Cryo-EM structure of the human Mas receptor reveals N-terminal occlusion of the orthosteric ligand binding pocket

• Cryo-EM structures of human MasR in complex with heterotrimeric Gq are determined at 2.89 Å and 3.08 Å resolution. • The receptor’s own N-terminal peptide (residues 2–11) occludes the orthosteric binding pocket as an endogenous pseudo-ligand. • N-terminal pocket occlusion is structurally stable but dispensable for constitutive Gq coupling. • A conserved Gq-coupling interface at ICL2 is shared across the Mrgpr receptor subfamily. The Mas receptor (MasR) is a class A G protein-coupled receptor (GPCR) that mediates the counter-regulatory arm of the renin–angiotensin system through the ACE2–angiotensin-(1-7)–MasR axis and represents a promising therapeutic target for cardiovascular and metabolic disease. Despite its physiological importance, the structural basis of MasR has remained unknown. Here we report cryo-EM structures of human MasR in complex with heterotrimeric Gq at resolutions of 2.9 Å and 3.1 Å, determined for the full-length receptor and an N-terminally truncated variant (del2-25), respectively. These structures reveal that the receptor’s own N-terminal peptide (residues 2–11) threads into and occludes the orthosteric binding pocket, functioning as an endogenous pseudo ligand. Functional mutagenesis and molecular dynamics simulations demonstrate that this N-terminal cap stably occupies the pocket but is dispensable for constitutive Gq coupling, distinguishing MasR from other N-terminal cap-forming GPCRs. Structural comparison with Mrgpr family members reveals a conserved Gq-coupling interface at the cytoplasmic face alongside divergent extracellular pocket architectures and identifies Y252 6.59 as a structural element that occludes a conserved sub-pocket present in Mrgpr paralogs. Molecular docking simulation of the MasR agonist AR234960 provides a structural template for orthosteric ligand design. Together, these findings establish the structural framework for MasR.