Fig. 6.

MT-stimulated ATPase and MT-depolymerizing activities of kinesin-13s. a, b Effect of DARPin on tubulin-stimulated ATPase activity of kinesin-13 proteins. ATP turnover rates of Kif2A-NM (orange) and Kif2A-MD (gray) in the presence of tubulin with the indicated concentrations of DARPin are shown in a. Equivalent data for Kif2C-NM (light orange) and Kif2C-MD (gray) are shown in b. c, d Effects of DARPin on MT-stimulated ATPase activities of kinesin-13 proteins: c ATP turnover rates by Kif2A-NM (red) and Kif2A-MD (gray) in the presence of 2 µM of taxol-stabilized MTs and the indicated concentrations of DARPin are shown. d Corresponding data sets of experiments for Kif2C-NM (light red) and Kif2C-MD (gray) as shown. Data from a and b represent averages from at least four independent experiments. Error bars, S.D.; ns: p > 0.05; *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001, by Student’s t-test. e Time course plots of inorganic phosphate release by Kif2A-NM under the same experimental conditions as in c. Averages from N = 3. Error bars, S.D. f Monitoring of MT polymers present at the 2.5 and 10-min time points of the experiment shown in c (right three pairs) and the corresponding controls without Kif2A-NM (left three pairs) by a centrifugation-based sedimentation assay. S supernatant fraction; P pellet fraction. Samples were resolved by SDS-PAGE and gels were stained with Coomassie blue. Representative gels are shown