Exploring Pitolisant Binding to Human Serum Albumin: Insights from Multi-Spectroscopic, Molecular Docking and Molecular Dynamics Studies

This study presents a comprehensive investigation of the molecular interactions between pitolisant and human serum albumin using a multi-technique approach. Advanced spectroscopic methods - including circular dichroism, UV–vis spectrophotometry, steady-state, three-dimensional, and synchronous fluorescence spectroscopy - were integrated with molecular docking and molecular dynamics simulations to elucidate the binding mechanism, structural implications, and stability of the pitolisant-HSA complex. Fluorescence quenching analysis revealed a static interaction mechanism, with thermodynamic parameters (ΔG < 0) confirming the spontaneity of the process, driven predominantly by hydrophobic interactions and a potential hydrogen bond formation. CD spectroscopy highlighted significant perturbations in HSA’s secondary structure upon pitolisant binding, notably a reduction in α-helical content, suggesting ligand-induced conformational reorganization. Molecular docking identified subdomain IIA (binding site I) as the primary interaction site. These findings were further validated by MD simulations, which demonstrated sustained complex stability over time. By delineating the structural and energetic determinants of pitolisant-HSA binding, this work advances the understanding of pitolisant’s pharmacokinetic behavior, particularly its distribution. The integrated experimental-computational framework also offers broader insights into ligand-protein interaction dynamics, with implications for rational drug design.