Extended Data Fig. 8: Workflow of cryo-EM data processing of PI3Kγ.
From: Molecular basis for Gβγ-mediated activation of phosphoinositide 3-kinase γ
(A) Representative micrograph (scale bar = 100 nm). About 5000 micrographs with similar outcomes were collected. (B) Representative 2D class averages (box size = 320 pixels where 1 pixel = 1.08 Å). (C) FSC plot, estimating 3.0 Å resolution using FSC at 0.143. (D) Cryo-EM reconstruction. In total, 269,357 particles were used for 3D map reconstruction. Map regions belonging to p110γ and p101 are roughly indicated by the black and red brackets. The CTD of p101 is colored magenta. (E) Local resolution estimation of the PI3Kγ cryo-EM map from (D). The overall resolution in most regions of p110γ is 2.5-3 Å with well-defined side chains, whereas the overall resolution of p101 is >3.5 Å. (F) Local refinement of PI3Kγ focused on the p101 NTD. The yellow circle indicates the fulcrum point used for local refinement. (G) Local refinement of PI3Kγ focused on the p101 C-terminal domain. The yellow circle indicates the fulcrum point used for local refinement. (H) Composite map generated from the three maps shown in panels (D), (F), and (G). (I) Local resolution estimation of the composite PI3Kγ cryo-EM map from (H). The overall resolution in most regions is 2.5-3 Å, whereas the p101 C-terminal domain has an improved resolution of 3-4 Å.
