Drosophila Ncd reveals an evolutionarily conserved powerstroke mechanism for homodimeric and heterodimeric kinesin-14s.
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 heterodimeric and homodimeric kinesin-14s share a common evolutionary structural and mechanochemical mechanism for force generation.
Department of Biological Sciences.
Pengwei
P
Zhang
Department of Chemical & Biological Engineering, and.
Wei
W
Dai
Department of Chemical & Biological Engineering, and Department of Biomedical Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY 12180 sgilbert@rpi.edu hahnj@rpi.edu.
Juergen
J
Hahn
Department of Biological Sciences, sgilbert@rpi.edu hahnj@rpi.edu.
Susan P
SP
Gilbert
D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D001696:Biomechanical Phenomena; D020285:Cryoelectron Microscopy; D019281:Dimerization; D029721:Drosophila Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000235:genetics*; D016547:Kinesins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008870:Microtubules / Q000378:metabolism*; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D011485:Protein Binding; D011487:Protein Conformation
D029721:Drosophila Proteins; C066696:ncd protein, Drosophila; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D016547:Kinesins
dynamic modeling; microtubules; presteady-state kinetics