Allostery between Distant Structural Regions Dictates Selectivity in GPCR:G Protein Coupling

Despite extensive structural and functional studies, the molecular mechanisms governing G-protein coupled receptor-G (GPCR-G) protein coupling selectivity remain unresolved. Here, using an interpretable machine learning Bayesian Network model with Molecular Dynamics simulations and experiments, we reveal the influence of distant residue communities within the Gα protein core on coupling selectivity. We observed distinct cooperative hotspot residues across different Gα protein subtypes, including key regions such as the N-terminus, h4s6 loop, and H5 helix. These results demonstrate the intricate allosteric dependencies between the core and the H5 helix in stabilizing selective interactions. The functional significance of these cooperative regions is validated through subtype-swapping mutations. By introducing targeted Gαq-like mutations in the Gαs core, we successfully altered the receptor coupling profile to signal through Gαq. Our findings emphasize that cooperative interactions in the Gα core are not only crucial for selectivity but can also be leveraged to engineer Gα proteins with tailored coupling preferences.