Fig. 2: Stomatin anchors to the membrane via N-terminal helices and the hydrophobic surface of SPFH1 and defines a mechanically distinct membrane domain.

a Side view of the cryo-EM density and atomic model of a Stomatin subunit showing its orientation relative to the membrane surface (solid line). Inset highlights the N-terminal helices (H1 and H2) and the membrane-facing hydrophobic region of the SPFH1 domain, both of which insert partially into the lipid bilayer. b Zoomed-in views of the membrane-anchoring interface. The N-terminal helices H1 and H2 (purple and blue) insert into the outer leaflet of the lipid bilayer, while the hydrophobic surface of the SPFH1 domain (cyan) also embeds into the membrane. The remaining surface of SPFH1 is hydrophilic and faces the cytosol, featuring stabilizing interactions such as a salt bridge between D89 and R67, and a positively charged residue, K55. Key residues involved in membrane interactions are labeled, including palmitoylation sites (C30, C53, C87), a membrane-anchoring residue (W51), and the topology-influencing proline (P47). The right panel shows a 180° rotation around the Y-axis to reveal the inner side of the complex. c Cryo-EM 3D reconstructions of vesicles containing the Stomatin complex, sorted by vesicle radius of curvature (Rv), ranging from 53.2 nm to 12.5 nm. In all cases, the Stomatin complex maintains a consistent cap-like architecture. d Cross-sectional views of the reconstructions show that the membrane beneath the Stomatin complex remains flat, even as the rest of the vesicle exhibits curvature. Black curved lines represent the vesicle membrane, while straight lines mark the flattened microdomain under the complex. e Superimposed atomic models from vesicles of different curvature (Rv = 30.8 nm, green; Rv = 12.5 nm, purple) show only a ~ 1.9 Å change in the membrane-facing diameter on each side. This minimal variation indicates that the Stomatin complex enforces a rigid, curvature-resistant membrane microdomain.