Identification and partial characterization of a somatostatin-14 binding protein on rat liver plasma membranes.
Binding of somatostatin-14 to rat liver plasma membranes was characterized with 125-labeled[tyr11] somatostatin-14. Binding at 24 degrees C reached a plateau at 50 min and was reversible by synthetic somatostatin-14. Scatchard analysis revealed a single class of binding sites (affinity constant = 2.4 +/- 0.2 nmol/L, binding capacity = 148 +/- 0.02 fmol/mg protein). Specificity for somatostatin-14 was demonstrated by the inhibition of 125I-[tyr11]somatostatin-14 binding by biologically active somatostatin analogs but not by a biologically inactive somatostatin analog or unrelated peptides. The radioiodinated binding site complex could be cross-linked with disuccinimidyl suberate. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel autoradiography revealed a 70,000-Da band. Dithiothreitol, a reducing reagent, did not alter the mobility of the band, and the band could be abolished in the presence of 10 mumol/L synthetic somatostatin-14. Covalently cross-linked, iodinated binding protein complexes could be solubilized by the nonreducing detergents Zwittergent 3-12 and 3-([3-cholamidopropyl] diethylammonio)-1-propanesulfonic acid (CHAPS). Solubilized complex bound to wheat-germ agglutinin-agarose columns and was eluted by N,N',N"-triacetylchitotriose. Binding to wheat-germ agglutinin agarose columns was lost after pretreatment with endo-beta-N-acetylglucosaminidase F. Binding studies with liver plasma membranes, 125I-labeled[tyrosine11]somatostatin-14 and guanine nucleotides showed inhibition of binding in the presence of guanine nucleotides. These results indicate that the purified rat liver plasma membranes contain a specific binding protein for somatostatin-14, the binding protein appears to be glycosylated and somatostatin-14 binding to rat liver plasma membranes may be regulated by G proteins.
Raper
S E
SE
Department of Surgery, University of Michigan Medical School, Ann Arbor 48109-0331.
Kothary
P C
PC
DelValle
J
J
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000818:Animals; D002352:Carrier Proteins / Q000032:analysis* / Q000302:isolation & purification; D002462:Cell Membrane / Q000737:chemistry; D002846:Chromatography, Affinity; D006160:Guanosine Triphosphate / Q000494:pharmacology; D008099:Liver / Q000737:chemistry*; D008297:Male; D051381:Rats; D011919:Rats, Inbred Strains; D013004:Somatostatin / Q000378:metabolism*
D002352:Carrier Proteins; D013004:Somatostatin; D000244:Adenosine Diphosphate; D006160:Guanosine Triphosphate