Preferential binding of steroids by anionic forms of rat glutathione S-transferase.
Rat liver glutathione S-transferases with isoelectric points near 6.7 were resolved from more basic forms of the protein. This anionic fraction represented about 30% of the total activity in liver with 1-chloro-2,4-dinitrobenzene and was the preponderant form utilizing trans-4-phenyl-3-butene-2-one as a substrate. The anionic transferases are dimeric proteins composed of two subunits designated as Yb and were distinguished from the cationic transferases on the basis of structural, immunological, and binding properties. Amino acid compositions and immunological properties of the anionic protein were similar to those of glutathione S-transferases A and C. The anionic forms had substantially less ordered secondary structure than cationic forms composed of subunits Ya and Yc. Stoichiometric ratios of two high affinity binding sites per dimer, also differentiated between the anionic and all of the cationic transferases which bind only a single mole of ligand. Affinity matrices composed of corticosterone or cholate, and circular dichroism methods, were used to demonstrate selective binding of steroids and bile acids to the anionic glutathione S-transferases. Glucocorticoids and progestins were shown to bind with high affinity whereas estrogens were bound at distinct lower affinity sites. In contrast to the cationic transferases, glutathione had no effect on binding of the steroids to the anionic forms, which suggested that these proteins have the capacity to bind these substances even in a milieu with high concentrations of glutathione.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D000838:Anions; D019826:Cholic Acid; D002793:Cholic Acids; D002846:Chromatography, Affinity; D002942:Circular Dichroism; D005982:Glutathione Transferase / Q000302:isolation & purification / Q000378:metabolism*; D005779:Immunodiffusion; D007525:Isoelectric Focusing; D007527:Isoenzymes / Q000302:isolation & purification / Q000378:metabolism*; D008099:Liver / Q000201:enzymology*; D008297:Male; D011485:Protein Binding; D011487:Protein Conformation; D051381:Rats; D011919:Rats, Inbred Strains; D013256:Steroids
D000596:Amino Acids; D000838:Anions; D002793:Cholic Acids; D007527:Isoenzymes; D013256:Steroids; D005982:Glutathione Transferase; D019826:Cholic Acid