PURIFICATION OF GLUCOCORTICOID RECEPTORS FROM RAT-LIVER CYTOSOL - PREPARATION OF ANTIBODIES AGAINST THE MAJOR RECEPTOR PROTEINS AND APPLICATION OF IMMUNOLOGICAL TECHNIQUES TO STUDY ACTIVATION AND TRANSLOCATION

Two dexamethasone binding components from rat liver cytosol have been purified by protamine sulfate precipitation, affinity chromatography on 11-deoxycorticosterone-Sepharose and ion-exchange chromatography. The two components elute from DEAE-cellulose column at 0.12 M and 0.20 M NaCl respectively. Dodecylsulfate polyacrylamide electrophoresis demonstrates that the 0.12 M NaCl eluted component contains predominantly a single polypeptide of 45,000 mol. wt. and the 0.2 M NaCl eluted component contains a single polypeptide of 90,000 mol. wt. Antibodies to these two components have been elicited in rabbits after removing the minor contaminants from them by preparative dodecylsulfate/ polyacrylamide electrophoresis. The antibodies to the 45,000 mol. wt. polypeptide cross react with the 90,000 mol. wt. component. Likewise the antibodies to the 90,000 mol. wt. protein precipitate the 45,000 mol. wt. polypeptide. Either of the antibody preparations immunoprecipitate the glucocorticoid receptor complex from rat liver and hepatoma tissue culture cytosols. Both these components produce similar peptide fragments after limited proteolytic digestion with staphilococcal aureus protease. © 1979.