Fig. 1: Cryo-EM structure of HCAR2–Gi1 signaling complex.

a Cryo-EM density map of HCAR2–Gi1 complex (represented by the 3-HB–bound complex). HCAR2 is depicted in blue, Gαi1 in yellow, Gβ in light pink, Gγ in dark purple, scFv16 in light green. b Cryo-EM densities and models of distinct agonists (Niacin, 3-HB, MK-6892 and compound 9n) from their activated complexes. Niacin in cyan, 3-HB in lavender, MK-6892 in beige, compound 9n in light green. c The compact conformation of the extracellular side of HCAR2 is stabilized by three unique pair disulfide bonds (Cys18^N-term–Cys183^ECL2, Cys19^N-term–Cys266^ECL3, Cys100^3.25–Cys177^ECL2). d Effects of mutations in the three disulfide bonds on Niacin-induced Gi1 dissociation signal as indicated by NanoBiT assay. Bars represent differences in calculated agonist potency (pEC50) for each mutant relative to the wild-type receptor (WT). Data are colored according to the extent of effect. nd not determined; ^****P < 0.0001 (one-way ANOVA followed by Dunnett’s multiple comparison test, compared with the response of WT, the detailed P value for each condition is P < 0.0001, P < 0.0001, P < 0.0001, and P < 0.0001, from left to right. Data are shown as the mean ± SEM from n = 3 independent experiments). Source data are provided as a Source Data file.