Exploring the activation pathway and Gi-coupling specificity of the μ-opioid receptor

Understanding the activation mechanism of the mu-opioid receptor (mu-OR) and its selective coupling to the inhibitory G protein (G(i)) is vital for pharmaceutical research aimed at finding treatments for the opioid overdose crisis. Many attempts have been made to understand the mechanism of the mu-OR activation, following the elucidation of new crystal structures such as the antagonist- and agonist-bound mu-OR. However, the focus has not been placed on the underlying energetics and specificity of the activation process. An energy-based picture would not only help to explain this coupling but also help to explore why other possible options are not common. For example, one would like to understand why mu-OR is more selective to G(i) than a stimulatory G protein (G(s)). Our study used homology modeling and a coarse-grained model to generate all of the possible end states of the thermodynamic cycle of the activation of mu-OR. The end points were further used to generate reasonable intermediate structures of the receptor and the G(i) to calculate two-dimensional free energy landscapes. The results of the landscape calculations helped to propose a plausible sequence of conformational changes in the mu-OR and G(i) system and for exploring the path that leads to its activation. Furthermore, in silico alanine scanning calculations of the last 21 residues of the C terminals of G(i) and G(s) were performed to shed light on the selective binding of Gi to mu-OR. Overall, the present work appears to demonstrate the potential of multiscale modeling in exploring the action of G protein-coupled receptors.