Insight into the interactions of proteinase inhibitor- alpha-2-macroglobulin with hypochlorite.

Hypochlorous acid (HOCl), an active bleaching agent is one of the major oxidant produced by neutrophils under physiological conditions. It is among one of the most potent reactive oxygen species (ROS) which causes oxidation of biomolecules. Treatment of proteins with hypochlorite results in direct oxidative damage to the protein. Alpha-2-macroglobulin (α2 M) is a major proteinase inhibitor that can inhibit proteinase of any kind regardless of their specificity and catalytic mechanism. The proteinase-antiproteinase balance plays an important role in mediating inflammation associated tissue destruction. In this paper, we intend to study hypochlorite induced modifications in proteinase inhibitor- α2 M via biophysical techniques such as absorption spectroscopy, fluorescence spectroscopy, circular dichroism (CD), fourier transform infrared spectrometry (FTIR) and isothermal titration calorimetry (ITC). It was found that hypochlorite decreases the anti-proteolytic potential and causes inactivation of sheep α2 M. It also causes structural and functional change in sheep α2 M as evident by UV-Visible absorption spectroscopy and fluorescence measurements. Change in secondary structure of α2 M was confirmed by CD and FTIR. Thermodynamics parameters such as entropy change (ΔS), enthalpy change (ΔH), Gibbs free energy change (ΔG) and the number of binding sites (N) of α2 M-HOCl binding in solutions were determined by ITC. Moreover, it was found that binding of HOCl with α2 M was exothermic in nature.