Forced unbinding of GPR17 ligands from wild-type and R255I mutant receptor models

Background: GPR17 is a “hybrid” GPCR that responds to two unrelated ligand families, extracellular nucleotides and cysteinyl-LTs. Its in vivo blockade reduces progression of cerebral ischemic damage, highlighting GPR17 as a novel therapeutic target for ischemia. Methods: To explore the binding mode of the ”purinergic” and “leukotrienic” components of the receptor, we study the binding and the forced unbinding of two GPR17 ligands, the endogenous agonist UDP and the receptor antagonist pranlukast, from the wild-type and a mutant (R255I) models of GPR17, by means of docking and molecular dynamics simulations (MD) techniques . Results: MD suggests that GPR17 nucleotides binding pocket is enclosed between the helical bundle and covered by EL2. The driving interaction involves R255 and the phosphate moieties of nucleotides. This hypothesis was also supported by steered MD experiments. These showed that the energy required to unbind UDP was higher for the WT receptor than for R255I. MD showed three possible binding sites for pranlukast. In one of its preferential docking conformation pranlukast places tetrazole group close to R255 and phenyl rings into a subpocket highly conserved among GPCRs. Pulling forces developed to break polar and aromatic interactions of pranlukast were comparable. No significant differences between wild-type and mutant receptors were found for the unbinding of pranlukast from receptor. Conclusion: MD suggest a crucial role for R255 in nucleotides binding for GPR17. Aromatic interactions were likely to play a predominant role in recognition of the leukotrienic ligand pranlukast, suggesting that two different binding sites are present in GPR17.