Extended Data Fig. 7: Interaction modes of DHEA with ADGRG2.
From: Structures of the ADGRG2–Gs complex in apo and ligand-bound forms
a, Schematic diagrams showing the sequence features of the ADGRG2 constructs used for the reconstitution of DHEA-ADGRG2-DNGs complex. The N-terminal Stachel sequence truncated ADGRG2-β subunit construct (ADGRG2-βT) was used. The C-tail was replaced with that of GPR120. b, Representative concentration dependent curve of the DHEA-induced cAMP accumulation in HEK293 cells overexpressing ADGRG2-βT or control (pcDNA3.1) using Glosensor assay. Values are mean ± SEM from three independent experiments performed in triplicates (n = 3). c-d, Orthogonal views of and structural models for the DHEA-ADGRG2-βT-120CT-DNGs-Nb35 (c) and DHEA-ADGRG2-AA-120CT-miniGs-Nb35 (d) complexes. The DHEA densities were highlighted with red dashed squares. e, Fitting of the DHEA (Right panel) or CHS (Left panel) into the EM density of the ligand binding pocket in the DHEA-ADGRG2-βT-DNGs complex structure. The DHEA can be fit into the EM density while CHS cannot. f, Representative concentration dependent curve of the DHEA-induced cAMP accumulation in ADGRG2-βT-overexpressed HEK293 cells in the absence or presence of 100 μM CHS. Values are mean ± SEM from three independent experiments performed in triplicates (n = 3). g, Diagram of DHEA interactions in the ligand binding pocket of ADGRG2. Hydrogen bonds are depicted as red dashed lines. h, Assignment of DHEA (yellow) in the ligand binding pocket of ADGRG2 (green). DHEA could be fit into the density in two conformational states. In the ‘upper’ state, DHEA forms a hydrogen bond (red dashed line) with a water molecule via its 3-hydroxyl group. i, Superposition of the structures of DHEA-ADGRG2-FL and DHEA-ADGRG2-βT complexes indicate that DHEA adopt a similar pose in the orthosteric ligand pocket of ADGRG2. For clarity, only the vertical cross section of ADGRG2-FL was shown. DHEA in DHEA-ADGRG2-FL is pink while in DHEA-ADGRG2-βT is yellow. j, Clear EM densities enabled unambiguous assignment of DHEA, a water molecule, and surrounding residues in the ligand binding pocket of ADGRG2-FL. The hydrogen bond formed between DHEA and water molecule is shown in red dashed line.
