title: Transformation of the 8-9 S molybdate-stabilized estrogen receptor from low-affinity to high-affinity state without dissociation into subunits.
abstract: The rate of dissociation of labeled estradiol from [3H] estradiol-8-9 S receptor complexes ([3H]E2-8-9 S ER) molybdate-stabilized was determined in the presence of either an excess of unlabeled hormone ("chase") or of charcoal/dextran suspension ("stripping"). Biphasic dissociation of the hormone was observed in both cases, but the fraction of the fast-dissociating component was dramatically reduced (5% instead of 60%) when stripping was used. As the dissociation patterns were independent of the degree of saturation of the receptor, the results do not favor the possibility of cooperative effects between binding sites in the 8-9 S ER. After pretreatment of cytosol by charcoal at 28 degrees C for 15 min, the dissociation studied by chase displayed only the slowly dissociating component (t1/2 approximately 65 min). This effect was dependent on temperature and influenced by the ligand bound to 8-9 S ER, being pronounced with estradiol (E2) and absent with [3H]4-hydroxytamoxifen. The slow-dissociating component obtained after charcoal treatment was reconverted to fast-dissociating state by adding dithiothreitol or by incubation with cytosol at 20 degrees C. The charcoal treatment did not change the sedimentation coefficient (approximately 9 S) and the Stokes radius (approximately 7 nm) of the [3H]E2-8-9 S ER, and the slow-dissociating form obtained did not bind to DNA-cellulose either in the presence or absence of molybdate ions. Thus there are likely small but functionally significant changes of structure in the 8-9 S ER which remain in a non-DNA-binding form, whereas the rate of estradiol dissociation is modified.
mesh_terms: D000818:Animals; D002417:Cattle; D004355:Drug Stability; D004958:Estradiol / Q000378:metabolism; D005260:Female; D007700:Kinetics; D046911:Macromolecular Substances; D008982:Molybdenum / Q000494:pharmacology*; D009994:Osmolar Concentration; D011959:Receptors, Estradiol / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D011960:Receptors, Estrogen / Q000378:metabolism*; D013816:Thermodynamics; D014599:Uterus / Q000378:metabolism*
chemical_list: D046911:Macromolecular Substances; D011959:Receptors, Estradiol; D011960:Receptors, Estrogen; C044659:molybdate; D004958:Estradiol; D008982:Molybdenum