Variant degree of saturation and geometry of the A/B ring fusion enables monohydroxysterol inhibition of BK channels. Representative single-channel recordings obtained after cbv1 incorporation into control (A), coprostanol- (B), or cholestanol-containing (C) POPE/POPS (3:1 wt/wt) bilayers. Sterols were introduced at 33 mol% in the lipid mix. For A–C, channel openings are shown as upward deflections; arrows on the top trace of each panel indicate the baseline. Channel activity (Po) was calculated as described in the legend to Fig. 2. (D) Scatter graph showing individual Po values in control and in the presence of cholesterol, coprostanol, or cholestanol in the lipid mixture. Each data point corresponds to one independent bilayer preparation. (E) Averaged Po data show that cholesterol, coprostanol, and cholestanol are all able to significantly reduce BK channel activity from control values. (F) Averaged degree of sterol-induced modification of cbv1 channel Po given as a percentage of inhibition from control bilayers. For D–F: *, significantly different from control (P < 0.05); **, significantly different from control (P < 0.01).