Characterization of a bombesin receptor on Swiss mouse 3T3 cells by affinity cross-linking

Journal Article · Thu Dec 01 04:00:00 EST 1988 · J. Cell. Biochem.; (United States)

DOI:https://doi.org/10.1002/jcb.240380403· OSTI ID:6101675

Sinnett-Smith, J; Zachary, I; Rozengurt, E

We have previously identified by chemical cross-linking a cell surface protein in Swiss 3T3 cells of apparent Mr 75,000-85,000, which may represent a major component of the receptor for peptides of the bombesin family in these cells. Because bombesin-like peptides may interact with other cell surface molecules, it was important to establish the correlation between receptor binding and functions of this complex and further characterize the Mr 75,000-85,000 cross-linked protein. Detailed time courses carried out at different temperatures demonstrated that the Mr 75,000-85,000 affinity-labelled band was the earliest cross-linked complex detected in Swiss 3T3 cells incubated with 125I-labelled gastrin-releasing peptide (125I-GRP). Furthermore, the ability of various nonradioactive bombesin agonists and antagonists to block the formation of the Mr 75,000-85,000 cross-linked complex correlated extremely well (r = 0.994) with the relative capacity of these peptides to inhibit 125I-GRP specific binding. Pretreatment with unlabelled GRP for up to 6 h caused only a slight decrease in both specific 125I-GRP binding and the affinity labelling of the Mr 75,000-85,000 protein. We also show that the cross-linked complex is a glycoprotein. First, solubilized affinity labelled Mr 75,000-85,000 complex applied to wheat germ lectin-sepharose columns was eluted by addition of 0.3 M N-acetyl-D-glucosamine. Second, treatment with endo-beta-N-acetylglucosaminidase F reduced the apparent molecular weight of the affinity-labelled band from 75,000-85,000 to 43,000, indicating the presence of N-linked oligosaccharide groups.

• Research Organization: Imperial Cancer Research Fund, London (England)
• OSTI ID: 6101675
• Journal Information: J. Cell. Biochem.; (United States), Journal Name: J. Cell. Biochem.; (United States) Vol. 38:4; ISSN JCEBD
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AFFINITY
ANIMALS
BETA DECAY RADIOISOTOPES
BIOCHEMICAL REACTION KINETICS
CELL CONSTITUENTS
CELL CULTURES
CELL MEMBRANES
CHEMICAL REACTIONS
CROSS-LINKING
DAYS LIVING RADIOISOTOPES
ELECTRON CAPTURE RADIOISOTOPES
ENZYMES
GLYCOPROTEINS
GLYCOSYL HYDROLASES
HYDROLASES
INTERMEDIATE MASS NUCLEI
IODINE 125
IODINE ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
MAMMALS
MEMBRANE PROTEINS
MEMBRANES
MICE
MOLECULAR WEIGHT
NUCLEI
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
PEPTIDES
POLYMERIZATION
PROTEINS
RADIOISOTOPES
REACTION KINETICS
RECEPTORS
RODENTS
TRACER TECHNIQUES
VERTEBRATES