EVIDENCE THAT CCK-58 HAS STRUCTURE THAT INFLUENCES ITS BIOLOGICAL-ACTIVITY

Many biologically active peptides exist in multiple molecular forms, b ut the functional significance of regions outside the region of bioact ivity is unknown. The biological and immunological data presented in t his study indicate that cholecystokinin-58 (CCK-58), unlike other form s of cholecystokinin, has structure that influences its bioactivity. C CK-58 was purified from acid extracts of canine intestinal mucosa unti l a single absorbance peak was obtained during reverse-phase chromatog raphy Amino acid analysis precisely determined the peptide concentrati ons of purified CCK-58 and synthetic CCK-8. Our hypothesis was that if the amino terminus of CCK-58 influences its bioactivity then its acti vity would be modified when this region was removed from the peptide. To evaluate the importance of the amino terminus of CCK-58 to influenc e its biological activity, the abilities of CCK-58 and CCK-8 to releas e amylase from pancreatic acini were compared before and after tryptic digestion. Tryptic digestion of CCK-58 decreased the half-maximal sti mulation (EC(50)) for amylase release from 96 to 28 pM. The EC(50) for digested CCK-58 was similar to that for CCK-8 (17 pM). These results suggest that CCK-58 has a structure that shields its bioactive carboxy l terminus. This is further supported by the finding that carboxyl fra gments generated from CCK-58 by trypsin or by partial acid hydrolysis were greater than twofold more immunoreactive than the intact CCK-58. The diminished activity of CCK-58 indicates that the amino terminus of CCK-58 shields the carboxyl terminus, which is important to its biolo gical and immunological activities.