Achappa Sharanappa, Aldabaan Nayef Abdulaziz, Alasmary Mohammed, Shaikh Ibrahim Ahmed, Mahnashi Mater H, Desai Shivalingsarj V, Muddapur Uday M, Khan Aejaz Abdullatif, Mannasaheb Basheerahmed Abdulaziz
Department of Biotechnology, KLE Technological University, Hubballi, Karnataka, India.
Department of Pharmacology, College of Pharmacy, Najran University, Najran, Saudi Arabia.
PLoS One. 2025 Jul 10;20(7):e0325613. doi: 10.1371/journal.pone.0325613. eCollection 2025.
Kyasanur Forest Disease Virus (KFDV) poses a significant public health threat due to the limited efficacy of existing vaccines, necessitating the development of effective antiviral therapeutics. The nonstructural protein 5 (NS5), essential for viral RNA synthesis and methylation, serves as a promising drug target. This study employs computational approaches to identify and evaluate potential NS5 inhibitors that may contribute to the development of antiviral compounds against KFDV. The 3D structure of NS5 was predicted using Robetta, SwissModel, and I-TASSER, with the Robetta model (ERRAT score: 96.40) selected for energy minimization. The globally minimized structure, obtained at 49.58 ns, had a potential energy of -416966.82 kcal/mol and was used for further studies. Active site residues were identified using template-based and structure-based methods (COACH-D, CASTp, PrankWeb) and were located within polymerase motif (A-G) of NS5 protein (residues 273-903 aa), which are essential for polymerase function, RNA synthesis, and viral replication. A total of 1523 compounds were identified using de novo, template-based design, pharmacophore modeling, and ligand screening. Virtual screening with PyRx 0.8 yielded 34 promising compounds, of which 11 were selected based on molecular docking (AutoDock 4.0) with binding energies of -8.86 kcal/mol (FDA-approved dasabuvir -L1), -8.28 kcal/mol (CNPO331352.1-L2), -7.94 kcal/mol (ZINC00103114410- L3), and -7.61 kcal/mol (CNPO202263.1-L4). MD simulations in triplicates under physiological conditions confirmed stability. with MM-GBSA binding free energy values of -52.28 ± 2.91 kcal/mol (NS5-Dasabuvur L1complex), -46.82 ± 4.31 kcal/mol (NS5-L2 complex), -50.72 ± 6.36 kcal/mol (NS5-L3 complex), and -57.03 ± 4.31 kcal/mol (NS5-L4 complex). The computational analysis suggests that compounds L2 and L4 have strong binding affinities comparable to dasabuvir (L1), indicating their potential as inhibitors of the KFDV NS5 protein. Further validation through in vitro assays would complement these in silico findings. These results provide a foundation for future drug development against KFDV, emphasizing the need for continued exploration of antiviral therapeutics.
由于现有疫苗的疗效有限，卡萨努尔森林病病毒（KFDV）对公众健康构成了重大威胁，因此有必要开发有效的抗病毒治疗方法。非结构蛋白5（NS5）对病毒RNA合成和甲基化至关重要，是一个有前景的药物靶点。本研究采用计算方法来识别和评估潜在的NS5抑制剂，这些抑制剂可能有助于开发针对KFDV的抗病毒化合物。使用Robetta、SwissModel和I-TASSER预测NS5的三维结构，选择Robetta模型（ERRAT评分：96.40）进行能量最小化。在49.58纳秒时获得的全局最小化结构，势能为-416966.82千卡/摩尔，并用于进一步研究。使用基于模板和基于结构的方法（COACH-D、CASTp、PrankWeb）识别活性位点残基，它们位于NS5蛋白的聚合酶基序（A-G）内（273-903氨基酸残基），这些基序对聚合酶功能、RNA合成和病毒复制至关重要。通过从头设计、基于模板的设计、药效团建模和配体筛选共鉴定出1523种化合物。使用PyRx 0.8进行虚拟筛选产生了34种有前景的化合物，其中11种基于分子对接（AutoDock 4.0）被选中，结合能分别为-8.86千卡/摩尔（FDA批准的达沙布韦-L1）、-8.28千卡/摩尔（CNPO331352.1-L2）、-7.94千卡/摩尔（ZINC00103114410-L3）和-7.61千卡/摩尔（CNPO202263.1-L4）。在生理条件下进行的三次重复分子动力学模拟证实了稳定性，MM-GBSA结合自由能值分别为-52.28±2.91千卡/摩尔（NS5-达沙布韦L1复合物）、-46.82±4.31千卡/摩尔（NS5-L2复合物）、-50.72±6.36千卡/摩尔（NS5-L3复合物）和-57.03±4.31千卡/摩尔（NS5-L4复合物）。计算分析表明，化合物L2和L4具有与达沙布韦（L1）相当的强结合亲和力，表明它们作为KFDV NS5蛋白抑制剂的潜力。通过体外试验进行进一步验证将补充这些计算机模拟结果。这些结果为未来抗KFDV药物开发奠定了基础，强调了持续探索抗病毒治疗方法的必要性。
