Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.6.1.3 (ATPase)
 65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Myosin has been purified from the principal pancreatic islet of catfish, hog salivary gland, and hog pituitary. Use of the protease inhibitor Trasylol (FBA Pharmaceuticals, New York) was essential in the isolation of pituitary myosin. Secretory tissue myosins were very similar to smooth muscle myosin, having a heavy chain of 200,000 daltons and light chains of 14,000 and 19,000 daltons. Salivary gland myosin cross-reacted with antibodies directed toward both smooth muscle myosin and fibroblast myosin, but not with antiskeletal muscel myosin serum. The specific myosin ATPase activity measured in 0.6 M KCl was present. Tissues associated with secretion of hormone granules contained substantial amounts of this ATPase, rat pancreatic islets having 4.5 times that of rat liver. Activation of low ionic strength myosin ATPase by actin could not be demonstrated despite adequate binding of the myosin to muscle actin and elution by MgATP. The myosins were located primarily in the cytoplasm as determined by cell fractionation and were quite soluble in buffers of low ionic strength.
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PMID:Myosins of secretory tissues. 15 Apr 27

Rabbit sarcoplasmic reticulum vesicles were fused into giant proteoliposomes in a medium of 0.1 M KCl, 10 mM Tris-maleate, pH 7.0, 10 micrograms ml-1 antipain, 10 micrograms ml-1 leupeptin, 25 IU per ml Trasylol, 3 mM NaN3, 3.75% PEG 1500 and 3% DMSO by brief exposure to 37 degrees C, followed by incubation for 4 h at 25 degrees C. Approximately 5-10% of the sarcoplasmic reticulum elements underwent fusion, forming single-walled spherical vesicles of 1-25 microns diameter, in which the polarity of the native membrane was preserved. The Ca(2+)-stimulated ATPase activity remained essentially unchanged after fusion. On exposure to decavanadate in a Ca(2+)-free medium the spherical vesicles assumed a corrugated appearance with the formation of long ridges separated by deep furrows that eventually pinched off longitudinally and separated into numerous long crystalline tubules of uniform (approximately 0.1 microns) diameter. The vanadate-induced transformation of giant vesicles into tubules implies that the geometry of the sarcoplasmic reticulum membrane is determined by the conformation of the Ca(2+)-ATPase.
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PMID:Giant sarcoplasmic reticulum vesicles: a study of membrane morphogenesis. 128 Nov 63

The Ca(2+)-ATPase crystals formed in detergent solubilized sarcoplasmic reticulum (SR) at 2 degrees C in a crystallization medium of 0.1 M KCl, 10 mM K-Mops (pH 6.0), 3 mM MgCl2, 3 mM NaN3, 5 mM DTT, 25 IU/ml Trasylol, 2 micrograms/ml 1,6-di-tert-butyl-p-cresol, 20% glycerol and 20 mM CaCl2 (J. Biol. Chem. 263, 5277 and 5287 (1988)) contain highly ordered sheets of ATPase molecules, that associate into large multilamellar stacks (greater than 100 layers). When the crystallization is performed in the same medium but in the presence of 40% glycerol at low temperature the stacking is reduced to 4-5 layers and the average diameter of the crystalline sheets is increased from less than 1 micron to 2-3 microns. Glycerol and low temperature presumably reduce stacking by interfering with the interactions between the hydrophilic headgroups of Ca(2+)-ATPase molecules in adjacent lamellae, while not affecting or promoting the ordering of ATPase molecules within the individual sheets. Electron diffraction patterns could be regularly obtained at 8 A and occasionally at 7 A resolution on crystals formed in 40% glycerol, either at 2 degrees C or at -70 degrees C. In the same media but in the absence of glycerol, polyethyleneglycol 1450, 3000 and 8000 (1-8%) induced the formation of ordered crystalline arrays containing 10-12 layers that were similar to those obtained in 40% glycerol. Replacement of 40% glycerol with 10-50% glucose or supplementation of the standard crystallization medium with polyethyleneglycol (PEG 3000 or 8000; 1, 2, 5 and 8%) had no beneficial effect on the order of crystalline arrays compared with media containing 40% glycerol.
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PMID:Effects of solutes on the formation of crystalline sheets of the Ca(2+)-ATPase in detergent-solubilized sarcoplasmic reticulum. 183 35

Conditions were developed for the long-term stabilization of Ca2+-ATPase in detergent-solubilized sarcoplasmic reticulum, purified Ca2+-ATPase, and purified-delipidated Ca2+-ATPase preparations. The standard storage medium contains 0.1 M KCl, 10 mM K-3-(N-morpholino)propanesulfonate, pH 6.0, 3 mM MgCl2, 20 mM CaCl2, 20% glycerol, 3 mM NaN3, 5 mM dithiothreitol, 25 IU/ml Trasylol, 2 micrograms/ml 1,6-di-tert-butyl-p-cresol, 2 mg/ml protein, and 2-4 mg of detergent/mg of protein. Preparations stored under these conditions at 2 degrees C in a nitrogen atmosphere retain significant Ca2+-stimulated ATPase activity for periods of 5-6 months or longer when assayed in the presence of asolectin. The same conditions are also conducive for the formation of three-dimensional microcrystals of Ca2+-ATPase. Of the 49 detergents tested for solubilization, optimal crystallization of Ca2+-ATPase was obtained in sarcoplasmic reticulum solubilized with octaethylene glycol dodecyl ether at a detergent/protein weight ratio of 2, and with Brij 36T, Brij 56, and Brij 96 at a detergent/protein ratio of 4. Similar Ca2+-induced crystals of Ca2+-ATPase were obtained with purified or purified delipidated ATPase preparations at lower detergent/protein ratios. The stabilization of the ATPase activity in the presence of detergents is the combined effect of high Ca2+ (20 mM) and a relatively high glycerol concentration (20%). Ethylene glycol, glucose, sucrose, or myoinositol can substitute for glycerol with preservation of ATPase activity for several weeks in the presence of 20 mM Ca2+.Ca2+-induced association between ATPase molecules may be an essential requirement for preservation of enzymatic activity, both in intact sarcoplasmic reticulum and in solubilized preparations.
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PMID:Stabilization and crystallization of Ca2+-ATPase in detergent-solubilized sarcoplasmic reticulum. 245 65

Crystalline arrays of Ca2+-ATPase molecules develop in detergent-solubilized sarcoplasmic reticulum during incubation for several weeks at 2 degrees C under nitrogen in a medium of 0.1 M KCl, 10 mM K-3-(N-morpholino)propanesulfonate, pH 6.0, 3 mM MgCl2, 20 mM CaCl2, 20% glycerol, 3 mM NaN3, 5 mM dithiothreitol, 25 IU/ml Trasylol, 2 micrograms/ml 1,6-di-tert-butyl-p-cresol, 2 mg/ml protein, and 2-4 mg of detergent/mg of protein. Electron microscopy of sectioned, negatively stained, freeze-fractured, and frozen-hydrated Ca2+-ATPase crystals indicates that they consist of stacked lamellar arrays of Ca2+-ATPase molecules. Prominent periodicities of ATPase molecules within the lamellae arise from a centered rectangular lattice of dimensions 164 x 55.5 A. The association of lamellae into three-dimensional stacks is assumed to involve interactions between the exposed hydrophilic headgroups of ATPase molecules, that is promoted by glycerol and 20 mM Ca2+. Similar Ca2+-induced crystals were observed with purified or purified and delipidated Ca2+-ATPase preparations at lower detergent/protein ratios. Cross-linking of Ca2+-ATPase crystals with glutaraldehyde protects the structure against conditions such as low Ca2+, high pH, elevated temperature, SH group reagents, high concentration of detergents, and removal of phospholipids by extraction with organic solvents that disrupt unfixed preparations.
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PMID:Electron microscope observations on Ca2+-ATPase microcrystals in detergent-solubilized sarcoplasmic reticulum. 296

Multilamellar 3-dimensional (Type I) microcrystals of detergent-solubilized crude microsomes or purified protein preparations of membrane-bound gastric (H+, K+)-ATPase from rabbit or hog stomachs develop in media consisting of 0.1 M KCl, 20 mM imidazole, 5 mM MgCl2, 3 mM NaN3, 5 mM DTT, 25 IU/ml Trasylol, 2 micrograms/ml DTBpC and 20-40% glycerol, using nonionic detergent of C12E8 or BRIJ 36 for solubilization. Crystals developed in a pH-range of 6.0-7.25, during 3-10 days of incubation, at 2 degrees C. For C12E8, the most effective detergent:protein ratio for crystallization varied between (1.8-2.0):1 for the microsomes and between (0.25-0.75):1 for the purified preparations. The results of biochemical and structural analysis of the (H+, K+)-ATPase crystals showed close resemblance to those of the sarcoplasmic reticulum Ca(2+)-ATPase from skeletal muscle and plasmamembrane (Na+, K+)-ATPase from kidney (J. Biol. Chem., 269, 10107-111, 1994). Based on these identities and the high (62%) overall sequential homology to the (Na+, K+)-ATPase, we conclude that the new crystals of the (H+, K+)-ATPase could also contain only the alpha-chain of the alpha beta heterodimers found in the native membrane. High-resolution electron microscopy of frozen-hydrated crystalline (H+, K+)-ATPase samples are in progress to give unit cell dimensions and molecular packing of the new crystals.
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PMID:Three-dimensional (type I) microcrystals of detergent-solubilized membrane-bound gastric (H+, K+)-ATPase enzyme from hog and rabbit stomachs. 778 47

Multilamellar 3-dimensional (Type I) crystals of detergent-solubilized, purified (Na+, K+)-ATPase enzyme of pig kidney grow in media consisting of 0.1 M KCl, 0.1 M NaCl, 20 mM imidazole pH: 7.5 (20 degrees C), 5 mM MgCl2, 5 mM DTT, 3 mM ZNaN3, 0.025 TIU/ml Aprotinin, 2 micrograms/ml BHT and 20-40% glycerol, using nonionic detergents of C12E8 or BRIJ 36 for solubilization. The refined crystallization protocol: the use of media containing 20% glycerol, the low detergent: protein ratio and preincubation at subzero temperature at the initial phase of crystallization resulted in a remarkable increase of the yield and overall dimension of the crystals (up to 3-4 microns), while the stacking of the crystalline sheets was dramatically reduced. Biochemical and structural analysis of these crystals revealed further similarities between the 3-d crystals of the (Na+, K+)-ATPase and the Ca2+ ATPase of skeletal muscle-SR (Taylor and Varga, J. Biol. Chem., 269, 10107-10111, 1994). Computer image processing of the electron micrographs of stacked crystalline sheets of (Na+,K+)-ATPase molecules gave unit cell dimensions: a = 166.2 +/- 3.8 A, b = 54.2 +/- 3.5 A, with an included angle of 90 degrees C. Based on the close identity of the filtered images in projection and of other data, we concluded that the 3-dimensional crystals of the (Na+, K+)-ATPase contain only the alpha catalytic subunits.
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PMID:Further characterization of the 3-dimensional crystals of detergent-solubilized (Na+,K+)-ATPase from pig kidney. 788 74