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

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

Kinesin superfamily proteins (KIFs) are probable motors in vesicular and non-vesicular transport along microtubular tracks. Since a variety of KIFs have been recently identified in the motile flagella of Chlamydomonas, we sought to ascertain whether KIFs are also associated with the connecting cilia of vertebrate rod photoreceptors. As the only structural link between the rod inner segment and the photosensitive rod outer segment, the connecting cilium is thought to be the channel through which all material passes into and out of the outer segment from the rod cell body. We have performed immunological tests on isolated sunfish rod inner-outer segments (RIS-ROS) using two antibodies that recognize the conserved motor domain of numerous KIFs (anti-LAGSE, a peptide antibody, and anti-Klp1 head, generated against the N terminus of Chlamydomonas Klp1) as well as an antibody specific to a neuronal KIF, KIF3A. On immunoblots of RIS-ROS, LAGSE antibody detected a prominent band at approximately 117 kDa, which is likely to be kinesin heavy chain, and Klp1 head antibody detected a single band at approximately 170 kDa; KIF3A antibody detected a polypeptide at approximately 85 kDa which co-migrated with mammalian KIF3A and displayed ATP-dependent release from rod cytoskeletons. Immunofluorescence localizations with anti-LAGSE and anti-Klp1 head antibodies detected epitopes in the axoneme and ellipsoid, and immunoelectron microscopy with the LAGSE antibody showed that the connecting cilium region was particularly antigenic. Immunofluorescence with anti-KIF3A showed prominent labelling of the connecting cilium and the area surrounding its basal body; the outer segment axoneme and parts of the inner segment coincident with microtubules were also labelled. We propose that these putative kinesin superfamily proteins may be involved in the translocation of material between the rod inner and outer segments.
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PMID:Localization of kinesin superfamily proteins to the connecting cilium of fish photoreceptors. 871 80

Calmodulin, a calcium modulated protein, regulates the activity of several proteins that control cellular functions. A cDNA encoding a unique calmodulin-binding protein, PKCBP, was isolated from a potato expression library using protein-protein interaction based screening. The cDNA encoded protein bound to biotinylated calmodulin and 35S-labeled calmodulin in the presence of calcium and failed to bind in the presence of EGTA, a calcium chelator. The deduced amino acid sequence of the PKCBP has a domain of about 340 amino acids in the C-terminus that showed significant sequence similarity with the kinesin heavy chain motor domain and contained conserved ATP- and microtubule-binding sites present in the motor domain of all known kinesin heavy chains. Outside the motor domain, the PKCBP showed no sequence similarity with any of the known kinesins, but contained a globular domain in the N-terminus and a putative coiled-coil region in the middle. The calmodulin-binding region was mapped to a stretch of 64 amino acid residues in the C-terminus region of the protein. The gene is differentially expressed with the highest expression in apical buds. A homolog of PKCBP from Arabidopsis (AKCBP) showed identical structural organization indicating that kinesin heavy chains that bind to calmodulin are likely to exist in other plants. This paper presents evidence that the motor domain has microtubule stimulated ATPase activity and binds to microtubules in a nucleotide-dependent manner. The kinesin heavy chain-like calmodulin-binding protein is a new member of the kinesin superfamily as none of the known kinesin heavy chains contain a calmodulin-binding domain. The presence of a calmodulin-binding motif and a motor domain in a single polypeptide suggests regulation of kinesin heavy chain driven motor function(s) by calcium and calmodulin.
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PMID:A plant kinesin heavy chain-like protein is a calmodulin-binding protein. 875 76

The kinesin motor proteins translocate toward either the plus or minus end of microtubules (MTs). Competitive microtubule binding assays were carried out with monomeric motor domains of the minus-end-directed nonclaret disjunctional (Ncd) and Kar3 and the plus-end-directed kinesin heavy chain (KHC) to determine whether motors of the same or opposite polarity compete for binding sites on MTs and to test the idea that motor polarity is determined by differences in binding sites on MTs of the motors. The stoichiometries of binding were approximately 1 motor:1 tubulin heterodimer for all three motors. Ncd and Kar3, both minus-end motors, severely inhibited the binding of one another to MTs, as predicted theoretically for binding of the two motors to the same site on MTs, indicating that the binding sites on MTs of Ncd and Kar3 are the same or overlap extensively. Motors of opposite polarity, KHC and Ncd or KHC and Kar3, showed partial or complete inhibition of binding to MTs under different experimental protocols. The differences in binding behavior could be due to experimental conditions or be inherent in the nature of motor binding to MTs. Alternatively, differences in KHC and Ncd or Kar3 binding sites on MTs may exist such that the motors bind to partially overlapping but nonidentical sites on MTs. These differences in binding sites may be related to the opposite polarity of translocation on MTs of the motors.
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PMID:Binding sites on microtubules of kinesin motors of the same or opposite polarity. 878 May 25

Previous work has shown that mutation of the gene that encodes the microtubule motor subunit kinesin heavy chain (Khc) in Drosophila inhibits neuronal sodium channel activity, action potentials and neurotransmitter secretion. These physiological defects cause progressive distal paralysis in larvae. To identify the cellular defects that cause these phenotypes, larval nerves were studied by light and electron microscopy. The axons of Khc mutants develop dramatic focal swellings along their lengths. The swellings are packed with fast axonal transport cargoes including vesicles, synaptic membrane proteins, mitochondria and prelysosomal organelles, but not with slow axonal transport cargoes such as cytoskeletal elements. Khc mutations also impair the development of larval motor axon terminals, causing dystrophic morphology and marked reductions in synaptic bouton numbers. These observations suggest that as the concentration of maternally provided wild-type KHC decreases, axonal organelles transported by kinesin periodically stall. This causes organelle jams that disrupt retrograde as well as anterograde fast axonal transport, leading to defective action potentials, dystrophic terminals, reduced transmitter secretion and progressive distal paralysis. These phenotypes parallel the pathologies of some vertebrate motor neuron diseases, including some forms of amyotrophic lateral sclerosis (ALS), and suggest that impaired fast axonal transport is a key element in these diseases.
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PMID:Kinesin mutations cause motor neuron disease phenotypes by disrupting fast axonal transport in Drosophila. 891 51

Kinesin-73 cDNA was shown to encode a kinesin heavy chain protein that contains an N-terminal motor domain and a long central region that lacks extensive coiled-coils. The amino acid sequence of the motor domain of kinesin-73 protein is most closely related to the motor domains of Caenorhabditis elegans unc-104 and mouse KIF1A. The central region of kinesin-73 protein also is related to unc-104 and KIF1A, but the homology is lower than that of the motor domain. The C-terminal region of kinesin-73 protein contains a cytoskeleton associated protein Gly-rich domain, which is a putative microtubule binding site that is present in some cytoskeleton or dynein-associated proteins. Kinesin-73 mRNA was shown by in situ hybridization to be maternally expressed and widely distributed in the syncytial blastoderm embryo. However, later in Drosophila embryo development, expression of the kinesin-73 gene becomes restricted mostly to the central and peripheral nervous systems.
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PMID:Kinesin-73 in the nervous system of Drosophila embryos. 903 10

The kinesin heavy chain consists of an N-terminal globular domain, referred to as the motor domain, a rod-like middle region, and a C-terminal domain. In this study, the human kinesin neck region, the region adjacent to the motor domain which promotes dimerization, has been investigated. First, we predicted coiled-coil regions including the neck region by our newly devised statistical method. The sequence (335-372) was predominated by a unique heptad amphipathy. A comparison of the bacterially expressed human kinesin heavy chain fragments, K349 (1-349), a monomeric motor domain, and K379 (1-379), a dimer, by circular dichroism (CD) spectroscopy showed that K379 had more alpha-helical content. Chemically synthesized peptides, (332-349), (350-379), and (332-369), gave CD spectra with an alpha-helix-rich pattern, but the spectra varied depending on the peptide concentration. Analysis of the molar ellipticity at 222 nm indicated that those peptides were in monomer-dimer equilibria, and the dissociation isotherms established dissociation constants of 9.6 mM. 60 microM, and 62 nM for the above peptides, respectively. Sedimentation equilibrium measurements verified that the peptide (332-369) existed as a dimeric form. These results strongly suggest that the sequence from 332 to 369 of the neck region forms an alpha-helical coiled coil. The differential peptide of K349 and K379, (350-379), did not show sufficient ability to make K379 dimeric. It is likely that the region (350-379) forms a stable alpha-helical coiled coil only together with the (332-349) region. Fluorescence energy transfer studies of [Cys363]-(332-369) labeled with a fluorescence donor and an acceptor revealed that the peptide formed a parallel coiled coil. This coiled coil was thermodynamically stable against urea and thermal denaturation, and peptide exchange of the coiled coil was undetectable, or extremely slow, at neutral pH. The dissociation free energy was estimated to be 57.7 kJ mol-1 at a peptide concentration of 22 microM. These results indicate that the neck region of kinesin forms a stable coiled coil which may be important for the motility of dimeric kinesin.
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PMID:Identification of kinesin neck region as a stable alpha-helical coiled coil and its thermodynamic characterization. 904 81

Granular/vesicular transport is thought to be supported by microtubule-based force-generating adenosine triphosphatases such as kinesin. Kinesin is a motor molecule that has been well studied in brain and other neuronal tissues. Although vesicular transport is important for pancreatic beta-cell secretory activities, the role of kinesin in beta-cell function has not been investigated. It is hypothesized that kinesin functions as a translocator that associates with both microtubules and insulin-containing granules in beta-cells and transports the secretory granules from deep within the cytoplasm, where insulin is synthesized and processed, to the surface of beta-cells upon secretory stimulation. To test this hypothesis, a mouse beta-cell kinesin heavy chain complementary DNA was cloned and sequenced. Kinesin expression in primary cultures of mouse beta-cells then was selectively suppressed by antimouse beta-cell kinesin heavy chain antisense oligonucleotide treatment. Analysis of insulin secretion determined that the basal level of insulin secretion from the treated cells was decreased by 50%. Furthermore, glucose-stimulated insulin release from treated beta-cells was reduced by almost 70% after suppression of kinesin expression by antisense treatment. The findings from this study provide the first direct evidence that kinesin, a microtubule-based motor protein, plays an important role in insulin secretion.
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PMID:Suppression of the expression of a pancreatic beta-cell form of the kinesin heavy chain by antisense oligonucleotides inhibits insulin secretion from primary cultures of mouse beta-cells. 911 96

A eubacterial homolog of a kinesin light chain gene has been isolated and characterized from the cyanobacterium Plectonema boryanum. Although the eubacterial and eukaryotic kinesin light chains are highly similar in amino acid sequence, the eubacterial sequence differs in several distinguishing structural features, including the absence of a putative PEST domain and the presence of additional highly conserved imperfect tandem repeats. Two soluble kinesin light chain antigens have been identified from whole-cell lysates by immunoblot analysis. Attempts to identify a canonical kinesin heavy-chain gene or protein were unsuccessful, suggesting that a kinesin heavy chain may be absent or unnecessary for kinesin light-chain function in this eubacterium. Our findings establish that certain basal elements of eukaryotic cellular transport appear to be resident in eubacteria. We discuss the possibility that the eukaryotic kinesin light chain was acquired by lateral gene transfer.
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PMID:Kinesin light chain in a eubacterium. 921 72

Kinesins comprise a large superfamily of microtubule-based motor proteins, individual members of which mediate specific types of motile processes. To identify kinesin family members (KIFs) that are critical to retinal function and thus to vision, a reverse transcriptase polymerase chain reaction (RT-PCR) cloning strategy was used to isolate putative KIFs expressed in the neural retina and retinal pigmented epithelium (RPE) of the striped bass, Morone saxatilus. Eleven fish KIFs (FKIFs) were isolated from neural retina and six of the same FKIFs were also isolated from RPE. One of the KIFs identified in this screen, FKIF2, was the most prevalent clone detected both in the retina (41% of clones) and RPE (72% of clones). Based on predicted amino acid sequence homology within the motor domain, seven of the FKIFs have been tentatively assigned to known kinesin families: the kinesin heavy chain family (FKIF1, 5 and 9), the unc104/KIF1 family (FKIF3 and 8), the KIF2 family (FKIF4), and the cKIF family (FKIF2). Northern blot analysis revealed that each detectable FKIF exhibited a unique tissue-specific expression pattern. Since FKIF2 was more highly expressed in retina than in any other tissue tested, including brain, and was the most abundant KIF message expressed in both retina and RPE, it was examined in more detail and the complete approximately 2.3 kb open reading frame for FKIF2 was cloned and sequenced. The predicted amino acid sequence indicates that FKIF2 has a C-terminal motor domain, and thus is a member of the cKIF family. FKIF2 is only 36.5% identical at the amino acid level to the most closely related cKIF in the database, suggesting that FKIF2 may be a novel member of this family. Antibodies raised against a unique peptide specific to FKIF2 recognize an approximately 80 kd protein in homogenates of retina, RPE, brain and kidney. The pronounced expression of FKIF2 in retina and RPE suggests that FKIF2 may play an important role in microtubule-dependent motile events in these two tissues.
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PMID:Multiple kinesin family members expressed in teleost retina and RPE include a novel C-terminal kinesin. 924 9

Treatment of proteins in vitro with sulfhydryl (SH)-reactive compounds has been used successfully to determine protein regions critical for normal function. To probe structure-function relationships in the microtubule (MT) motor kinesin, the motor was treated with two SH reactive compounds, n-ethylmaleimide and ethacrynic acid, and its function was assayed by motility and co-sedimentation techniques. In the motility assay, treatment of kinesin either before or after adsorption to the glass surfaces of a flow cell was found to inhibit the ability of coverslip-bound kinesin to bind to MTs. Inactivation of MT binding was slow, required high molar excess of the SH-reactive drug, and was very sensitive to temperature. Inhibition of MT binding occurred well after complete modification of kinesin light chain, but paralleled modification of the kinesin heavy chain. The results point to a model in which one critical cysteine per kinesin heavy chain is relatively inaccessible to solvent. Surprisingly, when the interaction between modified kinesin and MTs was examined by a co-sedimentation assay, kinesin retained the ability to bind MTs. These contrasting results may be due to conformational differences in the kinesin molecule that exist in the two assays.
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PMID:n-ethylmaleimide and ethacrynic acid inhibit kinesin binding to microtubules in a motility assay. 925 2


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