Abstract

Abstract Activation of FFA1 (GPR40), a member of GPCR family A, is mediated by medium and long chain fatty acids and leads to amplification of glucose stimulated insulin secretion, suggesting a potential role for FFA1 as a target for type 2 diabetes. It has been previously assumed that there is a single binding site for fatty acids and synthetic FFA1 agonists. However, using members of two chemical series of partial and full agonists that have been identified, radioligand binding interaction studies revealed that the full agonists do not bind to the same site as the partial agonists but exhibit positive heterotropic cooperativity. Analysis of functional data reveals positive functional cooperativity between the full agonists and partial agonists in various functional assays (in vitro and ex vivo) and also in vivo. Furthermore, the endogenous fatty acid, docosahexaenoic acid (DHA) shows negative or neutral cooperativity with members of both series of agonists in binding assays but displays positive cooperativity in functional assays. Another synthetic agonist is allosteric with members of both agonist series, but apparently competitive with DHA. Therefore there appear to be three allosterically-linked binding sites on FFA1 with agonists specific for each of these sites. Activation of FFAR1 by each of these agonists is differentially affected by mutations of two arginine residues, previously found to be important of FFAR1 binding and activation. The high potencies of these ligands and their strong positive functional cooperativity with endogenous fatty acids, demonstrated in vitro and in vivo, has the potential to deliver therapeutic benefits.