The Interaction of Caerulein with the Rat Pancreas. 1. Specific Binding of [3H]Caerulein on Plasma Membranes and Evidence for Negative Cooperativity

Abstract

1. The binding of [3H]caerulein (a stable, biologically active labeled analog of cholecystokininpancreozymin) to semi-purified rat pancreatic plasma membranes was investigated. The binding was dependent on time and temperature, as well as saturable, specific and reversible. This process was pH-dependent and optimal at pH 7.0. Cysteine and serine residues in plasma membranes were of importance for binding. Mg2 ' favored the binding. The acceleration of the dissociation of [3H]caerulein in the presence of an excess of native caerulein suggests that binding was characterized by a negative cooperativity. The fast dissociation state evoked by a high degree of occupancy by caerulein was inhibited by lowering the temperature, by decreasing the pH, or by the presence of wheat germ agglutinin.

The Interaction of Caerulein with the Rat Pancreas. 2. Specific Binding of [3H]Caerulein on Dispersed Acinar Cells

“…p-Chloromercuribenzoate, diisopropylfluorophosphate and glutaraldehyde, three protein-modifying reagents capable of reacting with thiol groups, serine residues, and NH2 groups, respectively, strongly inhibited the binding of [3H]caerulein to the outer surface of pancreatic acinar cells. These effects were comparable to those obtained on pancreatic plasma membranes [5] and occurred despite the presence of 1 "/, bovine serum albumin, which added extra amino acid residues to the incubation medium.…”

“…Increased binding was obtained by lowering the temperature (Fig. l), in sharp contrast with reduced binding observed with pancreatic plasma membranes exposed to similar changes in temperature [5]. Another characteristic of binding which was distinct from that of plasma membranes [5] was the lack of inhibitory effect of phospholipase A sug-gesting that this effect when present is exerted on the exposed internal face of the plasma membrane.…”

“…The source of most of the materials including [3H]caerulein and several unlabeled peptides has been indicated in the preceding paper [5]. Desulfated caerulein, bombesin, the C-terminal nonapeptide of bombesin (nonabombesin), and litorin were generous gifts from Dr R. Fig.…”

“…The interaction of monotritiated [3H]caeruleiii with rat pancreatic plasma membranes has been reported in the preceding paper [5]. A study with dispersed intact acinar cells was undertaken to observe the effect of extramembrane events on caerulein binding, e.g.…”

Study of the states and populations of the rat pancreatic cholecystokinin receptor using the full peptide antagonist JMV 179

“…Glu-Glu-Ala-Tyr(S0,H)-Gly-Trp-Met-Asp-Phe-NH, Heptapeptide of gastrin; (2) octapeptide of cholecystokinin; (3) [Thr28, Ahx31]CCK- (25)(26)(27)(28)(29)(30)(31)(32)(33); (4) JMV 179, (5) JMV 180. The peptide (3) is an agonist, (4) is a full antagonist and (5) has both agonist and antagonist activities.…”