Background: Rare ginsenosides (F2, Rg3, Gyp-XVII, and C-K) are pharmaceutically active components of Panax ginseng, which are derived from the conversion of major ginsenosides through various transformation methods. To date, most studies have failed to identify a competent bacterial strain and recombinant enzyme for converting protopanaxadiol (PPD)- and protopanaxatriol (PPT)-type ginsenosides to target minor ginsenosides. Method: Our study identified and employed nine sets of clones from different glycoside hydrolase bacterial strains for major ginsenoside bioconversion. Among these nine clones, only BglFc was selected based on its strong biotransformation ability and capacity to generate complete minor ginsenosides. bglFc was cloned and expressed in Escherichia coli using the pGEX-4T-1 vector system, and the recombinant enzyme was used for efficiently producing minor ginsenosides. Results: Recombinant BglFc is 2,394 bp and 798 amino acid residues long, with a predicted molecular mass of 78.8 kDa. BglFc belongs to the glycoside hydrolase family 3, and demonstrates a promising ability to convert major ginsenosides into minor ones. The K_m and V_max values of pNPG were 0.81 ± 0.06 and 4.0 ± 0.2 mM·min⁻¹·mg⁻¹ of protein, respectively. Under optimal conditions (37°C, pH 7.0), the ginsenoside transformation pathways for BglFc were as follows: Rb1→Rd→Rg3(S)→Rh2(S); GypXVII→GypLXXV→C-K; GypLXXV→C-K; F2→C-K; Rb2→C-O→C-Y; Rb3→C-Mx1→C-Mx; Rc→C-Mcl→C-Mc; Re→Rg2(S); and Rgl→Rhl(S), respectively. Conclusion: These results suggest that recombinant BglFc demonstrates a strong transformation activity for both PPD- and PPT-type major ginsenosides. Therefore, we conclude that BglFc can be used for gram unit production of various minor ginsenosides. Significance and Impact of Study: Previously, researchers have used a combination of enzymes for the production of minor ginsenosides. However, in this study, we found a favorable enzyme that can be used alone for the production of a different type of minor ginsenoside using the proposed conversion pathway.

背景：稀有的人参皂苷（F2，Rg3，Gyp-XVII和CK）是人参的药理活性成分，其是通过各种转化方法将主要的人参皂苷转化而来的。迄今为止，大多数研究都未能鉴定出能将原人参二醇（PPD）和原人参三醇（PPT）型人参皂甙转化为次要人参皂甙的有效细菌菌株和重组酶。方法：我们的研究鉴定并采用了来自不同糖苷水解酶细菌菌株的九套克隆，用于主要人参皂苷的生物转化。在这九个克隆中，仅基于BglFc的强大生物转化能力和生成完整次要人参皂苷的能力而选择了BglFc。克隆了bglFc并在大肠杆菌中表达使用pGEX-4T-1载体系统，并将重组酶用于有效生产次要人参皂苷。结果：重组BglFc为2394 bp，长798个氨基酸残基，预测分子量为78.8 kDa。BglFc属于糖苷水解酶家族3，并显示出将主要人参皂甙转化为次要人参皂甙的有前途的能力。p NPG的K _m和V _max值分别为0.81±0.06和4.0±0.2 mM min ^-1 mg ^-1蛋白质。在最佳条件下（37°C，pH 7.0），BglFc的人参皂甙转化途径如下：Rb1→Rd→Rg3（S）→Rh2（S）; GypXVII→GypLXXV→CK; GypLXXV→CK; F2→CK; Rb2→CO→CY; Rb3→C-Mx1→C-Mx; Rc→C-Mcl→C-Mc; Re→Rg2（S）; 和Rgl→Rhl（S）。结论：这些结果表明重组BglFc对PPD型和PPT型主要人参皂苷均具有很强的转化活性。因此，我们得出结论，BglFc可用于各种次要人参皂苷的克单位生产。研究的意义和影响：以前，研究人员使用多种酶来生产次要人参皂苷。但是，在这项研究中，我们发现了一种有利的酶，可以使用拟议的转化途径将其单独用于生产其他类型的次要人参皂苷。