Drosophila Ncd reveals an evolutionarily conserved powerstroke mechanism for homodimeric and heterodimeric kinesin-14s

• Published in: Proceedings of the National Academy of Sciences - PNAS 2015-05, Vol.112 (20), p.6359-6364
• Main Authors: Zhang, Pengwei, Dai, Wei, Hahn, Juergen, Gilbert, Susan P
• Format: Article
• Language: English
• Subjects: Adenosine Diphosphate - metabolism; Adenosine triphosphatase; Adenosine Triphosphate - metabolism; Animals; ATP; Biological Sciences; Biomechanical Phenomena; Chromosomes; Cryoelectron Microscopy; Dimerization; Drosophila melanogaster; Drosophila melanogaster - genetics; Drosophila Proteins - chemistry; Drosophila Proteins - genetics; Drosophila Proteins - metabolism; Insects; Kinesin - chemistry; Kinesin - genetics; Kinesin - metabolism; Kinetics; Metazoa; Microtubules - metabolism; Models, Molecular; Molecular Dynamics Simulation; Protein Binding; Protein Conformation; Saccharomyces cerevisiae; Yeast
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.1505531112

Significance Kinesin molecular motors couple ATP turnover to force production to generate microtubule-based movement and microtubule dynamics. Kinesin-14s are unique in that they are nonprocessive, bind to adjacent microtubule protofilaments rather than step along a single protofilament as observed for processive kinesins, and use a powerstroke mechanism to slide microtubules. Earlier studies proposed that only one head of the Ncd dimer interacts with the microtubule to drive the ATP-promoted powerstroke and therefore only one ATP turnover was required. The results presented here challenge the one head/one ATP turnover hypothesis and define a common pathway for Kar3Vik1, Kar3Cik1, and Ncd. These findings are significant because they reveal that the key principles for force generation by kinesin-14s are conserved from yeast to higher eukaryotes. Drosophila melanogaster kinesin-14 Ncd cross-links parallel microtubules at the spindle poles and antiparallel microtubules within the spindle midzone to play roles in bipolar spindle assembly and proper chromosome distribution. As observed for Saccharomyces cerevisiae kinesin-14 Kar3Vik1 and Kar3Cik1, Ncd binds adjacent microtubule protofilaments in a novel microtubule binding configuration and uses an ATP-promoted powerstroke mechanism. The hypothesis tested here is that Kar3Vik1 and Kar3Cik1, as well as Ncd, use a common ATPase mechanism for force generation even though the microtubule interactions for both Ncd heads are modulated by nucleotide state. The presteady-state kinetics and computational modeling establish an ATPase mechanism for a powerstroke model of Ncd that is very similar to those determined for Kar3Vik1 and Kar3Cik1, although these heterodimers have one Kar3 catalytic motor domain and a Vik1/Cik1 partner motor homology domain whose interactions with microtubules are not modulated by nucleotide state but by strain. The results indicate that both Ncd motor heads bind the microtubule lattice; two ATP binding and hydrolysis events are required for each powerstroke; and a slow step occurs after microtubule collision and before the ATP-promoted powerstroke. Note that unlike conventional myosin-II or other processive molecular motors, Ncd requires two ATP turnovers rather than one for a single powerstroke-driven displacement or step. These results are significant because all metazoan kinesin-14s are homodimers, and the results presented show that despite their structural and functional differences, the h