Extended Data Fig. 2: Data processing of OSCA1.2 in nanodiscs.
From: Mechanical activation opens a lipid-lined pore in OSCA ion channels
a, A representative cryo-EM image of OSCA1.2 in DOPC nanodiscs with a msp:lipid ratio of 1:50. Some particles are circled. Scale bar: 50 nm. b, Image-processing workflow of OSCA1.2-DOPC-1:50 with Relion and CryoSPARC. See Methods and Extended Data Table 1, 2 for details. c, Gold-standard FSC curves calculated between the independently refined half maps of OSCA1.2-DOPC-1:50 in the contracted state indicate a resolution of 3.59 Å (FSC = 0.143) and in the expanded state indicate a resolution of 3.63 Å (FSC = 0.143). d, Image-processing workflow of OSCA1.2 in DOPC nanodiscs reconstituted with a msp:lipid ratio of 1:20. Green box represents the contracted2 state. Salmon box represents contracted1 state. Blue box represents the expanded state. e, Gold-standard FSC curves calculated between the independently refined half maps for OSCA1.2-DOPC-1:20 in contracted2 state (left) and expanded state (right). f, The tentative density of TM6. A rod-shaped density (magenta) is located above the dilated pore in the contracted2 state of OSCA1.2. The map was low-pass filtered to facilitate the visualization of this density. This density is only visible at a high threshold, indicating its weak density level. Based on the ___location and orientation, this density is tentatively assigned as TM6. g, Dilation of the pore. The pore forming helices TM3-7 in closed (salmon) and activated (green) subunits are shown in ribbon. The residues on the ion permeation pathway are shown in magenta stick. h-i, Image-processing workflow of OSCA1.2 in DOPC (h) and azolectin (i) nanodiscs reconstituted with different msp:lipid ratios. Asterisk represents the class in which one protomer exhibits weaker density, but the overall conformation remains the same as that of the contracted1 state. j, The particle distribution of OSCA1.2 in different conformations was analyzed using the particles in green, salmon, and blue boxes shown in the image-processing workflow. k, The gel filtration profiles of nanodiscs-embedded OSCA1.2 reconstituted with different MSP:lipid ratios.
