Bovine hemoglobin thermal stability in the presence of naringenin: Calorimetric, spectroscopic and molecular modeling studies

Calorimetric, spectroscopic and dynamic light scattering experiments, together with complementary computational investigations afforded a detailed analysis of the thermal stability and conformation of the bovine hemoglobin (BHb)-naringenin (NAR) complex. Differential scanning microcalorimetry (mu DSC) revealed complex, multi-step aggregation processes above 330 K. There is a definite enhancement and alteration of the thermal signature of protein aggregation with increasing NAR concentration. Circular dichroism (CD) spectroscopy indicated minor conformational changes in the protein secondary structure upon ligand binding at 298 K. Temperature-dependent CD measurements evidenced steep variations of the alpha-helix (decrease) and unordered structures (increase), with only a slight increase of beta-structure as a result of unfolding and aggregation. UV-Vis absorbance and DLS measurements pointed to large size aggregates formation in the presence of ligand in high concentration. Molecular docking data identified the binding site of NAR, located in the internal cavity of BHb. Molecular dynamics simulations indicated hydrogen bonds, electrostatic and hydrophobic interactions as main contributions in the binding process; the bound ligand produced no sizable structural alterations of the protein molecule. Therefore, the clearly evidenced (mu DSC, CD, DLS) influence of the flavonoid on hemoglobin thermal aggregation is exerted as a free (not bound) component of the studied systems. (C) 2022 Elsevier B.V. All rights reserved.

Automated summary

Calorimetric, spectroscopic, and computational investigations were conducted to analyze the thermal stability and conformation of the bovine hemoglobin-naringenin complex. The results revealed that increasing naringenin concentration enhanced and altered the thermal signature of protein aggregation. Binding of naringenin caused minor conformational changes in the protein secondary structure. UV-Vis absorbance and dynamic light scattering measurements indicated the formation of large size aggregates in the presence of high naringenin concentration. Molecular docking and dynamics simulations showed that hydrogen bonds, electrostatic and hydrophobic interactions played significant roles in the binding process. The flavonoid exerted its influence on hemoglobin thermal aggregation as a free component of the system.