Molecular recognition of a Novel thiazolidinedione-morpholine-based ionic liquid salt by HSA: spectroscopic and...

Human serum albumin (HSA) is the most abundant transport protein in human plasma and plays a crucial role in the pharmacokinetic behavior of therapeutic agents. In the present study, the interaction between a newly synthesized thiazolidinedione-morpholine-based ionic liquid salt (TMH-IL) and HSA was systematically investigated using spectroscopic techniques in combination with molecular docking and molecular dynamics (MD) simulations. Steady-state fluorescence spectroscopy revealed concentration-dependent quenching of the intrinsic fluorescence of HSA. Temperature-dependent Stern-Volmer analysis supported a predominantly static quenching mechanism, as evidenced by decreases in both the Stern-Volmer constant (K _SV) and binding constant (K _a) with increasing temperature. The binding constants were on the order of 10⁴-10⁵ M^-1, indicating moderate affinity between TMH-IL salt and HSA. Thermodynamic analysis yielded negative values of ΔH (-73.08 kJ mol^-1), ΔS (-155 J mol^-1 K^-1), and ΔG, indicating a spontaneous and enthalpy-driven binding process. UV-vis absorption spectroscopy showed concentration-dependent absorption changes, providing complementary evidence for HSA-TMH-IL interaction. FT-IR deconvolution of the amide I region revealed only minor changes in HSA secondary structure, with the α-helical content decreasing from 62.03% to 60.16% upon ligand binding. Consistently, circular dichroism (CD) spectroscopy showed minor reductions in ellipticity at 208 and 222 nm while preserving the characteristic α-helical spectral profile. Three-dimensional fluorescence spectroscopy further indicated perturbation of the Trp214 microenvironment without substantial conformational disruption. Molecular docking predicted Sudlow's site I (subdomain IIA) as the most favorable binding region among the investigated HSA sites. The co-docked anion-cation system yielded the highest ChemPLP score, and interaction analysis revealed hydrophobic contacts and hydrogen-bonding interactions with key residues within the binding cavity. Comparative 150 ns MD simulations of free HSA and the HSA-TMH-IL complex showed that ligand binding did not substantially destabilize the protein or alter its overall compactness. RMSD, RMSF, R _g, SASA, secondary-structure, and hydrogen-bond analyses collectively indicated preservation of the overall HSA architecture, accompanied by limited local conformational rearrangements and persistent protein-ligand interactions. Overall, the combined spectroscopic and computational findings indicate a spontaneous association of moderate affinity between TMH-IL salt and HSA, with molecular docking predicting preferential recognition of Sudlow's site I, while the overall structural integrity of HSA is largely preserved upon ligand binding.