A DNA-BINDING (R-I) AND A NON-DNA-BINDING (R-II) ESTROGEN-RECEPTOR INTHE GOAT UTERINE NUCLEUS - PURIFICATION AND CHARACTERIZATION

Two forms of nuclear estrogen receptors have been isolated and purifie d from the goat uterus. The biochemical characteristics of the protein s imply that the receptors may be identified as the type I and type II nuclear estrogen receptors. Nevertheless, we felt a necessity to exer cise caution in using this nomenclature and, therefore, decided to ide ntify them instead as R-I and R-II, respectively. While R-I is the DNA -binding form, R-II is a non-DNA-binding protein. The two proteins are totally dissimilar in other physical characteristics like the Stokes radii (36 Angstrom for R-I and 21 Angstrom for R-II), sedimentation co efficients (4.8 S for R-I and 3.8 S for R-II), the K-d (1 nM for R-I a nd 2 nM for R-II), and the nature of the CNBr fragmentation of the pro teins. The two proteins, however, cross-react with polyclonal antibodi es raised against goat uterine estrogen receptor activation factor (E- RAF), a DNA-binding protein with no capacity to bind estradiol, origin ally discovered by T. N. R. V. Thampan and J. H. Clark (1981, Nature 2 90, 152-154). A major feature of the R-II isolation procedure is the c hromatography of the protein on a heat shock protein 90-Sepharose colu mn in the presence of molybdate ions and elution using a molybdate-fre e buffer. While estradiol-17 beta (E,) binding to R-II was inhibited b y the presence of dithiothreitol and quercetin in the medium, E(2)-R-I interaction remained unaffected by these exposures. (C) 1994 Academic Press, Inc.