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Deciphering the Agonist Binding Mechanism to the Adenosine A1 Receptor.

ABSTRACT: Despite being among the most characterized G protein-coupled receptors (GPCRs), adenosine receptors (ARs) have always been a difficult target in drug design. To date, no agonist other than the natural effector and the diagnostic regadenoson has been approved for human use. Recently, the structure of the adenosine A1 receptor (A₁R) was determined in the active, G_i protein complexed state; this has important repercussions for structure-based drug design. Here, we employed supervised molecular dynamics simulations and mutagenesis experiments to extend the structural knowledge of the binding of selective agonists to A₁R. Our results identify new residues involved in the association and dissociation pathway, they suggest the binding mode of N⁶-cyclopentyladenosine (CPA) related ligands, and they highlight the dramatic effect that chemical modifications can have on the overall binding mechanism, paving the way for the rational development of a structure-kinetics relationship of A₁R agonists.

SUBMITTER: Deganutti G

PROVIDER: S-EPMC7887845 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature

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Deciphering the Agonist Binding Mechanism to the Adenosine A1 Receptor.

Deganutti Giuseppe G Barkan Kerry K Preti Barbara B Leuenberger Michele M Wall Mark M Frenguelli Bruno G BG Lochner Martin M Ladds Graham G Reynolds Christopher A CA

ACS pharmacology & translational science 20210121 1

Despite being among the most characterized G protein-coupled receptors (GPCRs), adenosine receptors (ARs) have always been a difficult target in drug design. To date, no agonist other than the natural effector and the diagnostic regadenoson has been approved for human use. Recently, the structure of the adenosine A1 receptor (A<sub>1</sub>R) was determined in the active, G<sub>i</sub> protein complexed state; this has important repercussions for structure-based drug design. Here, we employed sup ...[more]

PMID: 33615181

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Project description:Background and purposeAdenosine may be generated by hydrolysis of extracellular nucleotides by ectonucleotidases, including ectonucleoside triphosphate diphosphohydrolase 1 (CD39), ecto-5'-nucleotidase (CD73), nucleotide pyrophosphatase phosphodiesterase 1 (NPP-1) and tissue non-specific alkaline phosphatase (TNAP). Previous work from our laboratory has uncovered a critical role for adenosine A1 receptors (A1 R) in osteoclastogenesis; blockade or deletion of these receptors diminishes osteoclast differentiation. Interestingly, selective A1 R agonists neither affect basal osteoclastogenesis nor do they reverse A1 R antagonist-mediated inhibition of osteoclastogenesis. In this study, we determined whether ectonucleotidase-mediated adenosine production was required for A1 R antagonist-mediated inhibition, and, when we saw no effect, determined whether A1 R was constitutively activated and the antagonist was acting as an inverse agonist to diminish osteoclast differentiation.Experimental approachOsteoclast formation derived from wild-type, CD39 knockout (KO), CD73 KO, NPP-1 KO and TNAP KO mice was examined by tartrate-resistant acid phosphatase staining of receptor activator of NF-κB ligand-macrophage colony-stimulating factor-stimulated osteoclasts and osteoclast gene expression (Ctsk, Acp5, MMP-9 and NFATc1). Intracellular cAMP concentration was determined by elisa.Key resultsRolofylline inhibited osteoclast formation in a dose-dependent manner (IC50 = 20-70 nM) in mice lacking all four of these phosphatases, although baseline osteoclast formation was significantly less in precursors from CD73 KO mice. Rolofylline (1 μM) stimulates cAMP production in bone marrow macrophages by 10.23 ± 0.89-fold.Conclusions and implicationsBased on these findings, we hypothesize that the A1 R is constitutively activated in osteoclast precursors, thereby diminishing basal AC activity, and that A1 R antagonists act as inverse agonists to release A1 R-mediated inhibition of basal AC activity and permit osteoclast differentiation. The constitutive activity of A1 R promotes osteoclast formation and down-regulation of this activity blocks osteoclast formation.

Dataset Information

Deciphering the Agonist Binding Mechanism to the Adenosine A1 Receptor.

Publications

Deciphering the Agonist Binding Mechanism to the Adenosine A1 Receptor.

Similar Datasets

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