Guided by the combined TRIM and CA lattice maps, 550 subvolumes encompassing entire TRIM hexagons were extracted (64 × 64 × 64 px) from 4× binned tomograms and assigned polar Euler angles in reference to the underlying CA lattice [which was modeled as “surface” in Dynamo (57)]. Azimuthal angles were randomized. One iteration of azimuthal refinement was performed to generate an initial model (fig. S6A). Classification was performed by multireference alignment in Dynamo, using as reference two copies of the initial model with random noise added, a cylindrical alignment mask that covered both the TRIM and CA densities, and a cylindrical classification mask that covered only the CA densities. The classification separated the particles into two classes (Classes 1 and 2) according to the rotation of the TRIM hexagon relative to the long axis of the tube (fig. S6, A and B). A second classification run was performed on the Class 2 particles, which further separated the particles into two subclasses (Classes 2a and 2b) that differed in translation of the TRIM hexagon relative to the underlying CA lattice (fig. S6, A and C). Class 1 particles from above (335 particles) were recropped from 2× binned tomograms (128 × 128 × 128 px) and refined for two additional iterations with progressively narrower angular and positional search ranges.

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