Structural insights into the structure-activity relationships of flavonoids targeting WSSV and LvGSTMu.

White spot syndrome virus (WSSV) represents one of the most severe threats to shrimp aquaculture worldwide, with no effective antiviral treatments currently available. In this study, the antiviral and antioxidant potential of the flavonoids amentoflavone, agathisflavone, and apigenin was investigated using an integrated in silico approach combining molecular docking, pharmacophore analysis, and Normal Mode Analysis (NMA)-based molecular dynamics. Docking simulations revealed that all flavonoids bind favorably to WSSV thymidylate synthase (wTS) and to the glutathione S-transferase from L. vannamei (LvGSTmu), with binding affinities superior to those of the co-crystallized ligands. Amentoflavone showed the highest affinity toward wTS, preserving key catalytic interactions and closely mimicking the binding mode of the native substrate. Pharmacophore analysis highlighted the importance of aromatic centers and hydrogen bond donor/acceptor groups in stabilizing ligand-protein complexes. NMA-based dynamics demonstrated that amentoflavone and apigenin confer increased structural stability to wTS and LvGSTmu, respectively, reducing conformational fluctuations without compromising protein integrity. Molecular dynamics simulations confirmed the stability of the complexes throughout the 200 ns simulation, while MM/GBSA analysis revealed favorable binding free energies, supporting stable interactions with wTS. Overall, these results suggest that flavonoids, particularly amentoflavone, represent promising scaffolds for the development of antiviral and antioxidant agents against WSSV, warranting further experimental validation.