Direct observation of single kinesin molecules moving along microtubules
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Kinesin
Organelle
Molecular motor
Abstract Molecular motor proteins, fueled by energy from ATP hydrolysis, move along actin filaments or microtubules, performing work in the cell. The kinesin microtubule motors transport vesicles or organelles, assemble bipolar spindles or depolymerize microtubules, functioning in basic cellular processes. The mechanism by which motor proteins convert energy from ATP hydrolysis into work is likely to differ in basic ways from man‐made machines. Several mechanical elements of the kinesin motors have now been tentatively identified, permitting researchers to begin to decipher the mechanism of motor function. The force‐producing conformational changes of the motor and the means by which they are amplified are probably different for the plus‐ and minus‐end kinesin motors. BioEssays 25:1212–1219, 2003. © 2003 Wiley Periodicals, Inc.
Kinesin
Molecular motor
Organelle
Chemical energy
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Kinesin
Molecular motor
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Biochemical, pharmacological and immunocytochemical studies have implicated the microtubule-activated ATPase, kinesin, in the movement of membrane bounded organelles in fast axonal transport. In vitro studies suggested that kinesin moves organelles preferentially in the anterograde direction, but data about the function and precise localization of kinesin in the living axon were lacking. The current study was undertaken to establish whether kinesin associates with anterograde or retrograde moving organelles in vivo. Peripheral nerves were ligated to produce accumulations of organelles moving in defined directions. Regions proximal (anterograde) and distal (retrograde) to the ligation were analyzed for kinesin localization by immunofluorescence, and by immunogold electron microscopy using ultracryomicrotomy. Substantial amounts of kinesin were associated with anterograde moving organelles on the proximal side, while significantly less kinesin was detected distally. Statistical analyses indicated that kinesin was mostly associated with membrane-bounded organelles. These observations indicate that axonal kinesin is primarily associated with anterograde moving organelles in vivo.
Kinesin
Organelle
Axoplasmic transport
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Intracellular transport based on molecular motors and its regulation are crucial to the functioning of cells. Filamentary tracks of the cells are abundantly decorated with nonmotile microtubule-associated proteins, such as tau. Motivated by experiments on kinesin-tau interactions [Dixit et al., Science 319, 1086 (2008)] we developed a stochastic model of interacting single-headed motor proteins KIF1A that also takes into account the interactions between motor proteins and tau molecules. Our model reproduces experimental observations and predicts significant effects of tau on bound time and run length which suggest an important role of tau in regulation of kinesin-based transport.
Kinesin
Molecular motor
Intracellular transport
Tau protein
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Kinesin
Molecular motor
Processivity
Microtubule-associated protein
Organelle
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Kinesins are molecular motors that transport various cargoes along microtubule tracks using energy derived from ATP hydrolysis. Although the motor domains of kinesins are structurally similar, the family contains members that move on microtubules in opposite directions. Recent biochemical and biophysical studies of several kinesins make it possible to identify structural elements responsible for the different directionality, suggesting that reversal of the motor movement can be achieved through small, local changes in the protein structure.
Kinesin
Directionality
Molecular motor
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A high-throughput molecular force sensor that uses single-stranded DNA as a force sensing molecule. It can measure pico-Newton level forces of multiple kinesin motors transporting a microtubule.
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Molecular motor
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ABSTRACT The human genome encodes 45 kinesins that drive cell division, cell motility, intracellular trafficking, and ciliary function. Determining the cellular function of each kinesin would be greatly facilitated by specific small molecule inhibitors, but screens have yielded inhibitors that are specific to only a small number of kinesins, likely due to the high conservation of the kinesin motor domain across the superfamily. Here we present a chemical-genetic approach to engineer kinesin motors that retain microtubule-dependent motility in the absence of inhibitor yet can be efficiently inhibited by small, cell-permeable molecules. Using kinesin-1 as a prototype, we tested two independent strategies to design inhibitable motors. First, we inserted the six amino acid tetracysteine tag into surface loops of the motor domain such that binding of biarsenic dyes allosterically inhibits processive motility. Second, we fused DmrB dimerization domains to the motor heads such that addition of B/B homodimerizer cross-links the two motor domains and inhibits motor stepping. We show, using cellular assays that the engineered kinesin-1 motors are able to transport artificial and natural kinesin-1 cargoes, but are efficiently inhibited by the addition of the relevant small molecule. Single-molecule imaging in vitro revealed that inhibitor addition reduces the number of processively moving motors on the microtubule, with minor effects on motor run length and velocity. It is likely that these inhibition strategies can be successfully applied to other members of the kinesin superfamily due to the high conservation of the kinesin motor domain. The described engineered motors will be of great utility to dynamically and specifically study kinesin function in cells and animals.
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Molecular motor
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Kinesin-1 is a motor protein that can step processively on microtubule by hydrolyzing ATP molecules, playing an essential role in intracellular transports. To better understand the mechanochemical coupling of the motor stepping cycle, numerous structural, biochemical, single molecule, theoretical modeling and numerical simulation studies have been undertaken for the kinesin-1 motor. Recently, a novel ultraresolution optical trapping method was employed to study the mechanics of the kinesin-1 motor and new results were supplemented to its stepping dynamics. In this commentary, the new single molecule results are explained well theoretically with one of the models presented in the literature for the mechanochemical coupling of the kinesin-1 motor. With the model, various prior experimental results for dynamics of different families of N-terminal kinesin motors have also been explained quantitatively.
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Molecular motor
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