Extended Data Fig. 2: Data processing pipeline, 3D classification and final global refinements.

Data processing pipeline, 3D classification and final global refinements. a, A low-pass filtered and motion corrected micrograph is shown as an example of the raw data. Particles are highlighted with red boxes. Image was exported from Cryolo boxmanager. 3D classification in this case was done with 6 classes (K = 6). The final reconstruction from RELION along with angular distribution of particles are displayed in the bottom right. Overall resolution is based on FSC cutoff of 0.143. b, Masks used for focused refinements in RELION. (c) and (d) are 3D classification and final global refinement results for the Ca²⁺/CHL dataset and the Ca²⁺/DMSO dataset respectively. Left panel shows Euler angular distribution of particles which contributed to the final 3D map. The height and color of bars represent the number of particles at that particular Euler angle with the smallest number in blue and the higher in red. The final corrected masked FSC curves for both Ca²⁺/CHL and Ca²⁺/DMSO datasets are displayed, with the FSC cutoff of 0.143 as reference (black dashed lines).