Dynamic residue communication networks underlie signal initiation at the mu-opioid receptor

The Mu-opioid receptor mediates physiological analgesia through endogenous opioid peptides, yet how structurally similar peptides generate distinct signaling outputs remains unclear. Here, we integrated computational and cellular approaches to resolve peptides-receptor interactions across structural and dynamic hierarchies. Although endogenous opioid peptides occupied a conserved orthosteric binding pocket, they differentially remodeled local hydrogen-bond interactions and intrareceptor dynamic coordination during signal propagation. These differences were further amplified at the network level, giving rise to distinct residue communication architectures associated with signal initiation. Mechanistically, D149^3.32 and W320^7.35 emerged as critical communication hubs that maintain network integrity required for efficient Gi-mediated cAMP signaling. Together, our findings reveal a hierarchical mechanism in which conserved peptide recognition is functionally separated from activation-specific network reorganization, providing a preliminary structural and dynamic framework for understanding ligand-specific signaling initiation in G protein-coupled receptors. It should be noted, however, that these computational predictions are based on 100 ns MD simulations and require further validation with extended timescales.