Importance of the C-terminal phenylalanine of gastrin for binding to the human CCK2 receptor

The importance, of the C-terminal Phe of gastrin and structural requirement s at position 17 for binding to the human CCK2 receptor were assessed using analogs of [Leu(15)]G(11-17). The following peptides were synthesized, Ac[ Leu(15)]G(11-17), Ac[Leu(15)]G(11-16)NH2, [Leu(15)]G(11-17), [Leu(15),Ala(1 7)]G(11-17), [Leu(15),Abu(17)]G(11-17), [Leu(15),Val(17)]G(11-17), [Leu(15) Leu(17)]G(11-17), [Leu(15),Cha(17)]G(11-17), [Leu(15),Trp(17)]G(11-17), [Le u(15),Tic(17)]G(11-17), [Leu(15), D-Phe(17)]G(11-17) and [Leu(15),p-X-Phe(1 7)]G(11-17), where X=F, Cl, Br, I, OH, CH3, NH2 and NO2. Competition bindin g experiments with [H-3]CCK-8 were performed using human CCK2 receptors sta bly expressed in CHO cells. Phe was shown to be important for binding. A hy drophobic side-chain larger than Leu is required at position 17 but aromati city does not appear to, be essential. Constraint of the aromatic side-chai n either in the g(+) or g(-) conformation, as in the case of Tic, results i n a significant decrease in affinity. In addition, the peptide conformation induced by incorporation Of D-Phe decreases binding. The size and electron withdrawing/donating properties of the para substituent are not important for interaction with the receptor. The current study shows that the use of des-Phe analogs of gastrin is not a viable strategy for development of anta gonists for the human CCK2 receptor.