Molecular Determinant Underlying Selective Coupling of Primary G-Protein by Class A GPCRs

G-protein-coupled receptors (GPCRs) transmit downstream signals predominantly via G-protein pathways. However, the conformational basis of selective coupling of primary G-protein remains elusive. Histamine receptors H₂R and H₃R couple with G_s- or G_i-proteins respectively. Here, three cryo-EM structures of H₂R-G_s and H₃R-G_i complexes are presented at a global resolution of 2.6-2.7 Å. These structures reveal the unique binding pose for endogenous histamine in H₃R, wherein the amino group interacts with E206^5.46 of H₃R instead of the conserved D114^3.32 of other aminergic receptors. Furthermore, comparative analysis of the H₂R-G_s and H₃R-G_i complexes reveals that the structural geometry of TM5/TM6 determines the primary G-protein selectivity in histamine receptors. Machine learning (ML)-based structuromic profiling and functional analysis of class A GPCR-G-protein complexes illustrate that TM5 length, TM5 tilt, and TM6 outward movement are key determinants of the G_s and G_i/o selectivity among the whole Class A family. Collectively, the findings uncover the common structural geometry within class A GPCRs that determines the primary G_s- and G_i/o-coupling selectivity.