Interaction site of the Ndc80/Nuf2 dimer on the microtubule lattice. (A and B) A model of a microtubule protofilament made from the tubulin dimer (PDB 1TUB) was docked into a single protofilament cut from the Ndc80/Nuf2 microtubule EM map. The crystal structure of an engineered, truncated Ndc80 complex was used for fitting into the remaining density in either a two-headed (A) or an alternating strong- or weak-binding (B) configuration. The red bracket demarcates an α/β-tubulin heterodimer based on the favored configuration in which the strong density overlaps the kinesin-binding site (red dashed line) at the interdimer interface. The + and − signs indicate microtubule polarity; β-tubulin of each α/β-tubulin heterodimer points toward the plus end. Lysines in Ndc80 and Nuf2, whose mutation to alanine or glutamate reduced microtubule-binding affinity (Ciferri et al., 2008), are indicated by yellow spheres; the black sphere marks the methionine preceding the first residue of Ndc80 (amino acid 80) in the crystal structure. For the two-headed model in A, the gray arrows labeled 1 and 2 highlight missing density and extra, unaccounted for density, respectively. For the strong or weak model in B, weak binding is schematized using light shading.