Purification of A-subtype pancreatic cholecystokinin receptor by immunoaffinity chromatography

Biochimie. 1992 Feb;74(2):149-51. doi: 10.1016/0300-9084(92)90039-h.

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Affinity Labels
  • Animals
  • Cell Membrane / chemistry
  • Cholecystokinin / isolation & purification*
  • Chromatography, Affinity*
  • Cross-Linking Reagents
  • Immunologic Techniques
  • Molecular Weight
  • Octoxynol
  • Pancreas / chemistry*
  • Photochemistry
  • Polyethylene Glycols
  • Rats
  • Solubility
  • Wheat Germ Agglutinins

Substances

  • Affinity Labels
  • Cross-Linking Reagents
  • Wheat Germ Agglutinins
  • Polyethylene Glycols
  • Octoxynol
  • Cholecystokinin
  • Nonidet P-40