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)

Steady-state ATPase activity, calcium binding, formation of phosphorylated enzyme intermediate with ATP in the presence of Ca2+, or with Pi in the absence of Ca2+, and association of ATPase molecules into bidimensional crystals, were studied using vesicular fragments of sarcoplasmic reticulum. The vesicles were exposed to increasing concentrations of urea in order to produce stepwise perturbations of protein structure and to test the effect of such perturbations on the partial reactions and crystallization pattern of sarcoplasmic reticulum ATPase. It was found that low concentrations of urea produce specific inhibition of Pi binding and enzyme phosphorylation with Pi (but not with ATP). Intermediate concentrations of urea reduce calcium binding affinity and cooperativity, while the ability of the enzyme to be phosphorylated with ATP and to form dimeric arrays is retained. These observations demonstrate that the sarcoplasmic reticulum ATPase is sensitive to physical perturbations producing specific and reversible changes in the Pi and calcium binding domains. These changes interfere with enzyme turnover, indicating that conformational effects related to binding and dissociation of Pi and calcium are tightly coupled to catalysis and energy transduction. Higher concentrations of urea produce irreversible denaturation, accompanied by total inhibition of calcium binding, enzyme phosphorylation with ATP, and association of ATPase chains in bidimensional crystals. Under these conditions, protein unfolding is manifested by a sharp reduction in the fluorescence of intrinsic tryptophan residues and of a covalently bound probe. These observations suggest that dimeric association and a tendency to form bidimensional crystals correspond to a basic property of the enzyme, which is linked to its native structure and whose character may change in the presence of ligands and/or during the catalytic cycle. On the other hand, the decavanadate-induced crystallization pattern cannot be interpreted in terms of a mechanistic relationship of ATPase dimerization with one of the intermediate states of the catalytic cycle.
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PMID:Effect of urea on the partial reactions and crystallization pattern of sarcoplasmic reticulum adenosine triphosphatase. 297 Aug 23

The Ca2+ binding site region of the Ca2+-ATPase of skeletal muscle sarcoplasmic reticulum was labeled with several fluorescent analogs of dicyclohexylcarbodiimide. As has been shown by Chadwick and Thomas, in the absence of Ca2+ in the medium, labeling with the naphthyl carbodiimide results in the inhibition of enzyme activity. Further, Ca2+ occupancy of the high affinity sites of the enzyme protects against incorporation into the site(s). The fluorescent carbodiimide has been used to determine the depth of the site of label incorporation relative to the aqueous-bilayer interfaces by quenching studies using spin-labeled fatty acid derivatives. The series of quenchers used have their spin-label moiety located at different positions along the fatty acid chain. It was found that after suitable correction for differences in partitioning of the various derivatives, the order of quenching efficiency was 16- greater than 12- greater than 10- greater than 7- greater than 5-NS, indicating that the naphthyl moiety is near the center of the bilayer. In contrast, quenching with the aqueous-restricted I- indicated that the label is accessible from the external milieu, likewise for a presumed aqueous quencher, acrylamide. The aqueous quenchers accessibilities were altered upon Ca2+ binding to the ATPase. Quenching of the intrinsic fluorescence with the x-NS derivatives indicates that the ATPase tryptophan residues are primarily localized at the aqueous-membrane interfaces, with the order of quenching being 5- greater than 7- greater than 10- greater than 12- greater than 16-NS. The trp residue(s) which changes its fluorescence upon Ca2+ binding is shown to be near the membrane surface.
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PMID:Molecular topography of the Ca2+-ATPase of sarcoplasmic reticulum. 297 13

Phosphorescence of protein tryptophan was analyzed in sarcoplasmic reticulum vesicles, and in the purified Ca2+ transport ATPase in deoxygenated aqueous solutions at room temperature. Upon excitation with light of 295 nm wavelength, the emission maxima of fluorescence and phosphorescence were at 330 nm and at 445 nm, respectively. The phosphorescence decay was multiexponential; the lifetime of the long-lived component of phosphorescence was approximately equal to 22 ms. ATP and vandate significantly reduced the phosphorescence in the presence of either Ca2+ or EGTA; ADP was less effective, while AMP was without effect. The quenching by ATP showed saturation consistent with the idea that the ATP-enzyme complex had a lower phosphorescence yield. Upon exhaustion of ATP, the phosphorescence returned to starting level. Significant quenching of phosphorescence with a decrease in phosphorescence lifetime was also caused by NaNO2, methylvinyl ketone and trichloroacetate, without effect on ATPase activity; this quenching did not show saturation and was therefore probably collisional in nature.
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PMID:Tryptophan phosphorescence of the Ca2+-ATPase of sarcoplasmic reticulum. 297 55

Ultraviolet irradiation above 300 nm of the stable MgADP-orthovanadate (Vi)-myosin subfragment 1 (S1) complex resulted in covalent modification of the S1 and in the rapid release of trapped MgADP and Vi. This photomodified S1 had Ca2+ATPase activity 4-5-fold higher than that of the non-irradiated control S1, while the K+EDTA-ATPase activity was below 10% of controls. There was a linear correlation between the activation of the Ca2+ATPase and the release of both ADP and Vi with irradiation time. Analysis of the total number of thiols and the ability of photomodified S1 to retrap MgADP by cross-linking SH1 and SH2 with various bifunctional thiol reagents indicated that the photomodification did not involve these reactive thiols. Irradiation of the S1-MgADP-Vi complex caused a large increase in absorbance of the enzyme at 270 nm which was correlated with the release of Vi from the active site, suggesting an aromatic amino acid(s) was (were) involved. However, analysis by three different methods showed no loss of tryptophan. All the irradiation-dependent phenomena could be prevented by replacing Mg2+ with either Co2+, Mn2+, or Ni2+. Unlike previous irradiation studies of Vi-dynein complexes [Lee-Eiford, A., Ow, R. A., & Gibbons, I. R. (1986) J. Biol. Chem. 261, 2337-2342], no peptide bonds were cleaved in photomodified S1. Photomodified S1 was able to retrap MgADP-Vi at levels similar to unmodified S1. Upon irradiation of the photomodified S1-MgADP-Vi complex, MgADP and Vi were again released from the active site, resulting in heavy chain cleavage to form NH2-terminal 21-kDa and COOH-terminal 74-kDa peptides. All evidence indicates that this new photomodification and subsequent chain cleavage occur specifically at the active site.
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PMID:UV-induced vanadate-dependent modification and cleavage of skeletal myosin subfragment 1 heavy chain. 1. Evidence for active site modification. 297 86

Changes in F-actin conformation in myosin-free single ghost fibers of rabbit skeletal muscle induced by the binding of skeletal and gizzard tropomyosin to F-actin were studied by measuring intrinsic tryptophan-polarized fluorescence of F-actin. It was found that skeletal and gizzard tropomyosin binding to F-actin initiate different conformational changes in actin filaments. Skeletal tropomyosin inhibits, while gizzard tropomyosin activates the Mg2+-ATPase activity of skeletal actomyosin. It is supposed that in muscle fibers tropomyosin modulates the ATPase activity of actomyosin via conformational changes in F-actin.
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PMID:[Tropomyosin from smooth and skeletal muscles initiates various conformational changes in skeletal F-actin]. 297 8

To investigate the mechanisms by which hydrostatic pressure inhibits (Na,K)-ATPase, we measured enzyme activity, as a function of pressure and temperature, of purified (Na,K)-ATPase from dog kidney and eel electroplax, and we monitored protein conformation, possible subunit interactions, and the fluidity of the membrane with fluorescent probes. The (Na,K)-ATPase and p-nitrophenylphosphatase activities were inhibited reversibly by pressures below 1.5 kilobars (eel enzyme) and 2.5 kilobars (dog kidney enzyme). Above these pressures, the enzymes were inactivated irreversibly. The plots of 1n(activity) versus pressure were curvilinear; this indicates that the reversible inhibition by pressure involves two or more rate-limiting steps. The calculated activation volumes varied with temperature and pressure and were larger for the (Na,K)-ATPase activity compared to the p-nitrophenylphosphatase activity. The fluorescence polarization of three hydrophobic probes decreased with increasing temperature (10-36 degrees C) and increased with increasing pressure (10(-3)-1.5 kilobars), reversibly, without any evidence of a lipid phase transition. Plots of enzyme activity versus fluorescence polarization of the lipid probes showed an inverse relationship; this indicates that enzyme activity was directly related to the fluidity of the membrane as measured by the lipid probes. Pressure had no effect on the fluorescence polarization of two cardiac glycoside probes nor on the efficiency of resonance energy transfer between either donor and acceptor cardiac glycosides specifically bound to the ouabain sites of different alpha-subunits, or tryptophan and the bound cardiac glycoside probe. These results suggest that dissociation of dimeric alpha-subunits is not related to the inhibition by pressure, and that the cardiac glycoside-complexed enzyme conformation is stabilized by pressure. It is concluded that increased pressure decreases the membrane fluidity which hinders conformational transitions associated with rate-limiting steps of the (Na,K)-ATPase reaction. It is proposed that ion-bound or -occluded forms of (Na,K)-ATPase are stabilized by pressure because they occupy a smaller volume.
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PMID:Mechanisms of inhibition of (Na,K)-ATPase by hydrostatic pressure studied with fluorescent probes. 299 10

The sequence of the 16,019 nucleotide-pair mitochondrial DNA (mtDNA) molecule of Drosophila yakuba is presented. This molecule contains the genes for two rRNAs, 22 tRNAs, six identified proteins [cytochrome b, cytochrome c oxidase subunits I, II, and III (COI-III), and ATPase subunits 6 and 8] and seven presumptive proteins (URF1-6 and URF4L). Replication originates within a region of 1077 nucleotides that is 92.8% A + T and lacks any open reading frame larger than 123 nucleotides. An equivalent to the sequence found in all mammalian mtCDNAs that is associated with initiation of second-strand DNA synthesis is not present in D. yakuba mtDNA. Introns are absent from D. yakuba mitochondrial genes and there are few (0-31) intergenic nucleotides. The genes found in D. yakuba and mammalian mtDNAs are the same, but there are differences in their arrangement and in the relative proportions of the complementary strands of the molecule that serve as templates for transcription. Although the D. yakuba small and large mitochondrial rRNA genes are exceptionally low in G and C and are shorter than any other metazoan rRNA genes reported, they can be folded into secondary structures remarkably similar to the secondary structures proposed for mammalian mitochondrial rRNAs. D. yakuba mitochondrial tRNA genes, like their mammalian counterparts, are more variable in sequence than nonorganelle tRNAs. In mitochondrial protein genes ATG, ATT, ATA, and in one case (COI) ATAA appear to be used as translation initiation codons. The only termination codon found in these genes is TAA. In the D. yakuba mitochondrial genetic code, AGA, ATA, and TGA specify serine, isoleucine, and tryptophan, respectively. Fifty-nine types of sense condon are used in the D. yakuba mitochondrial protein genes, but 93.8% of all codons end in A or T. Codon-anticodon interactions may include both G-A and C-A pairing in the wobble position. Evidence is summarized that supports the hypothesis that A and T nucleotides are favored at all locations in the D. yakuba mtDNA molecule where these nucleotides are compatible with function.
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PMID:The mitochondrial DNA molecular of Drosophila yakuba: nucleotide sequence, gene organization, and genetic code. 300 25

The effect of prenatal L-tryptophan supplementation on the serotonin (5-HT) synthesis and the activity of Na+,K+-ATPase in the cerebral cortex was studied during postnatal development, from birth up to day 30. A parallel and significant elevation of the serotonin content and the activity of tryptophan-5-hydroxylase was observed in the brain of infant rats born to mothers treated with L-tryptophan, as related to non-treated controls. The activity of Na+,K+-ATPase was also significantly elevated at the different ages studied throughout the developmental period, as related to controls. These results suggest an important role of L-tryptophan in the early regulation of the serotonin-synthesizing machinery, which lasts postnatally. Elevation of ATPase activity seems to be associated to the elevation in the activity of the 5-HT system.
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PMID:Brain serotonin synthesis and Na+,K+-ATPase activity are increased postnatally after prenatal administration of L-tryptophan. 300 68

Activities and some properties of microsomal ATPases have been studied in developing human placenta. The enzyme activities (Na+ + K+ + Mg2+, Mg2+, and Ca2+ dependent) in the placenta increase steadily with gestational age until the 18th to 21st week, and decrease in the second half of pregnancy. Mg2+-dependent and Na+ + K+ + Mg2+-dependent ATPases possess nearly the same Km (apparent) for ATP, while the Ca2+-dependent enzyme shows a different one. Mg2+-dependent ATPase shows higher substrate affinity than Ca2+-dependent ATPase, although the Vmax of the Mg2+-dependent enzyme is lower than that of the latter. However, for each enzyme, the Km remains almost constant and Vmax varies during ontogenic development. Vmax of the enzymes decline at term. The enzymes are heat-labile, unaffected by amino acids, namely, L-phenylalanine, L-leucine, and L-tryptophan, and deoxycholate inhibits the enzyme activities by about 50%.
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PMID:Characterization of microsomal ATPases from developing human placenta. 301 Oct 35

Mature rats were dosed with T3 by different routes and dose-levels at either 0.1 mg/kg for 14 days s.c. (Group A), 1 mg/kg for 3 alternative days i.p. (Group B), 5 mg/kg for 14 days p.o. (Group C), or with propylthiouracil (PTU 50 mg/day for 14 days p.o.-Group D). Measurement of cerebellar and striatal NA+,K+-ATPase activities showed that whereas Groups A, B and D were unaffected when compared with controls, there were 35-70% increases respectively in the activities of both molecular forms of the enzyme, alpha(+), high ouabain affinity, and alpha, low ouabain affinity, in Group C rat brains at the highest dose of T3 tested. Kidney Na+,K+-ATPase activity was also elevated (67% increase) in this group of animals showing significant changes in renal medullary tissue only. Acute elevation of brain dopamine levels by administration of an MAOI plus L-DOPA (50 mg/kg, 60 min) significantly elevated (20% increase) the activities of both molecular forms of Na+,K+-ATPase in corpus striatum. Treatment with L-tryptophan (50 mg/kg, 60 min) failed to produce any changes in the striatal activities. The possible relationship of increases in enzyme activities with T3 and increased brain monoamine function is discussed. Both plasma free T4(FT4) and total T4(TT4) were markedly depressed in all T3-treated rats. Although hypothalamic thyrotropin releasing hormone (TRH) concentrations were unaltered by any of the T3 treatments, pituitary thyroid stimulating hormone (TSH) concentrations were greatly diminished and it is thought that this may reflect a direct effect of T3 on TSH synthesis.
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PMID:Studies on the effect of chronic L-triiodothyronine (T3) treatment on brain Na+,K+-ATPase activity in the mature rat. 302 20


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