Fig. 1: Functional characterization of the TMEM16A blocker 1PBC.

a Chemical structure of 1PBC. The pKa values of ionizable groups were calculated with the chemistry package Chemicalize (ChemAxon, https://chemicalize.com/). b Steady-state current-voltage relationship of wild-type mouse TMEM16A at the indicated concentrations of 1PBC applied to the intracellular side of the membrane at 2 µM intracellular Ca²⁺. Data are averages of 6 biological replicates, errors are SEM. c Concentration-response relations of 1PBC at voltages from −140 to 140 mV, ΔV = 20 mV. Data are calculated from b, errors are SEM. Solid lines are fits to the Hill equation. d IC₅₀ values obtained from (c) at the indicated voltages. Data are best-fit values, errors are 95% CI. Concentration-response relations of 1PBC of mouse TMEM16B (e) and TMEM16F (f) at 15 and 300 µM intracellular Ca²⁺ respectively at −80 and 80 mV. Data are averages of 5 and 6 biological replicates respectively, errors are SEM. Solid lines are fits to the Hill equation. Dashed lines are the relations of TMEM16A. g Sequence alignment of the outer pore region of mouse TMEM16A (UniProt ID: Q8BHY3), mouse TMEM16B (UniProt ID: Q8CFW1), and mouse TMEM16F (UniProt ID: Q6P9J9). Sequence identity between TMEM16A and B, 60.5%; between TMEM16A and F, 39.5%. A conserved glycine in α3 is highlighted in gray and other colors indicate the type of the residues interacting with the blocker (yellow, hydrophobic; green, polar; blue, basic) in TMEM16A.