Purification and characterization of human platelet glutathione-S-transferase.
A glutathione-S-transferase was isolated and purified to homogeneity from human platelets. With a combination of ammonium sulfate fractionation and chromatographic methods, 0.2 mg of pure enzyme was obtained from 9 X 10(11) platelets with a 12% recovery. The purified enzyme had a specific activity of 7.5 U per milligram, representing an approximately 1,100-fold purification. The enzyme was found to be anionic, with an isoelectric point of 4.6. With reduced glutathione as a co-substrate, platelet glutathione-S-transferase was most active with the synthetic substrate, 1-chloro-2,4-dinitrobenzene, less active with 1,2-dichloro-4-nitrobenzene, and essentially inactive with nitroglycerin and 1,2-epoxy-3-(p-nitrophenoxy)-propane. The pH optimum for activity with glutathione and 1-chloro-2,4-dinitrobenzene was 7.0. Indomethacin (1-(p-chlorobenzoyl)-5-methoxy-2-methyindole-3-acetic acid), a chlorobenzene derivative, noncompetitively inhibited human platelet glutathione-S-transferase with an apparent KI of 0.23 mmol/L. This study represents the first complete purification and characterization of a glutathione-S-transferase from platelets. The presence of this enzyme in the platelet, within which high concentrations of reduced glutathione coexist, suggests the potential importance of the platelet in detoxification reactions and in the synthesis of the glutathione adducts of leukotriene metabolism.
D001792:Blood Platelets / Q000201:enzymology*; D004137:Dinitrochlorobenzene / Q000378:metabolism; D004852:Epoxy Compounds / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D005982:Glutathione Transferase / Q000302:isolation & purification*; D006801:Humans; D007213:Indomethacin / Q000494:pharmacology; D007526:Isoelectric Point; D007700:Kinetics; D009578:Nitrobenzenes / Q000378:metabolism; D005996:Nitroglycerin / Q000378:metabolism; D009596:Nitrophenols / Q000378:metabolism
D004137:Dinitrochlorobenzene; D004852:Epoxy Compounds; D009578:Nitrobenzenes; D009596:Nitrophenols; C007606:1,2-epoxy-3-(p-nitrophenoxy)propane; C028328:1,2-dichloro-4-nitrobenzene; D005982:Glutathione Transferase; D005996:Nitroglycerin; D005978:Glutathione; D007213:Indomethacin