Forced unbinding of GPR17 ligands from wild type and R255I mutant receptor models through a computational approach.
BACKGROUND
GPR17 is a hybrid G-protein-coupled receptor (GPCR) activated by two unrelated ligand families, extracellular nucleotides and cysteinyl-leukotrienes (cysteinyl-LTs), and involved in brain damage and repair. Its exploitment as a target for novel neuro-reparative strategies depends on the elucidation of the molecular determinants driving binding of purinergic and leukotrienic ligands. Here, we applied docking and molecular dynamics simulations (MD) to analyse the binding and the forced unbinding of two GPR17 ligands (the endogenous purinergic agonist UDP and the leukotriene receptor antagonist pranlukast from both the wild-type (WT) receptor and a mutant model, where a basic residue hypothesized to be crucial for nucleotide binding had been mutated (R255I) to Ile.


RESULTS
MD suggested that GPR17 nucleotide binding pocket is enclosed between the helical bundle and extracellular loop (EL) 2. The driving interaction involves R255 and the UDP phosphate moiety. To support this hypothesis, steered MD experiments showed that the energy required to unbind UDP is higher for the WT receptor than for R255I. Three potential binding sites for pranlukast where instead found and analysed. In one of its preferential docking conformations, pranlukast tetrazole group is close to R255 and phenyl rings are placed into a subpocket highly conserved among GPCRs. Pulling forces developed to break polar and aromatic interactions of pranlukast were comparable. No differences between the WT receptor and the R255I receptor were found for the unbinding of pranlukast.


CONCLUSIONS
These data thus suggest that, in contrast to which has been hypothesized for nucleotides, the lack of the R255 residue doesn't affect the binding of pranlukast a crucial role for R255 in binding of nucleotides to GPR17. Aromatic interactions are instead likely to play a predominant role in the recognition of pranlukast, suggesting that two different binding subsites are present on GPR17.
Department of Pharmacological Sciences, University of Milano, via Balzaretti 9, 20133 Milano, Italy.
Chiara
C
Parravicini
Maria P
MP
Abbracchio
Piercarlo
P
Fantucci
Graziella
G
Ranghino
D019943:Amino Acid Substitution; D001665:Binding Sites; D002867:Chromones / Q000737:chemistry; D019295:Computational Biology; D006801:Humans; D008024:Ligands; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D043562:Receptors, G-Protein-Coupled / Q000737:chemistry* / Q000235:genetics; D014530:Uridine Diphosphate / Q000737:chemistry
D002867:Chromones; C121670:GPR17 protein, human; D008024:Ligands; D043562:Receptors, G-Protein-Coupled; D014530:Uridine Diphosphate; C047681:pranlukast