High-throughput virtual screening and empirical validation of probable inhibitors of Plasmodium falciparum and vivax glutathione transferase using bromosulfophthalein as the benchmark ligand.

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Plasmodium falciparum Glutathione S-Transferase (PfGST) and Plasmodium vivax Glutathione S-Transferase (PvGST) play vital roles in detoxification and parasite survival, making them key targets for antimalarial drug development. These enzymes offer potential for creating therapies with improved efficacy, reduced resistance, and minimal toxicity. Natural compounds like flavonoids, known for their antiplasmodial properties, are promising scaffolds for new drug designs. This study computationally screened baicalin (BA) and 5,7,3'-Trihydroxy-6,4',5'-trimethoxyflavone (TTMF), synthesizable and affordable flavonoids from the MedChemExpress database, as potential inhibitors of PfGST and PvGST, outperforming BSP. Molecular dynamics simulations revealed that BA and TTMF stabilize enzyme interactions through hydrogen bonds and van der Waals forces, altering protein compactness and dynamics, suggesting non-competitive, allosteric inhibition. Empirical validation showed complete enzymatic inhibition by BA and TTMF with IC50 values of 1.69 and 1.71 μM, respectively, while minimizing human GST inhibition. Using 1-chloro-2,4-dinitrobenzene and reduced glutathione (GSH) as substrates, BA and TTMF demonstrated tight binding near the hydrophobic substrate-binding sites of PfGST and PvGST. Spectroscopic analysis using 8-anilino-1-naphthalenesulfonate (ANS) confirmed their ligandin effects and binding at the dimer interface. These findings highlight BA and TTMF as promising candidates for developing effective antimalarial therapies.