title: A mammalian two-hybrid system-based assay for small-molecular HIV fusion inhibitors targeting gp41.
abstract: 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.
authors.affiliation: College of Life Science, Hebei Normal University, Shijiazhuang 050016, China.
authors.forename: Xiaoxi
authors.initials: X
authors.lastname: Shui
authors.forename: Xinhua
authors.initials: X
authors.lastname: Lu
authors.forename: Yan
authors.initials: Y
authors.lastname: Gao
authors.forename: Chen
authors.initials: C
authors.lastname: Liu
authors.forename: Fengzhi
authors.initials: F
authors.lastname: Ren
authors.forename: Qin
authors.initials: Q
authors.lastname: Jiang
authors.forename: Hua
authors.initials: H
authors.lastname: Zhang
authors.forename: Baohua
authors.initials: B
authors.lastname: Zhao
authors.forename: Zhihui
authors.initials: Z
authors.lastname: Zheng
mesh_terms: D019380:Anti-HIV Agents / Q000302:isolation & purification / Q000494:pharmacology*; D004353:Drug Evaluation, Preclinical / Q000379:methods; D015700:HIV Envelope Protein gp41; D023581:HIV Fusion Inhibitors / Q000302:isolation & purification / Q000494:pharmacology*; D015497:HIV-1 / Q000187:drug effects*; D057166:High-Throughput Screening Assays / Q000379:methods; D006801:Humans; D008826:Microbial Sensitivity Tests / Q000379:methods; D012680:Sensitivity and Specificity; D020798:Two-Hybrid System Techniques
chemical_list: D019380:Anti-HIV Agents; D015700:HIV Envelope Protein gp41; D023581:HIV Fusion Inhibitors; C514787:gp41 protein, Human immunodeficiency virus 1