Transformation of the rat uterine estrogen receptor after partial purification

Warming crude rat uterine cytosol after the addition of ³H] estradiol accelerates the association of the 4-S estrogen-binding protein with a second macromolecule, resulting in the formation of the 5-S estrogen-binding protein. To determine whether the 5-S estrogen-binding protein consists of two similar or dissimilar subunits, uterine cytosol was subjected to a number of fractionation procedures that separate macromolecules by solubility, molecular gel sieving, sedimentation rate, ionic charge, and heat lability. Following each of these methods, the fraction containing the 4-S estrogen-binding protein was incubated at 28°C; each of these 4-S estrogen-binding protein-containing fractions retained its capacity to completely transform to the 5-S estrogen-binding protein. In samples subjected to partial purification procedures, it was necessary that the buffer contain 40 mM Tris, 60 mM KCI, 1–10 mM dithiothreitol, and 1 M urea at pH 7.4, in order to accomplish the 4-S to 5-S estrogen-binding protein transformation at 28°C. Formation of the 5-S estrogen-binding protein requires association of the 4-S estrogen-binding protein with a molecule identical to or very similar to itself.