Functional characterization of five different PRXamide receptors of the red flour beetle Tribolium castaneum with peptidomimetics and identification of agonists and antagonists.

The neuropeptidergic system in insects is an excellent target for pest control strategies. One promising biorational approach is the use of peptidomimetics modified from endogenous ligands to enhance biostability and bioavailability. In this study, we functionally characterized five different G protein-coupled receptors in a phylogenetic cluster, containing receptors for PRXamide in the red flour beetle Tribolium castaneum, by evaluating a series of 70 different peptides and peptidomimetics. Three pyrokinin receptors (TcPKr-A, -B, and -C), cardioacceleratory peptide receptor (TcCAPAr) and ecdysis triggering hormone receptor (TcETHr) were included in the study. Strong agonistic or antagonistic peptidomimetics were identified, and included beta-proline (β(3)P) modification of the core amino acid residue proline and also a cyclo-peptide. It is common for a ligand to act on multiple receptors. In a number of cases, a ligand acting as an agonist on one receptor was an efficient antagonist on another receptor, suggesting complex outcomes of a peptidomimetic in a biological system. Interestingly, TcPK-A was highly promiscuous with a high number of agonists, while TcPK-C and TcCAPAr had a lower number of agonists, but a higher number of compounds acting as an antagonist. This observation suggests that a target GPCR with more promiscuity will provide better success for peptidomimetic approaches. This study is the first description of peptidomimetics on a CAPA receptor and resulted in the identification of peptidomimetic analogs that demonstrate antagonism of CAPA ligands. The PRXamide receptor assays with peptidomimetics provide useful insights into the biochemical properties of receptors.