Fig. 3: S6I segment dynamics are associated with activation gate conformation.
a 3D variability analysis of the cryo-EM map after consensus refinement and viewed from the membrane plane. The first frame map (Frame 1) and the last frame map (Frame 14) are aligned. The relative positions of the S6I segments' density are marked. S6I density in Frame 14 is shorter as compared to Frame 1. b Intracellular view of Frame 1 and Frame 14. These maps highlight the shift of the S6I segment toward the AG, which correlates with a transition from an open to a more constricted, inactivated-like conformation. c The S6 segments (S6I-IV) fitted into the densities of Frame 1 and Frame 14 exhibit varying positions of S6I. d Superimposed intracellular views reveal a 13° inward tilt of the S6I segment in Frame 14 relative to Frame 1. e Cryo-EM densities for the S6I segments. The densities are contoured at 5σ for Cluster 1 and Cluster 2. Selected residues are labeled. f NTD and S6I segment interactions in Cluster 1 and Cluster 2. In Cluster 1, R433 of S6I forms a salt bridge with D66 of the NTD (inset ‘i’). In Cluster 2, K430 and R433 of S6I are positioned close to D57 of the NTD, forming a distinct salt bridge network (inset ‘ii’). g Pore radius profiles along the conduction pathway for Cluster 1 (salmon) and Cluster 2 (marine) show a narrower activation gate in Cluster 2. h Ion permeation paths (gray dots) overlaid with S6I and S6III segments. Regions corresponding to the SF, CC, and AG are indicated.
