Overlapping binding sites drive allosteric agonism and positive cooperativity in type 4 metabotropic glutamate receptors

Abstract

Type 4 metabotropic glutamate (mGlu 4 ) receptors are emerging targets for the treatment of various disorders. Accordingly, numerous mGlu 4 -positive allosteric modulators (PAMs) have been identified, some of which also display agonist activity. To identify the structural bases for their allosteric action, we explored the relationship between the binding pockets of mGlu 4 PAMs with different chemical scaffolds and their functional properties. By use of innovative mGlu 4 biosensors and second-messenger assays, we show that all PAMs enhance agonist action on the receptor through different degrees of allosteric agonism and positive cooperativity. For example, whereas VU0155041 and VU0415374 display equivalent efficacies [log(t B ) = 1.15 6 0.38 and 1.25 6 0.44, respectively], they increase the ability of L-AP4 to stabilize the active conformation of the receptor by 4 and 39 times, respectively. Modeling and docking studies identify 2 overlapping binding pockets as follows: a first site homologous to the pocket of natural agonists of class A GPCRs linked to allosteric agonism and a second one pointing toward a site topographically homologous to the Na + binding pocket of class A GPCRs, occupied by PAMs exhibiting the strongest cooperativity. These results reveal that intrinsic efficacy and cooperativity of mGlu 4 PAMs are correlated with their binding mode, and vice versa, integrating structural and functional knowledge from different GPCR classes.-Rovira, X., Malhaire, F., Scholler, P., Rodrigo, J., Gonzalez-Bulnes, P., Llebaria, A., Pin, J.-P., Giraldo, J., Goudet, C. Overlapping binding sites drive allosteric agonism and positive cooperativity in type 4 metabotropic glutamate receptors. FASEB J. 29, 116-130 (2015). www.fasebj.org

Allosteric Binding Site and Activation Mechanism of Class C G-Protein Coupled Receptors: Metabotropic Glutamate Receptor Family

“…5a to c, the three PAMs are located closely to the hydroxyl in Ser760 5.47 and Ser825 7.35 . So the hydrogen acceptor groups in the PAMs formed two hydrogen bonds with Ser760 5.47 and , respectively, indicating the importance of these two residues, which was consistent with former reports (18,26,64). Other residues also contributed to the binding and selectivity due to their hydrophobic nature, including (selective residues).…”

Allosteric modulation of metabotropic glutamate receptors by chloride ions

“…Interestingly, mGlu2 oligomers can transiently form if in their active state. Although the latter were too transient to detect FRET between dimers, their close contact could be trapped by cysteine cross-linking (38). Further studies are needed to explore these hypotheses and assess more precisely cooperativity phenomena in mGluRs.…”

“…This method allows the analysis of the effect of orthosteric and allosteric ligands on the receptor, notably the ability of PAMs to potentiate the action of agonists on the stabilization of the active conformation of the VFT dimer (38).…”

Molecular Insights into Metabotropic Glutamate Receptor Allosteric Modulation

“…Before this, researchers relied on family A 7TM templates to perform homology modeling and interpret site-directed mutagenesis data. Despite low sequence homology (,20%), these approaches proved fruitful, localizing allosteric binding pockets of mGlu 1 , mGlu 2, mGlu 4 , and mGlu 5 to a region analogous to that of the biogenic amine binding sites in family A GPCRs (Ott et al, 2000;Pagano et al, 2000;Malherbe et al, 2003a,b;Lundstrom et al, 2011;Molck et al, 2012;Gregory et al, 2013bGregory et al, , 2014Rovira et al, 2015). In light of the new crystal structures, although alignments and secondary structure predictions of TMs 5-7 deviated from that observed in the mGlu 1 and mGlu 5 structures, the general localization of the pocket was consistent with earlier reports.…”

“…Across the four subtypes multiple residues have repeatedly been reported as contributing to allosteric modulation. For all four subtypes, residues in positions 3.36, 5.43, and 6.48 are crucial for both positive and negative allosteric modulators (Pagano et al, 2000;Malherbe et al, 2003aMalherbe et al, , 2006Muhlemann et al, 2006;Fukuda et al, 2009;Lundstrom et al, 2011;Gregory et al, 2012Gregory et al, , 2013bGregory et al, , 2014Molck et al, 2012;Turlington et al, 2014;Wu et al, 2014;Rovira et al, 2015). Position 7.45 has also been implicated in mGlu 4 and mGlu 5 allosteric modulation (Molck et al, 2012;Gregory et al, 2013bGregory et al, , 2014Turlington et al, 2014;Rovira et al, 2015); there are no reports investigating the influence of this amino acid on selective allosteric modulators of mGlu 1 or mGlu 2 .…”