Purification of rabbit uterine cytosolic estrogen receptors by affinity chromatography
| Autor: | Benjamin S. Leung, Thelma C. Madhok |
|---|---|
| Rok vydání: | 1981 |
| Předmět: |
Chromatography
Heparin Chemistry Sepharose Uterus Size-exclusion chromatography Estrogen receptor Biochemistry High-performance liquid chromatography Chromatography Affinity Cytosol Endocrinology Receptors Estrogen Affinity chromatography Sephadex Chromatography Gel Animals Female Rabbits Receptor Polyacrylamide gel electrophoresis Chromatography High Pressure Liquid |
| Zdroj: | Journal of Steroid Biochemistry. 15:299-305 |
| ISSN: | 0022-4731 |
| DOI: | 10.1016/0022-4731(81)90288-0 |
| Popis: | Progress to elucidate the mechanism of estrogen action in uterine tissues has been hampered because of the formidable task of receptor purification. This report describes our recent success in the purification of ERc from estrogen-primed rabbit uterus. Crude cytosolic ERc are purified sequentially by affinity chromatography and gel filtration, using heparin-Sepharose 4B, 17β-estradiol-17-hemisuccinyl-oval-bumin-Sepharose 4B, and Sephadex G-50. This purified ERc sediments as a 4.5 S molecule on sucrose gradients in low-salt buffer, and exhibits a major radioactive peak in both polyacrylamide gel electrophoresis under nondenaturing conditions and in HPLC eluates. The radioactivity pattern in HPLC coincides with that of the u.v. scan, showing the elimination of other u.v. absorbing materials of the crude cytosol. These results show that the bulk of cytosolic proteins have been removed by this procedure and indicate the feasibility of using HPLC for the large scale purification and analysis of ERc. Attempts are initiated in this laboratory to produce antibodies against this purified receptor by hybridoma technique, and to further analyze the biochemical nature of the purified receptor. |
| Databáze: | OpenAIRE |
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