Studies on <i>in vitro</i> binding of parecoxib to p38MAPK using spectroscopic methods

Purpose: To investigate the interaction between parecoxib and p38MAPK under simulated physiological conditions.Methods: The interaction between parecoxib and p38MAPK was studied under simulated physiological conditions using spectroscopy-based methods. The effect of parecoxib on the microenvironment and conformation of p38MAPK chromophore was studied by synchronous fluorescence spectroscopy, three-dimensional fluorescence, time-resolved fluorescence spectroscopy, circular dichroism spectroscopy, and ultraviolet-visible absorption spectroscopy.Results: Synchronous fluorescence spectroscopy showed that addition of parecoxib changed the structure of p38MAPK and destroyed the original stable structure. Three-dimensional fluorescence spectroscopy showed that the hydrophilicity of the microenvironment in which the fluorescent chromophore is located was enhanced, and the polarity increased such that the serum protein macromolecules tend to be unfolded, and the alpha-helix content reduced. Time-resolved fluorescence spectroscopy showed that the presence of parecoxib hardly affected the fluorescence quenching of p38MAPK, and the combination of parecoxib and p38MAPK forms a stable complex (static quenching). Circular dichroism spectroscopy revealed the combined parecoxib change, the secondary structure of p38MAPK and reduced the alpha-helix content. Ultraviolet-visible absorption spectroscopy revealed changes in the microenvironment of the three amino acid residues as well as the tertiary structure of the protein.Conclusion: The results shows that parecoxib has a significant effect on the structure of p38MAPK. In addition to explicitly inhibiting COX-2 and blocking arachidonic acid synthesis of prostaglandins, it inhibits the pathway involved in p38MAPK.