Structure-activity relationships of the crustacean myotropic neuropeptide orcokinin

Orcokinin (OK, NFDEIDRSGFGFN) was recently identified from the crayfish, Orconectes limosus, as a potent hindgut-stimulating factor (14). To assess the importance of structural features of the peptide involved in effective ligand-receptor interactions, synthetic analogues of orcokinin were tested in the hindgut bioassay. Tests with N- and C-terminal-truncated analogues and the C-terminal-amidated analogue (OK-NH2) demonstrate that changes at the C-terminus interfere less with biological activity than changes at the N-terminus. Removal of more than one amino acid at the N-terminus resulted in a complete loss of activity, whereas the C-terminal deletion of three amino acids still produced an analogue with full intrinsic activity but with a drastic shift in threshold concentration of activity from 1 x 10(-10) to 1 x 10(-7) M. Deletion of four amino acids at the C-terminus resulted in a completely inactive analogue. The C-terminal hydroxyl group does not seem to be important because amidation (OK-NH2) resulted in almost no loss of activity. Replacing Arg7 with Ala produced an analogue almost equipotent to orcokinin. Replacement of Phe2 by Tyr resulted in considerable loss of activity. An important role of Phe2 is further suggested by the steep drop of activity after removal of this residue in the N-terminal-deleted analogues.