Extended Data Fig. 8: Cell-based GCaMP assay.
From: Cryo-EM structure of the insect olfactory receptor Orco
a, b, Relative fluorescence changes in HEK293 cells transfected with GCaMP plus wild-type (WT) or mutant Orco alone (a, titrated with VUAA1) or in the presence of A. gambiae OR65 (b, titrated with eugenol). Plots using the same symbols were collected on the same day. c, d, Fitted Hill equation parameters for Orco and Orco–OR65 (mean ± s.d.). The assays were repeated N independent times to obtain the maximum ΔF/Fo response, concentration for half-maximal response (EC50) and Hill coefficient (n). (ΔF/Fo)norm is the fitted maximum ΔF/Fo response relative to a wild-type control Orco or Orco–OR65 experiment from the same plate (for mutations that could not be accurately fit, the measured maximum ΔF/Fo response was used). Two one-way ANOVA tests were performed using these EC50 data followed by Tukey–Kramer multiple comparison tests. (1) Wild-type and mutant Orco homomers (P values are from comparisons to wild-type Orco). (2) Wild-type and mutant Orco co-transfected with OR65 (P values are from comparisons to wild-type Orco with OR65). *Fitted parameters for Orco T465A or S458A homomers + VUAA1 could not be obtained as the dose–response curves did not saturate: a lower-bound estimate for their EC50 is approximately 3 × 10−4 M. **No binding was observed for Orco Y466A in the presence of OR65.
