Effects of Substitution of Hydrophobic Amino Acids by Tryptophan on Receptor Binding and Biological Activity of Neuropeptide Nociceptin

Nociceptin is a neuropeptide that binds to and activates the opioid receptor-like ORL1 receptor. In order to explore the structural elements necessary for receptor recognition and activation, we designed and synthesized a series of nociceptin analogues, in which nonpolar amino acid residues such as Gly6, Ala7, Ala11, Leu14, and Ala15were substituted respectively by Trp. [Trp6]- and [Trp7]nociceptins exhibited rather weak activities (5—15% of nociceptin), and [Trp11]- and [Trp15]nociceptins also showed reduced activities (40—50%). These results suggested that the space available for these particular residues are relatively restricted. By contrast, the Trp/Leu-substitution at position 14 retained full receptor binding activity, and the resulting [Trp14]nociceptin exhibited an increased biological activity in the functional assay using [35S]GTPγS. This suggested that the receptor residue interacting with nociceptin-Leu14 is the aromatic amino acid.