Deciphering the molecular interaction between Vitamin D3 and pepsin by in vitro and in silico perspectives

The current study explored the molecular interaction between Vitamin D3 (Vit D3) and pepsin using multispectroscopic, molecular dynamic simulation (MDS), and molecular docking. The fluorescence emission spectra discovered Vit D3 interacted with pepsin in a static quenching manner due to the formation of the steadystate complex. Thermodynamic data revealed the spontaneous binding of Vit D3 on pepsin. The formation of the Pepsin-Vit D3 complex was also validated by circular dichroism (CD) spectroscopy. The fluorescence and CD spectroscopy results revealed Vit D3 altered the tertiary and secondary structure of pepsin, respectively. Meanwhile, FTIR spectroscopy results revealed a hypochromic shift in the amide I and II peaks. Kinetic parameters showed Vit D3 inhibited the activity of pepsin by the uncompetitive process. Applied spectroscopic methods disclosed that Vit D3 binding to pepsin caused microenvironmental modifications around the aromatic residues of protein and changed its structure and function. Moreover, MD simulation and molecular docking were done to analyze the formation of Pepsin-Vit D3 complexes. Molecular docking findings demonstrated the interaction of Vit D3 with pepsin mainly involved van der Waals forces and hydrogen bonds that were in good agreement with the fluorescence results. Finally, MDS findings including RMSD, RMSF, and RG confirmed all the experimental data.