Fig. 4.

HsCKK, but not NgCKK, induces tilting of whole protofilaments. a Raw and Fourier filtered images of 13- and 14- protofilament MTs decorated by NgCKK and HsCKK. In each set of three panels: left, raw image; centre, filtered image to include data at, and adjacent to, the origin and the 1/8 nm layer line; right, filtered image to include data at, and adjacent to, the origin highlights the MT moiré pattern; blue arrows indicate variations in the moiré pattern that arise from protofilament skew. b Protofilament skew for a 16 MT subset from each dataset plotted as the average rotation angle around the MT axis (PHI) change per dimer moving axially towards the MT plus-end. HsCKK-decorated MTs compared to kinesin decorated paclitaxel-stabilized MTs (13-protofilament kinesin-3 data from⁴⁰; 14-protofilament kinesin-1 data from⁴⁰). All data points are plotted and bars represents mean ± SD. HsCKK 13-protofilament (tSA201 tubulin) vs kinesin-3 13-protofilament, p < 0.0001, NgCKK 13-protofilament vs HsCKK 13-protofilament, p < 0.0001, NgCKK 13-protofilament vs kinesin-3 13-protofilament, not significant (p = 0.164), HsCKK 14-protofilament (tSA201 tubulin) vs kinesin-1 14-protofilament, p < 0.001, NgCKK 14-protofilament vs HsCKK 14-protofilament, p < 0.0001, NgCKK 14-protofilament vs kinesin-1 14-protofilament, not significant (p = 0.889), one-way ANOVA with Tukey’s multiple comparisons test; source data are provided as a Source Data file. c Schematic of influence of NgCKK/HsCKK on protofilament skew; d left, MT protofilaments fitted into aligned HsCKK and NgCKK C1 reconstructions were overlaid; the bottom dimer corresponds to the point at which the density was aligned; divergence between NgCKK and HsCKK models increases from this point; right, RMSD of backbone positions in panel (i) depicted on a NgCKK protofilament; e RMSDs between NgCKK and HsCKK protofilaments calculated when the bottom dimers in the models themselves are directly aligned, where RMSD does not decrease significantly with distance from the superimposed dimer; f mechanisms of protofilament skew induction: left, no skew change; middle, protofilament skew arising from interdimer subunit stagger, e.g. from tubulin GTPase; right, protofilament skew arising from whole-protofilament tilting e.g. due to changes in protofilament number or HsCKK binding