title: Simple procedures for purification and stabilization of human serum paraoxonase-1.
abstract: Human paraoxonases-1 is one of the most important detoxifying enzymes. In this study using simple chromatographic procedures human paraoxonases-1 was purified from human pooled plasma. The enzyme was purified using DEAE Sephadex an anion exchanger and G-200 a gel filtration chromatographic media. Results showed a single band of approximately 43 KD proteins in SDS-PAGE, corresponding to the human PON1. Using paraoxon as the substrate the activity was related to the concentration of calcium and sodium ions (Km=1.2+/-0.2 mM). Phenyl acetate hydrolyzing activity was independent of sodium and calcium ions (Km=0.78+/-0.08 mM). Keeping at 25 degrees C for 20 days 75% of the enzyme original activity was restored in 20% (v/v) glycerol. EDTA and zinc chloride both inhibited the enzyme activity. In conclusion the applied procedures can be used for large scale purification. It would greatly facilitate their structural and functional characterization and permit examination of their weak, yet potentially most biologically relevant activities, in the complete absence of other serum proteins.
authors.affiliation: Departments of Biochemistry, Faculty of Medicine and Research center for Molecular Biology, Baqiyatallah Medical Sciences University, Tehran, Iran.
authors.forename: Leila
authors.initials: L
authors.lastname: Golmanesh
authors.forename: Hossein
authors.initials: H
authors.lastname: Mehrani
authors.forename: Mohammad
authors.initials: M
authors.lastname: Tabei
mesh_terms: D043303:Aryldialkylphosphatase / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism*; D004492:Edetic Acid / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D006358:Hot Temperature; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics
chemical_list: D004791:Enzyme Inhibitors; D004492:Edetic Acid; D043303:Aryldialkylphosphatase; C468406:PON1 protein, human