Received for publication August 29, 2006.
Revised December 12, 2006.
Accepted for publication December 12, 2006.

Amino-pyrrolidine tricarboxylic acids (APTCs) give new insight into group III metabotropic glutamate receptor activation mechanism

Abstract

Like most class-C G-protein coupled receptors, metabotropic glutamate (mGlu) receptors possess a large extracellular domain where orthosteric ligands bind. Crystal structures revealed that this domain, called Venus Fly-trap (VFT), adopts a closed vs. open conformation upon agonist or antagonist binding, respectively. Recently, we described amino-pyrrolidine tricarboxylic acids (APTCs), including FP0429, as new selective group III mGlu agonists. Whereas FP0429 is an almost full mGlu4 agonist, it is a weak and partial agonist of the closely-related mGlu8 subtype. To get more insight into the activation mechanism of mGlu receptors, we aimed at elucidating why FP0429 behaves differently at these two highly homologous receptors, by focusing on two residues within the binding site that differ between mGlu4 and mGlu8. Site-directed mutagenesis of S157 and G158 of mGlu4 into their mGlu8 homologues (Ala) turned FP0429 into a weak partial agonist. Conversely, introduction of Ser and Gly residues into mGlu8 increased FP0429 efficacy. Docking of FP0429 in mGlu4 VFT 3D model helped us characterize the role of each residue. Indeed, mGlu4 S157 seems to have an important role in FP0429 binding, while G158 may allow a deeper positioning of this agonist in the cavity of lobe I, thereby ensuring optimal interactions with lobe-II residues in the fully closed state of the VFT. In contrast, the presence of a methyl group in mGlu8 (Ala instead of Gly) weakens the interactions with the lobe-II residues. This probably results in a less stable or a partially closed form of the mGlu8 VFT, leading to partial receptor activation.

Key words: Metabotropic glutamate, Gi family, Structure-activity relationships and modeling, Func. analysis receptor/ion channel mutants, Mutagenesis/Chimeric approaches

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