Fig. 2. ZP3780 binding and activation at GCGR.

(A) The N terminus of ZP3780 (red) is required for full ligand efficacy and penetrates deep in the receptor core to make H-bonds (dotted lines) with residues in TM1, TM3, TM7, and ECL3 (cyan). (B) The difference in receptor recognition by full-agonist ZP3780 (red) and partial-agonist NNC1702 (dark blue) that lacks H1 and has a D9E mutation is shown. NNC1702-bound GCGR is shown in orange (PDB 5YQZ) (17), and the ZP3780-bound GCGR-G_s complex structure is shown in cyan. The polar interactions are shown as dotted lines colored according to the GCGR structures bound to the two ligands, respectively (ZP3780, cyan; NNC1702, orange). (C) Comparison of the structures of the partial agonist NNC1702 (blue)–bound GCGR (orange, PDB 5YQZ) (17) and the full-agonist ZP3780 (red)–bound GCGR (cyan) reveal conformational differences in ECL3 and TM5, TM6, and TM7. The presence of H1 in ZP3780 ensures interaction with Q232^3.37 and may induce rearrangement of residues in TM5. The interaction of D9 seems to stabilize TM7 and ECL3 displacement, which might trigger GCGR activation. (D) Mutation of Q232^3.37, D370^ECL3, R378^7.35, and D385^7.42 to alanine has a large effect on GCGR-mediated cAMP signaling. All mutants were expressed to similar levels as that of the WT receptor. For (D), data represent mean ± SEM from at least three independent experiments, performed in triplicates. Superscripts are Wootten numbering.