Glutathione S-transferase: Purification and Characterization from Quail (Coturnix coturnix japonica) Liver and the Impact of Some Metal Ions on Enzyme Activity

The objective of this study was to examine the inhibitory effect of some heavy metals on the glutathione S-transferase (GST) enzyme, which is one of the major enzymes of glutathione metabolism purified from quail liver tissues. The quail liver GST enzyme was purified with 15.86 EU/mg specific activity, in a yield of 12.36% and 46.1 purification fold by ammonium sulfate precipitation and glutathione-agarose affinity chromatography. The molecular weight of subunits of the enzyme and the purity were determined by SDS-PAGE. In the characterization studies, the optimum pH of the GST enzyme was determined to be pH = 8.0 in Tris/HCl buffer. Optimum ionic strength was determined to be 140 mM in Tris/HCI buffer. Stable pH was found to be pH = 8.5 in Tris/HCl buffer. Optimum temperature was found to be 50 degrees C. K-M and V-max values for substrates 1-chloro-2,4-dinitrobenzene (CDNB) and GSH of the enzyme were determined to be K-M 0.048 mM V-max 0.479 EU/mL and K-M 0.114 mM V-max 0.672 EU/mL, respectively. In vitro inhibition effects of metal ions, including Ag+, Ni2+, Cd2+, Fe2+, Pb2+, Co2+ Zn2+, and Al3+, were investigated on the GST enzyme activity. The results showed that Ag+, Cd2+, Ni2+ Zn2+, and Al3+ metal ions inhibited GST enzyme (IC50 values 0.239, 0.250, 0.265, 0.320, 0.594 mM, respectively), while Fe2+, Pb2+, and Co2+ metal ions activated the enzyme. Finally, K-i values and inhibition types for these substances were determined by Lineweaver-Burk graphs.