Extended Data Fig. 3: Cryo-EM data processing of VAChTACh.
From: Binding mechanism and antagonism of the vesicular acetylcholine transporter VAChT
a. Flow chart for cryo-EM data processing of VAChTACh. A total of 6,600 micrographs were motion-corrected using patch motion correction, and CTF parameters were determined using patch CTF estimation in CryoSPARC. A representative motion-corrected micrograph of this dataset is shown here (scale bar = 30 nm). The particles were picked using template picker, and 2D classification was performed to remove junk particles. After three rounds of heterogeneous refinements, followed by ab-initio reconstruction and non-uniform refinement, a 3.8-Å resolution map was obtained. The 201,774 good particles were used as the “seed” and subjected to seed-facilitated 3D classification to further improve the resolution. Details of data processing can be found in Method. b. The angular distribution of the final reconstruction. c. Local resolution distribution of VAChTACh. d. Fourier Shell Correlations (FSC) of the final map of the VAChTACh, calculated between two independently refined half-maps before (blue) and after (red) post-processing, overlaid with an FSC curve calculated between the cryo-EM map and the structural model shown in black. e. Representative overlay of cryo-EM density and transmembrane helices of VAChTACh. The cryo-EM maps are shown as grey surface.
