315-24. doi: 10.1007/s00253-013-4828-7. Epub 2013 Mar 17.
A novel β-glucosidase from Penicillium aculeatum was purified as a single 110.5-kDa band on SDS-PAGE with a specific activity of 75.4 U mg⁻¹ by salt precipitation and Hi-Trap Q HP and Resource Q ion exchange chromatographies. The purified enzyme was identified as a member of the glycoside hydrolase 3 family based on its amino acid sequence. The hydrolysis activity for p-nitrophenyl-β-D-glucopyranoside was optimal at pH 4.5 and 70 °C with a half-life of 55 h. The enzyme hydrolyzed exo-, 3-O-, and 6-O-β-glucosides but not 20-O-β-glucoside and other glycosides of ginsenosides. Because of the novel specificity, this enzyme had the transformation pathways for ginsenosides: Rb₁ → Rd → F₂ → compound K, Rb₂ → compound O → compound Y, Rc → compound Mc₁ → compound Mc, Rg₃ → Rh₂ → aglycone protopanaxadiol (APPD), Rg₁ → F₁, and Rf → Rh₁ → aglycone protopanaxatriol (APPT). Under the optimum conditions, the enzyme converted 0.5 mM Rb₂, Rc, Rd, Rg₃, Rg₁, and Rf to 0.49 mM compound Y, 0.49 mM compound Mc, 0.47 mM compound K, 0.23 mM APPD, 0.49 mM F₁, and 0.44 mM APPT after 6 h, respectively.
从细青霉中纯化得到一种新型β-葡萄糖苷酶，在 SDS-PAGE 上显示为单一的 110.5 kDa 条带，比活为 75.4 U·mg⁻¹，经盐沉淀和 Hi-Trap Q HP 和 Resource Q 离子交换层析纯化。根据其氨基酸序列，该酶被鉴定为糖苷水解酶 3 家族的成员。对 p-硝基苯-β-D-吡喃葡萄糖苷的水解活性在 pH 4.5 和 70°C 时最佳，半衰期为 55 小时。该酶水解外切、3-O-和 6-O-β-葡萄糖苷，但不水解 20-O-β-葡萄糖苷和其他人参皂苷糖苷。由于其新颖的特异性，该酶具有人参皂苷的转化途径：Rb₁→Rd→F₂→化合物 K，Rb₂→化合物 O→化合物 Y，Rc→化合物 Mc₁→化合物 Mc，Rg₃→Rh₂→原型原人参二醇（APPD），Rg₁→F₁，和 Rf→Rh₁→原型原人参三醇（APPT）。在最佳条件下，该酶将 0.5 mM 的 Rb₂、Rc、Rd、Rg₃、Rg₁ 和 Rf 在 6 小时内分别转化为 0.49 mM 的化合物 Y、0.49 mM 的化合物 Mc、0.47 mM 的化合物 K、0.23 mM 的 APPD、0.49 mM 的 F₁ 和 0.44 mM 的 APPT。
