Effect of subunit interactions on enzymatic activity of glutathione S-transferases: a radiation inactivation study.
The glutathione S-transferases are a family of dimeric enzymes. Three isozymes from the alpha family, termed YaYa, YaYc, and YcYc, and three from the mu family, termed Yb1Yb1, Yb1Yb2, and Yb2Yb2, were purified from rat liver. Binding studies were performed by equilibrium dialysis using a radiolabeled product, S(-)[14C](dinitrophenyl)glutathione. Each isozyme contained two independent binding sites which had equal affinity for the ligand. The presence of two independent active sites per enzyme dimer suggests that each subunit contains a complete active site. This conclusion was examined further using radiation inactivation which also allowed for assessment of the importance of subunit interactions in catalytic activity. The activity target size of YaYa (47 kDa) was significantly larger than the protein monomer target size (31 kDa); similarly the activity target size of YaYc was that of the dimer (54 kDa). In contrast, the activity target sizes of Yb1Yb1 and Yb2Yb2 were the same, being 35 and 29 kDa, respectively, and the protein monomer target size of Yb1Yb1 also was similar, being 32 kDa. These data indicate that interactions between subunits are critical for the maintenance of enzymatic activity of alpha class enzymes whereas each subunit of the two mu class proteins is capable of independent catalytic activity.
Boyer
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Division of Digestive Diseases, Emory University School of Medicine, Atlanta, Georgia 30322.
Kempner
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D001665:Binding Sites; D002250:Carbon Radioisotopes; D003956:Dialysis; D004789:Enzyme Activation / Q000528:radiation effects; D005978:Glutathione / Q000031:analogs & derivatives / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism* / Q000528:radiation effects*; D007527:Isoenzymes / Q000378:metabolism* / Q000528:radiation effects*; D007700:Kinetics; D046911:Macromolecular Substances
D002250:Carbon Radioisotopes; D007527:Isoenzymes; D046911:Macromolecular Substances; C030190:S-(2,4-dinitrophenyl)glutathione; D005982:Glutathione Transferase; D005978:Glutathione