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Biopolymers Aug 2016Dyneins are multiprotein complexes that move cargo along microtubules in the minus end direction. The largest individual component of the dynein complex is the heavy... (Review)
Review
Dyneins are multiprotein complexes that move cargo along microtubules in the minus end direction. The largest individual component of the dynein complex is the heavy chain. Its C-terminal 3500 amino-acid residues form the motor domain, which hydrolyses ATP in its ring of AAA+ (ATPases associated with diverse cellular activities) domains to generate the force for movement. The production of force is synchronized with cycles of microtubule binding and release, another important prerequisite for efficient motility along the microtubule. Although the large scale conformational changes that lead to force production and microtubule affinity regulation are well established, it has been largely enigmatic how ATP-hydrolysis in the AAA+ ring causes these rearrangements. The past five years have seen a surge of high resolution information on the dynein motor domain that finally allowed unprecedented insights into this important open question. This review, part of the "ATP and GTP hydrolysis in Biology" special issue, will summarize our current understanding of the dynein motor mechanism with a special emphasis on the recently obtained crystal and EM structures. © 2016 Wiley Periodicals, Inc. Biopolymers 105: 557-567, 2016.
Topics: Adenosine Triphosphate; Animals; Dyneins; Humans; Hydrolysis; Microtubules; Movement; Protein Domains
PubMed: 27062277
DOI: 10.1002/bip.22856 -
Annual Review of Biophysics and... 1985Dynein and myosin show several important similarities in design as well as some interesting differences in detail. Both ATPases function as crossbridges that undergo... (Review)
Review
Dynein and myosin show several important similarities in design as well as some interesting differences in detail. Both ATPases function as crossbridges that undergo microscopic movements to drive the sliding of filaments, which results in macroscopic movements. They share a common design employing globular heads attached to flexible strands. Each head contains one ATP-binding site and one filament-binding site, and the binding of ATP induces an extremely rapid dissociation of the crossbridge-filament "rigor" complex. Following ATP hydrolysis, which is readily reversible, the crossbridge reassociates with the filament and returns to its original state with the release of products. Thus, the nucleotide-induced changes in conformation are effectively used to couple the hydrolysis of ATP to the dissociation and reassociation of the crossbridge in order to produce a force for net movement according to the Lymn-Taylor-Eisenberg model. The utilization of nucleotide-binding energy to induce a change in conformation can be rationalized in terms of our understanding of enzyme catalysis in general, whereby substrate binding energy is used to induce a change in conformation that stabilizes the transition state for catalysis. In these crossbridge ATPases, the substrate-induced change in conformation also serves to weaken the crossbridge-filament interaction. The pathway is symmetrical, with a return to the tight (filament) binding state coupled to product release. The ball on a string design may provide a reasonable basis to explain how a unidirectional force is obtained from a symmetrical cycle; opposite changes in conformation with the binding and release of the nucleotide produce a significant force only when pulling on the flexible strand. Moreover, the very rapid dissociation of the crossbridge following ATP binding limits the time that a negative force is in effect and also prevents a rigor crossbridge from retarding the sliding movements generated by other crossbridges. Myosin and dynein exhibit nearly identical kinetic constants governing ATP binding and the ATP-induced dissociation of the crossbridge. These appear as invariant steps that may reflect the basic principles of enzyme catalysis as applied to the mechanochemical cycle. The rates of ATP hydrolysis and synthesis by myosin and dynein differ slightly, but in each case the reactions are readily reversible with an equilibrium constant less than one. Steps involving the loss and rebinding of products occur at rates two to three orders of magnitude faster for dynein than for myosin.(ABSTRACT TRUNCATED AT 400 WORDS)
Topics: Actomyosin; Adenosine Triphosphatases; Adenosine Triphosphate; Animals; Dyneins; Kinetics; Macromolecular Substances; Microscopy, Electron; Microscopy, Electron, Scanning; Microtubules; Models, Biological; Models, Molecular; Protein Conformation; Structure-Activity Relationship; Vanadates; Vanadium
PubMed: 3159394
DOI: 10.1146/annurev.bb.14.060185.001113 -
Annual Review of Biophysics May 2021Dyneins make up a family of AAA+ motors that move toward the minus end of microtubules. Cytoplasmic dynein is responsible for transporting intracellular cargos in... (Review)
Review
Dyneins make up a family of AAA+ motors that move toward the minus end of microtubules. Cytoplasmic dynein is responsible for transporting intracellular cargos in interphase cells and mediating spindle assembly and chromosome positioning during cell division. Other dynein isoforms transport cargos in cilia and power ciliary beating. Dyneins were the least studied of the cytoskeletal motors due to challenges in the reconstitution of active dynein complexes in vitro and the scarcity of high-resolution methods for in-depth structural and biophysical characterization of these motors. These challenges have been recently addressed, and there have been major advances in our understanding of the activation, mechanism, and regulation of dyneins. This review synthesizes the results of structural and biophysical studies for each class of dynein motors. We highlight several outstanding questions about the regulation of bidirectional transport along microtubules and the mechanisms that sustain self-coordinated oscillations within motile cilia.
Topics: Animals; Biological Transport; Cilia; Dyneins; Humans; Intracellular Space; Microtubules
PubMed: 33957056
DOI: 10.1146/annurev-biophys-111020-101511 -
The Journal of Biological Chemistry Oct 2000Cytoplasmic dynein is a microtubule-associated motor that utilizes ATP hydrolysis to conduct minus-end directed transport of various organelles. Dynactin is a...
Cytoplasmic dynein is a microtubule-associated motor that utilizes ATP hydrolysis to conduct minus-end directed transport of various organelles. Dynactin is a multisubunit complex that has been proposed to both link dynein with cargo and activate dynein motor function. The mechanisms by which dynactin regulates dynein activity are not clear. In this study, we examine the role of dynactin in regulating dynein ATPase activity. We show that dynein-microtubule binding and ATP-dependent release of dynein from microtubules are reduced in dynactin null mutants, Deltaro-3 (p150(Glued)) and Deltaro-4 (Arp1), relative to wild-type. The dynein-microtubule binding activity, but not the ATP-dependent release of dynein from microtubules, is restored by in vitro mixing of extracts from dynein and dynactin mutants. Dynein produced in a Deltaro-3 mutant has approximately 8-fold reduced ATPase activity relative to dynein isolated from wild-type. However, dynein ATPase activity from wild-type is not reduced when dynactin is separated from dynein, suggesting that dynein produced in a dynactin mutant is inactivated. Treatment of dynein isolated from the Deltaro-3 mutant with lambda protein phosphatase restores the ATPase activity to near wild-type levels. The reduced dynein ATPase activity observed in dynactin null mutants is mainly due to altered affinity for ATP. Radiolabeling experiments revealed that low molecular mass proteins, particularly 20- and 8-kDa proteins, that immunoprecipitate with dynein heavy chain are hyperphosphorylated in the dynactin mutant and dephosphorylated upon lambda protein phosphatase treatment. The results suggest that cytoplasmic dynein ATPase activity is regulated by dynactin-dependent phosphorylation of dynein light chains.
Topics: Adenosine Triphosphate; Animals; Brain; Cattle; Centrifugation, Density Gradient; Cytoplasm; Dynactin Complex; Dyneins; Enzyme Activation; Fungal Proteins; Kinetics; Microtubule-Associated Proteins; Microtubules; Molecular Sequence Data; Molecular Weight; Mutation; Neurospora crassa; Phosphorylation; Precipitin Tests; Protein Binding; Protein Tyrosine Phosphatases; Receptor-Like Protein Tyrosine Phosphatases, Class 2
PubMed: 10921911
DOI: 10.1074/jbc.M000449200 -
Trends in Biochemical Sciences Jan 2016Cytoplasmic dynein, a member of the AAA (ATPases Associated with diverse cellular Activities) family of proteins, drives the processive movement of numerous... (Review)
Review
Cytoplasmic dynein, a member of the AAA (ATPases Associated with diverse cellular Activities) family of proteins, drives the processive movement of numerous intracellular cargos towards the minus end of microtubules. Here, we summarize the structural and motile properties of dynein and highlight features that distinguish this motor from kinesin-1 and myosin V, two well-studied transport motors. Integrating information from recent crystal and cryoelectron microscopy structures, as well as high-resolution single-molecule studies, we also discuss models for how dynein biases its movement in one direction along a microtubule track, and present a movie that illustrates these principles.
Topics: Animals; Dyneins; Humans; Microtubules; Models, Molecular
PubMed: 26678005
DOI: 10.1016/j.tibs.2015.11.004 -
Nature Communications Nov 2020Cytoplasmic dynein is the primary motor for microtubule minus-end-directed transport and is indispensable to eukaryotic cells. Although each motor domain of dynein...
Cytoplasmic dynein is the primary motor for microtubule minus-end-directed transport and is indispensable to eukaryotic cells. Although each motor domain of dynein contains three active AAA+ ATPases (AAA1, 3, and 4), only the functions of AAA1 and 3 are known. Here, we use single-molecule fluorescence and optical tweezers studies to elucidate the role of AAA4 in dynein's mechanochemical cycle. We demonstrate that AAA4 controls the priming stroke of the motion-generating linker, which connects the dimerizing tail of the motor to the AAA+ ring. Before ATP binds to AAA4, dynein remains incapable of generating motion. However, when AAA4 is bound to ATP, the gating of AAA1 by AAA3 prevails and dynein motion can occur. Thus, AAA1, 3, and 4 work together to regulate dynein function. Our work elucidates an essential role for AAA4 in dynein's stepping cycle and underscores the complexity and crosstalk among the motor's multiple AAA+ domains.
Topics: AAA Domain; Adenosine Triphosphate; Cytoplasmic Dyneins; Hydrolysis; Microtubules; Movement; Mutagenesis; Optical Tweezers; Protein Binding; Protein Conformation; Protein Multimerization; Saccharomyces cerevisiae
PubMed: 33230227
DOI: 10.1038/s41467-020-19477-3 -
Trends in Cell Biology Dec 1998Three classes of cytoskeletal motor protein have been identified--myosins, kinesins and dyneins. Together, these proteins are now thought to be responsible for the... (Review)
Review
Three classes of cytoskeletal motor protein have been identified--myosins, kinesins and dyneins. Together, these proteins are now thought to be responsible for the remarkable variety of movements that occur in eukaryotic cells and that are essential for reproduction and survival. Crystallographic analysis of the myosin and kinesin motor domains at atomic resolution has provided insight into their mechanism of force production. However, because of its relative intractability to molecular manipulation, definition of the dynein motor domain, let alone progress in understanding how it works, has been slower. Evidence now indicates that the microtubule-binding domain of dynein is spatially isolated from the ATPase domain at the tip of a projecting coiled coil. As proposed here, this curious arrangement might serve to accommodate multiple copies of the outsized and functionally complex motor heads on the microtubule surface.
Topics: Animals; Dyneins; Microtubules; Signal Transduction
PubMed: 9861671
DOI: 10.1016/s0962-8924(98)01379-8 -
The Journal of Cell Biology Sep 1980Two distinctly different ATPases have been reported to be endogenous to the mitotic apparatus: a Mg2+-ATPase resembling axonemal dynein, and a Ca2+-ATPase postulated to...
Two distinctly different ATPases have been reported to be endogenous to the mitotic apparatus: a Mg2+-ATPase resembling axonemal dynein, and a Ca2+-ATPase postulated to be bound in membranes. To examine the nature of the Mg2+-ATPase, we isolated membrane-free mitotic spindles from Stronglylocentrotus droebachiensis embryos by rapidly lysing these in a calcium-chelating, low-ionic-strength buffer (5 mM EGTA, 0.5 mM MgCl2, 10 mM PIPES, pH 6.8) that contained 1% Nonidet P-40. The fibrous isolated mitotic spindles closely resembled spindles in living cells, both in general morphology and in birefringence. In electron micrographs, the spindles were composed primarily of microtubules, free from membranes and highly extracted of intermicrotubular cytoplasmic ground substance. As analyzed by SDS-polyacrylamide gel electrophoresis (SDS-PAGE), the pelleted spindles contain 18% tubulin, variable amounts of actin (2-8%), and an unidentified protein of 55 kdaltons in a constant weight ratio to tubulin (1:2.5). The isolated spindles also contained two polypeptides, larger than 300 kdaltons, that comigrated with egg dynein polypeptides, and ATPase activity (0.02 mumol Pi/mg . min) that closely resembled both flagellar and egg dynein. The spindle Mg2+-ATPase showed a ratio of Ca2+-/Mg2+-ATPase = 0.85, had minimal activity in KCl and EDTA, and cleaved GTP at 35% of the rate of ATP. The Mg2+-ATPase was insensitive to ouabain or oligomycin. The spindle Mg2+-ATPase was inhibited by sodium vanadate but, like egg dynein, was less sensitive to vanadate than flagellar dynein. The spindle Mg2+-ATPase does not resemble the mitotic Ca2+-ATPase described by others. We propose that the spindle Mg2+-ATPase is egg dynein. Bound carbohydrate on the two high-molecular-weight polypeptides of both egg dynein and the spindle enzyme suggest that these proteins may normally associate with membranes in the living cell.
Topics: Adenosine Triphosphatases; Animals; Calcium-Transporting ATPases; Dyneins; Flagella; Microtubules; Mitosis; Molecular Weight; Sea Urchins; Substrate Specificity; Tubulin
PubMed: 6447705
DOI: 10.1083/jcb.86.3.738 -
Journal of Cell Science Oct 2010Cytoplasmic dynein in filamentous fungi accumulates at microtubule plus-ends near the hyphal tip, which is important for minus-end-directed transport of early endosomes....
Cytoplasmic dynein in filamentous fungi accumulates at microtubule plus-ends near the hyphal tip, which is important for minus-end-directed transport of early endosomes. It was hypothesized that dynein is switched on at the plus-end by cargo association. Here, we show in Aspergillus nidulans that kinesin-1-dependent plus-end localization is not a prerequisite for dynein ATPase activation. First, the Walker A and Walker B mutations in the dynein heavy chain AAA1 domain implicated in blocking different steps of the ATPase cycle cause different effects on dynein localization to microtubules, arguing against the suggestion that ATPase is inactive before arriving at the plus-end. Second, dynein from ΔkinA (kinesin 1) mutant cells has normal ATPase activity despite the absence of dynein plus-end accumulation. In ΔkinA hyphae, dynein localizes along microtubules and does not colocalize with abnormally accumulated early endosomes at the hyphal tip. This is in contrast to the colocalization of dynein and early endosomes in the absence of NUDF/LIS1. However, the Walker B mutation allows dynein to colocalize with the hyphal-tip-accumulated early endosomes in the ΔkinA background. We suggest that the normal ability of dyenin to interact with microtubules as an active minus-end-directed motor demands kinesin-1-mediated plus-end accumulation for effective interactions with early endosomes.
Topics: Aspergillus nidulans; Dyneins; Endosomes; Fungal Proteins; Kinesins; Microtubule-Associated Proteins; Microtubules
PubMed: 20876661
DOI: 10.1242/jcs.075259 -
Chemical Communications (Cambridge,... Jan 2021Along with various experimental methods, a combination of theoretical and computational methods is essential to explore different length-scale and time-scale processes... (Review)
Review
Along with various experimental methods, a combination of theoretical and computational methods is essential to explore different length-scale and time-scale processes in the biological system. The functional mechanism of a dynein, an ATP-fueled motor protein, working in a multiprotein complex, involves a wide range of length/time-scale events. It generates mechanical force from chemical energy and moves on microtubules towards the minus end direction while performing a large number of biological processes including ciliary beating, intracellular material transport, and cell division. Like in the cases of other conventional motor proteins, a combination of experimental techniques including X-crystallography, cryo-electron microscopy, and single molecular assay have provided a wealth of information about the mechanochemical cycle of a dynein. Dyneins have a large and complex structural architecture and therefore, computational modeling of different aspects of a dynein is extremely challenging. As the process of dynein movement involves varying length and timescales, it demands, like in experiments, a combination of computational methods covering such a wide range of processes for the comprehensive investigation of the mechanochemical cycle. In this review article, we will summarize how the use of state-of-the-art computational methods can provide a detailed molecular understanding of the mechanochemical cycle of the dynein. We implemented all-atom molecular dynamics simulations and hybrid quantum-mechanics/molecular-mechanics simulations to explore the ATP hydrolysis mechanisms at the primary ATPase site (AAA1) of dynein. To investigate the large-scale conformational changes we employed coarse-grained structure-based molecular dynamics simulations to capture the domain motions. Here we explored the conformational changes upon binding of ATP at AAA1, nucleotide state-dependent regulation of the mechanochemical cycle, and inter-head coordination by inter-head tension. Additionally, implementing a phenomenological theoretical model we explore the force-dependent detachment rate of a motorhead from the microtubule and the principle of multi-dynein cooperation during cargo transport.
Topics: Adenosine Triphosphate; Dyneins; Hydrolysis; Molecular Dynamics Simulation
PubMed: 33332489
DOI: 10.1039/d0cc05857b