Gastrin receptor characterization: affinity cross-linking of the gastrin receptor on canine gastric parietal cells.
We applied affinity cross-linking methods to label the gastrin receptor on isolated canine gastric parietal cells in order to elucidate the nature of its chemical structure. 125I-labeled Leu15-gastrin and 125I-labeled gastrin2(-17) bound to intact parietal cells and their membranes with equal affinity, and half-maximal inhibition of binding was obtained at an incubation concentration of 3.2 X 10(-10) M unlabeled gastrin. 125I-gastrin2(-17) was cross-linked to plasma membranes or intact parietal cells by incubation in disuccinimidyl suberate. The membrane pellets were solubilized with or without dithiothreitol and applied to electrophoresis on 7.5% sodium dodecyl sulfate polyacrylamide gels. Autoradiograms revealed a band of labeling at Mr 76,000 and labeling of this band was inhibited in a dose-dependent fashion by addition of unlabeled gastrin to the incubation mixture. Dithiothreitol in concentrations as high as 100 mM did not alter the electrophoretic mobility of the labeled band. After taking into account the molecular weight of 125I-gastrin2(-17), our results suggest that the gastrin receptor on parietal cells is a single protein of Mr 74,000 without disulfide-linked subunits.
D000818:Animals; D001667:Binding, Competitive; D002462:Cell Membrane / Q000378:metabolism; D003432:Cross-Linking Reagents; D004285:Dogs; D005748:Gastric Fundus / Q000166:cytology / Q000378:metabolism*; D005753:Gastric Mucosa / Q000166:cytology / Q000378:metabolism*; D005755:Gastrins / Q000378:metabolism; D007700:Kinetics; D008970:Molecular Weight; D011949:Receptors, Cholecystokinin / Q000302:isolation & purification / Q000378:metabolism*
D003432:Cross-Linking Reagents; D005755:Gastrins; D011949:Receptors, Cholecystokinin