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Physicochemical characterization of the androgen receptor from hyperplastic human prostate
Author(s) -
Murthy L. R.,
Chang C. H.,
Rowley D. R.,
Scardino P. T.,
Tindall D. J.
Publication year - 1984
Publication title -
the prostate
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.295
H-Index - 123
eISSN - 1097-0045
pISSN - 0270-4137
DOI - 10.1002/pros.2990050602
Subject(s) - epitestosterone , androgen receptor , chromatofocusing , chemistry , endocrinology , medicine , pmsf , androgen , receptor , chromatography , size exclusion chromatography , hormone , biology , biochemistry , cancer , enzyme , prostate cancer
In this manuscript we have characterized the androgen receptor from human benign prostatic hypertrophied (BPH) tissue. BPH tissue was obtained fresh from surgery, ground, and frozen in liquid nitrogen. Tissue was homogenized in 10 mM Tes buffer (pH 7.4 at 25°C) containing 0.25 M sucrose, 20 mM Na 2 MoO 4 , 12 mM monothioglycerol, and 1.5 mM EDTA. Cytosol was labeled with ( 3 H)‐R1881 (0.5–10 nM) ± a 100‐fold excess of R1881 and a 500‐fold excess of triamcinolone acetonide to identify specific androgen receptor sites. An exchange assay was developed, whereby maximum binding was achieved by raising the temperature to 30°C for 15 min. A high‐affinity binding protein was detected (Kd = 1.3 ± 0.8 nM) which had a binding capacity of 15 ± 7 fmol/mg protein. Binding was specific for androgens with the following Ki (nM) values: R1881 (0.09), dihydrotestosterone (6), testosterone (50), progesterone (92), estradiol (100), epitestosterone (860), and cortisol (> 1,000). Under high ionic conditions the sedimentation coefficient of the receptor was 4.1 S. Higher S values were not observed in the presence of the following protein inhibitors: leupeptin (20 or 150 μM), iodoacetamide (10 mM), DFP (5 mM), PMSF (1 mM), bacitracin (0.1 mM), antipain (0.1 mM), aprotinin (1 TIU/ml). Gel filtration analysis revealed a Stokes radius of 6.5 nm. These data indicate a M r of 120,000 and a f/fo of 2.01. The receptor eluted from a chromatofocusing column at a pH of 6.8. The receptor bound to phosphocellulose and DNA cellulose after being heat‐treated for 15 min at 30°C. These results suggest that BPH tissue contains an androgen receptor which is similar to receptors in other androgen‐responsive tissues.