Rapid screening and identification of active ingredients in licorice extract interacting with V3 loop region of HIV-1 gp120 using ACE and CE-MS
Rapid screening and identification of active ingredients in licorice extract interacting with V3 loop region of HIV-1 gp120 using ACE and CE-MS
复制标题
使用 ACE 和 CE-MS 快速筛选和鉴定甘草提取物中与 HIV-1 gp120 V3 环区相互作用的活性成分
DOI：
10.1016/j.jpba.2015.02.021
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发表时间：
2015-07-10
影响因子：
3.4
通讯作者：
Ling, Xiaomei
中科院分区：
文献类型：
--
作者：
Li, Zhongjie;Zhao, Yiran;Ling, Xiaomei
在人类免疫缺陷病毒进入宿主细胞的过程中，囊膜蛋白gp120与神经鞘糖脂的结合是非常重要的。这一步骤尤其发生在V3环区。将gp120中V3环的保守核心序列命名为R15K。以R15K为靶点的抗HIV药物可避免HIV-1基因多样性引起的耐药性。首次采用亲和毛细管电泳联用(CE-MS)技术，建立了一种简便、快速、有效的甘草提取物(抗HIV)的筛选方法，避免了复杂的分离纯化过程。用R15K、3‘-唾液酸乳糖(阳性对照)和n-半乳糖(阴性对照)建立和验证ACE方法。经ACE证实R15K与甘草提取物相互作用后，用固相萃取分离相关活性成分，用CE-ESI-MS在线确证其结构。在本研究中，从甘草提取物中提取的两种混合物被发现具有活性。通过电喷雾电离质谱仪和液质联用仪对甘草酸苷和甘草皂苷G2进行了鉴定，结果表明甘草酸苷和甘草皂苷G2的含量分别为74.23%和9.52%。经Scatchard分析，甘草苷/R15K络合物的结合常数最高，为(1.69+/-0.08)×10~7 L/摩尔。细胞水平的生物活性实验进一步证实了甘草酸的抗HIV活性。这一策略可能会为在天然产品中寻找HIV-1抑制剂提供一个高通量的筛选和鉴定平台。(C)2015爱思唯尔B.V.保留所有权利。
The binding of envelope protein gp120 to glycosphingolipids is very important during the human immunodeficiency virus entering into the host cell. This step occurs in the V3 loop region in particularly. The conserved core sequence of V3 loop in gp120 was named R15K. Anti-HIV drug targeting to R15K would avoid the drug-resistance caused by HIV-1 genetic diversity. Here, for the first time, affinity capillary electrophoresis (ACE) and capillary electrophoresis-mass spectrometry (CE-MS) were used for establishing a simple, rapid and effective method of screening the licorice extract for biological activity (anti-HIV), which avoided the complicated isolation and purification process. R15K, 3'-sialyllactose (the positive control), and n-galactose (the negative control) were used for the development and validation of ACE method. After the interaction between licorice extract and R15K was confirmed by ACE, the relative active ingredients were isolated by SPE and their structures were determined by CE-ESI-MS online. In this research, two mixtures from licorice extract were found to be active. Furthermore, glycyrrhizin and licorice saponin G2 were verified as the main ingredients that significantly interacted with RISK via CEMS and LC MS. The results of quantitative assays showed that the active mixture contained glycyrrhizin of 74.23% and licorice saponin G2 of 9.52%. Calculated by Scatchard analysis method, glycyntizin/R15K complex had the highest binding constant (1.69 +/- 0.08) x 107 L/mol among 27 compounds isolated from licorice extract. The anti-HIV activity of glycyrrhizin was further confirmed by bioactive experiment of cellular level. This strategy might provide a high throughput screening and identifying platform for seeking HIV-1 inhibitors in natural products. (C) 2015 Elsevier B.V. All rights reserved.