College of Life Science, Hebei Normal University, Shijiazhuang 050016, China.
Antiviral Res. 2011 Apr;90(1):54-63. doi: 10.1016/j.antiviral.2011.02.006. Epub 2011 Feb 23.
gp41 is a major component of the envelope glycoprotein of human immunodeficiency virus type 1 (HIV-1) responsible for fusion of the viral envelope with the target cellular membrane. The formation of the trimer-of-hairpins core structure of gp41, via the interaction between its N-terminal heptad repeat (NHR) and its C-terminal heptad repeat (CHR) plays a key role in the membrane fusion process. Hence, inhibitors of trimer-of-hairpins formation have become a promising new class of HIV therapeutics. In the present study, based on the mammalian two-hybrid system, we developed a cell-based assay for detecting small-molecular HIV-1 fusion inhibitors targeting gp41. The optimized assay can be adapted to high-throughput screening in 96- and 384-well microplates with high signal-to-background ratios and acceptable Z' factors. The known small-molecular gp41 inhibitors, ADS-J1, XTT formazan and tannin acid, tested positive in this assay, with half-maximal inhibitory concentration (IC50) values of 4.9 μM, 5.6 μM and 0.8 μM, respectively. These data suggested that this novel assay is robust, sensitive and specific for identifying small-molecular HIV-1 gp41 inhibitors.
gp41 是人类免疫缺陷病毒 1 型（HIV-1）包膜糖蛋白的主要成分，负责病毒包膜与靶细胞膜的融合。gp41 的三聚体发夹核心结构的形成，通过其 N 端七肽重复（NHR）与其 C 端七肽重复（CHR）之间的相互作用，在膜融合过程中发挥关键作用。因此，三聚体发夹形成的抑制剂已成为一种有前途的新型 HIV 治疗药物。在本研究中，我们基于哺乳动物双杂交系统，开发了一种用于检测针对 gp41 的小分子 HIV-1 融合抑制剂的基于细胞的测定法。该优化的测定法可适应 96 孔和 384 孔微孔板的高通量筛选，具有高信号与背景比和可接受的 Z' 因子。在该测定法中，已知的小分子 gp41 抑制剂 ADS-J1、XTT 甲臜和鞣酸呈阳性，其半最大抑制浓度（IC50）值分别为 4.9 μM、5.6 μM 和 0.8 μM。这些数据表明，该新型测定法对于鉴定小分子 HIV-1 gp41 抑制剂是稳健、灵敏和特异的。
