Supplementary Figure 3: Biological heterodimer versus asymmetric-unit heterodimer.
From: Structural and mechanistic insight into Holliday-junction dissolution by Topoisomerase IIIα and RMI1
a) A crystal packing diagram showing the coexistence of two TopIIIα-RMI1 interfaces. To determine which interface is biologically relevant, we performed EM and single-particle analysis of negatively stained TopIIIα-RMI complex sample used for growing crystals. b) Example of a negatively stained micrograph and individual particles marked with white squares. The scale bar corresponds to 50 nm. c) Reference-free class averages (top), reprojections of the 3D reconstruction (middle row) and reprojections of the crystal structure filtered at 30 Å resolution (bottom line) as matched by cross-correlation. d) The angular distribution for the TopIIIα-RMI reconstruction mapped on a half sphere. The size of the spot is proportional to the number of raw images contributing to the class. (e) Fourier shell correlation (FSC) showing a resolution of 34 Å calculated using the 0.5 threshold criterion (dashed lines). f) Fitting of the two interfaces (ASU interface and biological interface) found in the crystal lattice in the calculated EM map. In the alternative orientation found in the ASU the orientation and location of RMI1 does not agree with the EM envelope demonstrating that the ASU interface is not the biological interface in solution. g) Different views of the fit between the crystal structure and the calculated EM map.
