Highly Potent Nociceptin Analog Containing the Arg-Lys Triple Repeat

Abstract One of the structural characteristics of a neuropeptide nociceptin is the existence of Arg-Lys (RK) residues at positions 8–9 and 12–13; both RKs have been suggested to bind to the acidic amino acid cluster in the second extracellular loop of the seven transmembrane domain receptor ORL1. With a design strategy of attempting to obtain an analog that binds more strongly to the receptor's acidic cluster, we synthesized a series of nociceptin analogs in which the RK dipeptide unit was placed at positions 6–7, 10–11, or 14–15 adjacent to the parent RKs. Among these nociceptin analogs containing the RK triple repeat, [Arg-Lys 6–7 ]- and [Arg-Lys 10–11 ]nociceptins exhibited weak activities (6–9 and 60–90% of nociceptin, respectively) both in the receptor binding assay and in the [ 35 S]GTPγS binding functional assay. In contrast, [Arg-Lys 14–15 ]nociceptin was found to be very potent in both assays (3-fold in binding and 17-fold in GTPγS functional assay). [Arg-Lys 14–15 ]nociceptin was the first peptide analog found to be stronger than the parent nociceptin, and structure–activity studies have suggested that the incorporated Arg-Lys 14–15 interacts with either the receptor acidic amino acid cluster or the receptor aromatic amino acid residues.