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

Abstract

Metabotropic glutamate receptors (mGluR) are mainly expressed in the central nervous system (CNS) and contain eight receptor subtypes, named mGluR1 to mGluR8. The crystal structures of mGluR1 and mGluR5 that are bound with the negative allosteric modulator (NAM) were reported recently. These structures provide a basic model for all class C of G-protein coupled receptors (GPCRs) and may aid in the design of new allosteric modulators for the treatment of CNS disorders. However, these structures are only combined with NAMs in the previous reports. The conformations that are bound with positive allosteric modulator (PAM) or agonist of mGluR1/5 remain unknown. Moreover, the structural information of the other six mGluRs and the comparisons of the mGluRs family have not been explored in terms of their binding pockets, the binding modes of different compounds, and important binding residues. With these crystal structures as the starting point, we built 3D structural models for six mGluRs by using homology modeling and molecular dynamics (MD) simulations. We systematically compared their allosteric binding sites/pockets, the important residues, and the selective residues by using a series of comparable dockings with both the NAM and the PAM. Our results show that several residues played important roles for the receptors' selectivity. The observations of detailed interactions between compounds and their correspondent receptors are congruent with the specificity and potency of derivatives or compounds bioassayed in vitro. We then carried out 100 ns MD simulations of mGluR5 (residue 26-832, formed by Venus Flytrap domain, a so-called cysteine-rich domain, and 7 trans-membrane domains) bound with antagonist/NAM and with agonist/PAM. Our results show that both the NAM and the PAM seemed stable in class C GPCRs during the MD. However, the movements of "ionic lock," of trans-membrane domains, and of some activation-related residues in 7 trans-membrane domains of mGluR5 were congruent with the findings in class A GPCRs. Finally, we selected nine representative bound structures to perform 30 ns MD simulations for validating the stabilities of interactions, respectively. All these bound structures kept stable during the MD simulations, indicating that the binding poses in this present work are reasonable. We provided new insight into better understanding of the structural and functional roles of the mGluRs family and facilitated the future structure-based design of novel ligands of mGluRs family with therapeutic potential.

Fig. 1

Binding pockets of mGluR family. The binding pocket of a mGluR1 and b mGluR5 in group I. The binding pocket of c mGluR2 and d mGluR3 in group II. The binding pocket of e mGluR4, f mGluR6, g mGluR7, and h mGluR8 in group III

Fig. 2

Detailed interactions between mGluR1 (group I) and its selective NAM. a The detailed interactions between mGluR1 and YM-202074 (K _i = 4.8 ± 0.37 nM), b the detailed interactions between mGluR1 and YM-230888, and c the binding pose of YM-202074 in mGluR5, comparing the selectivity between mGluR1 and mGluR5. Gln660^3.32 and Thr815^7.32 played important roles for the bindings in mGluR1. Val664^3.36, Ser668^3.40, Thr815^7.32, and Ala818^7.35 in mGluR1 played key roles for its selectivity

Fig. 3

Detailed interactions between mGluR5 (group I) and its NAMs and PAMs. a The binding of MTEP (NAM) in mGluR5, b the binding of Basimglurant (NAM) in mGluR5, c the detailed interactions of CDPPB (PAM) in mGluR5, and d the detailed interactions of VU0403602 (PAM) in mGluR5. Several important residues may form hydrogen bonds with the ligands, including Tyr659^3.44, Asn747^5.47, Ser805^7.35, and Ser809^7.39. A region formed by Gly624^2.45, Ile625^2.46, Gly628^2.49, Pro655^3.40, and Ser809^7.39 played important roles for recognizing the active compounds of mGluR5

Fig. 4

Comparisons of the binding modes and the selective residues between mGluR2 and mGluR3 in group II. The binding poses of an analog of BINA in mGluR2/3 a and c and the binding modes of RO-4491533 in mGluR2/3 b and d. Hydrogen bond interactions between the ligands and residues were shown as a dashed red line

Fig. 5

The detailed interactions of three highly selective PAMs: a VUP155041, b ADX88178, and c VU0364770 in the allosteric binding pocket of mGluR4, and d VU0364770 in mGluR6

Fig. 6

The detailed interactions of highly selective PAM a AMN082 in mGluR7 and two potential binding modes (b and c) of selective PAM AZX88178 in mGluR8

Fig. 7

Conformational change of a antagonist-LY-344545 and b agonist-glutamate in the Venus Flytrap domain (VFTD) during 100 ns MD simulation. The structure and residues before MD are highlighted in green, the structure and residues in mGluR5 bound with antagonist and NAM are highlighted in blue, while the structure and residues in mGluR5 bound with agonist and PAM are highlighted in purple

Fig. 8

Conformational changes of mGluR5 (formed by VFTD, CRD, and 7TMD). a The RMSD of 7TMD in mGluR5, b the RMSF of 7TMD in mGluR5, c conformation changes of mavoglurant (NAM), d conformation changes of VU0405386 (PAM), e “ionic lock” in mGluR5-mavoglurant (NAM), f “ionic lock” in mGluR5-VU0405386 (PAM), g conformation comparisons of 7TMD (viewed from intracellular side), and h conformational comparisons of activation-related residues, including Trp785^6.50, Lys821^7.51, and Tyr823^7.53