Abstract
The classical model of G protein-coupled receptor (GPCR) activation has evolved from a simple two-state toggle to a complex landscape of multiple active conformations. Ligand-biased agonism, or functional selectivity, describes the ability of specific ligands to stabilize distinct receptor conformations that preferentially engage one intracellular effector (e.g., G proteins) over another (e.g., β-arrestins). This review synthesizes recent structural breakthroughs, primarily driven by high-resolution cryo-electron microscopy (cryo-EM) and advanced molecular dynamics, to elucidate the molecular determinants of coupling specificity. We examine how subtle variations in the orthosteric binding pocket are transduced through conserved motifs—such as the DRY, NPxxY, and PIF motifs—to the intracellular coupling interface. Recent studies on the β2-adrenergic receptor and opioid receptors highlight the role of transmembrane helix 6 (TM6) displacement and intracellular loop (ICL) dynamics in governing effector recruitment. Furthermore, we discuss the integration of the GPCR-IPL score, a multilevel featurization of ligand-interaction patterns, in predicting biased activation. By comparing G-protein-biased and arrestin-biased structural ensembles, we identify key residues that act as 'molecular switches' for signal divergence. These insights provide a roadmap for the rational design of pathway-selective therapeutics with reduced side-effect profiles, moving beyond traditional agonist/antagonist paradigms towards precision pharmacology. The integration of kinetic allostery and conformational plasticity remains central to understanding the spatio-temporal regulation of GPCR signaling in complex biological systems.