Direct cross-linking of 125I-labeled glucagon to its membrane receptor by UV irradiation.
125I-labeled glucagon was directly crosslinked to its receptor in isolated liver plasma membranes and on the surface of intact hepatocytes, by using a UV irradiation procedure. This investigation resulted in the identification of a glucagon-receptor complex of apparent Mr 62,000. The specificity of labeling was shown by the interference of unlabeled hormone at physiological concentration with incorporation of radioactive glucagon into the 62,000 Mr species. The receptor behaved as a typical integral membrane protein: it was not released by extraction with lithium diiodosalicylate or at basic pH but was solubilized by digitonin treatment. Reduction of the receptor polypeptide with dithiothreitol resulted in a decrease in its electrophoretic mobility, suggesting the presence of intramolecular disulfide bonds. Soluble glucagon-receptor complexes adsorbed to Con A-Sepharose and could be eluted with methyl alpha-D-mannoside, indicating that the receptor molecule is a glycoprotein. Treatment of glucagon-labeled liver plasma membrane with endoglycosidase F resulted in the appearance of four intermediate species, indicating that glucagon receptor contains at least four N-linked oligosaccharide chains.
Iwanij
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D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D004229:Dithiothreitol / Q000494:pharmacology; D005934:Glucagon / Q000378:metabolism / Q000528:radiation effects*; D008099:Liver / Q000378:metabolism; D008297:Male; D008970:Molecular Weight; D051381:Rats; D011956:Receptors, Cell Surface / Q000378:metabolism / Q000528:radiation effects*; D018027:Receptors, Glucagon; D014466:Ultraviolet Rays
D011956:Receptors, Cell Surface; D018027:Receptors, Glucagon; D005934:Glucagon; D004229:Dithiothreitol