Partial purification of paraoxonase from human serum and its detoxication effect

AIM: To explore the possibility of paraoxonase as exogeneous antidote against G type organophosphate nerve agents. METHODS: Paraoxonase is partially purified by affinity gel column chromatography. The K_m value of paraoxonase of different substrate is determined. The effect of calcium ion on the enzyme activity is measured. Paraoxonase activities from different species are compared. Effect of paraoxonase on different substrates is studied by in vitro hydrolysis and in vivo poisoning. RESULTS: The human serum paraoxonase was partially purified using affinity gel column chromatography, it was found that the partially purified paraoxonase was able to well catalyze the hydrolysis of paraoxon and soman. The K_m while using paraoxon as the substrate, came to 0.22 mmol · L^-1 with a specific activity of 375 μmol · min^-1 · g^-1, whereas using soman as substrate, the K_m came to 0.60 mmol · L^-1 with a specific activity of 434 μmol · min^-1 · g^-1. The difference of paraxonase activity in various animal species is ascribed to the difference of susceptibility to organophosphate poisoning of various animals. In the presence of 1 mmol · L^-1 CaCl₂, the partially purified paraoxonase manifested well the enzyme activity. However, the enzyme activity was profoundly inhibited by the addition of EDTA, when soman or paraoxon was used as substrate. It suggests that human serum paraoxonase be a calcium ion-dependent enzyme. The pI₅₀ of EDTA came to about 1.8 mmol · L^-1. The human serum paraoxonase could efficiently catalyze the hydrolysis of soman, sarin, tabun, paraoxon and dichlorvos, however, it is inefficient to degrade VX and dimethoate. It indicates that human serum paraoxonase selectively hydrolyzes the P-F, P-CN and P-O bonds but not the P-S bond. CONCLUSION: Paraoxonase is a potential antidote against G type nerve agents.