Glow discharged carbon coated grids

3 μl of the purified complex diluted to 0.04 mg/ml was applied to glow-discharged carbon-coated grids (Agar Scientific). Triton X-100 was added at a final concentration of 0.1 % in order to increase the number of side views. The sample was negatively stained with 2 % uranyl acetate and visualized in a FEI Tecnai F20 FEG microscope operating at a voltage of 200 kV under low dose conditions (~25 e/Ų). Images were recorded as described above at a magnification of 68,200 (2.2 Å/pixel) and a defocus range of 0.8-2.0 μm. Images were corrected for the CTF effect as described above and 1981 particles selected manually using BOXER. Images were normalised, band-pass filtered and centered using EMAN. They were then subjected to a reference-free classification using EMAN. The images corresponding to end and side views were extracted into two separate subsets containing 638 and 368 single particle images, respectively. End views were further aligned and classified using MLalign2D (XMIPP)³⁰ . The final classes were checked for symmetry by calculating their rotational auto-correlation function as described previously^{7 (link)}. A plot of the rotational auto-correlation function revealed 14 peaks reflecting the C14 symmetry of the core complex (Extended Data Fig. 2d).