high-affinity oestrogen-binding protein in cockerel liver cytosol

• Published in: Biochemical journal 1979-05, Vol.180 (2), p.347-353
• Main Authors: Lazier, C.B, Haggarthy, A.J
• Format: Article
• Language: English
• Subjects: animal breeding; animal science; Animals; Chickens; Chromatography, Gel; Cytosol - metabolism; Estradiol - metabolism; Estradiol - pharmacology; In Vitro Techniques; Liver - drug effects; Liver - metabolism; livestock; Male; Protein Binding; Receptors, Estrogen - metabolism
• ISSN: 0306-3275; 0264-6021; 0306-3283; 1470-8728
• DOI: 10.1042/bj1800347

In contrast with several earlier reports, cytosol from cockerel liver contains a significant concentration of a protein that binds oestradiol with high affinity. To demonstrate the activity, certain alterations in the conventional method of preparation of cytosol must be made. Homogenization in sucrose-containing buffer at pH 8.4 in the presence of proteinase inhibitors and rapid fractionation of the cytosol with (NH4)2SO4 enables demonstration of a single class of oestradiol-binding sites with a Kd of about 1 nM and specificity only for oestrogens. The concentration is about 300 sites per cell in liver from 2-week-old cockerels. Oestradiol treatment in vivo decreases the number of exchangeable cytosol oestradiol-binding sites by about 80% for 1--4h, after which time it is gradually restored. Gel filtration of the cytosol preparation in the presence of high salt concentrations reveals that most of the oestradiol-binding activity is in high-molecular-weight aggregates, but a mild trypsin treatment generates a specific binding protein with an approximate mol.wt. of 40 000. This protein may be an oestrogen receptor.