title: Purification of the pancreatic cholecystokinin receptor.
abstract: We have previously shown that the pancreatic cholecystokinin (CCK) receptor can be solubilized in 1% digitonin. In this study, digitonin-solubilized CCK receptors from rat pancreas were purified using sequential affinity chromatography on ricin-II agarose and on AffiGel-CCK. Electrophoresis of the radioiodinated purified receptors on SDS-polyacrylamide gels followed by autoradiography revealed two proteins: a major band of Mr = 80,000-90,000, and a minor band of Mr = 55,000. Through the purification procedure, the receptors preserved their agonist specificity (CCK-8 less than CCK-33 less than desulfated CCK-8 less than CCK-4) and binding affinity. Scatchard transformations of binding data for the purified receptor preparation were best fit by linear plots compatible with a single class of binding sites with Kd = 9.4 nM. The estimated purification was about 80,000 fold and consistent with the expected Bmax for a pure Mr = 80,000 protein binding one CCK molecule. This two-step purification procedure opens the possibility for molecular studies of the CCK receptor.
authors.affiliation: Cell Biological Laboratory, Mount Zion Hospital and Medical Center, San Francisco, CA 94120.
authors.forename: J
authors.initials: J
authors.lastname: Szecowka
authors.forename: G
authors.initials: G
authors.lastname: Hallden
authors.forename: I D
authors.initials: ID
authors.lastname: Goldfine
authors.forename: J A
authors.initials: JA
authors.lastname: Williams
mesh_terms: D000818:Animals; D001667:Binding, Competitive; D002846:Chromatography, Affinity; D007700:Kinetics; D008297:Male; D008970:Molecular Weight; D010179:Pancreas / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D011949:Receptors, Cholecystokinin / Q000302:isolation & purification* / Q000378:metabolism
chemical_list: D011949:Receptors, Cholecystokinin