Steric control of signaling bias in the immunometabolic receptor GPR84

Biased signaling in G protein-coupled receptors offers therapeutic promise, yet rational design of biased ligands remains challenging due to limited mechanistic understanding. Here, we report a molecular framework for controlling signaling bias at the immunometabolic receptor GPR84. We identified three structurally-matched ligands (OX04529, OX04954, and OX04539) with varying steric profiles that exhibit comparable Gi protein activation but dramatically different β-arrestin recruitment capacities. A high-resolution cryo-EM structure of GPR84-Gi in complex with OX04529, complemented by molecular dynamics simulations and targeted mutagenesis, revealed that steric interactions between ligand substituents and Leu3366.52 and Phe1875.47 indirectly disrupt a critical polar network involving Tyr3326.48, Asn1043.36 and Asn3627.45 essential for β-arrestin recruitment. Based on these insights, we developed a steric-dependent model that enabled rational design of G protein-biased agonists with predictable β-arrestin recruitment profiles. This mechanistic framework provides a blueprint for designing biased agonists with customized signaling profiles at GPR84 and potentially other class A GPCRs.