The carboxyl terminus of Vps32 inhibits the lateral association of filaments within spirals. (A) Analysis of the spacing between filaments in Vps32 spirals. (left) A representative cryopreserved Vps32 spiral with its trajectory highlighted in purple is shown. The ends of the spiral are indicated (S [starting point] and E [ending point]). Bar, 25 nm. (middle) A portion of this spiral was straightened, and its distance to the adjacent filament within the same structure was calculated along its entire length. (right) The average distance between the centers of each filament is plotted. Error bars represent means ± SEM. (B) Analysis of the spacing between filaments in a negatively stained spiral composed of a truncated form of Vps32 (amino acids 1–190) was conducted using electron tomography. Representative raw images, tomograms, and trajectories are shown for a spiral pair (top) and a large-diameter spiral (bottom). (right) The average distance between the centers of each filament is plotted. Boxed regions highlight the areas that were magnified (shown in the images to the right). Bars: (23,000× magnification) 50 nm; (59,000× magnification, tomogram, and trajectory) 20 nm. The differences in distances are statistically significant (P < 0.001, compared with wild type, one-tail t test). Error bars represent means ± SEM.