title: Isolation of the human plasma corticotrophin-releasing factor-binding protein.
abstract: We report the purification of human corticotrophin-releasing factor-binding protein (hCRF-BP) using repeated affinity chromatography (with the aid of synthetic CRF immobilized on Sepharose 4B solid phase) followed by gel filtration. Presence of the binding protein was tracked throughout the procedure by its ability to inhibit binding of 125I-labelled hCRF to an hCRF antiserum; normal human plasma exhibits 80% inhibition in this system whereas sheep plasma, which does not contain an hCRF-BP, has no effect. Affinity cross-linking of 125I-labelled hCRF to the purified hCRF-BP was performed using disuccinimidyl suberate (1 mmol/l). SDS electrophoresis of the purified CRF cross-linked binding protein followed by radioautography resulted in one major band of Mr 37,000 which corresponded to our original molecular weight estimate based on the elution position of the binding protein on Sephacryl S-200. A 10(7)-fold purification of the hCRF-BP resulted in a preparation estimated to be 95% pure, with an overall yield of 5%.
authors.affiliation: Department of Biochemistry & Physiology, University of Reading, Whiteknights.
authors.forename: D P
authors.initials: DP
authors.lastname: Behan
authors.forename: E A
authors.initials: EA
authors.lastname: Linton
authors.forename: P J
authors.initials: PJ
authors.lastname: Lowry
mesh_terms: D000324:Adrenocorticotropic Hormone / Q000378:metabolism*; D002352:Carrier Proteins / Q000302:isolation & purification*; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D003346:Corticotropin-Releasing Hormone / Q000378:metabolism*; D004586:Electrophoresis; D006801:Humans
chemical_list: D002352:Carrier Proteins; D000324:Adrenocorticotropic Hormone; D003346:Corticotropin-Releasing Hormone