Binding and molecular dynamics studies of 7-hydroxycoumarin derivatives with human serum albumin and its pharmacological importance.

Human serum albumin (HSA) is one of the most widely studied proteins and is an important plasma protein responsible for binding and transport of many exogenous and endogenous drugs. Coumarin derivatives play a critical role as anticancer, antidiabetic, anticoagulant, and analgesic agents. Here we have studied the cytotoxic activity of 7-hydroxycoumarin derivatives (7HC-1, 7HC-2, and 7HC-3) on mouse macrophage (RAW 264.7) cell lines. These studies revealed that 7-hydroxycoumarin derivatives caused an increased inhibition in growth of inflamed macrophages in a concentration-dependent manner with an IC50 of 78, 63, and 50 μM. Further studies, using fluorescence, circular dichroism spectroscopy, molecular docking, and molecular dynamics methods, show binding of 7HC (umbelliferone) derivatives with HSA at physiological pH 7.2. The binding constant of 7HC derivatives with HSA obtained from fluorescence emission was found to be K7HC-1 = 4.6 ± 0.01 × 10(4) M(-1), K7HC-2 = 1.3 ± 0.01 × 10(4) M(-1), and K7HC-3 = 7.9 ± 0.01 × 10(4) M(-1) which corresponds to -6.34 kcal/mol, -5.58 kcal/mol, and -6.65 kcal/mol of free energy. In contrast, the binding of these coumarin derivatives (7HC-1, 7HC-2, and 7HC-3) was almost negligible with α-1-glycoprotein (AGP). Circular dichroism (CD) studies revealed a decreased α-helix content with an increase in the β-sheets and random coils in HSA upon interaction with coumarin derivatives, suggesting a partial unfolding of the HSA secondary structure. Site probe studies with phenylbutazone (Site I) and ibuprofen (Site II) indicated that 7HC derivatives specifically bind to sub domains IIIA and IIIB of HSA which is further corroborated by molecular dynamics and docking studies suggesting that binding is specific in nature. The values of free energies and binding constants coincide for both experimental and in silico analysis and suggest that there are hydrophobic interactions when coumarin derivatives bind to HSA. Molecular dynamics studies showed that the HSA-coumarin complex reaches an equilibration state at around 3.5 ns which indicates that the HSA-coumarin complexes were stable. Thus these interactions play a central role in development of coumarin derivative-inspired drugs.