Exploring the binding of cinchonine with human transferrin: combined experimental and computational approaches

Neurodegenerative diseases (NDs), like Alzheimer's disease (AD), present immense global health challenges, marked by progressive and irreversible neuronal loss. While many studies have reported the neuroprotective potential of various phytochemicals, the neurotherapeutic relevance of alkaloids, like cinchonine remains largely unexplored. This study for the first time investigates cinchonine, a natural Cinchona alkaloid with reported antioxidant, anti-inflammatory, and amyloid-inhibitory properties, for its interaction with human transferrin (hTf), a glycoprotein central to iron homeostasis and neuroprotection employing a combination of computational and experimental approaches. UV-Vis spectroscopy revealed significant changes in hTf's absorbance upon cinchonine binding, confirming stable protein-ligand complex formation, with a binding constant (K) of 0.7 × 10⁵ M^{- 1}. Fluorescence binding assay further validated the formation of a stable protein-ligand complex. Cinchonine binds with hTf with a binding constant (K) of 0.4 × 10⁶ M^{- 1}, signifying the strength of interaction. Molecular docking pinpointed cinchonine's specific binding site on hTf with a binding affinity of - 6.9 kcal/mol and its interactions with critical residues like Thr392. These findings were reinforced by molecular dynamics (MD) simulations and MM-PBSA, which showcased the stability and conformational integrity of the hTf-cinchonine complex over time. Additionally, hydrogen bonding and free energy analyses provided deeper insights into the molecular basis of the protein-ligand complex. All the findings imply the formation of a stable hTf-cinchonine complex. This study underscores cinchonine's potential as a therapeutic lead, generating hypotheses for future experimental validation of its efficacy in preventing or mitigating NDs.