Bilirubin binding affects the structure and function of alpha-2-macroglobulin

Bilirubin is an endogenous antioxidant that is a metabolite of the heme in red blood cells (RBC). In blood, bilirubin is associated with albumin to form a water-soluble complex, known as unconjugated bilirubin. Alpha-2-macroglobulin (α₂M) is a proteinase inhibitor found in the plasma of vertebrates. In the present study, we have investigated the interaction of photo-illuminated bilirubin with serum α₂M using various biophysical and thermodynamic techniques. The binding of bilirubin to α₂M leads to various functional and structural changes in α₂M protein. The result of ultraviolet (UV) and fluorescence spectroscopy suggests that binding of bilirubin to α₂M induces a conformational change in the secondary structure of protein which was corroborated by circular dichroism (CD) and Fourier-transform infrared spectroscopy (FT-IR). This binding leads to the conversion of β-sheet into α-helical conformation and subsequently loss in protein activity. The thermodynamic parameters of bilirubin-α₂M binding indicate that the binding is exothermic, and the reaction spontaneous. Our studies show that binding of bilirubin with α₂M in the presence of light induces structural and functional modifications in the protein. Bilirubin possesses multiple biological activities, including immunomodulatory property which has not been extensively explored and which may be of interest for further study.