Computational Analysis of <i>Leishmania donovani</i> Secreted Metalloprotease GP‐63 as an Inhibitor of the Retinoic Acid Synthesis Pathway

Abstract This study explores the potential mechanism by which Leishmania donovani downregulates retinoic acid (RA) synthesis to promote survival within host macrophages. Parasites were treated with varying concentrations of RA, and secreted proteins were identified via LC–MS analysis. Among these, GP‐63, a metalloprotease known to aid parasite survival by degrading host proteins was investigated for its interaction with retinaldehyde dehydrogenase 2 (RALDH2), a key enzyme in RA biosynthesis. Molecular docking revealed a strong and stable interaction between GP‐63 and RALDH2, with a highly favorable binding energy of –950.3 kJ/mol. Molecular dynamics simulations over 200 ns indicated structural stability, evidenced by an average RMSD of 1.78 Å and a consistent radius of gyration (42 Å). MM/GBSA calculations yielded a total binding energy (ΔG bind ) of –80.59 kJ/mol, while interaction entropy analysis showed an entropic contribution (–TΔS) of 88.8 kcal/mol, resulting in a net binding energy of –1.72 kcal/mol. These results suggest that GP63 directly targets RALDH2, potentially inhibiting RA synthesis and thereby enhancing parasite persistence within macrophages. This work provides molecular‐level insights into the GP63–RALDH2 interaction, offering a foundation for the development of targeted therapeutics against visceral leishmaniasis.