Mahajan Supreeti, Kumar Ravi, Singh Ankur, Dhaka Preeti, Pareek Akshay, Kumar Pravindra, Tomar Shailly
Department of Biosciences and Bioengineering, Indian Institute of Technology Roorkee, Roorkee, Uttarakhand, India, 247667.
Department of Biosciences and Bioengineering, Indian Institute of Technology Roorkee, Roorkee, Uttarakhand, India, 247667.
Virology. 2025 Jul;608:110551. doi: 10.1016/j.virol.2025.110551. Epub 2025 Apr 22.
The molecular interactions between Chikungunya virus (CHIKV) non-structural protein 3 (nsP3) and the host GTPase Activating SH3 Domain Binding Protein 1 (G3BP1) are critical for CHIKV replication. The C-terminus hypervariable domain (HVD) of nsP3 protein binds to the nuclear transport factor 2 (NTF2)-like domain of G3BP1 through two tandem FGDF motifs, aiding in the disruption of stress granule (SG) formation. Given G3BP1's role in the antiviral response, it presents an attractive target for antiviral drug development. In this study, seven potential small molecules targeting the FGDF motif binding pocket of G3BP1 were identified using a structure-based virtual screening approach. The binding modes of these molecules were further investigated through molecular docking and simulations. Surface Plasmon Resonance (SPR) and Isothermal Titration Calorimetry (ITC) experiments confirmed their binding to purified G3BP1 with micromolar (μM) affinity. The antiviral efficacy of these molecules was assessed using in vitro cell culture-based assays, revealing that L-7, WIN, SB2, NAL, DHD, GSK, and FLU effectively inhibited CHIKV replication with EC values of 1.99, 0.40, 5.38, 1.52, 7.39, 3.66, and 0.61 μM, respectively. Additionally, CHIKV-infected cells treated with these compounds exhibited fewer virus-induced SGs compared to untreated controls without affecting SG formation under oxidative stress conditions. These findings indicate that identified inhibitors successfully block G3BP1-nsP3 interactions and suppress CHIKV replication. This is one of the first reports of small antiviral molecules targeting G3BP1, a host protein essential for stress granule formation in the antiviral cellular response and CHIKV replication.
基孔肯雅病毒（CHIKV）非结构蛋白3（nsP3）与宿主GTP酶激活SH3结构域结合蛋白1（G3BP1）之间的分子相互作用对CHIKV复制至关重要。nsP3蛋白的C末端高变域（HVD）通过两个串联的FGDF基序与G3BP1的核转运因子2（NTF2）样结构域结合，有助于破坏应激颗粒（SG）的形成。鉴于G3BP1在抗病毒反应中的作用，它是抗病毒药物开发的一个有吸引力的靶点。在本研究中，使用基于结构的虚拟筛选方法鉴定了七种靶向G3BP1的FGDF基序结合口袋的潜在小分子。通过分子对接和模拟进一步研究了这些分子的结合模式。表面等离子体共振（SPR）和等温滴定量热法（ITC）实验证实它们以微摩尔（μM）亲和力与纯化的G3BP1结合。使用基于体外细胞培养的试验评估了这些分子的抗病毒效力，结果表明L-7、WIN、SB2、NAL、DHD、GSK和FLU分别以1.99、0.40、5.38、1.52、7.39、3.66和0.61 μM的EC值有效抑制CHIKV复制。此外，与未处理的对照相比，用这些化合物处理的CHIKV感染细胞显示出较少的病毒诱导的SGs，且在氧化应激条件下不影响SG的形成。这些发现表明，鉴定出的抑制剂成功阻断了G3BP1-nsP3相互作用并抑制了CHIKV复制。这是首批针对G3BP1的小型抗病毒分子的报告之一，G3BP1是抗病毒细胞反应中应激颗粒形成和CHIKV复制所必需的宿主蛋白。
