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)

Our efforts have been directed towards characterizing amino acid uptake, metabolism and release in bulk-isolated glia and neuronal perikarya studied in parallel with nerve-endings, especially as it concerns the transmitter amino acids and the participation of glia in the clearing of the synpatic space during impulse conduction. A possible neuromodulator role for the glia at the synapse is also suggested by K+-stimulated release. Our most definitive conclusions have been based so far on studies with GABA, although we are also beginning to accumulate data for glutamate related to glutamate-glutamine compartmentation. Glia preferentially accumulate potassium and amino acids compared to neuronal perikarya, have higher Na+/K+-ATPase activity, possess high-affinity, sodium-dependent uptake systems for GABA and glutamate similar to the ones in synaptosomes, and release amino acid in response to a potassium pulse by a calcium-independent process. Low neuronal uptake could be due to loss of dendrites. Unidirectional GABA-flux from the synaptosomal to glial compartment is supported by high GAD in nerve endings compared to high GABA-T in glia. Glutamine may be a transmitter glutamate-precursor in nerve-endings since glutaminase activity is high in nerve-endings, but low in glia where glutamine is presumably made. Glutamine uptake in both glia and synaptosomes obeys low-affinity kinetics in contrast to glutamate, consistent with the inability of glutamine to excite the neuronal membrane. The studies with GABA, which are considerably more extensive, are supported by related work using glia in tissue-culture and autoradiography. There appears to be a suggested difference in the behavior of amines which were poorly taken up by the glial system. Glia, synaptosomes and neuronal perikarya, in general behaved similarly with respect to requirements for uptake and release, except in the case of Ca++, which exerted opposite effects on glial and synaptosomal uptake of GABA. We believe that work along these lines tends to firmly establish a direct role for glial cells as modulators of neuronal excitability and represents a convergence between transmitter amino acid neuropharmacology and cellular biochemistry. This not only deepens and enlarges the vocabulary of synaptic biochemistry but also undoubtedly will have major clinical applications in the fields of epilepsy and behavior.
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PMID:Amino acid transport in isolated neurons and glia. 0 26

The activities of renal lactate and malate dehydrogenases, glutaminase, and Na-K-ATPase were determined in aging male C57BL/6 mice. Urine concentrating ability in these mice and renal response to metabolic acidosis were also studied. Total enzyme activities were measured in vitro in tissue homogenates from mice that were 120, 400, 500, 600, 700, and 800 days old. Urine concentrating ability was determined in these mice prior to sacrifice. Lactate and malate dehydrogenase activities decreased between 120 and 700 days with only male dehydrogenase activity increasing between 700 and 800 days. Age did not affect glutaminase or Na-K-ATPase activities and urine concentrating ability was decreased only at 700 days. Both urine ammonia excretion and renal glutaminase activity increased at 120 and 600 days in response to metabolic acidosis. However, only 5 of 12 animals tested at 600 days survived the acid stress for a full 7 days.
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PMID:Effects of age on renal function and enzyme activity in male C57BL/6 mice. 12 7

The ATP analogue 5'-[p-(fluorosulfonyl)benzoyl]adenosine (FSBA) was used to chemically modify the ATP binding sites of the carbamyl phosphate synthetase domain of CAD, the multifunctional protein that catalyzes the first steps in mammalian pyrimidine biosynthesis. Reaction of CAD with FSBA resulted in the inactivation of the ammonia- and glutamine-dependent CPSase activities but had no effect on its glutaminase, aspartate transcarbamylase, or dihydroorotase activities. ATP protected CAD against inactivation by FSBA whereas the presence of the allosteric effectors UTP and PRPP afforded little protection, which suggests that the ATP binding sites were specifically labeled. The inactivation exhibited saturation behavior with respect to FSBA with a K1 of 0.93 mM. Of the two ATP-dependent partial activities of carbamyl phosphate synthetase, bicarbonate-dependent ATPase was inactivated more rapidly than the carbamyl phosphate dependent ATP synthetase, which indicates that these partial reactions occur at distinct ATP binding sites. The stoichiometry of [14C]FSBA labeling showed that only 0.4-0.5 mol of FSBA/mol of protein was required for complete inactivation. Incorporation of radiolabeled FSBA into CAD and subsequent proteolysis, gel electrophoresis, and fluorography demonstrated that only the carbamyl phosphate synthetase domain of CAD is labeled. Amino acid sequencing of the principal peaks resulting from tryptic digests of FSBA-modified CAD located the sites of FSBA modification in regions that exhibit high homology to ATP binding sites of other known proteins. Thus CAD has two ATP binding sites, one in each of the two highly homologous halves of the carbamyl phosphate domain which catalyze distinct ATP-dependent partial reactions in carbamyl phosphate synthesis.
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PMID:Identification of the ATP binding sites of the carbamyl phosphate synthetase domain of the Syrian hamster multifunctional protein CAD by affinity labeling with 5'-[p-(fluorosulfonyl)benzoyl]adenosine. 168

Carbamoyl phosphate synthetase from Escherichia coli catalyzes the formation of carbamoyl phosphate from ATP, bicarbonate, and glutamine. The amidotransferase activity of this enzyme is catalyzed by the smaller of the two subunits of the heterodimeric protein. The roles of four conserved histidine residues within this subunit were probed by site-directed mutagenesis to asparagine. The catalytic activities of the H272N and H341N mutants are not significantly different than that of the wild-type enzyme. The H353N mutant is unable to utilize glutamine as a nitrogen source in the synthetase reaction or the partial glutaminase reaction. However, binding to the glutamine active site is not impaired in the H353N enzyme since glutamine is found to activate the partial ATPase reaction by 40% with a Kd of 54 microM. The H312N mutant has a Michaelis constant for glutamine that is 2 orders of magnitude larger than the wild-type value, but the maximal rate of glutamine hydrolysis is unchanged. These results are consistent with His-353 functioning as a general acid/base catalyst for proton transfers while His-312 serves a critical role for the binding of glutamine to the active site.
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PMID:Role of the four conserved histidine residues in the amidotransferase domain of carbamoyl phosphate synthetase. 186 65

The inhibitory effects of omeprazole and omeprazole-derived metabolites were studied on Escherichia coli glutaminase activity at pH 2.5 which might represent the conditions present at the target enzyme (K+/H+-ATPase) in the secretory membrane of the intact parietal cell. Omeprazole and the omeprazole-derived cyclic sulphenamide inhibited glutaminase at pH 2.5 with identical potency (IC50 36 microM). The substrate, glutamine as well as the mercaptane, dithiothreitol, protect the enzyme. Furthermore, dithioerythritol was found to reverse inhibition. This indicates that an SH-group localized in the substrate binding center of glutaminase is most likely involved in the reaction leading to enzyme inhibition. Glutaminase inhibition by both compounds was less pronounced at pH 5.0. Omeprazole radical, the metabolite generated from the cyclic sulphenamide at more neutral pH values, failed to affect the enzyme. These findings were in contrast with the properties of the omeprazole-derived cyclic sulphenamide and radical at the K+/H+-ATPase preparation. This enzyme was inhibited by both compounds at pH 7.5 with a high potency, and reversal experiments with dithiothreitol demonstrate that these agents interfere with SH-groups of the K+/H+-ATPase. From these data it is suggested that the cyclic sulphenamide and the radical interfere by different reaction pathways with enzymatic SH-groups.
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PMID:Studies on the mechanism of action of the omeprazole-derived cyclic sulphenamide. 283 Aug 83

Formylglycinamidine ribonucleotide (FGAM) synthetase, which catalyzes the conversion of formylglycinamide ribonucleotide (FGAR), glutamine, and ATP to FGAM, ADP, glutamate, and Pi, has been purified to homogeneity (sp act. 0.20 mumol min-1 mg-1) from chicken liver by an alternative procedure to that of Buchanan et al. [Buchanan, J. M., Ohnoki, S., & Hong, B. S. (1978) Methods Enzymol. 51, 193-201] (sp act. 0.12 mumol min-1 mg-1). A variety of new analogues of formylglycinamide ribonucleotide have been prepared in which the formylglycinamide arm (R = CH2NHCHO) has been replaced by R = CH3, CH2OH, CH2Cl, CH2NH3, CH2NHCOCH3, CH2NHCOCH2Cl, CH2NHCO2CH2Ph, and L-CHC-H3NHCHO. These compounds have been characterized by 1H and 13C NMR spectroscopy. With compounds R = CH3, CH2OH, and CH2NHCOCH3 and ATP, in the presence or absence of glutamine, FGAM synthetase catalyzes the production of Pi at 4.5, 48, and 20%, respectively, the rate of production of Pi from formylglycinamide ribonucleotide. Only R = CH2NHCOCH3 causes glutaminase activity as well as ATPase activity and has been shown to be converted to the amidine analogue. Both FGAR (R = CH2NHCHO) and the FGAR analogue (R = CH2NHCHOCH3) in the presence of ATP and FGAM synthetase and in the absence of glutamine form a complex isolable by Sephadex G-50 chromatography. FGAM synthetase is thus highly specific for its formylglycine side chain. [18O]-beta-FGAR was prepared biosynthetically, and FGAM synthetase was shown by 31P NMR spectroscopy to catalyze the transfer of amide 18O to inorganic phosphate.
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PMID:Substrate specificity of formylglycinamidine synthetase. 293 78

In order to evaluate the possible role of sodium- and potassium-activated adenosine triphosphatase in the active transport of sodium by the renal tubules, we examined the effect of large changes in the tubular reabsorptive load of sodium on the Na-K-ATPase activity of rat kidney homogenates. Glomerular filtration and tubular reabsorption of sodium per gram of kidney tissue increased progressively after contralateral uninephrectomy. This was paralleled by an increase in Na-K-ATPase per milligram of protein in a microsomal fraction of kidney cortex. The importance of this change is underlined by the absence of simultaneous increases in other microsomal enzymes such as glucose-6-phosphatase and Mg(++)-dependent ATPase, or in succinic dehydrogenase or glutaminase. Similar increases in Na-K-ATPase were observed when the net tubular reabsorption of sodium was increased by feeding the animals a high-protein diet or after injection of methylprednisolone. On the other hand, Na-K-ATPase was lowered when tubular transport of sodium was reduced by bilateral adrenalectomy. The results of these experiments show that renal Na-K-ATPase changes in an adaptive way when renal reabsorption of sodium is chronically increased or diminished and support the hypothesis that this enzyme system is involved in the process by which sodium is actively transported across the renal tubule.
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PMID:The role of sodium-potassium-activated adenosine triphosphatase in the reabsorption of sodium by the kidney. 429 72

Phosphate depletion (PD) in vivo causes a sundry of abnormalities in pancreatic islets including a rise in cytosolic calcium, low ATP content, reduced Ca2+ ATPase and Na(+)-K+ ATPase activity, and impaired insulin secretion in response to glucose or potassium. L-Leucine is a strong secretagogue that triggers insulin secretion by deamination to alpha-ketoisocaproic acid (KIC) and the subsequent metabolism of the latter to ATP and by the activation of glutamate dehydrogenase (GLDH), which acts on glutamate to generate alpha-ketoglutarate, the metabolism of which results in ATP production. The generation of ATP triggers events that lead to insulin secretion. It is not known whether PD impairs leucine-induced insulin secretion, and the cellular derangements that are involved in such an abnormality are not defined. These issues were studied in PD rats and in pair-weighed normal animals as controls. D-Leucine uptake by islets from PD rats is normal, but both leucine- and KIC-induced insulin secretions are impaired and the activity of branched-chain keto acid dehydrogenase, which facilitates the metabolism of KIC, is reduced. Both leucine and 2-aminobicyclo (2-2-1) haptene failed to stimulate GLDH and to augment the generation of alpha-ketoglutarate in the islets of PD rats. Also, the concentration of basal alpha-ketoglutarate was significantly higher in the islets of PD rats, suggesting that its metabolism is impaired. In addition, the activity of glutaminase is significantly reduced, an abnormality that would result in decreased production of glutamate, the substrate for GLDH. The data show that PD impairs leucine-induced insulin secretion.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Phosphate depletion impairs leucine-induced insulin secretion. 787 37

Streptococcus bovis deaminated glutamine by a mechanism that did not involve glutaminase. Since pyroglutamate and ammonia were the only end-products, it appeared that glutamine deamination was catalyzed by a cyclotransferase reaction. Stationary S. bovis cells had essentially no intracellular ATP or membrane potential (delta psi), however, when they were provided with glutamine, intracellular ATP and delta psi increased to 0.52 mM and 158 mV, respectively. When glutamine-energized cells were treated with N,N-dicyclohexylcarbodiimide (DCCD, 150 microM), there was an even greater increase in intracellular ATP (> 5-fold) and the delta psi was dissipated. Because toluene-treated cells produced ATP from ADP and Pi, it did not appear that the cell membrane was directly involved in glutamine-dependent ATP generation. The rate of ammonia production was directly proportional to the glutamine concentration, but the stoichiometry of ATP to ammonia was always 1 to 1. Based on these results, it appeared that glutamine was deaminated by glutamine cyclotransferase which was coupled to ATP formation. The membrane bound ATPase then used the ATP to create a delta psi.
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PMID:The glutamine cyclotransferase reaction of Streptococcus bovis: a novel mechanism of deriving energy from non-oxidative and non-reductive deamination. 840 35

Microbial carbamoyl phosphate synthetases (CPS) use glutamine as nitrogen donor and are composed of two subunits (or domains), one exhibiting glutaminase activity, the other able to synthesize carbamoyl phosphate (CP) from bicarbonate, ATP, and ammonia. The pseudodimeric organization of this synthetase suggested that it has evolved by duplication of a smaller kinase, possibly a carbamate kinase (CK). In contrast to other prokaryotes the hyperthermophilic archaeon Pyrococcus furiosus was found to synthesize CP by using ammonia and not glutamine. We have purified the cognate enzyme and found it to be a dimer of two identical subunits of Mr 32,000. Its thermostability is considerable, 50% activity being retained after 1 h at 100 degrees C or 3 h at 95 degrees C. The corresponding gene was cloned by PCR and found to present about 50% amino acid identity with known CKs. The stoichiometry of the reaction (two ATP consumed per CP synthesized) and the ability of the enzyme to catalyze at high rate a bicarbonate-dependent ATPase reaction however clearly distinguish P. furiosus CPS from ordinary CKs. Thus the CPS of P. furiosus could represent a primeval step in the evolution of CPS from CK. Our results suggest that the first event in this evolution was the emergence of a primeval synthetase composed of subunits able to synthesize both carboxyphosphate and CP; this step would have preceded the duplication assumed to have generated the two subdomains of modern CPSs. The gene coding for this CK-like CPS was called cpkA.
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PMID:The carbamate kinase-like carbamoyl phosphate synthetase of the hyperthermophilic archaeon Pyrococcus furiosus, a missing link in the evolution of carbamoyl phosphate biosynthesis. 937 56


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