biostudies-literatureRothman AMKWellcome Trust2603-2620.e18https://www.ebi.ac.uk/biostudies/studies/S-EPMC12087876biostudies-literatureUnknown188(10)Targeting ubiquitin E3 ligases is therapeutically attractive; however, the absence of an active-site pocket impedes computational approaches for identifying inhibitors. In a large, unbiased biochemical screen, we discover inhibitors that bind a cryptic cavity distant from the catalytic cysteine of the homologous to E6-associated protein C terminus domain (HECT) E3 ligase, SMAD ubiquitin regulatory factor 1 (SMURF1). Structural and biochemical analyses and engineered escape mutants revealed that these inhibitors restrict an essential catalytic motion by extending an α helix over a conserved glycine hinge. SMURF1 levels are increased in pulmonary arterial hypertension (PAH), a disease caused by mutation of bone morphogenetic protein receptor-2 (BMPR2). We demonstrated that SMURF1 inhibition CellTherapeutic potential of allosteric HECT E3 ligase inhibition.PMC12087876206632/Z/17/ZOttl JQuigley CRoger JSutton DShaw DBinmahfooz SKKhvalevsky OJarai GGuth-Gundel STroxler TYariv EHemmig RAvishid RThomas MJWeiler SPrag GKordonsky AThaker MHuang SMFlorentin ABonneau OJain RRathi RCobb JSOstermann NRejtar TZhang JHRothman AMKMorrell NWSrivastava SSmith NKleifeld OCotesta SBeckwith REJTallarico JRowlands DJHachey AZink FHumbert MfalseTherapeutic potential of allosteric HECT E3 ligase inhibition.Targeting ubiquitin E3 ligases is therapeutically attractive; however, the absence of an active-site pocket impedes computational approaches for identifying inhibitors. In a large, unbiased biochemical screen, we discover inhibitors that bind a cryptic cavity distant from the catalytic cysteine of the homologous to E6-associated protein C terminus domain (HECT) E3 ligase, SMAD ubiquitin regulatory factor 1 (SMURF1). Structural and biochemical analyses and engineered escape mutants revealed that these inhibitors restrict an essential catalytic motion by extending an α helix over a conserved glycine hinge. SMURF1 levels are increased in pulmonary arterial hypertension (PAH), a disease caused by mutation of bone morphogenetic protein receptor-2 (BMPR2). We demonstrated that SMURF1 inhibition 2025-01-01T00:00:00Z2025 May2026-06-01T06:15:19.159Z2026-04-08T09:50:17.93ZS-EPMC120878764017988510.1016/j.cell.2025.03.001