Extended Data Fig. 7: Protein environment of the non-annular lipid along the S6 helix.

(a) Cryo-EM density of SthK in 3:1 DOPC:POPA nanodiscs with the protein in light blue, lipids in orange. Lipids that have not been investigated in this study are transparent. Zoom shows the non-annular lipid along S6 at the center of this study. (b) Non-annular lipid along S6 (orange, spheres) embedded in the protein (electrostatic surface representation calculated in Chimera X). (c) S5 and S6 helices of adjacent subunits are shown as cartoon (light and dark blue) with Arg136 on S5 and Asp226 on S6 in ball and stick representation and the lipid along the S6 helix of one subunit as density obtained from cryo-EM. Three different lipids are modelled into this density and shown in ball and stick representation (from left to right: POPA, POPG, DOPC) revealing that the PA headgroup fits best in the experimental density. (d) Cryo-EM density of SthK in 3:1 DOPC:POPA nanodiscs (grey, unsharpened map) showing continuous density between Arg136 and the exposed phosphate group of POPA. (e) Lipid (orange, ball and stick representation) along the S6 helix between two adjacent subunits (dark and light blue, cartoon, only S5 and S6 are shown for clarity). The lipid is intertwined between S5 and S6 of neighboring subunits and is in contact with the bundle crossing and the pore helix/ selectivity filter.