Faizan Rabia, Naveed Muhammad, Estevez Inmaculada Bellido, Hanif Nimra, Arshad Arooj, Aziz Tariq, Alamri Abdulhakeem S, Alsanie Walaa F, Alhomrani Majid
Department of Pharmacology and Clinical Therapeutics, School of Medicine, University of Malaga, Spain.
Department of Biotechnology, Faculty of Science and Technology, University of Central Punjab, Lahore 54000, Pakistan.
Pathol Res Pract. 2025 Oct;274:156184. doi: 10.1016/j.prp.2025.156184. Epub 2025 Aug 21.
Naegleria fowleri, a thermophilic, free-living amoeba, is the causative agent of Primary Amoebic Meningoencephalitis (PAM), a rare but nearly always fatal brain infection. The rising number of PAM cases in Karachi, Pakistan, particularly linked to a unique local strain, underscores the urgent need for effective therapeutic interventions. In this study, a computational approach was employed to identify potential natural inhibitors targeting toxin-producing proteins from the N. fowleri Karachi strain. Eight exons encoding toxin proteins were retrieved from the Soft Berry Fgenesh 2.6 database, annotated using Gene Ontology tools, and subjected to physicochemical characterization. Hypothetical protein 4 was prioritized for molecular docking in the NF001 Karachi strain of Naegleria fowleri because it was identified through comparative mapping with previously known strains. Its function was predicted based on sequence alignment, suggesting that it may serve as a promising target for drug docking studies. Protein structures were predicted via AlphaFold2 and validated using MolProbity and Ramachandran plot analysis. Virtual screening of phytochemicals was conducted using PyRx, identifying himbacine as the most promising ligand with a binding affinity of -8.7 kcal/mol against hypothetical protein 4. Binding interactions were further confirmed using CB-Dock2, which revealed key binding residues involved in hydrogen bonding and hydrophobic interactions. ADMET profiling indicated that himbacine possesses favorable pharmacokinetics, non-toxicity, and high gastrointestinal absorption. Density Functional Theory (DFT) analysis showed a small HOMO-LUMO energy gap, indicating high reactivity and binding potential. Molecular dynamics simulations confirmed the structural stability of the protein-ligand complex over time. These findings suggest that himbacine, a plant-derived compound, holds promise as a safe and effective inhibitor against N. fowleri infections. However, further in vitro and in vivo studies are essential to validate its therapeutic potential.
福氏耐格里阿米巴是一种嗜热的自由生活阿米巴，是原发性阿米巴脑膜脑炎（PAM）的病原体，这是一种罕见但几乎总是致命的脑部感染。巴基斯坦卡拉奇PAM病例数量的上升，特别是与一种独特的本地菌株有关，凸显了对有效治疗干预措施的迫切需求。在这项研究中，采用了一种计算方法来识别针对福氏耐格里阿米巴卡拉奇菌株产生毒素的蛋白质的潜在天然抑制剂。从Soft Berry Fgenesh 2.6数据库中检索出八个编码毒素蛋白的外显子，使用基因本体论工具进行注释，并进行理化特性分析。假设蛋白4在福氏耐格里阿米巴NF001卡拉奇菌株的分子对接中被优先考虑，因为它是通过与先前已知菌株的比较图谱鉴定出来的。根据序列比对预测了它的功能，表明它可能是药物对接研究的一个有前景的靶点。通过AlphaFold2预测蛋白质结构，并使用MolProbity和拉马钱德兰图分析进行验证。使用PyRx对植物化学物质进行虚拟筛选，确定himbacine是最有前景的配体，对假设蛋白4的结合亲和力为-8.7 kcal/mol。使用CB-Dock2进一步证实了结合相互作用，该软件揭示了参与氢键和疏水相互作用的关键结合残基。ADMET分析表明，himbacine具有良好的药代动力学、无毒性和高胃肠道吸收性。密度泛函理论（DFT）分析显示HOMO-LUMO能隙较小，表明具有高反应性和结合潜力。分子动力学模拟证实了蛋白质-配体复合物随时间的结构稳定性。这些发现表明，himbacine这种植物衍生化合物有望成为一种安全有效的抗福氏耐格里阿米巴感染抑制剂。然而，进一步的体外和体内研究对于验证其治疗潜力至关重要。
