Extended Data Fig. 2: Conformational flexibility of the CTD domain of FANCD2 and its associated DNA.

a, The CTD of FANCD2 and its associated DNA are evident in the consensus reconstruction before temperature-factor sharpening. Because the DNA has higher temperature factors than the protein (Extended Data Table 1), the temperature-factor calculated from the overall map degrades the continuity of the DNA density. The cartoon representation of the refined model is coloured as follows: FANCI, cyan; FANCD2, pink; DNA, gold. The schematic of the ICL DNA is shown to the right of the map, with the deoxycytidine bases that are crosslinked by a triazole coloured red. The 20-nt ssDNA arms consist of (dT)₂₀ to minimize secondary structure. b, 3D classification of the particles showing the conformational flexibility of FANCD2. The 3D classes are arranged starting with the most compact conformation in which the CTD of FANCD2 is closer to its NTD. Also shown is the refined consensus model rigid-body-fitted into each class and coloured as in a. The conformational flexibility starts within the helical domain (starting around residue 645). c, The five 3D classes are superimposed by aligning the FANCI portion of each map (top), or of each PDB structure (bottom) coloured according to their map in b. d, Cryo-EM reconstructions using particles from the top component of the principal component analysis (PCA) of the multi-body refinement angles, separated into five bins. This component accounts for 21.5% of the variance in the relative orientation of the CTD of FANCD2 (Supplementary Video 1). Left, five ID–ICL DNA models refined in real-space with PHENIX (overall solvent-corrected resolution ranging from 3.7 to 3.9 Å) against maps reconstructed with particles derived from five bins of eigenvalues for the top eigenvector. This PCA component corresponds to a rotation of up to 16° (curved arrow) about an axis running through the helical domain that is roughly perpendicular to the plane of the figure (grey stick). The helical axes of the individual duplexes are shown as black sticks. Right, the corresponding maps, without temperature-factor sharpening, starting with the conformation (pink map) in which the CTD of FANCD2 is closest to the CTD of FANCI. e, The second component from the PCA accounts for 17% of the variance in the relative orientation of the CTD of FANCD2 (Supplementary Video 2). It corresponds to a rotation of up to 10° (curved arrow) about an axis (grey stick) that is roughly parallel to the plane of the figure. Left, the refined models; right, the maps as in d.