Extended Data Figure 6. The N-terminal ECD of the CTR.

a, Rigid body fitting of the structure of CTR ECD bound to sCT (PDB: 5II0)²² into the corresponding regions of the cryo-EM map revealed additional density (close to residue 130) that may be attributed to glycosylation. b-d, Asp mutation of four consensus glycosylation residues (N28D, N73D, N125D and N130D) reveals little role of glycosylation on cell surface expression (b), determined via a cell surface ELISA to the N-terminal epitope tag. c, Competition radioligand binding studies for sCT in competition with the radiolabelled ligand ¹²⁵I-sCT(8-32) revealed reduced affinity for N130D, and to a lesser extent N125D compared to the WT CTR. d, Concentration response curves for cAMP accumulation for mutant receptors relative to WT show that N130D and to a lesser extent N125D also reduce the potency of sCT in functional experiments. All data are + SEM of five independent experiments, conducted in duplicate or triplicate.