Peptide YY (PYY) not only has distinct sequence homology with neuropeptide Y (NPY) and avian pancreatic polypeptide (APP), but it also exhibits both NPY- and APP-like biological activities. We synthesized two analogues of PYY, A1 and A2, with modified N-terminal regions, and compared their chemical and biological properties to those of PYY and the C-terminal fragment of PYY, (13-36)PYY. This study shows that there is a good correlation between the stability of amphiphilic alpha-helical structure and the biological activity of these peptides. A CD study of (13-36)PYY in mixed H2O and trifluoroethanol (TFE) solutions indicated a significant increase in alpha-helical segments (26-79%) with increasing TFE proportions. Since the fragment (13-36)PYY had potent activity in the rat vas deferens (RVD) assay, the secondary structure is possibly required on the RVD cell surface receptors. The analogues, A1 and A2, were designed to increase the stability of the alpha-helical structures by incorporation of modified N-terminal regions. The CD studies and the RVD assays of A1 and A2, suggest that the amphiphilic alpha-helical structures are stabilized by intramolecular hydrophobic interactions with the N-terminal regions and/or by intermolecular hydrophobic interactions in the self-association process, and subsequently potentiate the activities of the peptides compared to those of PYY and (13-36)PYY.