Specific binding and degradation of somatostatin by membrane vesicles from pig gut

Plasma membrane vesicles were prepared from the basolateral face of pig small intestinal epithelial cells and were enriched in the activity of Na⁺-K⁺-ATPase (9-fold relative to the cell homogenate) and ranged in size from 0.15 to 0.40 μm diam. Incubation of somatostatin-14 and [¹²⁵I-Tyr¹¹]-somatostatin-14 with the vesicles at 37°C resulted in rapid proteolytic degradation of the peptides. Metabolites were isolated by reverse-phase high-performance liquid chromatography and identified by amino acid composition. Cleavages between Ala¹-Gly², Phe⁶-Phe⁷-Trp⁸, and Thr¹⁰-Phe¹¹ were observed, indicative of aminopeptidase and endopeptidase action. Degradation was inhibited by 1,10-phenanthroline and by bacitracin, and in the presence of these inhibitors and at 21°C binding of [¹²⁵I-Tyr¹¹]somatostatin-14 to the vesicles was observed. Binding was inhibited in a concentration-dependent manner by somatostatin-14 (half-maximal inhibition at 2.0 ± 0.1 nM) and by somatostatin-28 (0.8 ± 0.1 nM) but not by structurally unrelated peptides. The rate of degradation of [¹²⁵I-Leu⁸, D-Trp²², Tyr²⁵]somatostatin-28 by basolateral membrane was < 20 fold that of [¹²⁵I-Tyr¹¹]somatostatin-14 and a two- to three-fold enhanced binding to the vesicles was observed. Analysis of the inhibition of binding of this analogue by somatostatin-28 indicates the presence of single class of binding site with K(d) = 1.3 ± 0.3 nM. Rapid degradation but no specific binding of somatostatin-14 by brush-border membranes was observed. Binding of somatostatins by epithelial cell membranes may be related to the regulation of nutrient absorption by the gut, but the rapid rate of degradation suggests that binding may also serve to limit the release of the peptides into the circulation.