Extended Data Fig. 1: Purification and cryo-EM data processing of native-source eisosome filaments.
From: Cryo-EM architecture of a near-native stretch-sensitive membrane microdomain
a, Helical reconstruction data processing strategy. b, Unrolled and aligned helical structures of native-source eisosome filaments of different diameters show nearly identical lattice pattern. c, Data processing strategy for symmetry expansion of helical reconstructions d, Total intensity of Pil1 and Lsp1 peptides in mass spectrometry analysis. Intensity ratio of Pil1:Lsp1 is 3.1:1. e, Electrostatic surface prediction of Pil1 model with potentials ranging from −10 kcal*mol−1e−1 (red) to +10 kcal*mol−1e−1 (blue). f, Two representative aligned micrographs with native-source MCC–eisosome tubules and other putative contaminants visible. 2827 micrographs were collected for this dataset. g, Example 2D class averages with varying filament diameters. h, Representative Coomassie staining of protein gel of Bit61-TAP purification of MCC–eisosome tubules. Clear bands for TORC2 complex components Tor2, Avo3, Avo1 and Bit61-TAP are visible. The faint band at ~40 kDa is likely to correspond to Pil1/Lsp1. This protein purification was repeated n > 20 times yielding similar results. See Supplementary Data 4 for raw gel image.
