Lin Yingjia, Li Yang, Song Zhi-Guang, Zhu Hongyan, Jin Ying-Hua
Key Laboratory for Molecular Enzymology and Engineering of the Ministry of Education, College of Life Science, Jilin University, Changchun, Jilin, China.
College of Chemistry, Jilin University, Changchun, China.
J Ginseng Res. 2017 Jul;41(3):330-338. doi: 10.1016/j.jgr.2016.06.005. Epub 2016 Jul 1.
Ginsenoside Rh2 (G-Rh2) is a ginseng saponin that is widely investigated because of its remarkable antitumor activity. However, the molecular mechanism by which (20S) G-Rh2 triggers its functions and how target animals avoid its cytotoxic action remains largely unknown.
Phage display was used to screen the human targets of (20S) G-Rh2. Fluorescence spectroscopy and UV-visible absorption spectroscopy were used to confirm the interaction of candidate target proteins and (20S) G-Rh2. Molecular docking was utilized to calculate the estimated free energy of binding and to structurally visualize their interactions. MTT assay and immunoblotting were used to assess whether human serum albumin (HSA), bovine serum albumin (BSA), and bovine serum can reduce the cytotoxic activity of (20S) G-Rh2 in HepG2 cells.
In phage display, (20S) G-Rh2-beads and (20R) G-Rh2-beads were combined with numerous kinds of phages, and a total of 111 different human complementary DNAs (cDNA) were identified, including HSA which had the highest rate. The binding constant and number of binding site in the interaction between (20S)-Rh2 and HSA were 3.5 × 10 M and 1, and those in the interaction between (20S) G-Rh2 and BSA were 1.4 × 10 M and 1. The quenching mechanism is static quenching. HSA, BSA and bovine serum significantly reduced the proapoptotic effect of (20S) G-Rh2.
HSA and BSA interact with (20S) G-Rh2. Serum inhibited the activity of (20S) G-Rh2 mainly due to the interaction between (20S) G-Rh2 and serum albumin (SA). This study proposes that HSA may enhance (20S) G-Rh2 water solubility, and thus might be used as nanoparticles in the (20S) G-Rh2 delivery process.
人参皂苷Rh2（G-Rh2）是一种人参皂苷，因其显著的抗肿瘤活性而受到广泛研究。然而，（20S）G-Rh2发挥其功能的分子机制以及靶动物如何避免其细胞毒性作用在很大程度上仍不清楚。
采用噬菌体展示技术筛选（20S）G-Rh2的人类靶点。利用荧光光谱和紫外可见吸收光谱来确认候选靶蛋白与（20S）G-Rh2之间的相互作用。运用分子对接来计算结合的估计自由能并从结构上可视化它们的相互作用。采用MTT法和免疫印迹法评估人血清白蛋白（HSA）、牛血清白蛋白（BSA）和牛血清是否能降低（20S）G-Rh2对肝癌细胞系HepG2细胞的细胞毒性活性。
在噬菌体展示中，（20S）G-Rh2磁珠和（20R）G-Rh2磁珠与多种噬菌体结合，共鉴定出111种不同的人类互补DNA（cDNA），其中HSA的出现率最高。（20S）-Rh2与HSA相互作用的结合常数和结合位点数分别为3.5×10 M和1，（20S）G-Rh2与BSA相互作用的结合常数和结合位点数分别为1.4×10 M和1。猝灭机制为静态猝灭。HSA、BSA和牛血清显著降低了（20S）G-Rh2的促凋亡作用。
HSA和BSA与（20S）G-Rh2相互作用。血清抑制（20S）G-Rh2的活性主要是由于（20S）G-Rh2与血清白蛋白（SA）之间的相互作用。本研究提出HSA可能增强（20S）G-Rh2的水溶性，因此可能在（20S）G-Rh2递送过程中用作纳米颗粒。
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