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:3.6.1.3 (ATPase)
65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Calmodulin (CaM) mediates the Ca(2+)-dependent activation of many enzyme systems in accordance with its cellular localization. We have described previously a muscarinic receptor-mediated translocation of CaM from membranes into the cytosol of SK-N-SH human neuroblastoma cells. To explore the potential targets (CaM-binding proteins, CaMBP) for CaM upon translocation, a photoreactive CaM derivative was introduced into living SK-N-SH cells using a scrape-loading technique. Scrape-loading incorporated rhodamine isothiocyanate-labeled CaM with an efficiency of 38%. CaM-diazopyruvamide (CaM-DAP), a Ca(2+)-dependent and CaM-specific probe, was also introduced into the cells. The muscarinic agonist carbachol stimulated a translocation of CaM from membranes into cytosol in CaM-DAP-loaded SK-N-SH cells. Upon photochemical cross-linking, cross-linked adducts of CaM-CaMBP were detected by immunoblotting with anti-CaM antibody. Carbachol stimulated increased photoaffinity labeling of three proteins with relative adduct molecular masses of 70, 120, and 180 kDa. The time course of labeling for the 70- and 120-kDa adducts showed maximal increased by 15-30 min. The 180-kDa adduct displayed a slower time course of maximal labeling, with increases maintained for 2-4 h. Subtracting the molecular mass of CaM, carbachol stimulated binding to CaMBPs of 55, 105, and 163 kDa. Predominant cellular CaMBP were identified using a biotinylated CaM overlay procedure. Western blot analysis indicated the expression of specific CaM-dependent enzymes such as calcineurin, phosphodiesterase, the beta-isoform (rat brain) of CaM kinase II, and Ca(2+)-ATPase. Numerous cytoskeletal CaMBP were expressed such as microtubule-associated protein-2, spectrin, tubulin, caldesmon, adducin, and neuromodulin. Of the CaMBP expressed, phosphodiesterase, calcineurin, caldesmon, and adducin cross-linked with CaM-DAP in the loaded SK-N-SH cells. Carbachol stimulated the time-dependent CaM-DAP labeling of calcineurin and adducin. This study demonstrates the novel incorporation of a photoreactive CaM derivative into living cells, as well as muscarinic receptor-activated CaM-DAP interaction with several cellular CaMBP. We postulate that carbachol-stimulated CaM translocation in SK-N-SH cells may affect the activity of CaM-dependent enzymes and may alter aspects of cytoskeletal function.
...
PMID:Carbachol stimulates binding of a photoreactive calmodulin derivative to calmodulin-binding proteins in intact SK-N-SH human neuroblastoma cells. 155 1

Fast axonal transport is manifested at the sub-cellular level as the anterograde or retrograde movement of membrane-bounded organelles along microtubules. Earlier work implicated the protein kinesin as the motor for anterograde axonal transport. More recent work indicates that a brain microtubule-associated protein, MAP 1C, is responsible for retrograde transport. Of additional interest, MAP 1C has been found to be a cytoplasmic form of the ciliary and flagellar ATPase dynein, indicating a much more general functional role for this enzyme in cells than had been suspected.
...
PMID:The role of dynein in retrograde axonal transport. 246 13

We have examined the relationship of the ubiquitous 68-70-kDa cytoskeletal-associated protein beta-internexin (Napolitano, E. W., Pachter, J. S., Chin, S. S. M., and Liem, R. K. H. (1985) J. Cell Biol. 101, 1323-1331) to heat-shock cognate 70 (hsc70), the major constitutive member of the mammalian heat-shock protein 70 (hsp70) family of stress proteins. We purify beta-internexin from rat brain microtubules and confirm its identity with hsc70 and the clathrin-uncoating ATPase by the following criteria: 1) The partial sequence of a cyanogen bromide-derived peptide from beta-internexin matches the inferred amino acid sequence of the cDNA clone pRC62 encoding hsc70 from rat brain (O'Malley, K., Mauron, A., Barchas, J. D., and Kedes, L. (1985) Mol. Cell. Biol. 5, 3476-3483). 2) Mixing experiments followed by two-dimensional gel analyses reveal the precise co-migration of beta-internexin, the clathrin-uncoating ATPase, and the in vitro translation product of cDNA clone pHSP-4 encoding rat brain hsc70. 3) beta-Internexin is recognized by a monoclonal antibody reactive against the class of hsp70 proteins. 4) beta-Internexin purified from a microtubule-associated protein-enriched fraction of rat brain by virtue of high affinity binding to ATP-agarose possesses clathrin cage-specific ATPase activity.
...
PMID:Beta-internexin is a microtubule-associated protein identical to the 70-kDa heat-shock cognate protein and the clathrin uncoating ATPase. 252 48

Concurrent exposures to organophosphorus insecticide leptophos and the industrial solvents n-hexane and toluene were implicated in causing an outbreak of neuropathy in workers. Although both leptophos and n-hexane produce central-peripheral distal axonopathy, the morphology and distribution of neuropathic lesions are distinct, reflecting different modes of action. The molecular mechanisms of organophosphorus compound-induced delayed neurotoxicity (OPIDN) and aliphatic hexacarbon-induced neurotoxicity have been investigated utilizing various biochemical techniques, (i.e. one- and two-dimensional gel electrophoresis, immunoblotting, peptide mapping). Oral administration of tri-o-cresyl phosphate (TOCP) produced delayed neurotoxicity and increased in vitro Ca2+ and calmodulin-dependent kinase protein phosphorylation of cytoskeletal proteins in brain, spinal cord, and sciatic nerve of chickens. This enhanced protein phosphorylation correlated well with the following characteristics of OPIDN: test chemical, whether an OPIDN-producing or not; dose-dependence and time course of the effect; and the animal sex sensitivity, age selectivity, and species susceptibility. The proteins that showed an increased phosphorylation were identified to be; alpha- and beta-tubulin, microtubule-associated protein-2 (MAP-2), and the 3 neurofilament proteins 70 kDa, 160 kDa, and 210 kDa. Further studies suggested that the increased protein phosphorylation is not related to an effect on protein phosphatase or ATPase activity, but rather to altered Ca2+-calmodulin kinase II activity. Aliphatic hexacarbon-induced neurotoxicity is characterized by an accumulation of 10 nm neurofilaments above the nodes of Ranvier in the spinal cord and peripheral nerve. Treatment of rats with 2,5-hexanedione, the active neurotoxic metabolite of n-hexane, produced protein crosslinking in a dose-dependent manner. This treatment also decreased protein phosphorylation of neurofilament proteins as well as MAP-2. These studies demonstrate the involvement of cytoskeletal proteins in the molecular pathogenesis of chemical-induced neurotoxicity.
...
PMID:Cytoskeletal proteins as targets for organophosphorus compound and aliphatic hexacarbon-induced neurotoxicity. 283 76

Previous work has indicated that following the rapid adenosine 5'-triphosphate (ATP) induced dissociation of the microtubule-dynein complex, the rate-limiting step in the ATPase cycle is product release [Johnson, K. A. (1983) J. Biol. Chem. 258, 13825-13832], which occurs at a rate of approximately 2-6 s-1. In this report we complete the analysis of the ATPase cycle by examining the effect of microtubules on the rate of product release. For these studies we used repolymerized Tetrahymena axonemal microtubules and microtubule-associated protein (MAP) free bovine brain microtubules which were shown to be free of any measureable ATPase activity. Tetrahymena 22S dynein bound to these microtubules predominantly by the ATP-sensitive site and at a rate giving an apparent second-order rate constant of (0.2-1) X 10(6) M-1 s-1, which is 50-fold greater than the rate observed with brain microtubules containing MAPs. ATP induced the rapid dissociation of the microtubule-dynein complex with an apparent second-order rate constant vs. ATP concentration equal to 1.6 X 10(6) M-1 s-1; this value is only slightly lower than that observed in the presence of MAPs. After the ATP-induced dissociation, the dynein reassociated with the microtubules following a lag period due to the time required to hydrolyze the ATP. The duration of the lag time for reassociation decreased with increasing microtubule concentration, suggesting that microtubules increased the rate of ATP turnover. Direct measurements at steady state showed that the specific activity of the dynein increased with increasing microtubule concentration.(ABSTRACT TRUNCATED AT 250 WORDS)
...
PMID:Activation of the dynein adenosinetriphosphatase by microtubules. 293 50

We observe that one of the high molecular mass microtubule-associated proteins (MAPs) from brain exhibits nucleotide-dependent binding to microtubules. We identify the protein as MAP IC, which was previously described in this laboratory as a minor component of standard microtubule preparations (Bloom, G.S., T. Schoenfeld, and R.B. Vallee, 1984, J. Cell Biol., 98:320-330). We find that MAP 1C is enriched in microtubules prepared in the absence of nucleotide. Kinesin is also found in these preparations, but can be specifically extracted with GTP. A fraction highly enriched in MAP 1C can be prepared by subsequent extraction of the microtubules with ATP. Two activities cofractionate with MAP 1C upon further purification, a microtubule-activated ATPase activity and a microtubule-translocating activity. These activities indicate a role for the protein in cytoplasmic motility. MAP 1C coelectrophoreses with the beta heavy chain of Chlamydomonas flagellar dynein, and has a sedimentation coefficient of 20S. Exposure to ultraviolet light in the presence of vanadate and ATP results in the production of two large fragments of MAP 1C. These characteristics suggest that MAP 1C may be a cytoplasmic analogue of axonemal dynein.
...
PMID:MAP 1C is a microtubule-activated ATPase which translocates microtubules in vitro and has dynein-like properties. 295 82

The inner and outer arms of the flagellar axoneme generate the forces needed for flagellar movement; these arms contain ATPases called dyneins. To date, there has been no method for studying the mechanochemical transducing activity of isolated dyneins. Recently, it was found that the brain microtubule-associated protein (MAP) 1C is a microtubule-activated ATPase with the structural and force-producing properties of dynein. MAP 1C translocates microtubules in an in vitro gliding assay, suggesting that such an assay could also be used with axonemal dyneins. Here, we demonstrate that outer-arm dynein isolated from sea urchin (Strongylocentrotus purpuratus) sperm and adsorbed to a glass coverslip can translocate calf-brain microtubules along the surface of the coverslip. Our results conclusively demonstrate that outer-arm dynein by itself is capable of generating shearing forces. The ability to examine the force-generating properties of flagellar dynein in vitro should greatly facilitate studies of the mechanism of action of this important mechanochemical transducer.
...
PMID:Isolated flagellar outer arm dynein translocates brain microtubules in vitro. 296 Sep 3

Dynein, an ATPase, is the force-generating protein in cilia and flagella. It has long been speculated that cytoplasmic microtubules contain a related enzyme involved in cell division or in intracellular organelle transport. A 'cytoplasmic dynein' has been described in sea urchin eggs, but because the egg stockpiles precursors for both cytoplasmic and ciliary microtubules, the role of this enzyme in the cell has remained unresolved. We recently found that the microtubule-associated protein (MAP) 1C (ref. 6) from brain is a microtubule-activated ATPase that produces force in the direction corresponding to retrograde organelle transport in the cell. MAP 1C has several similar properties to ciliary and flagellar dynein. Here we show directly, using scanning transmission electron microscopy, that MAP 1C is structurally equivalent to the ciliary and flagellar enzyme and is the long-sought cytoplasmic analogue of this enzyme.
...
PMID:Microtubule-associated protein 1C from brain is a two-headed cytosolic dynein. 296 91

We recently found that the brain cytosolic microtubule-associated protein 1C (MAP 1C) is a microtubule-activated ATPase, capable of translocating microtubules in vitro in the direction corresponding to retrograde transport. (Paschal, B. M., H. S. Shpetner, and R. B. Vallee. 1987b. J. Cell Biol. 105:1273-1282; Paschal, B. M., and R. B. Vallee. 1987. Nature [Lond.]. 330:181-183.). Biochemical analysis of this protein (op. cit.) as well as scanning transmission electron microscopy revealed that MAP 1C is a brain cytoplasmic form of the ciliary and flagellar ATPase dynein (Vallee, R. B., J. S. Wall, B. M. Paschal, and H. S. Shpetner. 1988. Nature [Lond.]. 332:561-563). We have now characterized the ATPase activity of the brain enzyme in detail. We found that microtubule activation required polymeric tubulin and saturated with increasing tubulin concentration. The maximum activity at saturating tubulin (Vmax) varied from 186 to 239 nmol/min per mg. At low ionic strength, the Km for microtubules was 0.16 mg/ml tubulin, substantially lower than that previously reported for axonemal dynein. The microtubule-stimulated activity was extremely sensitive to changes in ionic strength and sulfhydryl oxidation state, both of which primarily affected the microtubule concentrations required for half-maximal activation. In a number of respects the brain dynein was enzymatically similar to both axonemal and egg dyneins. Thus, the ATPase required divalent cations, calcium stimulating activity less effectively than magnesium. The MgATPase was inhibited by metavandate (Ki = 5-10 microM for the microtubule-stimulated activity), 1 mM NEM, and 1 mM EHNA. In contrast to other dyneins, the brain enzyme hydrolyzed CTP, TTP, and GTP at higher rates than ATP. Thus, the enzymological properties of the brain cytoplasmic dynein are clearly related to those of other dyneins, though the brain enzyme is unique in its substrate specificity and in its high sensitivity to stimulation by microtubules.
...
PMID:Characterization of the microtubule-activated ATPase of brain cytoplasmic dynein (MAP 1C). 297 Oct 69

Microtubules in the cytoplasm of rat Sertoli cell stage VI-VIII testicular seminiferous epithelium were studied morphometrically by electron microscopy. The Sertoli cell microtubules demonstrated axonal features, being largely parallel in orientation and predominantly spaced one to two microtubule diameters apart, suggesting the presence of microtubule-bound spacer molecules. Testis microtubule-associated proteins (MAPs) were isolated by a taxol, salt elution procedure. Testis MAPs promoted microtubule assembly, but to a lesser degree than brain MAPs. High molecular weight MAPs, similar in electrophoretic mobilities to brain MAP-1 and MAP-2, were prominent components of total testis MAPs, though no shared immunoreactivity was detected between testis and brain high molecular weight MAPs using both polyclonal and monoclonal antibodies. Unlike brain high molecular weight MAPs, testis high molecular weight MAPs were not heat stable. Testis MAP composition, studied on postnatal days 5, 10, 15, and 24 and in the adult, changed dramatically during ontogeny. However, the expression of the major testis high molecular weight MAP, called HMW-2, was constitutive and independent of the development of mature germ cells. The Sertoli cell origin of HMW-2 was confirmed by identifying this protein as the major MAP found in an enriched Sertoli cell preparation and in two rat models of testicular injury characterized by germ cell depletion. HMW-2 was selectively released from testis microtubules by ATP and co-purified by sucrose density gradient centrifugation with MAP-1C, a neuronal cytoplasmic dynein. The inhibition of the microtubule-activated ATPase activity of HMW-2 by vanadate and erythro-(2-hydroxy-3-nonyl)adenine and its proteolytic breakdown by vanadate-dependent UV photocleavage confirmed the dynein-like nature of HMW-2. As demonstrated by this study, the neuronal and Sertoli cell cytoskeletons share morphological, structural and functional properties.
...
PMID:Sertoli cell processes have axoplasmic features: an ordered microtubule distribution and an abundant high molecular weight microtubule-associated protein (cytoplasmic dynein). 297 29


1 2 3 4 5 Next >>

---