Microtubule cross-linking triggers the directional motility of kinesin-5

Kapitein, Lukas C.; Kwok, Benjamin H.; Weinger, Joshua S.; Schmidt, Christoph F.; Kapoor, Tarun M.; Peterman, Erwin J. G.

doi:10.1083/jcb.200801145

Cited by 137 publications

(181 citation statements)

References 29 publications

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“…Whereas processive motors take hundreds of steps before they detach from a track, nonprocessive motors detach after a single step. Some motors such as kinesin-5, kinesin-14, and cytoplasmic dynein exhibit low or variable processivity depending on the local environment, including modified microtubule (MT) lattice and the arrangement of MT tracks (6)(7)(8)(9). All types of motors have been shown to increase the force or run length of cargos, which are generally larger than can be achieved by single motors (10)(11)(12), and many studies have suggested that the number of motors in these teams regulates the transport (3,5,10).…”

mentioning

confidence: 99%

Measuring collective transport by defined numbers of processive and nonprocessive kinesin motors

Furuta

Toyoshima

Amino

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

216

223

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Intracellular transport is thought to be achieved by teams of motor proteins bound to a cargo. However, the coordination within a team remains poorly understood as a result of the experimental difficulty in controlling the number and composition of motors. Here, we developed an experimental system that links together defined numbers of motors with defined spacing on a DNA scaffold. By using this system, we linked multiple molecules of two different types of kinesin motors, processive kinesin-1 or nonprocessive Ncd (kinesin-14), in vitro. Both types of kinesins markedly increased their processivities with motor number. Remarkably, despite the poor processivity of individual Ncd motors, the coupling of two Ncd motors enables processive movement for more than 1 μm along microtubules (MTs). This improvement was further enhanced with decreasing spacing between motors. Force measurements revealed that the force generated by groups of Ncd is additive when two to four Ncd motors work together, which is much larger than that generated by single motors. By contrast, the force of multiple kinesin-1s depends only weakly on motor number. Numerical simulations and single-molecule unbinding measurements suggest that this additive nature of the force exerted by Ncd relies on fast MT binding kinetics and the large drag force of individual Ncd motors. These features would enable small groups of Ncd motors to crosslink MTs while rapidly modulating their force by forming clusters. Thus, our experimental system may provide a platform to study the collective behavior of motor proteins from the bottom up.

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mentioning

confidence: 99%

Measuring collective transport by defined numbers of processive and nonprocessive kinesin motors

Furuta

Toyoshima

Amino

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

216

223

View full text Add to dashboard Cite

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“…Positively charged coverslips with DETA were prepared according to ref. 32. Flow chambers were assembled by joining a DETA-treated coverslip and a nontreated coverslip using double stick tape.…”

Section: Methodsmentioning

confidence: 99%

A mobile kinesin-head intermediate during the ATP-waiting state

Asenjo

Sosa

2009

Proc. Natl. Acad. Sci. U.S.A.

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Kinesin1 is a motor protein that uses the energy from ATP hydrolysis to move intracellular cargoes along microtubules. It contains 2 identical motor domains, or heads, that coordinate their mechano-chemical cycles to move processively along microtubules. The molecular mechanism of coordination between head domains remains unclear, partly because of the lack of structural information on critical intermediates of the kinesin1 mechano-chemical cycle. A point of controversy has been whether before ATP binding, in the so called ATP-waiting state, 1 or 2 motor domains are bound to the microtubule. To address this issue, here we use ensemble and single molecule fluorescence polarization microscopy (FPM) to determine the mobility and orientation of the kinesin1 heads at different ATP concentrations and in heterodimeric constructs with microtubule binding impaired in 1 head. We found evidence for a mobile head during the ATP-waiting state. We incorporate our results into a model for kinesin translocation that accounts well for many reported experimental results.cytoskeleton ͉ fluorescence ͉ microtubule ͉ single-molecule ͉ polarization K inesin1 is a motor protein that uses the energy of ATP hydrolysis to generate force and movement along microtubules (1). It contains 2 identical motor domains or heads where microtubule binding and ATP hydrolysis occur. It takes 8-nm steps between binding sites along the microtubule (2), hydrolyzing 1 ATP molecule per step (3-5). A stepping event is thought to occur when ATP binds to one of the heads inducing a disorder-to-order transition (docking) of a region called the neck linker, a short segment of Ϸ15 residues located at the C-terminal end of each head (6-8). Kinesin1 is a processive motor that can walk for more than an micrometer, going through hundreds of ATP hydrolysis cycles without dissociating from the microtubule (9). Processivity is achieved by an asymmetric hand-over-hand walking mechanism, in which the 2 heads coordinate their activities to alternate leading and trailing positions at each step (10-13).The molecular mechanism of coordination between kinesin1 motor domains is still unclear. Kinetic studies have identified ADP release and nucleotide binding as one of the coordinating steps. In solution, kinesin1 binds ADP tightly and interaction with the microtubule results in rapid release of 1 ADP molecule per kinesin head dimer. The second ADP molecule is released much more slowly, unless there is ATP present (or a nonhydrolysable ATP analogue) that accelerates the release of the second ADP molecule (14-16). This phenomenon constitutes a coordinating step or gate, as 1 kinesin head has to wait for the partner head to complete a particular step (ATP binding) to proceed (17). Several possible models have been proposed to explain this coordinating gate (18). One model proposes that in the ATPwaiting state the ADP containing head is kept away from the microtubule by interactions with the microtubule-bound head (19). Another model proposes that both motor domains can bind to th...

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“…The screen yielded a compound which inhibited Eg5, a member of the kinesin-5 family of motor proteins. Eg5, also known as kinesin spindle protein (KSP), is a plus-end directed motor which acts as a homotetramer to cross-link and slide anti-parallel microtubules at the onset of mitosis, thereby pushing apart the centrosomes to form the bipolar spindle [12]. In the absence of Eg5 function, the centrosomes remain tightly clustered yielding a monopolar spindle; consequently, the inhibitor was christened Monastrol [11].…”

Section: Ksp Inhibitors-bringing the Antimitotic Concept Into The 21smentioning

confidence: 99%