The Isolation and Characterization of the Soluble Glutathione S-Transferases of Rat Basophil Leukemia-1 Cells
Peck, Rebecca E.
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The family of proteins known as glutathione S-transferases are a group of multifunctional enzymes catalyzing the conjugation of glutathione with a variety of substrates possessing a sufficiently electrophilic center. The enzymes occur in relatively high conceentrations in mammalian liver, where they play an important role in the detoxification of potentially harmful electrophilic compounds. Interest in the role of the transferases in detoxification has led to the isolation and purification of the enzymes from rat liver and from human liver. At least six catalytically different forms of the enzyme are known to occur in rat liver cytosol. In a different physiological context, glutathione S-transferases are found in mast cells and basophils, where a particulate form of the enzyme is involved in the formation of leukotrienes, also known as slow-reacting substance of anaphylaxis (SRS-A). Leukotrienes are potent spasmogens which exacerbate the overall effects of an acute allergic reaction. Pharmacologic modulation of leukotriene formation may be beneficial in the treatment of allergy, asthma, and other diseases of hypersensitivity. One approach is the inhibition of the glutathione S-transferase-catalyzed step. However, the glutathione S-trasferases have not been well characterized in leukotriene-generating cells, such as the rat basophilic leukemia (RBL-l) cell line which is commonly used as a model system. It was the purpose of this study to isolate the soluble glutathione S-transferases from RBL-1 cells and to characterize the various forms of the enzyme with respect to substrate specificity and inhibitor profile. Using a recently described chromatographic technique known as chromatofocusing, the glutathione S-transferases were separated on the basis of their isoelectric points. Ten peaks of activity were eluted from the chromatofocusing column, which were then further characterized according to substrate specificity and inhibitor profile. On the basis of isoelectric point, and quantitative and qualitative response to various substrates and inhibitors, there are at least six and possibly more catalytically different forms of the enzyme in RBL cells.