Affinity labeling of the rat pancreatic cholecystokinin (CCK) receptor with decapeptide probes has identified an Mr = 85,000-95,000 protein, distinct from the Mr = 80,000 component previously labeled with 125I-Bolton Hunter-CCK-33. We have characterized the carbohydrate composition of this novel protein labeled with 125I-D-Tyr-Gly-[(Nle28,31)-CCK-26-33] and disuccinimidyl suberate by using chemical and enzymatic deglycosylation and lectin chromatography. The Mr = 85,000-95,000 component was demonstrated to be an N-linked sialoglycoprotein based on neuraminidase digestion to Mr = 75,000-85,000 and endo-beta-N-acetylglucosaminidase F (Endo F) digestion to Mr = 42,000. This was distinct from the Mr = 65,000 product of Endo F digestion of the protein labeled with 125I-Bolton Hunter-CCK-33. Lack of an effect of endo-beta-N-acetylglucosaminidase H demonstrated the absence of N-linked simple oligosaccharides, while products of chemical deglycosylation with hydrogen fluoride and endo-alpha-N-acetylgalactosaminidase supported the absence of O-linked carbohydrate. The presence of at least four oligosaccharide chains on the core protein was suggested by Endo F digestion of the Mr = 85,000-95,000 protein using limiting enzyme conditions. This glycoprotein was retained on wheat germ agglutininagarose and eluted by N,N',N"-triacetylchitotriose. Identification of the Mr = 85,000-95,000 component on the ectodomain of the plasmalemma of intact pancreatic acini confirmed this to be the fully processed form of the CCK-binding protein.
|Original language||English (US)|
|Number of pages||7|
|Journal||The Journal of biological chemistry|
|State||Published - Oct 5 1987|
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology