Abstract
Activation of free fatty acid receptor 1 (GPR40) by synthetic partial and full agonists occur via distinct allosteric sites. A crystal structure of GPR40-TAK-875 complex revealed the allosteric site for the partial agonist. Here we report the 2.76-Å crystal structure of human GPR40 in complex with a synthetic full agonist, compound 1, bound to the second allosteric site. Unlike TAK-875, which acts as a Gαq-coupled partial agonist, compound 1 is a dual Gαq and Gαs-coupled full agonist. compound 1 binds in the lipid-rich region of the receptor near intracellular loop 2 (ICL2), in which the stabilization of ICL2 by the ligand is likely the primary mechanism for the enhanced G protein activities. The endogenous free fatty acid (FFA), γ-linolenic acid, can be computationally modeled in this site. Both γ-linolenic acid and compound 1 exhibit positive cooperativity with TAK-875, suggesting that this site could also serve as a FFA binding site.
Highlights
Activation of free fatty acid receptor 1 (GPR40) by synthetic partial and full agonists occur via distinct allosteric sites
Hypothesizing functional and structural similarities between endogenous free fatty acid (FFA) and the literature GPR40 agonists led to the discovery of benzofurane acid derivatives as potent and highly selective GPR40 A2 agonists with a unique pharmacology (Supplementary Discussion). compound 1 does not bind the A1 site as TAK-875; but rather, it competes with AM-1638 for the A2 site (Fig. 1a)
The absence of increased insulin levels by compound 1 in GPR40 knock-out (KO) mice indicates that the ability of compound 1 to increase insulin plasma level is GPR40-mediated (Fig. 2c). compound 1 exhibits similar dose-dependent reduction of blood glucose in oral glucose tolerance test (OGTT) that is GPR40-mediated (Fig. 2e–g)
Summary
Activation of free fatty acid receptor 1 (GPR40) by synthetic partial and full agonists occur via distinct allosteric sites. A crystal structure of GPR40-TAK-875 complex revealed the allosteric site for the partial agonist. We report the 2.76-Å crystal structure of human GPR40 in complex with a synthetic full agonist, compound 1, bound to the second allosteric site. AM-1638 was discovered among a novel series of agonists that exerts full agonism on GPR40 and stimulates incretin secretion[15] This class of full agonists acts allosterically with the endogenous FFAs, but binds to a topographically distinct site from TAK-87516. We report the 2.76-Å crystal structure of human GPR40 complexed with compound 1 bound in a second structurally distinct allosteric site (A2), located at the receptor side facing the membrane lipophilic environment. Positive functional cooperativity is observed between TAK-875 and compound 1 as the reported TAK-875 activity augmentation by γ-linolenic acid (γ-LA)13. γ-LA can be modeled into the A2 site using docking and free energy calculations, raising the possibility that site A2 could serve as a free fatty acid binding site
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