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Query: EC:3.6.3.14 (ATP synthase)
 7,042 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

STUDY OBJECTIVE - The aim of the study was to measure variations in ATP synthase capacity in cultured cardiomyocytes under conditions of metabolic stimulation. DESIGN - ATP synthase activity was measured in cultured rat cardiomyocytes using a procedure which allowed rapid measurement of mitochondrial function during changes in metabolic state. EXPERIMENTAL MATERIAL - Calcium tolerant cardiomyocytes were prepared from male Wistar rats, weight 250-300 g, n = 6-22 per experiment. MEASUREMENTS AND MAIN RESULTS - Electrical stimulation of cardiomyocytes led to an approximate doubling of ATP synthase capacity within 1-2 min, and was rapidly reversible. Activation was reduced when extracellular calcium was lowered and abolished in presence of the calcium entry blocker ruthenium red. Exposure of cardiomyocytes to isoprenaline or to an inhibitor of phosphodiesterase III also led to a large increase in ATP synthase capacity, which was abolished in presence of ruthenium red. However, the response of cells to isoprenaline depended on their pretreatment: activation of ATP synthase was abolished after 20 min anoxia prior to isoprenaline treatment but regained after a subsequent 30 min reoxygenation. This may reflect down regulation of beta receptors on the cell surface during anoxia. CONCLUSIONS - ATP synthase is directly controlled in vivo by a non-allosteric mechanism. Activation of ATP synthase is a response to intramitochondrial Ca2+ concentration.
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PMID:Control of mitochondrial ATP synthase in heart cells: inactive to active transitions caused by beating or positive inotropic agents. 169 47

Fuel stimulation of insulin secretion from pancreatic beta-cells is thought to be mediated by metabolic coupling factors that are generated by energized mitochondria, including protons, adenine nucleotides, and perhaps certain amino acids (AA), as for instance aspartate, glutamate, or glutamine (Q). The goal of the present study was to evaluate the role of such factors when insulin release (IR) is stimulated by glucose or AA, alone or combined, using (31)P, (23)Na and (1)H NMR technology, respirometry, and biochemical analysis to study the metabolic events that occur in continuously superfused mouse beta-HC9 cells contained in agarose beads and enhanced by the phosphodiesterase inhibitor IBMX. Exposing beta-HC9 cells to high glucose or 3.5 mM of a physiological mixture of 18 AA (AAM) plus 2 mM glutamine caused a marked stimulation of insulin secretion associated with increased oxygen consumption, cAMP release, and phosphorylation potential as evidenced by higher phosphocreatine and lower P(i) peak areas of (31)P NMR spectra. Diazoxide blocked stimulation of IR completely, suggesting involvement of ATP-dependent potassium (K(ATP)) channels in this process. However, levels of MgATP and MgADP concentrations, which regulate channel activity, changed only slowly and little, whereas the rate of insulin release increased fast and very markedly. The involvement of other candidate coupling factors was therefore considered. High glucose or AAM + Q increased pH(i). The availability of temporal pH profiles allowed the precise computation of the phosphate potential (ATP/P(i) x ADP) in fuel-stimulated IR. Intracellular Na+ levels were greatly elevated by AAM + Q. However, glutamine alone or together with 2-amino-2-norbornanecarboxylic acid (which activates glutamate dehydrogenase) decreased beta-cell Na levels. Stimulation of beta-cells by glucose in the presence of AAM + Q (0.5 mM) was associated with rising cellular concentrations of glutamate and glutamine and strikingly lower aspartate levels. Methionine sulfoximine, an inhibitor of glutamine synthetase, blocked the glucose enhancement of AMM + Q-induced IR and associated changes in glutamine and aspartate but did not prevent the accumulation of glutamate. The results of this study demonstrate again that an increased phosphate potential and a functional K(ATP) channel are essential for metabolic coupling during fuel-stimulated insulin release but illustrate that determining the identity and relative importance of all participating coupling factors and second messengers remains a challenge largely unmet.
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PMID:Metabolic and ionic coupling factors in amino acid-stimulated insulin release in pancreatic beta-HC9 cells. 1726 32

The involvement of extracellular nucleotides and adenosine in an array of cell-specific responses has long been known and appreciated, but the integrative view of purinergic signalling as a multistep coordinated cascade has emerged recently. Current models of nucleotide turnover include: (i) transient release of nanomolar concentrations of ATP and ADP; (ii) triggering of signalling events via a series of ligand-gated (P2X) and metabotropic (P2Y) receptors; (iii) nucleotide breakdown by membrane-bound and soluble nucleotidases, including the enzymes of ecto-nucleoside triphosphate diphosphohydrolase (E-NTPDase) family, ecto-nucleotide pyrophosphatase/phosphodiesterase (E-NPP) family, ecto-5'-nucleotidase/CD73, and alkaline phosphatases; (iv) interaction of the resulting adenosine with own nucleoside-selective receptors; and finally, (v) extracellular adenosine inactivation via adenosine deaminase and purine nucleoside phosphorylase reactions and/or nucleoside uptake by the cells. In contrast to traditional paradigms that focus on purine-inactivating mechanisms, it has now become clear that "classical" intracellular ATP-regenerating enzymes, adenylate kinase, nucleoside diphosphate (NDP) kinase and ATP synthase can also be co-expressed on the cell surface. Furthermore, data on the ability of various cells to retain micromolar ATP levels in their pericellular space, as well as to release other related compounds (adenosine, UTP, dinucleotide polyphosphates and nucleotide sugars) gain another important insight into our understanding of mechanisms regulating a signalling cascade. This review summarizes recent advances in this rapidly evolving field, with particular emphasis on the nucleotide-releasing and purine-converting pathways in the vasculature.
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PMID:Nucleotide- and nucleoside-converting ectoenzymes: Important modulators of purinergic signalling cascade. 1830 42

Extracellular nucleotides and nucleosides mediate diverse signaling effects in virtually all organs and tissues. Most models of purinergic signaling depend on functional interactions between distinct processes, including (i) the release of endogenous ATP and other nucleotides, (ii) triggering of signaling events via a series of nucleotide-selective ligand-gated P2X and metabotropic P2Y receptors as well as adenosine receptors and (iii) ectoenzymatic interconversion of purinergic agonists. The duration and magnitude of purinergic signaling is governed by a network of ectoenzymes, including the enzymes of the nucleoside triphosphate diphosphohydrolase (NTPDase) family, the nucleotide pyrophosphatase/phosphodiesterase (NPP) family, ecto-5'-nucleotidase/CD73, tissue-nonspecific alkaline phosphatase (TNAP), prostatic acid phosphatase (PAP) and other alkaline and acid phosphatases, adenosine deaminase (ADA) and purine nucleoside phosphorylase (PNP). Along with "classical" inactivating ectoenzymes, recent data provide evidence for the co-existence of a counteracting ATP-regenerating pathway comprising the enzymes of the adenylate kinase (AK) and nucleoside diphosphate kinase (NDPK/NME/NM23) families and ATP synthase. This review describes recent advances in this field, with special emphasis on purine-converting ectoenzymes as a complex and integrated network regulating purinergic signaling in such (patho)physiological states as immunomodulation, inflammation, tumorigenesis, arterial calcification and other diseases. The second part of this review provides a comprehensive overview and basic principles of major approaches employed for studying purinergic activities, including spectrophotometric Pi-liberating assays, high-performance liquid chromatographic (HPLC) and thin-layer chromatographic (TLC) analyses of purine substrates and metabolites, capillary electrophoresis, bioluminescent, fluorometric and electrochemical enzyme-coupled assays, histochemical staining, and further emphasizes their advantages, drawbacks and suitability for assaying a particular catalytic reaction.
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PMID:Enzymes involved in metabolism of extracellular nucleotides and nucleosides: functional implications and measurement of activities. 2541 35