Peptide/Receptor Evolution Explains the Lipolytic Function of the Neuropeptide TLQP-21

Functional diversity of peptide substrates and their cognate G-Protein Coupled Receptors (GPCR) are often the result of a long evolutionary process shaped by accumulation of mutations and natural selection. Even small changes in amino acid sequence can drastically alter their binding affinity and receptor activation. Since GPCR are often investigated as molecular targets for pharmacotherapies, it is fundamentally essential to experimentally compare peptide/receptor interaction when translating promising pre-clinical therapeutic efficacy into humans. Here, we focused on the role of the Complement 3a Receptor (C3aR1) activated by the VGF-derived neuropeptide TLQP-21 in rodent and human cell system, and used molecular modelling and evolutionary analysis to infer the exact mechanism of ligand/receptor interaction. After having characterized the mechanism of TLQP-21 potentiation of beta-Adrenergic Receptor (bARs)-induced lipolysis in rodent adipocytes, we used structural data-driven homology modeling and molecular dynamics simulation to identify the TLQP-21 binding motif and its corresponding C3aR1 binding site. We found that the mouse/rat TLQP-21 had enhanced binding affinity at the human C3aR1, when compared to the human TLQP-21. Similarly, the mouse/rat TLQP-21, but not the human TLQP-21, potentiate lipolysis in human adipocytes. This surprising finding led us to reconstruct the evolution of sequence motifs within the C3aR1 binding site and the TLQP-21 pharmacophore in the Murinae sub-family of rodents, to identify changes which explain the pharmacological efficacy by virtue of enhanced hydrophobicity, and which represent stark enhancement of function for a neuropeptide that is thus rendered a potent agonist at C3aR1. Since we further demonstrated that changes in gene expression of critical nodes in TLQP-21/C3aR1-mediated lipolysis are conserved in obesity for humans and mice, and that the rodent peptide is a potent agonist at the human C3aR1, our discovery can pave the way to investigate a peptide/receptor mechanism and to develop novel pharmacotherapies for obesity and other disease conditions in which C3aR1 has been implicated.