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)

In contrast with wild-type Salmonella typhimurium LT2, strain HfrA did not have ATP-driven energy-dependent transhydrogenase activity, although ATP-dependent quenching of atebrin fluorescence was normal. Respiration-dependent and energy-independent transhydrogenase, and Ca2+-activated ATPase (adenosine triphosphatase) activities were similar in both strains. Purified ATPases from the two strains had similar specific activities, similar subunit polypeptides, and were equally effective in restoring energy-dependent transhydrogenase activities to membrane particles of strain LT2 from which the ATPase had been stripped. The purified ATPases from both strains could restore respiration-dependent but not ATP-dependent transhydrogenation to stripped particles of strain HfrA. Both strains grew aerobically equally well on salts media containing glucose, malate, succinate, citrate, acetate, pyruvate, fumarate, lactate or aspartate as substrates. Growth on glucose under anaerobic conditions was similar. Strains LT2 and HfrA were equally effective in the accumulation under both aerobic and anaerobic conditions of the amino acids proline, phenylalanine, histidine, lysine, isoleucine and aspartic acid. Inhibition of amino acid accumulation by KCN and dicyclohexylcarbodi-imide occurred to the same extent in both strains. The complete inhibition by dicyclohexylcarbodi-imide of amino acid uptake under anaerobic conditions suggested that ATP could drive amino acid uptake in both strains. The ability of strain HfrA to carry out ATP-dependent transport or quenching of atebrin fluorescence but not ATP-dependent transhydrogenation is different from the wild-type strain and from any previously described energy-coupling mutant. It is difficult to reconcile the properties of this mutant with the chemiosmotic hypothesis.
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PMID:Salmonella typhimurium HfrA, a mutant in which adenosine triphosphate can drive amino acid transport but not energy-dependent nicotinamide nucleotide transhydrogenation. 12 57

We described previously the existence of a soluble ATPase activity in rat liver mitochondria [1]. The purification and catalytic properties have been described [2]. In a continuation of these experiments, we have studied the immunologic and structural properties of one molecular form of this enzyme : ATPase I. We have prepared the antiserum anti-ATPase I and demonstrated the purity of our enzyme preparation by immunodiffusion and immunoelectrophoresis. An immunohistochemical method also confirmed the localization of ATPase I in the soluble fraction of mitochondria. The molecular weight of ATPase I was measured by G 100 Sephadex gel filtration and was found to be 18,400; electrophoresis on polyacrylamide gels gave a value of 18,600. The pHi of ATPase I was found to be 7,2. Amino acid analysis showed high amounts of aspartic acid, glutamic acid, serine and glycine. The molecular weight calculated from the total amino acid residues was found to be 17,000. Alanine is the NH2 terminal amino acid. The peptide maps obtained after degrading ATPase I with cyanogen bromide or trypsin are in accordance with the methionine, lysine and arginine residues we found in the ATPase I molecule. ATPase I does not appear to be a glycoprotein.
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PMID:Studies of soluble rat liver mitochondrial acid ATPases. II. Structural and immunological properties of ATPase 1. 15 69

Arg-210 of the a subunit of the Escherichia coli F0F1-ATPase has been proposed previously as a component of the proton pore. A mutant in which lysine was substituted for Arg-210 was generated and was found to be unable to translocate protons. A plasmid carrying this mutation, along with wild-type genes encoding the c and b subunits, was unusual in that it failed to complement a chromosomal c-subunit mutation on succinate minimal medium. Three revertants on succinate minimal medium contained plasmids that showed complementation with chromosomal c-subunit but not with a-subunit mutations. One of these had a deletion in the a subunit. The other two were point mutations, resulting in the substitution of aspartic acid by Gly-53 and of arginine for Leu-211. The Gly-53 to aspartic acid change implied that Gly-53 and Arg-210 are normally in close proximity. To test this idea further, a series of mutants in which aspartic acid was placed in helix I at positions ranging from 42 to 57 was generated. Full complementation was regained only when the aspartic acid residue was present on the same side of a putative helix as Gly-53 over a span of three turns of the alpha-helix. These results and others suggest modifications of a previously proposed model for the transmembrane helices of the F0 portion of the F0F1-ATPase. The implications of these modifications for the mechanism of proton translocation are discussed.
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PMID:Second-site revertants of an arginine-210 to lysine mutation in the a subunit of the F0F1-ATPase from Escherichia coli: implications for structure. 140 2

Yeast mutants in which genes encoding subunits of the vacuolar H(+)-ATPase were interrupted were assayed for their vacuolar ATPase and proton-uptake activities. The vacuoles from the mutants lacking subunits A (72 kDa), B (57 kDa), or c (proteolipid, 16 kDa) were completely inactive in these reactions. Immunological studies revealed that in the absence of each one of those subunits the catalytic sector was not assembled. Labeling with N,N'-[14C]dicyclohexylcarbodiimide showed the presence of the proteolipid in vacuoles of mutants in which genes encoding subunits of the catalytic sectors were interrupted. No labeling was detected in the mutant in which the gene encoding the proteolipid was interrupted. We conclude that of all the ATPase subunits only the proteolipid is assembled independently and it serves as a template for the assembly of the other subunits. Site-specific mutations were generated in the gene encoding the proteolipid. All of the drastic changes and replacements gave inactive proteins. About half of the single amino acid replacements gave active proteins. Replacing glutamic acid-137 by any of several amino acids, except for aspartic acid, abolished the activity of the enzyme. Other amino acids that may function in proton conductance were changed. It was found that glycine residues may replace amino acids with exchangeable protons.
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PMID:Mutational analysis of yeast vacuolar H(+)-ATPase. 182 30

The Ca2+/Mg2+ ATPase, which is activated by millimolar concentrations of Ca2+ or Mg2+, was solubilized from rat heart plasma membrane by employing lysophosphatidylcholine, CHAPS, NaI, EDTA and Tris-HCl at pH 7.4. The enzyme was purified by sucrose density gradient, Affi-Gel Blue column and Sepharose 6B column chromatography. The purified enzyme was seen as a single peptide band in the sodium dodecyl sulfate polyacrylamide gel electrophoresis with a molecular weight of about 90,000. The apparent molecular weight of the holoenzyme as determined under non-dissociating conditions by gel filtration on Sepharose 6B column was about 180,000 indicating two subunits. The enzyme was insensitive to ouabain, verapamil, vanadate, oligomycin, N,N-dicyclohexylcarbodiimide and NaN3, but was markedly inhibited by 20 microM gramicidin S and 50 microM trifluoperazine. Analysis of the purified Ca2+/Mg2+ ATPase revealed the presence of 17 amino acids where leucine, glutamic acid and aspartic acid were the major components and histidine, cysteine and methionine were the minor components. The purified enzyme was associated with 19.7 mumol phospholipid/mg protein which was 60 times higher than the phospholipid content in plasma membrane. The cholesterol content in the purified enzyme preparation was 0.75 mumol/mg protein and this represented an 8-fold enrichment over plasma membrane. The glycoprotein nature of the enzyme was evident from the positive periodic acid-Schiff staining of the purified Ca2+/Mg2+ ATPase in the sodium dodecyl sulfate polyacrylamide gel. The polysaccharide content of the enzyme was enriched 8-fold over plasma membrane; neuraminidase treatment decreased the polysaccharide content. Concanavalin A prevented the ATP-dependent inactivation of the purified Ca2+/Mg2+ ATPase and was found to bind to the purified enzyme with a KD of 576 nM and Bmax of 4.52 nmol/mg protein. The results indicate that Ca2+/Mg2+ ATPase is a glycoprotein and contains a large amount of lipids.
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PMID:Purification and composition of Ca2+/Mg2+ ATPase from rat heart plasma membrane. 183 89

A synthetic, 28-residue peptide derived from the calmodulin-binding sequence of the plasma membrane Ca2+ pump (C28W) inhibits the ATPase activity of a calpain-produced, truncated fragment of the enzyme. The fragment, which has lost the calmodulin-binding domain, has a molecular mass of 124 kDa and is fully active in the absence of calmodulin. Replacement of Trp-8 in the peptide by an Ala decreases the overall inhibitory activity, while replacement with a Tyr increases it. However, at very low peptide concentrations the effect of Tyr replacement disappears. The synthetic peptide has been made photoactivatable by replacing Phe in position 9 with a synthetic phenylalanine analogue containing a diazirine group and was radioactively labeled by coupling a [3H]acetyl function to its N terminus. After cross-linking with the derivatized peptide, the 124-kDa fragment has been proteolyzed with either Lys-C, Asp-N, or V8 proteases, and the fragment(s) have been separated. Partial sequencing of the cross-linked, radioactive peptides has identified a site of the pump located C terminally to the phosphoenzyme-forming aspartic acid, spanning residues 537-544 of the hPMCA4 isoform of the enzyme. It is concluded that this sequence is part of a site which binds the calmodulin-binding domain of the pump.
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PMID:The plasma membrane Ca2+ pump contains a site that interacts with its calmodulin-binding domain. 184 39

In a model proposed for the structure of the a-subunit of the Escherichia coli F0F1-ATPase (Howitt, S.M., Gibson, F. and Cox, G.B. (1988) Biochim. Biophys. Acta 936, 74-80), a cluster of charged residues, including one arginine and four aspartic acid residues, lie on the periplasmic side of the membrane. On the cytoplasmic side, three pairs of lysine residues and an arginine residue are present. Site-directed mutagenesis was used to investigate the roles of these residues. It was found that none was directly involved in the proton pore. However, the substitutions of Asp-124 or Asp-44 by asparagine or Arg-140 by glutamine had similar effects in that the membranes from such mutants from which the F1-ATPase was removed were proton-impermeable. A combination of the Asp-44 mutation with either the Asp-124 or Arg-140 mutations in the same strain resulted in complete loss of oxidative phosphorylation. It was tentatively concluded that Asp-124 and Arg-140 form a salt bridge, as did Asp-44 with an unknown residue, and these salt bridges were concerned with the maintenance of correct a-subunit structure. Further support for this conclusion was obtained when second site revertants of a Glu-219 to histidine mutant were found to have either histidine or leucine replacing Arg-140. Thus, the lack of the Asp-124/Arg-140 salt bridge might enable repositioning of the helices of the a-subunit such that His-219 becomes a functional component of the proton pore.
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PMID:Mutational analysis of the function of the a-subunit of the F0F1-APPase of Escherichia coli. 213 15

The isoelectric points of most proteins from the extremely halophilic archaebacteria are between 4.0 and 4.65 which agrees with the generally high content of glutamic and aspartic acid in proteins from halobacteria. The subunits from two purified halobacterial membrane enzymes (ATPase and nitrate reductase) behaved differently with respect to isoelectric focusing, silver staining and interaction with ampholytes. Differential behavior was also observed in whole cell proteins from Halobacterium saccharovorum regarding resolution in two-dimensional gels and silver staining. We propose that these differences reflect the existence of two classes of halobacterial proteins, one resembling non-halophilic proteins, and the other possessing unique properties that may be related to salt dependence.
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PMID:Electrophoresis and isoelectric focusing of whole cell and membrane proteins from the extremely halophilic archaebacteria. 253 17

A material with inhibitory action to Na+/K+ ATPase was found in the lens of the ICR/f rat, a recessive hereditary cataractous rat. The material also induced lens opacification in vitro. From the results of amino acid analysis and by secondary ion mass spectroscopy, it was suggested that the material might contain approximately equimolar amounts of four amino acids, ie, aspartic acid, serine, glutamic acid and glycine, and that the molecular weight was 444. These facts suggested that this material with Na+/K+ ATPase inhibitory action might be a peptide. However, there is not yet any corroborating evidence to show whether this peptide is only a single material or not. The peptide significantly increased with aging in the lens of the ICR/f rat until approximately 90 days, when cataract became manifest, but its content decreased thereafter. This study suggests that one of the causes of cataractogenesis in the ICR/f rat might be this peptide, which is transformed in the lens with aging, and also that the peptide might accelerate lens opacification after cataractogenesis.
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PMID:Characterization of peptide inducing cataractogenesis in lens of hereditary cataractous rat (ICR/f RAT). 255 1

Site-specific mutagenesis was used to study the function of a conserved, extracellular aspartic acid residue from the sheep Na,K-ATPase alpha subunit. This amino acid, Asp-121, is the penultimate residue of the first extracellular domain of the alpha subunit. The border residues of this particular extracellular loop of the alpha subunit have been shown to be determinants of ouabain sensitivity (Price, E. M., and Lingrel, J. B. (1988) Biochemistry 27, 8400-8408). In order to determine if Asp-121 is involved in ouabain binding, five different amino acid substitutions at this position were generated. Four of the five mutant alpha subunits, containing either Asn, Ala, Glu, or Ser in place of Asp-121, conferred ouabain resistance to HeLa cells when expressed in those cells. Cloned sublines of cells selected in ouabain were characterized in terms of ouabain-inhibitable cell growth and Na,K-ATPase activity. The cells expressing the mutant Na,K-ATPase alpha subunit containing either Asn, Ala, Glu, or Ser in place of Asp-121 contained a component of Na,K-ATPase activity that was nearly 100-times more resistant to ouabain than the endogenous HeLa (human) or sheep enzyme. Apparently, conservative (Glu for Asp), isosteric (Asn for Asp), and nonconservative (Ala or Ser for Asp) substitutions all significantly decreased ouabain sensitivity. These data suggest that Asp-121 of the sheep Na,K-ATPase alpha subunit participates in the binding interaction between the enzyme and ouabain.
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PMID:Site-directed mutagenesis of a conserved, extracellular aspartic acid residue affects the ouabain sensitivity of sheep Na,K-ATPase. 255 44


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