Structural basis for the tethered peptide activation of adhesion GPCRs

Abstract

Adhesion G-protein-coupled receptors (aGPCRs) are important for organogenesis, neurodevelopment, reproduction and other processes^1-6. Many aGPCRs are activated by a conserved internal (tethered) agonist sequence known as the Stachel sequence^7-12. Here, we report the cryogenic electron microscopy (cryo-EM) structures of two aGPCRs in complex with G_s: GPR133 and GPR114. The structures indicate that the Stachel sequences of both receptors assume an α-helical-bulge-β-sheet structure and insert into a binding site formed by the transmembrane domain (TMD). A hydrophobic interaction motif (HIM) within the Stachel sequence mediates most of the intramolecular interactions with the TMD. Combined with the cryo-EM structures, biochemical characterization of the HIM motif provides insight into the cross-reactivity and selectivity of the Stachel sequences. Two interconnected mechanisms, the sensing of Stachel sequences by the conserved 'toggle switch' W^6.53 and the constitution of a hydrogen-bond network formed by Q^7.49/Y^7.49 and the P^6.47/V^6.47φφG^6.50 motif (φ indicates a hydrophobic residue), are important in Stachel sequence-mediated receptor activation and G_s coupling. Notably, this network stabilizes kink formation in TM helices 6 and 7 (TM6 and TM7, respectively). A common G_s-binding interface is observed between the two aGPCRs, and GPR114 has an extended TM7 that forms unique interactions with G_s. Our structures reveal the detailed mechanisms of aGPCR activation by Stachel sequences and their G_s coupling.