Mechanisms Underlying Allosteric Molecular Switches of Metabotropic Glutamate Receptor 5

Abstract

The metabotropic glutamate 5 (mGlu5) receptor is a class C G protein-coupled receptor (GPCR) that is implicated in several CNS disorders making it a popular drug discovery target. Years of research have revealed allosteric mGlu5 ligands showing an unexpected complete switch in functional activity despite only small changes in their chemical structure, resulting in positive allosteric modulators (PAM) or negative allosteric modulators (NAM) for the same scaffold. Up to now, the origins of this effect are not understood, causing difficulties in a drug discovery context. In this work, experimental data was gathered and analyzed alongside docking and Molecular Dynamics (MD) calculations for three sets of PAM and NAM pairs. The results consistently show the role of specific interactions formed between ligand substituents and amino acid side chains that block or promote local movements associated with receptor activation. The work provides an explanation for how such small structural changes lead to remarkable differences in functional activity. While this work can greatly help drug discovery programs avoid these switches, it also provides valuable insight into the mechanisms of class C GPCR allosteric activation. Furthermore, the approach shows the value of applying MD to understand functional activity in drug design programs, even for such close structural analogues.

Allosteric activation of metabotropic glutamate receptor 5

“…In addition, residues enclosing the allosteric site, namely, S654 3.39c and Y659 3.44c , both close to the aromatic substituent causing functional switch of the ligands, change their conformation that leads to an increased and more polar chamber. Interestingly, the role of intrahelical waters has not been investigated in the recent studies (Llinas del Torrent, Casajuana-Martin, Pardo, Tresadern, & P erez-Benito, 2019). We found that the TM bundle becomes less tightly packed and a continuous water channel is formed among the TM helices in the intracellular side below the PAM, analogously to the formation of a water channel upon activation of class A GPCRs (Angel et al, 2009;Yuan et al, 2014).…”

“…The structural and functional importance of these interactions is underlined by several findings. The water molecule mediating the H-bond between Y659 3.44c and T781 6.46c is identified in all available X-ray structures of mGluR5-NAM complexes (Christopher et al, 2015(Christopher et al, , 2019Dor e et al, 2014) and the Hbond was also observed in MD simulations of the mGluR2 (P erez-Benito et al, 2017) and mGluR5 receptors in complex with NAMs (Llinas del Torrent, Casajuana-Martin, et al, 2019). Moreover, it was shown that mutations of Y659 3.44c and T781 6.46c in mGluR5 significantly impact the activity of allosteric ligands by either reducing (Gregory et al, 2014;Malherbe et al, 2006) or inverting (Turlington et al, 2013) their functional effect.…”

“…Previous computational studies on the activation of mGluR receptors include MD simulations for mGluR2 that confirmed the role of the trigger switch amino acids (F643 3.40c , N735 5.47c and W773 6.50c ) in allosteric modulation (Doornbos et al, 2016;P erez-Benito et al, 2017). MD simulations with mGluR5 NAM and PAM complexes revealed the critical role of the transmission switch residues (S658 3.343c , Y659 3.44c and T781 6.46c ) in allosteric activation (Llinas del Torrent, Casajuana-Martin, Pardo, Tresadern, & P erez-Benito, 2019). In a recent MD study, dual binding mode of the investigated PAM was found and although elements of the activation mechanism in comparison with class A GPCRS were analysed, limited differences between the hydration of the allosteric sites of active and inactive receptors were observed (Cong, Ch eron, Golebiowski, Antonczak, & Fiorucci, 2019).…”

Exploring the Activation Mechanism of the mGlu5 Transmembrane Domain

“…by "a" according to Ballesteros-Weinstein scheme (Ballesteros and Weinstein, 1995)] for class A GPCRs or followed by "c" according to Pin et al (2003) scheme for class C GPCRs). Despite previous studies have proposed functional similarities between class A GPCR micro-switches and certain non-homologous class C GPCR residues (Perez-Benito et al, 2017; Llinas Del Torrent et al, 2019) their respective conformational changes have been observed only in MD simulations of mGlu receptor homology models, but not in mGlu receptor crystal structures (Dore et al, 2014;Christopher et al, 2015;Koehl et al, 2019) or in MD simulations starting from those (Feng et al, 2015;Dalton et al, 2017;Perez-Benito et al, 2017;Llinas Del Torrent et al, 2019). Furthermore, while the outward movement of TM6 is a hallmark of class A GPCR activation, the extent of this movement is noticeably lower in mGlu5 even when enhanced sampling methods are employed (Cong et al, 2019).…”

“…Significant efforts have been made to decipher the mGlu5 receptor structure-activity translation machinery (Kaae et al, 2012;Dalton et al, 2014Dalton et al, , 2017Gregory et al, 2014;Gomez-Santacana et al, 2017;Cong et al, 2019;Jojart et al, 2019;Llinas Del Torrent et al, 2019). In general, a delicate moleculargear receptor system whose mechanism of action is not yet well understood has become apparent.…”