Proteins associated with untransformed estrogen receptor in vitro. Perturbation of hydrophobic interactions induces alterations in quaternary structure and exposure of the DNA-binding site.
Estrogen receptors from calf uteri have been analyzed by high-performance size-exclusion chromatography, chromatofocusing, and DNA affinity chromatography using conditions designed to evaluate the relative contribution of hydrophobic interactions between the steroid-binding subunit and other receptor-associated proteins. The single large (untransformed) species of soluble estrogen-receptor consistently (n = 9) found in calf uteri displayed a rapid change in Stokes radius from 8.0 to 3.5 nm upon exposure to elevated ionic strengths (0.4 M KCl). However, equilibration of the estrogen-receptor complex into urea (up to 6 M) did not dissociate the untransformed receptor into the 3.5-nm receptor form (subunit) observed in hypertonic (0.4 M KCl) buffers. Exposure to 6 M urea did result in conversion of the untransformed receptor (8.0 nm) to a 6.0-6.5-nm receptor form not previously observed in either hypotonic or hypertonic buffers. In the presence of both 6 M urea and 0.4 M KCl, the untransformed estrogen-receptor complex was converted to a smaller receptor form intermediate in apparent size (4.5-5.0 nm) to that observed in 6 M urea or 0.4 M KCl alone. The formation of this 4.5-5.0-nm receptor form was partially estrogen dependent as determined by parallel analyses of unliganded receptor in urea/KCl buffer. The urea-induced change in apparent size (8 nm to 6.0-6.5 nm) at low ionic strength was accompanied by little or no detectable change in net surface charge as determined by chromatofocusing but a complete exposure of the DNA-binding site as evidenced by nearly quantitative interaction with DNA-agarose.(ABSTRACT TRUNCATED AT 250 WORDS)
Hutchens
T W
TW
Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston, Texas 77030.
Li
C M
CM
Besch
P K
PK
D000818:Animals; D002417:Cattle; D002846:Chromatography, Affinity / Q000379:methods; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D003600:Cytosol / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D004958:Estradiol / Q000378:metabolism; D005260:Female; D046911:Macromolecular Substances; D011189:Potassium Chloride / Q000494:pharmacology; D011487:Protein Conformation; D011960:Receptors, Estrogen / Q000302:isolation & purification / Q000378:metabolism*; D014508:Urea / Q000494:pharmacology; D014599:Uterus / Q000378:metabolism*
D046911:Macromolecular Substances; D011960:Receptors, Estrogen; D004958:Estradiol; D011189:Potassium Chloride; D014508:Urea; D004247:DNA