Characterization and Purification of Glutathione S-Transferase from the Liver and Gill Tissues of Agri Balik Lake Trout Salmo trutta labrax and the Effects of Heavy Metal Ions on Its Activity

摘要

Glutathione S-transferase (GST) from the liver and gill tissues of Agri Balik Lake Trout (also known as Black Sea Trout) Salmo trutta labrax was characterized and purified, and the toxic effects of some heavy metal ions on the enzyme's activity were analyzed. Liver GST was purified 930 times, resulting in 56% yield using glutathione-agarose affinity chromatography and a specific activity of 60.87 endotoxin units (EU)/mg protein. Using the same procedure, gill GST was purified 576 times, resulting in a 60% yield and specific activity of 46.8 EU/mg protein. The purity check of the purified enzymes was determined with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Optimal pH, ionic strength, and stable pH were found for each tissue, and separate K-M and V-max values were determined for reduced glutathione and 2,4-dinitrochlorobenzene substrates. Heavy metal ions that have toxic effects on living organisms and are known to contribute to environmental pollution were selected, and their in vitro effects on enzyme activity were analyzed. The IC50 values and K-i constants of those metal ions showing an inhibitory effect on GST activity were determined.