title: Purification of A-subtype pancreatic cholecystokinin receptor by immunoaffinity chromatography.
abstract: So far, no efficient affinity chromatography for CCK receptor purification has been reported that prevented obtention of sequenceable amounts of purified receptor. In this work, 10% of plasma membrane receptor sites were specifically cross-linked with the photoreactive cleavable agonist 125I-ASD-[Thr28, Ahx31]-CCK-25-33, solubilized by NP-40, chromatographied on immobilized wheat germ agglutinin and further immunopurified using anti-CCK antibodies to an overall rate of 3000-3600-fold. Analysis of eluted material demonstrated a protein migrating at Mr 85,000-100,000 and the absence of 35S-labeled impurity. This single and efficient affinity chromatography should provide enough homogeneous receptor protein for microsequence determination and leads to consider immunoaffinity chromatography on immobilized anti-ligand antibodies as a potential tool for purification of membrane receptors.
authors.affiliation: INSERM U151, CHU Rangueil, Toulouse, France.
authors.forename: M
authors.initials: M
authors.lastname: Dufresne
authors.forename: S
authors.initials: S
authors.lastname: Poirot
authors.forename: J
authors.initials: J
authors.lastname: Jimenez
authors.forename: N
authors.initials: N
authors.lastname: Vaysse
authors.forename: D
authors.initials: D
authors.lastname: Fourmy
mesh_terms: D000345:Affinity Labels; D000818:Animals; D002462:Cell Membrane / Q000737:chemistry; D002766:Cholecystokinin / Q000302:isolation & purification*; D002846:Chromatography, Affinity; D003432:Cross-Linking Reagents; D007158:Immunologic Techniques; D008970:Molecular Weight; D017830:Octoxynol; D010179:Pancreas / Q000737:chemistry*; D010777:Photochemistry; D011092:Polyethylene Glycols; D051381:Rats; D012995:Solubility; D014909:Wheat Germ Agglutinins
chemical_list: D000345:Affinity Labels; D003432:Cross-Linking Reagents; D014909:Wheat Germ Agglutinins; D011092:Polyethylene Glycols; D017830:Octoxynol; D002766:Cholecystokinin; C010615:Nonidet P-40