Two populations of the estrogen receptor separated and characterized using aqueous two-phase partitioning.
Two populations of the rat uterine estrogen receptor (ER) were separated and characterized using aqueous two-phase partitioning. Countercurrent distribution of rat uterine cytosolic ER allowed rapid and efficient separation of two populations, one population partitioned preferentially into the top phase (T, K(obs) = 3-6) and the other into the bottom phase (B, K(obs) = 0.01-0.03). The majority of unoccupied cytosolic ER is in the T population. Upon estrogen binding and/or heating to 30 degrees C in vitro the T population is converted to the B population. The transition from T to B does not exclusively involve loss of heat shock protein 90 and does not alter the ligand binding ability of the steroid binding domain. Using the human ER steroid binding domain overproduced in Escherichia coli and the steroid binding domain generated by partial trypsinization of the rat uterine ER, we demonstrate that the characteristic distinguishing T and B populations is not localized to this domain alone but may be associated with the amino terminal half of the ER (the A/B and DNA binding domains). The T to B transition of the ER also occurs in human MCF-7 breast cancer cells upon treatment with estrogen at 37 degrees C.
Fritsch
M
M
National Institutes of Health, National Cancer Institute, Laboratory of Pathology, Bethesda, Maryland 20892, USA.
Anderson
I
I
Leary
C M
CM
Gorski
J
J
D000818:Animals; D001943:Breast Neoplasms; D002499:Centrifugation, Density Gradient; D003377:Countercurrent Distribution; D003600:Cytosol / Q000737:chemistry; D004967:Estrogens / Q000378:metabolism; D005260:Female; D006801:Humans; D010446:Peptide Fragments; D051381:Rats; D011960:Receptors, Estrogen / Q000235:genetics / Q000302:isolation & purification*; D011994:Recombinant Proteins; D014407:Tumor Cells, Cultured; D014599:Uterus / Q000737:chemistry*
D004967:Estrogens; D010446:Peptide Fragments; D011960:Receptors, Estrogen; D011994:Recombinant Proteins