Extended Data Fig. 1: The data-processing workflow of sea urchin SLC9C1 in GDN.

a. Size-exclusion chromatography trace of Sea urchin SLC9C1 after nanobody GFP-affinity based purification in GDN and on-column TEV cleavage and removal; eleven independent purifications were performed with similar results. The arrow indicates the collected SLC9C1 protein peak collected for blotting as analysed by SDS-page and Coomassie staining (inset). b. The data-processing workflow of sea urchin SLC9C1 in GDN. a. The dataset contained 16,072 videos that were corrected by patch motion correction and patch CTF estimation in cryoSPARC⁵⁴. After reference-based auto-picking, 2,039,463 particles were picked. Several rounds of 2D classification were performed, yielding 312,610 particles, which were subjected to 3D classification. One of the 3D classes was selected, and it contained 195,956 particles. After repeated heterogenous and local refinement led to a final resolution of 3.23 Å in C2 symmetry at gold-standard FSC (0.143), with a local resolution range of 2.5–6.5 Å. To improve the density of the VSDs, symmetry expansion in C1 and masked refinement was further applied (light-blue maps) with the model-to-map fitting shown in Extended Data Fig. 3 and Supplementary Video 1. A final resolution of 3.22 Å was achieved at gold-standard FSC (0.143), with a local resolution range of 2.0–6.0 Å.