Interaction of mercury and copper on papain and their combined inhibitive determination

Abstract

Influence and interaction of mercury ion (Hg2+) and copper ion (Cu2+) on papain activity in casein hydrolysis were investigated. Single Hg2+ or Cu2+ at low concentrations induced an increase in papain activity, but decreased it at high concentrations, confirming a typical hormesis phenomenon. The interaction of Hg2+ and Cu2+ at various concentration combinations showed that the binary interaction of 10−8mol/L Cu2+ and 10−6mol/L Hg2+ (Binary union S) buffer was of synergistic nature, while 10−4mol/L Cu2+ and 10−4mol/L Hg2+ (Binary union I) buffer was of competitive inhibition. The conformational changes in papain structure due to the interaction of binary metal ions were studied by ATR-FTIR, UV–vis and intrinsic fluorescence spectroscopies, also the changes of papain catalytic behavior were studied through kinetic analysis. Decreasing of α-helix content with increasing in intermolecular β-sheet aggregates content in Binary union I buffer resulted in an inactivation of papain activity by 57.2% and lower affinity for casein. On the contrary, papain activity increased with α-helix content increasing and intermolecular β-sheet aggregates content decreasing in Binary union S buffer. The competitive interaction between Cu2+ and Hg2+ on papain activity was found at higher concentrations (≥10−4mol/L), and the inhibition of the binary metal ions on papain was of a noncompetitive type.