This is a proposal to examine the physiological role of cholecystokinin (CCK)-58, the main circulating form of CCK. The applicant and his coworkers have succeeded recently in synthesizing canine and rat CCK-58 and have found that rat CCK-58 is substantially more potent than CCK-8 for stimulation of pancreatic enzyme and fluid secretion in rats. NMR analysis shows that CCK-58 has a different solution conformation than CCK-8 and that the conformation is stable enough to allow CCK-58 to bind more potently to CCK-A receptors on pancreatic acinar cells. Based on these results, the PI has formulated a hypothesis whereby processing of CCK-58 to CCK-8 changes the conformation of the active carboxyl terminus, which in turn modifies the intracellular signals and results in altered bioactivity. The following studies are proposed to test this hypothesis: (1) establish an assay capable of detecting functional differences in the conformation of the carboxyl terminus of CCK-58; (2) demonstrate that the conformation of the carboxyl terminus of CCK-58 is stabilized by interactions between this and other regions of CCK-58; and (3) examine how the stable conformation of the carboxyl terminus of CCK-58 alters the expression of its bioactivity.
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