Journal of Clinical Endocrinology & Metabolism, Vol 57, 741-748, Copyright © 1983 by Endocrine Society

ARTICLES

Analysis of the estrogen receptor from human uterus and breast tumor tissue by isoelectric focusing

SN Thibodeau, WP Sullivan and NS Jiang

Cytosolic estrogen receptors from human breast carcinoma (approximately 50 patients) and uterine myometrium tissue (approximately 20 patients) were analyzed by isoelectric focusing on agarose gels, by their ability to bind to isolated nuclei, and by ultracentrifugation, under diverse conditions. Through the use of diisopropylfluorophosphate (DFP) and sodium molybdate (Mo), two in vitro processes were examined by these techniques: receptor activation and receptor degradation. These agents are capable of inhibiting these processes selectively so that the resulting stabilized forms of the estrogen receptor can be characterized in more detail. The predominant molecular forms, as determined by the parameters stated above, vary dramatically depending on the presence of DFP or Mo. At least six distinct species of the estrogen receptor were resolved by isoelectric focusing. Of these, four appear to be products of proteolysis. These had isoelectric points of 6.9, 6.8, 6.4, and 6.3, with a combined sedimentation coefficient of approximately 3.2S. The nonactivated form of the receptor, prepared in the presence of both DFP and Mo, focused at pH 4.8 and had a sedimentation coefficient of approximately 9.5S. Finally, the activated form detected in the presence of DFP sedimented at 8.0S and had an apparent pI of 5.4.