Characterization of the binding interaction between bovine hemoglobin and riboflavin sodium phosphate: multi-spectroscopic analysis and molecular docking studies

Proteins are essential bimolecular substances that play a crucial role in the maintaining life and are closely associated with its the origin, evolution, and metabolism. Through the use of molecular docking, synchronous fluorescence spectroscopy, ultraviolet-visible absorption, fluorescence, and Fourier transform infrared (FT-IR) spectroscopy, this study sought to clarify the relationship between bovine hemoglobin (BHb) and riboflavin sodium phosphate (RSP) at the normal biological condition. The fluorescence quenching experiment indicated that RSP can cause a static quenching mechanism that quenches BHb's natural fluorescence. Thermodynamic measurements demonstrated that the hydrogen bonding molecular force and hydrophobic contacts caused negative enthalpy and entropy changes during RSP binding to BHb. Using Förster resonance energy transfer, the binding distance between RSP and the BHb tryptophan residues was determined to be 3.11 nm. Fourier transform infrared spectroscopy, synchronous fluorescence, and UV-visible studies revealed that the secondary structure of BHb was considerably altered due to interaction with RSP. The molecular docking simulation revealed that, in addition to hydrophobic interactions, the hydrogen bonds were involved in the interaction of BHb-RSP complex. This study aims to enhance our understanding of the molecular interactions between BHb and RSP, which is significant for elucidating the biochemical pathways involved in metabolic processes.