Structure–activity relationship of GPR15L peptide analogues and investigation of their interaction with the GPR15 receptor

Abstract

The 57‐mer full‐length GPR15L(25‐81) peptide has been identified as the principal endogenous agonist of the G protein‐coupled receptor GPR15. Its main activity resides in the C‐terminal 11‐mer GPR15L(71‐81), which has full efficacy but ~40‐fold lower potency than the full‐length peptide. Here, we systematically investigated the structure–activity relationship of GPR15L(71‐81) by truncations/extensions, alanine‐scanning, and N‐ and C‐terminal capping. The synthesized peptide analogues were tested at GPR15 stably expressed in HEK293A cells using a homogenous time‐resolved Förster resonance energy transfer‐based Gi cAMP functional assay. We show that the C‐terminal α carboxyl group and the residues Leu78, Pro75, Val74, and Trp72 are critical for receptor interaction and contribute significantly to the peptide potency. Furthermore, we tested the ability of GPR15L(71‐81), C‐terminally amidated GPR15L(71‐81), and GPR15L(25‐81) to activate the three GPR15 receptor mutants in a bioluminescence resonance energy transfer‐based G protein activation assay. The results demonstrate that the Lys192 and Glu272 residues in GPR15 are important for the potency of the GPR15L peptide. Overall, our study identifies critical residues in the peptide and receptor sequences for future drug design.

Molecular recognition of the atypical chemokine-like peptide GPR15L by its cognate receptor GPR15

“…Its uniqueness lies in the C-terminal region, serving as the activation domain, distinguishing it from traditional chemokines 4 . Specifically, the 11 amino acids at the C-terminus of GPR15L (GPR15L C11 ) have been shown to exhibit potent efficacy in activating GPR15 5 .…”

“…While the full-length ligand demonstrates superior activation potency compared to GPR15L C11 (Supplementary Fig. S4a ), prior research has highlighted the pivotal role of the 11 amino acids located in the C-terminus for receptor recognition and activation 3 , 5 , 15 . We hypothesized that the N-terminal portion might participate in interactions with regions outside the orthosteric pocket of the receptor (pertaining to the interaction between chemokine C-termini and their receptors), potentially influencing ligand activation potency to some extent.…”