pH-dependence of warfarin binding to alpha 1-acid glycoprotein (orosomucoid)

The binding of warfarin to alpha 1-acid glycoprotein (AAG) was found to increase with decreasing pH. The u.v.-visible difference spectra generated upon binding to AAG at pH 5.0 or 7.4 showed warfarin to bind as the anion. Warfarin-binding data were satisfactorily fitted to a model that incorporates the effect of pH and discriminates the association constants of the non-protonated and protonated binding site of the protein. It was shown that AAG-binding site in the protonated form had a markedly higher affinity for warfarin than the non-protonated form, with a pK value of 7.7 +/- 0.1, which is likely to be a histidine residue. Among other possible interactions, it is suggested that ligand binding to AAG involves a reinforced hydrogen bond.