Fig. 2: IST2⁷¹⁶ structure.

a, Cryo-EM density of IST2⁷¹⁶ at 2.84 Å viewed from within the membrane. The relationship between views is indicated. Subunits are colored in yellow and violet, with residual density presumably corresponding to detergent or lipids in gray. b, Ribbon representation of IST2⁷¹⁶ with colors and orientation as in a. Membrane boundaries are indicated and selected regions of the structure are labeled. c, Ribbon representation of the IST2 subunit at indicated orientations, with the NTD colored in red. Transmembrane helices are labeled. d, Cryo-EM density of IST2⁷¹⁶ with residual density corresponding to detergent and lipids colored in violet. A lowpass-filtered map at low contour showing the distortion in the detergent micelle is superimposed. ‘#’ and ‘*’ indicate sites of deformation. e, Subunit cavity viewed from within the membrane. Shown are α3–α8 from a single subunit. Superposition of IST2⁷¹⁶ (yellow) and the Ca²⁺-bound open state of nhTMEM16 (left, brown; PDB 6QM9), IST2⁷¹⁶ (center) and nhTMEM16 (right). The molecular surface in the region of the subunit cavity is displayed and colored according to the properties of contacting residues (polar, green; acidic, red; basic, blue). f, Region of IST2 corresponding to the regulatory Ca²⁺-binding site of the TMEM16 family. Left, cryo-EM density superimposed on the model. Right, ribbon with the corresponding side chains of IST2 displayed. g, Comparison of the regulatory region between nhTMEM16 and IST2. Left, nhTMEM16 with two Ca²⁺ ions bound, Right, superposition of equivalent regions of IST2 and nhTMEM16. Bottom, sequence alignment of relevant regions in fungal and murine TMEM16 proteins. The numbering refers to IST2. h, Superposition of transmembrane α6 and α7 and their connecting region in IST2 and nhTMEM16. Inset, zoomed-in view of the interacting cytoplasmic helices α6_i and α6’. In e–g, Ca²⁺ ions bound to nhTMEM16 are displayed as blue spheres. i, Conformation of α4 and α6 lining the edge of the subunit cavity in a superposition of the subunits of IST2 and nhTMEM16 in the presence (open; PDB 6QM9) and absence (closed; PDB 8TPM) of Ca²⁺. The protein is shown as a ribbon with the equivalent positions of α4 occupied by glycine residues in nhTMEM16 marked as spheres.