Drug Discovery and Development Division, Patanjali Research Institute, Haridwar, 249405, Uttarakhand, India.
Department of Allied and Applied Sciences, University of Patanjali, Haridwar, 249405, Uttarakhand, India.
Drug Des Devel Ther. 2021 Mar 11;15:1111-1133. doi: 10.2147/DDDT.S292805. eCollection 2021.
SARS-CoV-2 engages human ACE2 through its spike (S) protein receptor binding domain (RBD) to enter the host cell. Recent computational studies have reported that withanone and withaferin A, phytochemicals found in , target viral main protease (M) and host transmembrane TMPRSS2, and glucose related protein 78 (GRP78), respectively, implicating their potential as viral entry inhibitors. Absence of specific treatment against SARS-CoV-2 infection has encouraged exploration of phytochemicals as potential antivirals.
This study aimed at in silico exploration, along with in vitro and in vivo validation of antiviral efficacy of the phytochemical withanone.
Through molecular docking, molecular dynamic (MD) simulation and electrostatic energy calculation the plausible biochemical interactions between withanone and the ACE2-RBD complex were investigated. These in silico observations were biochemically validated by ELISA-based assays. Withanone-enriched extract from was tested for its ability to ameliorate clinically relevant pathological features, modelled in humanized zebrafish through SARS-CoV-2 recombinant spike (S) protein induction.
Withanone bound efficiently at the interacting interface of the ACE2-RBD complex and destabilized it energetically. The electrostatic component of binding free energies of the complex was significantly decreased. The two intrachain salt bridge interactions (K31-E35) and the interchain long-range ion-pair (K31-E484), at the ACE2-RBD interface were completely abolished by withanone, in the 50 ns simulation. In vitro binding assay experimentally validated that withanone efficiently inhibited (IC=0.33 ng/mL) the interaction between ACE2 and RBD, in a dose-dependent manner. A withanone-enriched extract, without any co-extracted withaferin A, was prepared from leaves. This enriched extract was found to be efficient in ameliorating human-like pathological responses induced in humanized zebrafish by SARS-CoV-2 recombinant spike (S) protein.
In conclusion, this study provided experimental validation for computational insight into the potential of withanone as a potent inhibitor of SARS-CoV-2 coronavirus entry into the host cells.
SARS-CoV-2 通过其刺突（S）蛋白受体结合域（RBD）与人类 ACE2 结合，进入宿主细胞。最近的计算研究表明，植物化学物质 withanone 和 withaferin A 分别靶向病毒主要蛋白酶（M）和宿主跨膜 TMPRSS2 以及葡萄糖相关蛋白 78（GRP78），这表明它们具有作为病毒进入抑制剂的潜力。由于缺乏针对 SARS-CoV-2 感染的特异性治疗方法，人们鼓励探索植物化学物质作为潜在的抗病毒药物。
本研究旨在通过计算机模拟探索，以及植物化学物质 withanone 的体外和体内抗病毒功效验证。
通过分子对接、分子动力学（MD）模拟和静电能计算，研究了 withanone 与 ACE2-RBD 复合物之间可能的生化相互作用。这些计算机模拟观察结果通过基于 ELISA 的测定进行了生化验证。从 中提取的富含 withanone 的提取物用于测试其改善通过 SARS-CoV-2 重组刺突（S）蛋白诱导的人源化斑马鱼中临床相关病理特征的能力。
Withanone 有效地结合在 ACE2-RBD 复合物的相互作用界面上，并在能量上使其不稳定。复合物结合自由能的静电分量显着降低。ACE2-RBD 界面上的两个链内盐桥相互作用（K31-E35）和链间远程离子对（K31-E484）完全被 withanone 消除，在 50 ns 模拟中。体外结合实验证实，withanone 以剂量依赖的方式有效地抑制了 ACE2 和 RBD 之间的相互作用（IC=0.33ng/mL）。从 叶中提取了一种富含 withanone 的提取物，没有任何共提取的 withaferin A。发现这种富含提取物能够有效改善 SARS-CoV-2 重组刺突（S）蛋白诱导的人源化斑马鱼中引起的类似人类的病理反应。
总之，本研究为计算机模拟研究提供了实验验证，表明 withanone 作为 SARS-CoV-2 冠状病毒进入宿主细胞的有效抑制剂具有潜力。
