Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

---

Query: EC:3.6.4.4 (kinesin)
5,033 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Studies of immobilized kinesin have shown that a single dimeric molecule can maintain contact with and drive sliding of a microtubule. In solution, however, native kinesin binds microtubules too weakly and hydrolyses ATP too slowly to produce the high sliding velocities seen in motility assay. This apparent inhibition in solution appears to be caused by the binding of kinesin's tail domains to its motor (head) domains in a folded conformation. DKH392, a construct containing two heads but no tails, has been shown to display both tight binding to microtubules and high ATPase rates. Furthermore, it retains one molecule of ADP per dimer when bound to microtubules, which could facilitate a 'hand-over-hand' mechanism for processive motion. Here we show that DKH392 hydrolyses more than 100 ATP molecules per diffusional encounter with a microtubule, even in the high-salt conditions encountered physiologically. This provides direct evidence that kinesin's activity is highly processive, with the motor remaining attached to a microtubule through many cycles of ATP hydrolysis.
...
PMID:Highly processive microtubule-stimulated ATP hydrolysis by dimeric kinesin head domains. 756 25

Kinesin is a 'motor' molecule, consisting of two head domains, an alpha-helical coiled coil rod, and a tail part that binds to its cargo. When expressed in a bacterial system, the head domain is functional, and can bind to microtubules with the stoichiometry of one head per tubulin dimer. Kinesin moves along microtubules by means of a cyclic process of nucleotide binding, hydrolysis and product release. We have used negative-stain electron microscopy and image analysis to study the structures of microtubules and tubulin sheets decorated with the motor domain (head) of kinesin in three states: in the presence of an unhydrolysable ATP analogue, 5'-adenylylimidodiphosphate (AMP-PNP); without nucleotides; and with adenosine 5'-diphosphate (ADP). A single kinesin head bound to a microtubule has a pear-shaped structure, with the broader end towards the 'plus' end of the microtubule under all conditions; the reverse motor, ncd, is similarly oriented. Three-dimensional maps reveal that kinesin heads have a spike that is assumed to form the attachment to the tail of a complete kinesin molecule. This spike is perpendicular to the microtubule axis in the presence of ADP, but points towards the plus end (approximately 45 degrees) in the presence of AMP-PNP or absence of nucleotides. Our results provide direct evidence for a conformational change of the kinesin motor domain during the ATPase cycle.
...
PMID:Nucleotide-dependent angular change in kinesin motor domain bound to tubulin. 761 26

In the nematode Caenorhabditis elegans, mutants in osm-3 gene are known to be defective in osmotic avoidance, chemotaxis and dauer formation behaviours. To study the molecular basis of these pleiotropic defects we have cloned the osm-3 gene by germline transformation of osm-3 (p802) mutants through microinjection of the wild type genomic DNA. Northern analysis reveals a 3.0 kb transcript corresponding to osm-3. DNA sequencing of the transforming 4.3 kb fragment revealed a kinesin heavy chain-like protein, which contains conserved ATPase and microtubule binding domains. Our results are consistent with the previous EM data on osm-3 (p802) mutants that show an accumulation of dense matrix material in the amphid sheath cytoplasm and a shortened distal segment of the amphid channel cilium. These data suggest a kinesin-like role of the osm-3 product in axonal transport.
...
PMID:C. elegans osm-3 gene mediating osmotic avoidance behaviour encodes a kinesin-like protein. 769 Feb 65

We have established pathway of the kinesin ATPase by direct measurement of each step in the pathway. Kinesin binds to microtubules with an 8-nm repeat and a stoichiometry of one kinesin monomer unit per tubulin dimer. Thus, the dimeric kinesin binds with both heads attached to the microtubule and on adjacent tubulin subunits. In the steady state, kinesin has a low ATPase activity that is limited by the rate of ADP release (< 0.01 s-1) in the absence of microtubules and is activated 2000-fold by the addition of microtubules to achieve a maximum rate of approximately 20 s-1. Transient-state kinetic analysis has provided direct measurement of individual steps of the reaction to define the pathway of the microtubule-kinesin ATPase. These studies establish that the rate-limiting step in the ATPase pathway is the release of the kinesin-product complex (K.ADP.P) from the microtubule following ATP hydrolysis. After phosphate release, the rebinding of kinesin-ADP to the microtubule is fast, accounting for the high activation of the ATPase at low microtubule concentration. This ATPase cycle explains the phenomenological differences between myosin and kinesin observed in motility assays. Kinesin remains associated with a microtubule through multiple rounds of hydrolysis, because it spends only a small fraction of its duty cycle in the dissociated state. The discussion of this paper will focus on the new data, their interpretation, and significance for mechanisms of force production. The ATPase coupling mechanism will be compared with dynein and myosin.
...
PMID:Pathway of the microtubule-kinesin ATPase. 778 62

The diffusion-limited rate for association of the ADP complex of dimeric DKH392 kinesin head domains with a microtubule was estimated to be 2-3 x 10(7) M-1 s-1 based on approximation of a microtubule as a highly elongated prolate ellipsoidal adsorber of 100% efficiency. This theoretical bimolecular rate is approximately 100-fold smaller than the experimental rate, kcat/KMT0.5, for DKH392 that was determined from the stimulation of the steady-state ATPase rate by microtubules. The large difference between these two estimates of the bimolecular rate indicates that it is likely that dimeric DKH392 hydrolyzes multiple ATP molecules during each diffusional encounter with a microtubule.
...
PMID:Implications of diffusion-controlled limit for processivity of dimeric kinesin head domains. 778 88

The oligomeric structure was determined for four recombinant kinesin derivatives containing N-terminal fragments of the kinesin alpha-subunit. Some of the proteins were dimeric (two-headed) molecules with mechanochemical properties similar to those of intact kinesin. Comparison of the primary and quaternary structures of the derivatives with those of intact kinesin suggests that structures distinct from the long alpha-helical coiled-coil rod domain contribute to subunit self-association. Three of the proteins contain a single engineered site for post-translational biotination in vivo; this facilitates analysis of motility in experiments in which the proteins are specifically bound to streptavidin-conjugated microscopic plastic beads. One of the derivatives is monomeric (one-headed); like the two-headed derivatives, it is functional in the motility assay and is a microtubule-dependent ATPase. Unlike intact kinesin and the two-headed derivatives, the one-headed enzyme fails to track microtubule protofilaments. This confirms a prediction of proposed "hand-over-hand" mechanisms of kinesin movement. The ability of molecules with a one-headed solution structure to generate movement is consistent with a translocation-generating conformational change internal to the kinesin head. A simple set of coupling rules can be used to formulate consistent mechano-chemical mechanisms that explain movement by both one- and two-headed kinesin molecules.
...
PMID:Structural and functional features of one- and two-headed biotinated kinesin derivatives. 778 90

We have measured the ATPase activity of squid optic lobe kinesin bound to polystyrene beads in the presence of microtubules. We find that there is a substantial increase (> 10-fold) in the microtubule-activated ATPase activity for bead-bound kinesin over free kinesin. We tentatively attribute such cargo-activated ATPase activity to the presence of a self-inhibited form of kinesin in solution, which becomes activated when bound to a bead in the presence of alpha-casein. Further experiments are underway to unravel this phenomenon and, in addition, to associate the traveling distance of beads with the observed ATPase rate to determine the average number of ATP consumed per kinesin-bead per micron of travel along microtubule.
...
PMID:Cargo-activated ATPase activity of kinesin. 778 91

A 100-kDa polypeptide with microtubule-interacting properties was identified in a Golgi vesicle-enriched fraction from Corylus avellana pollen. The k71s23 antibody (directed to the kinesin heavy chain from bovine brain) [Tiezzi et al., 1992: Cell Motil. Cytoskeleton 21:132-137] localized the polypeptide on the external surface of membrane-bounded organelles. Some 100-kDa-containing vesicles copelleted with microtubules (polymerized from purified bovine brain tubulin) either in presence or absence of 5 mM AMPPNP, but they could be released by 10 mM ATP or 0.5 M KCl. The pollen microtubule-interacting protein, salt-extracted from membranes and partially purified by gel filtration, exhibited an ATPase activity (16.2 nmolPi/mg/min) which could be stimulated about 2-fold (32.5 nmolPi/mg/min) by addition of bovine brain microtubules. We suppose that the 100-kDa polypeptide is part of a molecular complex showing properties of the kinesin class.
...
PMID:Kinesin-related polypeptide is associated with vesicles from Corylus avellana pollen. 782 Aug 65

Cytoplasmic dynein and ncd, a kinesin-related protein from Drosophila, are motor proteins that move toward the minus ends of microtubules, while kinesin moves to the microtubule plus end. In previous work, we examined the nucleotide dependence of motility and enzymatic activity by kinesin [Shimizu, T., Furusawa, K., Ohashi, S., Toyoshima, Y. Y., Okuno, M., Malik, F., & Vale, R. D., (1991) J. Cell Biol. 112, 1189-1197]. In this study, we examined these activities of the cytoplasmic dynein from bovine brain and ncd in order to explore what enzymatic features might be shared by these two minus-end-directed motors. Both ncd and cytoplasmic dynein demonstrated an activation of ATPase activity upon the addition of microtubules (30-fold and 6-fold, respectively). A significant difference between ncd and cytoplasmic dynein was their relative sensitivity to vanadate and to aluminum fluoride. In contrast to cytoplasmic dynein, ncd polypeptide was not cleaved by UV-vanadate treatment, and its ATPase and motility were unaffected by vanadate (up to 0.1 mM). When the nucleotide requirement for movement as examined using a battery of 20 nucleotides and nucleotide analogues, cytoplasmic dynein was found to exhibit a specificity very similar to that of axonemal dyneins from Tetrahymena. Surprisingly, however, the nucleotide specificities of in vitro motility produced by ncd or its construct, GST/MC1 (a fusion protein of glutathione S-transferase and 210-700 of the predicted ncd amino acid sequence), were quite distinct from that of kinesin. Thus, the nucleotide specificity profiles of members of the kinesin motor superfamily do not appear to be identical.
...
PMID:Comparison of the motile and enzymatic properties of two microtubule minus-end-directed motors, ncd and cytoplasmic dynein. 784 16

Direct measurement of the kinetics of kinesin dissociation from microtubules, the release of phosphate and ADP from kinesin, and rebinding of kinesin to the microtubule have defined the mechanism for the kinesin ATPase cycle. The processivity of ATP hydrolysis is ten molecules per site at low salt concentration but is reduced to one ATP per site at higher salt concentration. Kinesin dissociates from the microtubule after ATP hydrolysis. This step is rate-limiting. The subsequent rebinding of kinesin-ADP to the microtubule is fast, so kinesin spends only a small fraction of its duty cycle in the dissociated state. These results provide an explanation for the motility differences between skeletal myosin and kinesin.
...
PMID:Pathway of processive ATP hydrolysis by kinesin. 785 36


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---