Abstract
Metabotropic glutamate (mGlu) receptors play a crucial role in synaptic transmission through homodimeric or heterodimeric assemblies. Despite their dimeric nature, only one subunit within the mGlu dimer engages with G proteins during activation, and the biased activation can be further controlled by allosteric modulators. Considering the related molecular mechanisms remain elusive, we employed Gaussian accelerated molecular dynamics (GaMD) simulations to investigate the regulated mechanisms in mGlu(2)-mGlu(4) heterodimers. Our results demonstrate that the G(i) protein exhibits a higher binding affinity for mGlu(4) compared to mGlu(2) within the mGlu(2)-mGlu(4) heterodimer. Meanwhile, when the positive allosteric modulator (PAM) binds to G(i)-coupled subunits-whether mGlu(2) or mGlu(4)-it can enhance the binding affinity between the G(i) protein and the subunits of the mGlu(2)-mGlu(4) heterodimer. However, if the PAM binds to mGlu(2) while the G(i) protein is coupled to mGlu(4), the binding affinity may be reduced. Additionally, our results highlight the crucial role of the ICL2 region and the perturbation of the residue-residue coupling network involved in the regulatory pathways in mediating the PAM-induced modulation of G(i) protein preference. In conclusion, these findings provide novel insights into the molecular mechanism underpinning the G(i) protein's preference for mGlu(4) within the mGlu(2)-mGlu(4) heterodimers and the regulatory influence of PAM on G(i) protein binding, advancing our understanding of their functional mechanisms.