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

Purified skeletal muscle myosin (EC 3.6.1.3) has been covalently bound to Sepharose 4B by the cyanogen bromide procedure. The resulting complex, Sepharose-Myosin, possesses adenosine triphosphatase activity and is relatively stable for long periods of time. Under optimal binding conditions, approximately 33% of the specific ATPase activity of the bound myosin is retained. Polyacrylamide gel electrophoresis of polypeptides released from denatured Sepharose-Myosin indicates that 85% of the myosin is attached to the agarose beads through the heavy chains and the remainder through the light chains, in agreement with predictions of binding and release based upon either the lysine contents or molecular weights of themyosin subunits. The adenosine triphosphatase of the immobilized myosin has been investigated under conditions of varying pH, ionic strength, and cation concentration. The ATPase profiles of immobilized myosin are quite similar to those for free myosin, however subtle differences are found. The Sepharose-Myosin ATPase is not as sensitive as myosin to alterations in salt concentration and the apparent KM is approximately two-fold higher than that of myosin. These differences are probably due to chemical modification in the region of the attachment site(s) to the agarose beads and hydration and diffusion limitations imposed by the polymeric agarose matrix.
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PMID:Preparation and characterization of an enzymatically active immobilized derivative of myosin. 0 72

1. The terminal phosphate of (gamma-32P)ATP is rapidly incorporated into cardiac sarcoplasmic reticulum membranes (0.7--1.3 mumol/g protein) in the presence of calcium and magnesium. Cardiac sarcoplasmic reticulum membranes catalize an ATP-ADP phosphate exchange in the presence of calcium and magnesium. 2. Half-maximum activation of the phosphoprotein formation and ATP-ADP phosphate exchange is reached at an ionized calcium concentration of about 0.3 muM. The Hill coefficients are 1.3. 3. Transphosphorylation and ATP-ADP phosphate exchange require magnesium and are maximally activated at magnesium concentrations close to or equal to the ATP concentration. 4. The phosphoprotein level is reduced to about 45% at an ADP/ATP ratio of 0.1. The rate of calcium-dependent ATP splitting declines, whilst the rate of the calcium-dependent ATP-ADP phosphate exchange increases when the ADP/ATP ratio is varied from 0.1 to 1. The sum of both, the rate of ATP splitting and the rate of ADP-ATP phosphate exchange remains constant. 5. Phosphoprotein formation and ATP-ADP phosphate exchange are not affected by azide, dinitrophenol, dicyclohexyl carbodiimide and oubain, whilst both activities are reduced by blockade of -SH groups localized on the outside of the sarcoplasmic reticulum membrane. 6. The isolated phosphoprotein is acid stable. The trichloroacetic acid denatured 32P-labelled membrane complex is dephosphorylated by hydroxylamine, which might indicate that the phosphorylated protein is an acyl-phosphate. 7. Polyacrylamide gel elctrophoresis (performed with phenol/acetic acid/water) of phosphorylated sarcoplasmic reticulum fractions demonstrates that the 32P-incorporation occurs into a protein of about 100000 molecular weight. 8. It is suggested that the phosphoprotein represents a phosphorylated intermediate of the calcium-dependent ATPase which formation occurs as an early step in the reaction sequence of calcium translocation by cardiac sarcoplasmic reticulum similar as in skeletal muscle.
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PMID:Characterization of cardiac sarcoplasmic reticulum ATP-ADP phosphate exchange and phosphorylation of the calcium transport adenosine triphosphatase. 0 67

H+-Translocating ATPase, which catalyzes ATP synthesis in biomembranes, is composed of a head piece (F1) and a membrane moiety (F0). Using highly-purified F0 from a thermophilic bacterium PS3 (TF0), the following results were obtained. 1. Inhibition by N,N'-dicyclohexylcarbodiimide (DCCD) of H+ conduction through TF0 followed pseudo-first-order kinetics. The second-order rate constant for inhibitor-enzyme interaction was 5 times 10(3) M(-1)-min(-1). 2. H+ conductivity blocked by DCCD was proportional to the amount of DCCD incorporated in the band 8 protein of TF0. When only one-third of the band 8 protein was labeled with DCCD, TF0 hardly transported any H+. 3. By extracting TF0 with chloroform-methanol, the band 8 protein was obtained as a proteolipid. Polyacrylamide gel electrophoresis with dodecyl sulfate and urea showed that the molecular weight was about 6,000. 4. The amino acid composition of band 8 protein indicated that this protein contained an extremely high percentage of hydrophobic amino acids (0.29 in polarity) and was devoid of histidine, tryptophan, cysteine, and lysine. Its minimum molecular weight was 6,500. 5. The role of band 8 protein (DCCD-binding protein) in H+ conduction through TF0 is discussed on the basis of these results.
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PMID:Carbodiimide-binding protein of H+-translocating ATPase and inhibition of H+ conduction by dicyclohexylcarbodiimide. 3 78

The Mg2+ -dependent ATPase (EC 3.6.I.3) of Proteus L-form membrane has been solubilized according to various procedures (Tris - HCL shock-wash with or without MG2+, EDTA, Triton X-100). The best results were obtained by the same 33mM Tris-HCL (pH 7.5) shock-wash without MG2+ used for ATPase of protoplasts from Streptococcus faecalis. The solubilized enzyme after 105 000 times g centrifugation was purified on acrylamide/agarose. The molecular weight was established to be 360 000 by gel filtration and by sedimentation coefficient (12.5 S). Polyacrylamide disc-gel electrophoresis in sodium dodecylsulphate revealed two classes or subunit of mol. wt. 64 000 (alpha) and 58 000 (beta), associated in ratio 1:1. We propose a formula alpha-3beta-3 for the native ATPase of Proteus L-forms. Structural similarities to ATPase of various origins are discussed.
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PMID:Membrane ATPase of Proteus L-forms. Solubilization and molecular properties. 12 72

Recovery of calcium transport and calcium-activated ATPase activity was studied in relation to the retention of protein components in sarcoplasmic reticulum reconstituted after solubilization with deoxycholate and centrifugation, followed by removal of the detergent from the supernatant by dialysis. Control sarcoplasmic reticulum was similarly treated except for omission of deoxycholate. Maximum capacity for oxalate- and phosphate-supported calcium uptake was increased 2- to 3-fold in reconstituted sarcoplasmic reticulum compared to original and control. Calcium uptake velocity of the reconstituted sarcoplasmic reticulum was approximately 80% that of original and 90% of control sarcoplasmic reticulum. Calcium uptake/ATP hydrolysis ratio was approximately 2 in the original sarcoplasmic reticulum and decreased to approximately 1 in the control and reconstituted sarcoplasmic reticulum. Calcium storage in the absence of calcium-precipitating anion was approximately 85% in control and 70% in reconstituted sarcoplasmic reticulum, compared to the original sarcoplasmic reticulum. Ethylene glycol bis(beta-aminoethyl ether)-N,N'-tetraacetic acid-induced calcium release after phosphate-supported calcium uptake was slower in reconstituted sarcoplasmic reticulum than in original or control sarcoplasmic reticulum. Polyacrylamide gel electrophoresis of original and control sarcoplasmic reticulum showed similar amounts of protein components of approximately 93,000, 59,000, 50,000, 30,000 to 37,000, and 20,000 to 26,000 daltons. Reconstituted sarcoplasmic reticulum, however, lost over 85% of the 50,000- and 20,000- to 26,000-dalton proteins while retaining most of its calcium transport functions.
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PMID:Reconstitution of an active calcium pump in sarcoplasmic reticulum. 13 3

Purified right-side-out (RSO) and inside-out (IO) plasma membrane vesicles release 35% of the total plasma membrane proteins after EDTA treatment. After such a treatment both types of vesicles exhibited the same total activity of (Na+ + K+)-stimulated Mg2+ adenosinetriphosphatase (ATPase; ATP phosphohydrolase, EC 3.6.1.3) as in their native state. The EDTA treatment increases the enzyme sensitivity to ouabain by 350-fold in IO vesicles while being without any effect RSO vesicles. Thus, proteins released only from the IO vesicles led to a change in ouabain sensitivity of the (Na+ + K+)-stimulated Mg2+ ATPase. Moreover, only proteins released from IO vesicles, when added to treated IO vesicles with divalent cations, were able to restore the original resistance of the enzyme to ouabain; released proteins from RSO vesicles failed to make such a reconstitution. Thus, we assume that these proteins detach from the inner face of the plasma membrane upon EDTA treatment and are distinct from the enzyme. Polyacrylamide gel electrophoresis indicates that these inner face plasma membrane proteins are approximately 30,000 daltons.
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PMID:Roles of proteins from inner face of plasma membranes in susceptibility of of (Na+ + K+)-stimulated Mg2+ adenosinetriphosphatase to ouabain. 13 8

Purified plasma membranes of Schizosaccharomyces pombe were obtained by precipitation at pH 5.2 of a crude particulate fraction, followed by differential centrifugations and isopycnic centrifugation in a discontinuous sucrose gradient. The specific activity of the Mg2+-requiring plasma membrane ATPase activity (EC 3.6.1.3) was enriched from 0.3 mumol min-1 x mg-1 of protein in the homogenate to 26 in the purified membranes. The optimal conditions for solubilization of the ATPase activity by lysolecithin were found to be: 2 mg/ml of lysolecithin, a lysolecithin to protein ratio of 8 at pH 7.5, and 15 degrees C in the presence of 1 mM ATP and 1 mM ethylenediaminetetraacetic acid. A 6- to 7-fold purification of the solubilized ATPase activity was obtained by centrifugation of the lysolecithin extract in sucrose gradient. Part of the ATPase activity which was inactivated during the centrifugation in the sucrose gradient could be restored by addition of a micellar solution of 50 microgram of lysolecithin/ml during the assay. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate of the purified enzyme showed only one band of Mr = 105,000 stained with Coomassie blue. Another ATPase component of apparent molecular weight lower than 10,000 was stained by periodic Schiff reagent but not colored by Coomassie blue. The purified enzyme was 85% inhibited by 50 micrometer N,N'-dicyclohexylcarbodiimide and 94% inhibited by 53 microgram of Dio-9/ml.
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PMID:Solubilization by lysolecithin and purification of the plasma membrane ATPase of the yeast Schizosaccharomyces pombe. 15 Oct 99

The costa is a rod-shaped intracellular organelle lying in the cytoplasm immediately below the undulating membrane. In certain large species of Trichomonas (T. gigantea, T. termopsidis and an undescribed species from the termite Porotermes adamsoni) this organelle is motile. Bending waves are transmitted along the length of the costa, in either direction. It is shown that the bending is actively produced by the costa itself. The form of the movements is described in detail. The costa is birefringent. At the point at which bending occurs there is a sharply localized reduction in birefringence. Electron microscopy shows that the costa is composed of longitudinally running lamellae, 2--3 nm thick and spaced 12 nm apart. These are connected to transversely running elements spaced at intervals of about 37 nm. The lamellae occur in two alternative configurations: straight and zig-zag. Bending probably results from a local, transient change from the straight to the zig-zag configuration. This would account for the local change in birefringence which accompanies bending. Polyacrylamide gel electrophoresis of isolated costas shows them to contain a number of protein components, of which the principal one has a molecular weight of about 90,000. Preliminary cytochemical evidence is presented for ATPase activity in the costa. The costa is a type of motile system distinct from any hitherto described.
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PMID:Structure, protein composition and birefringence of the costa: a motile flagellar root fibre in the flagellate Trichomonas. 15 23

1. Isolation of ATPase from rat liver submitochondrial particles by chloroform treatment requires the presence of ATP or ADP during enzyme solubilization. In the absence of adenine nucleotides the enzyme activity is very low although all protein components of F1-ATPase are released. The low concentrations of ATP or ADP required (5 microM) indicate that the high affinity nucleotide-binding sites are involved in enzyme stabilization. Other nucleotides tested (ITP, GTP, UTP, CTP) were found to be less effective. 2. Polyacrylamide gel electrophoresis and immunodiffusion in agar plates revealed that in the absence of adenine nucleotides a fraction of F1-ATPase released by chloroform treatment is split into fragments. The part of the dissociated enzyme molecule has a molecular weight identical with that of a beta-subunit of F1-ATPase. 3. Dissociation of the F1-ATPase molecule could also be prevented by aurovertin. 4. Crude F1-ATPase solubilized by chloroform treatment can be further purified by Sepharose 6B gel filtration. Specific ATPase activity of the purified enzyme was 90 mumol Pi/min per mg protein and the enzyme was composed of five protein subunits (alpha, beta, gamma, delta, epsilon) with molecular weights 58 000, 55 000, 28 000, 13 000 and 8000, respectively. 5. Chloroform-released F1-ATPase from rat liver mitochondria displayed immunochemical cross-reactivity with that isolated from beef heart mitochondria.
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PMID:Stabilization of rat liver mitochondrial F1-adenosine triphosphatase during chloroform-induced solubilization. 15 60

Bacillus megaterium KM dormant spore inner membrane ATPase exhibits a ten-fold increase in specific activity during the first 10 min of germination in the absence of protein synthesis. During this time period both in the presence and absence of chloramphenicol extensive proteolysis of spore inner membrane takes place, which results in degradation of approximately half of the membrane protein. Polyacrylamide gel electrophoresis reveals the extent and selectivity of this degradation of inner membrane protein.
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PMID:Membrane changes during germination of Bacillus megaterium KM spores. 16 97


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