Solubilization of somatostatin receptors in hamster pancreatic beta cells. Characterization as a glycoprotein interacting with a GTP-binding protein.
Somatostatin receptors of plasma membranes from beta cells of hamster insulinoma were covalently labelled with 125I-[Leu8,D-Trp22,Tyr25]somatostatin-28 (125I-somatostatin-28) and solubilized with the non-denaturing detergent Triton X-100. Analysis by SDS/PAGE and autoradiography revealed three specific 125I-somatostatin-28 receptor complexes with similar molecular masses (228 kDa, 128 kDa and 45 kDa) to those previously identified [Cotroneo, P., Marie, J.-C. & Rosselin, G. (1988) Eur. J. Biochem. 174, 219-224]. The major labelled complex (128 kDa) was adsorbed to a wheat-germ-agglutinin agarose column and eluted by N-acetylglucosamine. Also, the binding of 125I-somatostatin-28 to plasma membranes was specifically inhibited by the GTP analog, guanosine-5'-O-(3-thiotriphosphate) (GTP[S]) in a dose-dependent manner. Furthermore, when somatostatin-28 receptors were solubilized by Triton X-100 as a reversible complex with 125I-somatostatin-28, GTP[S] specifically dissociated the bound ligand to a larger extent from the soluble receptors than from the plasma-membrane-embedded receptors, the radioactivity remaining bound after 15 min at 37 degrees C being 30% and 83% respectively. After pertussis-toxin-induced [32P]ADP-ribosylation of pancreatic membranes, a 41-kDa [32P]ADP-ribose-labelled inhibitory guanine nucleotide binding protein coeluted with the 128-kDa and 45-kDa receptor complexes. The labelling of both receptor proteins was sensitive to GTP[S]. The labelling of the 228-kDa band was inconsistent. These results support the conclusion that beta cell somatostatin receptors can be solubilized as proteins of 128 kDa and 45 kDa. The major labeled species corresponds to the 128-kDa band and is a glycoprotein. The pancreatic membrane contains a 41-kDa GTP-binding protein that can complex with somatostatin receptors.
Marie
J C
JC
Unité 55, Institut National de la Santé et de la Recherche Médicale, Paris, France.
Cotroneo
P
P
de Chasseval
R
R
Rosselin
G
G
D000818:Animals; D002478:Cells, Cultured; D002850:Chromatography, Gel; D006224:Cricetinae; D004591:Electrophoresis, Polyacrylamide Gel; D006160:Guanosine Triphosphate / Q000378:metabolism; D007340:Insulinoma / Q000378:metabolism; D007457:Iodine Radioisotopes; D007515:Islets of Langerhans / Q000378:metabolism; D010190:Pancreatic Neoplasms / Q000378:metabolism; D017981:Receptors, Neurotransmitter / Q000378:metabolism*; D017481:Receptors, Somatostatin; D012995:Solubility; D013004:Somatostatin / Q000378:metabolism*; D053561:Somatostatin-28
D007457:Iodine Radioisotopes; D017981:Receptors, Neurotransmitter; D017481:Receptors, Somatostatin; D013004:Somatostatin; D053561:Somatostatin-28; D006160:Guanosine Triphosphate