Extended Data Fig. 6: PTH1R kinetic experiments.
From: Insights into G-protein coupling preference from cryo-EM structures of Gq-bound PTH1R
a. Control fluorescence time-lapse images of TMR-FiBiT on LgBiT-AT1R-expressing cells (upper panels) with and without perfusion flow and of TMR-PTH/PTHrP on mock transfected cells (lower panels) with flow. Images show representative data from 10-14 cells across two independent experiments. b, c. Fluorescence time-lapse images of TMR-PTH (b) and PTHrP (c) dissociation from wild-type and mutant PTH1R-expressing cells. The images of wild-type and the triple mutant are the same as Fig. 2e for comparison. The white bar indicates 5 μm (a–c). d. Time-course changes in fluorescence intensity normalized at the start of perfusion (red: PTH, blue: PTHrP). Lines and error bands represent mean and SEM, respectively, from 8-20 cells done in two independent experiments (d). e, f. AIC comparison between double- and triple-exponential fitting for PTH (e) and PTHrP (f). Dots with a line show the AICs calculated from the same cell data. Red lines show mean AIC of all cells plotted. Statistical analysis was performed with paired t-test. g, h. Logarithmic time constants (g) and percentage (h) of three kinetic components (upper: fast, middle: medium, lower: slow). Mean and SEM are plotted as bar graphs (n = 8-20 cells). The exact sample sizes are provided in Supplementary Tables 2. Statistical analysis was performed with two-way ANOVA, followed by Sidak’s post-hoc test for PTH and PTHrP comparison, and Dunnett’s post-hoc test for wild-type and mutants. In all panels, p-values are shown as follows: ns, p > 0.05; *, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001. The exact p-values are provided in source data file.
