Abstract
Thirteen forms of glutathione S-transferase were isolated from human liver in high yields by glutathione-affinity chromatography and chromatofocusing. Apparent isoelectric points ranged from 4.9 to 8.9 and included neutral forms. All 13 forms appeared to be identical immunochemically in a quantitative enzyme-linked immunosorbent assay. These forms were immunochemically distinct from the major acidic glutathione S-transferase found in placenta and erythrocyte and were immunochemically distinct from two forms of higher molecular weight glutathione S-transferase found in some but not all liver samples. The 13 forms exhibited similar activities with 1-chloro-2,4-dinitro-benzene as substrate, specific activities of 33-94 mumol/min/mg. Likewise, these forms all exhibited glutathione peroxidase activity with cumene hydroperoxide, specific activities of 1.5-8.3 mumol/min/mg. All 13 forms bound bilirubin with subsequent conformational changes leading to states devoid of transferase activity, a process prevented by the presence of foreign proteins. As hematin-binding proteins, however, these multiple transferases exhibited a very broad range of binding extending from nonbinding to high-affinity binding (KD approximately 10(-8) M). Hematin binding was noncompetitive with transferase activity and did not involve the bilirubin-binding site, suggesting the existence of unique heme-binding sites on these proteins. The two forms of the immunochemically distinct glutathione S-transferases transferases found in some liver samples also exhibited both transferase and peroxidase activities. In addition, they also have separate sites for binding bilirubin and hematin.
Highlights
Most work on the glutathione S-transferases has concenisolated from human liver in high yields by glutathi- trated on the role of theseproteins in protection against one-affinitychromatographyand chromatofocusing. potentially toxic chemicals, which is undoubtedly one of the Apparent isoelectric points ranged from4.9 to 8.9 and in vivo functions of the glutathione S-transferases
There are neutral forms, one of which appears to be unique and is found in only some petitive with transferase activity and did not involve livers [25], and others that show some cross-reactivity with the bilirubin-binding site, suggesting the existenceof the basic enzymes [26]
Isolation of Multiple Glutathione S-Transferases from Human Liver-A sample of human liver, 100 g, was obtained at autopsy of an adult female who died from accidental causes
Summary
Most work on the glutathione S-transferases has concenisolated from human liver in high yields by glutathi- trated on the role of theseproteins in protection against one-affinitychromatographyand chromatofocusing. potentially toxic chemicals, which is undoubtedly one of the Apparent isoelectric points ranged from4.9 to 8.9 and in vivo functions of the glutathione S-transferases. All 13 forms appeared to be concentrations of the glutathione S-transferases in liver and identical immunochemically in a quantitative enzyme- their high affinities for bilirubin support a role for these linked immunosorbent assay.
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