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Sample records for er cta4 atpase

  1. Inhibition of the sarco/endoplasmic reticulum (ER) Ca2+-ATPase by thapsigargin analogs induces cell death via ER Ca2+ depletion and the unfolded protein response

    DEFF Research Database (Denmark)

    Sehgal, Pankaj; Szalai, Paula; Olesen, Claus

    2017-01-01

    Calcium (Ca2+) is a fundamental regulator of cell signaling and function. Thapsigargin (Tg) blocks the sarco/endoplasmic reticulum (ER) Ca2+-ATPase (SERCA), disrupts Ca2+ homeostasis, and causes cell death. However, the exact mechanisms whereby SERCA-inhibition induces cell death are incompletely...... extensive drainage of the ER Ca2+ stores. This Ca2+ depletion was followed by markedly reduced cell proliferation rates and morphological changes that developed over 2–4 days and culminated in cell death. Interestingly, these changes were not accompanied by bulk increases in cytosolic Ca2+ levels. Moreover...... and their detrimental effects on cell viability. Furthermore, caspase activation and cell death were associated with a sustained unfolded protein response (UPR). We conclude that ER Ca2+ drainage and sustained UPR activation are key for initiation of apoptosis at low concentrations of Tg and Tg analogs, whereas high...

  2. Final Report for DE-FG02-04ER15626: P-type ATPases in Plants – Role of Lipid Flippases in Membrane Biogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Harper, Jeffrey F. [Univ. of Nevada, Reno, NV (United States)

    2015-02-24

    The long-range goal of the research is to understand the structure and biological functions of different P-type ATPases (ion pumps) in plant cells, and to use that knowledge to enhance the production of bioenergy from plants, or plant-research inspired technologies. Ptype ATPases include ion pumps that specifically transport H+, Ca2+, Zn2+, Cu2+, K+, or Na+, as well as at least one unusual subfamily that appears to function as lipid flippases, flipping specific lipids from one side of a membrane bilayer to the other. As a group, P-type ATPases are thought to consume more than 1/3 of the cellular ATP in typical eukaryotic cells. Recent research in the Harper lab focused on understanding the biochemical and biological functions of P-type ATPases that flip lipids. These flippases belong to the P4 subfamily of P-type ATPases. The activity of lipid flippases is thought to induce membrane curvature and/or create an asymmetry in which certain lipid head groups are preferential exposed to one surface or the other. In Arabidopsis thaliana there are 12 members of this family referred to as Aminophospholipid ATPase (ALA) 1 to ALA12. Using genetic knockouts, the Harper lab has established that this unusual subfamily of P-type ATPases are critical for plants to cope with even modest changes in temperature (e.g., down to 15°C, or up to 30°C). In addition, members of one subclade are critical for cell expansion, and loss of function mutants result in severe dwarfism. Other members of this same sub-clade are critical for pollen tube growth, and loss of function mutants are sterile under conditions of hot days and cold nights. While the cellular processes that depend on lipid flippases are still unclear, the genetic analysis of loss of function mutants clearly show they are of fundamental importance to plant growth and response to the environment.

  3. P4-ATPases

    DEFF Research Database (Denmark)

    Lopez Marques, Rosa Laura; Theorin, Lisa; Palmgren, Michael Broberg

    2014-01-01

    ) comprises lipid flippases that catalyze the translocation of phospholipids from the exoplasmic to the cytosolic leaflet of cell membranes. While initially characterized as aminophospholipid translocases, recent studies of individual P4-ATPase family members from fungi, plants, and animals show that P4......Cellular membranes, notably eukaryotic plasma membranes, are equipped with special proteins that actively translocate lipids from one leaflet to the other and thereby help generate membrane lipid asymmetry. Among these ATP-driven transporters, the P4 subfamily of P-type ATPases (P4-ATPases...... to include the regulation of membrane traffic, cytoskeletal dynamics, cell division, lipid metabolism, and lipid signaling. In this review, we will summarize the basic features of P4-ATPases and the physiological implications of their lipid transport activity in the cell....

  4. Towards defining the substrate of orphan P5A-ATPases

    DEFF Research Database (Denmark)

    Sørensen, Danny Mollerup; Holen, Henrik Waldal; Holemans, Tine

    2015-01-01

    leads to broad and unspecific phenotypes related to the impairment of basic ER functions such as protein folding and processing. Genetic interactions in Saccharomyces cerevisiae point to a role of the endogenous P5A-ATPase Spf1p in separation of charges in the ER, in sterol metabolism, and in insertion...... significance Identification of the substrate of P5A-ATPases would throw light on an important general process in the ER that is still not fully understood. This article is part of a Special Issue entitled Structural biochemistry and biophysics of membrane proteins....

  5. Iron overload impact on P-ATPases.

    Science.gov (United States)

    Sousa, Leilismara; Pessoa, Marco Tulio C; Costa, Tamara G F; Cortes, Vanessa F; Santos, Herica L; Barbosa, Leandro Augusto

    2018-03-01

    Iron is a chemical element that is active in the fundamental physiological processes for human life, but its burden can be toxic to the body, mainly because of the stimulation of membrane lipid peroxidation. For this reason, the action of iron on many ATPases has been studied, especially on P-ATPases, such as the Na + ,K + -ATPase and the Ca 2+ -ATPase. On the Fe 2+ -ATPase activity, the free iron acts as an activator, decreasing the intracellular Fe 2+ and playing a protection role for the cell. On the Ca 2+ -ATPase activity, the iron overload decreases the enzyme activity, raising the cytoplasmic Ca 2+ and decreasing the sarco/endoplasmic reticulum and the Golgi apparatus Ca 2+ concentrations, which could promote an enzyme oxidation, nitration, and fragmentation. However, the iron overload effect on the Na + ,K + -ATPase may change according to the tissue expressions. On the renal cells, as well as on the brain and the heart, iron promotes an enzyme inactivation, whereas its effect on the erythrocytes seems to be the opposite, directly stimulating the ATPase activity, or stimulating it by signaling pathways involving ROS and PKC. Modulations in the ATPase activity may impair the ionic transportation, which is essential for cell viability maintenance, inducing irreversible damage to the cell homeostasis. Here, we will discuss about the iron overload effect on the P-ATPases, such as the Na + ,K + -ATPase, the Ca 2+ -ATPase, and the Fe 2+ -ATPase.

  6. Effect of ionizing radiation on Ca2+-ATPase and Mg2+-ATPase: the role of ligands

    International Nuclear Information System (INIS)

    Dreval', V.I.

    1994-01-01

    The change of Ca 2+ -ATPase and Mg 2+ -ATPase activity in plasma membranes of thymocytes irradiated with doses of 10 2 , 10 3 and 10 4 Gy in the presence of Ca 2+ , Mg 2+ and ATP was studied. Stabilizing effect of Ca 2+ and Mg 2+ on Ca 2+ -ATPase and ATP on Mg 2+ -ATPase under irradiation was established

  7. On archaebacterial ATPase from Halobacterium saccharovorum

    Science.gov (United States)

    Kristjansson, H.; Ponnamperuma, C.; Hochstein, L.; Altekar, W.

    1984-01-01

    The energy transducing ATPase from Halobacterium saccharovorum was studied in order to define the origin of energy transducing systems. The ATPase required high salt concentration (4M NaCl) for activity; activity was rapidly lost when NaCl was below 1 Molar. At low salt concentration, the membrane bound ATPase activity could be stabilized in presence of spermine. However, following solubilization spermine was ineffective. Furthermore, F1 ATPase activity was stabilized by ammonium sulfate even when the NaCl concentration was less than 1 Molar. These studies suggest that stabilization by hydrophobic interactions preceded ionic ones in the evolution of the energy transducing ATPases.

  8. Myocardial Na,K-ATPase: Clinical aspects

    OpenAIRE

    Kjeldsen, Keld

    2003-01-01

    The specific binding of digitalis glycosides to Na,K-ATPase is used as a tool for Na,K-ATPase quantification with high accuracy and precision. In myocardial biopsies from patients with heart failure, total Na,K-ATPase concentration is decreased by around 40%; a correlation exists between a decrease in heart function and a decrease in Na,K-ATPase concentration. During digitalization, around 30% of remaining pumps are occupied by digoxin. Myocardial Na,K-ATPase is also influenced by other drugs...

  9. Evolution of plant P-type ATPases

    Directory of Open Access Journals (Sweden)

    Christian N.S. Pedersen

    2012-02-01

    Full Text Available Five organisms having completely sequenced genomes and belonging to all major branches of green plants (Viridiplantae were analyzed with respect to their content of P-type ATPases encoding genes. These were the chlorophytes Ostreococcus tauria and Chlamydomonas reinhardtii, and the streptophytes Physcomitrella patens (a moss, Selaginella moellendorffii (a primitive vascular plant, and Arabidopsis thaliana (a model flowering plant. Each organism contained sequences for all five subfamilies of P-type ATPases. Our analysis demonstrates when specific subgroups of P-type ATPases disappeared in the evolution of Angiosperms. Na/K-pump related P2C ATPases were lost with the evolution of streptophytes whereas Na+ or K+ pumping P2D ATPases and secretory pathway Ca2+-ATPases remained until mosses. An N-terminally located calmodulin binding domain in P2B ATPases can only be detected in pumps from Streptophytae, whereas, like in animals, a C-terminally localized calmodulin binding domain might be present in chlorophyte P2B Ca2+-ATPases. Chlorophyte genomes encode P3A ATPases resembling protist plasma membrane H+-ATPases and a C-terminal regulatory domain is missing. The complete inventory of P-type ATPases in the major branches of Viridiplantae is an important starting point for elucidating the evolution in plants of these important pumps.

  10. Phylogenetic analysis of P5 P-type ATPases, a eukaryotic lineage of secretory pathway pumps

    DEFF Research Database (Denmark)

    Møller, Annette; Asp, Torben; Holm, Preben Bach

    2008-01-01

    prokaryotic genome. Based on a protein alignment we could group the P5 ATPases into two subfamilies, P5A and P5B that, based on the number of negative charges in conserved trans-membrane segment 4, are likely to have different ion specificities. P5A ATPases are present in all eukaryotic genomes sequenced so......Eukaryotes encompass a remarkable variety of organisms and unresolved lineages. Different phylogenetic analyses have lead to conflicting conclusions as to the origin and associations between lineages and species. In this work, we investigated evolutionary relationship of a family of cation pumps...... exclusive for the secretory pathway of eukaryotes by combining the identification of lineage-specific genes with phylogenetic evolution of common genes. Sequences of P5 ATPases, which are regarded to be cation pumps in the endoplasmic reticulum (ER), were identified in all eukaryotic lineages but not in any...

  11. Calcineurin Interacts with PERK and Dephosphorylates Calnexin to Relieve ER Stress in Mammals and Frogs

    OpenAIRE

    Bollo, Mariana; Paredes, R. Madelaine; Holstein, Deborah; Zheleznova, Nadezhda; Camacho, Patricia; Lechleiter, James D.

    2010-01-01

    Background The accumulation of misfolded proteins within the endoplasmic reticulum (ER) triggers a cellular process known as the Unfolded Protein Response (UPR). One of the earliest responses is the attenuation of protein translation. Little is known about the role that Ca2+ mobilization plays in the early UPR. Work from our group has shown that cytosolic phosphorylation of calnexin (CLNX) controls Ca2+ uptake into the ER via the sarco-endoplasmic reticulum Ca2+-ATPase (SERCA) 2b. Methodology...

  12. Politik er ikke lykken

    DEFF Research Database (Denmark)

    Steenbuch, Johannes Aakjær

    2011-01-01

    Der er ikke længere nogen højere sandhed i livet end den, flertallet bestemmer sig for – og dermed ingen del af livet, der ikke er politisk. Højre- og venstrefløjen er i bund og grund enige - enige om, at det er politikernes opgave at forære os det gode liv. Dermed bliver demokratiet totalitært. ...

  13. Man er aldrig alene

    DEFF Research Database (Denmark)

    Hoffmeyer, Jesper

    2013-01-01

    Nu ved vi, at der er mange slags DNA i vores krop, og at samarbejdet mellem de organismer, som bærer alt dette DNA, er essentielt for vores overlevelse" … "Kroppen er en slags økosystem, hvor alle slags samarbejde hen ad vejen bliver afprøvet"...

  14. Novel aspects of Na+,K+-ATPase

    OpenAIRE

    Aizman, Oleg

    2002-01-01

    Na,K-ATPase, an integral membrane protein expressed in each eukaryotic cell, serves as the major determinant of intracellular ion composition. In the current study we investigated novel aspects of Na,K-ATPase function and regulation. It is well established that Na,K-ATPase activity is regulated by reversible phosphorylation. New findings in this study are: 1) the level of intracellular Ca 2. concentration determines the functional effects of PKA and PKC-mediated Na,K-ATP...

  15. AAA-ATPases in Protein Degradation

    Directory of Open Access Journals (Sweden)

    Ravikiran S. Yedidi

    2017-06-01

    Full Text Available Proteolytic machineries containing multisubunit protease complexes and AAA-ATPases play a key role in protein quality control and the regulation of protein homeostasis. In these protein degradation machineries, the proteolytically active sites are formed by either threonines or serines which are buried inside interior cavities of cylinder-shaped complexes. In eukaryotic cells, the proteasome is the most prominent protease complex harboring AAA-ATPases. To degrade protein substrates, the gates of the axial entry ports of the protease need to be open. Gate opening is accomplished by AAA-ATPases, which form a hexameric ring flanking the entry ports of the protease. Protein substrates with unstructured domains can loop into the entry ports without the assistance of AAA-ATPases. However, folded proteins require the action of AAA-ATPases to unveil an unstructured terminus or domain. Cycles of ATP binding/hydrolysis fuel the unfolding of protein substrates which are gripped by loops lining up the central pore of the AAA-ATPase ring. The AAA-ATPases pull on the unfolded polypeptide chain for translocation into the proteolytic cavity of the protease. Conformational changes within the AAA-ATPase ring and the adjacent protease chamber create a peristaltic movement for substrate degradation. The review focuses on new technologies toward the understanding of the function and structure of AAA-ATPases to achieve substrate recognition, unfolding and translocation into proteasomes in yeast and mammalian cells and into proteasome-equivalent proteases in bacteria and archaea.

  16. AAA-ATPases in Protein Degradation.

    Science.gov (United States)

    Yedidi, Ravikiran S; Wendler, Petra; Enenkel, Cordula

    2017-01-01

    Proteolytic machineries containing multisubunit protease complexes and AAA-ATPases play a key role in protein quality control and the regulation of protein homeostasis. In these protein degradation machineries, the proteolytically active sites are formed by either threonines or serines which are buried inside interior cavities of cylinder-shaped complexes. In eukaryotic cells, the proteasome is the most prominent protease complex harboring AAA-ATPases. To degrade protein substrates, the gates of the axial entry ports of the protease need to be open. Gate opening is accomplished by AAA-ATPases, which form a hexameric ring flanking the entry ports of the protease. Protein substrates with unstructured domains can loop into the entry ports without the assistance of AAA-ATPases. However, folded proteins require the action of AAA-ATPases to unveil an unstructured terminus or domain. Cycles of ATP binding/hydrolysis fuel the unfolding of protein substrates which are gripped by loops lining up the central pore of the AAA-ATPase ring. The AAA-ATPases pull on the unfolded polypeptide chain for translocation into the proteolytic cavity of the protease. Conformational changes within the AAA-ATPase ring and the adjacent protease chamber create a peristaltic movement for substrate degradation. The review focuses on new technologies toward the understanding of the function and structure of AAA-ATPases to achieve substrate recognition, unfolding and translocation into proteasomes in yeast and mammalian cells and into proteasome-equivalent proteases in bacteria and archaea.

  17. Vreden er over os

    DEFF Research Database (Denmark)

    Mehlsen, Camilla

    2006-01-01

    Urolige elever, cyklister - der giver fuck-finger, aggressive demonstranter. Samtiden er på vej ind i en æra domineret af vrede, siger rektor Lars-Henrik Schmidt, der er aktuel med bogen 'Om vreden'. Udgivelsesdato: Juni......Urolige elever, cyklister - der giver fuck-finger, aggressive demonstranter. Samtiden er på vej ind i en æra domineret af vrede, siger rektor Lars-Henrik Schmidt, der er aktuel med bogen 'Om vreden'. Udgivelsesdato: Juni...

  18. Hvad f er meningen?

    DEFF Research Database (Denmark)

    Rydén, Pernille; Ringberg, Torsten; Wilke, Ricky

    En forskningsrapport fra CBS om danske lederes opfattelse af sociale medier i detail- og servicebranchen. Rapporten er udarbejdet i regi af Service Platform.......En forskningsrapport fra CBS om danske lederes opfattelse af sociale medier i detail- og servicebranchen. Rapporten er udarbejdet i regi af Service Platform....

  19. Fluxus-øer

    DEFF Research Database (Denmark)

    van der Meijden, Peter Alexander

    2008-01-01

    "Fluxus-øer" er en introduktion til Fluxus med udgangspunkt i den tyske galleri-ejer René Blocks samling, som udstillingen "Food for Thought" i Sukkerfabrikken i Stege (Møn) præsenterede et udvalg af. Artiklen beskriver Fluxus som et heterotopi som beskrevet af Michel Foucault i "Of Other Spaces"...

  20. Decoding P4-ATPase substrate interactions.

    Science.gov (United States)

    Roland, Bartholomew P; Graham, Todd R

    Cellular membranes display a diversity of functions that are conferred by the unique composition and organization of their proteins and lipids. One important aspect of lipid organization is the asymmetric distribution of phospholipids (PLs) across the plasma membrane. The unequal distribution of key PLs between the cytofacial and exofacial leaflets of the bilayer creates physical surface tension that can be used to bend the membrane; and like Ca 2+ , a chemical gradient that can be used to transduce biochemical signals. PL flippases in the type IV P-type ATPase (P4-ATPase) family are the principle transporters used to set and repair this PL gradient and the asymmetric organization of these membranes are encoded by the substrate specificity of these enzymes. Thus, understanding the mechanisms of P4-ATPase substrate specificity will help reveal their role in membrane organization and cell biology. Further, decoding the structural determinants of substrate specificity provides investigators the opportunity to mutationally tune this specificity to explore the role of particular PL substrates in P4-ATPase cellular functions. This work reviews the role of P4-ATPases in membrane biology, presents our current understanding of P4-ATPase substrate specificity, and discusses how these fundamental aspects of P4-ATPase enzymology may be used to enhance our knowledge of cellular membrane biology.

  1. MIT HJEM ER HVOR MIT HJERTE ER

    DEFF Research Database (Denmark)

    Høst, Jeppe Engset

    2014-01-01

    Ideen om at arbejde med det man kalder de stedbundne ressourcer på en ny måde, er relevant på Bornholm hvor events som blandt andet festivalen ’Wonderfestiwall’, strandfesten ’Vang Pier Beach Party’, karnevallet ’Svaneke Beach Party, kokkekonkurrencen ’Sol over Gudhjem’ og filmfestivalen ’Bornsho...

  2. Endoplasmic reticulum calcium transport ATPase expression during differentiation of colon cancer and leukaemia cells

    International Nuclear Information System (INIS)

    Papp, Bela; Brouland, Jean-Philippe; Gelebart, Pascal; Kovacs, Tuende; Chomienne, Christine

    2004-01-01

    The calcium homeostasis of the endoplasmic reticulum (ER) is connected to a multitude of cell functions involved in intracellular signal transduction, control of proliferation, programmed cell death, or the synthesis of mature proteins. Calcium is accumulated in the ER by various biochemically distinct sarco/endoplasmic reticulum calcium transport ATPase isoenzymes (SERCA isoforms). Experimental data indicate that the SERCA composition of some carcinoma and leukaemia cell types undergoes significant changes during differentiation, and that this is accompanied by modifications of SERCA-dependent calcium accumulation in the ER. Because ER calcium homeostasis can also influence cell differentiation, we propose that the modulation of the expression of various SERCA isoforms, and in particular, the induction of the expression of SERCA3-type proteins, is an integral part of the differentiation program of some cancer and leukaemia cell types. The SERCA content of the ER may constitute a new parameter by which the calcium homeostatic characteristics of the organelle are adjusted. The cross-talk between ER calcium homeostasis and cell differentiation may have some implications for the better understanding of the signalling defects involved in the acquisition and maintenance of the malignant phenotype

  3. Inactivation of mitochondrial ATPase by ultraviolet light

    International Nuclear Information System (INIS)

    Chavez, E.; Cuellar, A.

    1984-01-01

    The present work describes experiments that show that far-ultraviolet irradiation induce the inhibition of ATPase activity in both membrane-bound and soluble F1. It was also found that ultraviolet light promotes the release of tightly bound adenine nucleotides from F1-ATPase. Experiments carried out with submitochondrial particles indicate that succinate partially protects against these effects of ultraviolet light. Titration of sulfhydryl groups in both irradiated submitochondrial particles and soluble F1-ATPase indicates that a conformational change induced by photochemical modifications of amino acid residues appears involved in the inactivation of the enzyme. Finally, experiments are described which show that the tyrosine residue located in the active site of F1-ATPase is modified by ultraviolet irradiation

  4. Hvor anvendelig er PKI?

    OpenAIRE

    Nielsen, Jon Magne

    2006-01-01

    Denne oppgaven ser på bruken av elektronisk ID i statlige etater i Norge i dag. Det ses spesielt på om bruken av tekologien PKI er en god løsning på etatenes behov på dette området. Som utgangspunkt for analysen er det sett spesielt på to statlige etater. Disse etatenes behov og bruk av elektronisk ID generelt og PKI spesielt blir undersøkt. Det er videre gjort rede for hvilke lover, forskrifter og andre førende dokumenter som danner de formelle rammebetingelsene for etaters bruk av PKI. ...

  5. Mg,Ca-ATPase activity under irradiation

    International Nuclear Information System (INIS)

    Ladutin, V.V.; Orlova, V.V.; Lob, P.A.; Gerasiminko, I.V.; Mack, E.I.

    2003-01-01

    Full text: The influence of different doses irradiation at the Mg,Ca-ATPase activity at the rat brain has been investigated. The analyses were made at the apparatus of LKB and Carl-Ceis-Jena firm with help of reagents of Sigma and Boehringer firm. Rats decapitated after 1, 3, 6, 24 and 48 h after action of irradiation. Dose 0.206 C/kg. Erythrocytes. 1 and 3h after irradiation influence- decrease of Mg,Ca-ATPase activity to 86-87% relatively control level, 24 and 48 h - increase of activity to the control level. Dose 0.312 C/kg. Large hemispheres. 1h - decrease of ATPase activity to 90% relatively control, 3h - increase to control level, 24h - fall to 86%, after 48h small increase to 93% relatively control. Dose 9.287 C/kg. Large hemispheres. 1h - sharp fall of Mg, Ca-ATPase activity to 67 % relatively control, increase of activity to 96% after 3h and sharp fall of activity to 64% 6h after action of irradiation. Dose 9.287 C/kg. Cerebellum. 1h - sharp decrease of ATPase activity to 80%. After 3h -sharp increase to 160% relatively control level and sharp fall of ATPase activity to 47% relatively control after 6h. The mechanism of radiation pathology of active ion transport has been discussed

  6. Innovation er brugerdreven!

    DEFF Research Database (Denmark)

    Helms, Niels Henrik

    2008-01-01

    Brugerdreven innovation er blevet svaret på mange af de udfordringer, som vores moderne samfund står overfor.Det er skrevet ind i såvel regeringsgrundlaget som i de forskellige tiltag, som skal ruste Danmark i forhold til globaliseringen. Vi har ifølge argumentationen her enrække særlige forudsæt....... Udgivelsesdato: marts 2008...

  7. Metabolomics er fremtiden

    DEFF Research Database (Denmark)

    Pedersern, Birger

    2010-01-01

    Forskningen i fødevarer har fået et potent redskab i hånden. Metabolomics er vejen frem, mener professor Søren Balling Engelsen fra Københavns Universitet......Forskningen i fødevarer har fået et potent redskab i hånden. Metabolomics er vejen frem, mener professor Søren Balling Engelsen fra Københavns Universitet...

  8. Er KU et mobbeuniversitet?

    DEFF Research Database (Denmark)

    Olden-Jørgensen, Sebastian

    2009-01-01

    Ansatte på KU mobber hverken mere eller mindre end på andre danske arbejdspladser. Mediernes dækning af APV-undersøgelsens resultater mht. mobning er sensationalistisk og delvis vildledende.......Ansatte på KU mobber hverken mere eller mindre end på andre danske arbejdspladser. Mediernes dækning af APV-undersøgelsens resultater mht. mobning er sensationalistisk og delvis vildledende....

  9. Er jeres ledere 'likeable'?

    DEFF Research Database (Denmark)

    Nielsen, Rikke Kristine

    2013-01-01

    Færdigheder: Oftest er lederne langtfra de første til at kaste sig over ny teknologi. It-kundskaberne på chefgangen trænger til en opgradering.......Færdigheder: Oftest er lederne langtfra de første til at kaste sig over ny teknologi. It-kundskaberne på chefgangen trænger til en opgradering....

  10. er 1999

    DEFF Research Database (Denmark)

    Jensen, J. P.; Søndergaard, M.; Jeppesen, E.

    små cladoceer og hjuldyr, og især er maksimumsforekom-sterne af calanoide vandlopper og de små og store cladoceer og daf-nier gået tilbage. Den gennemsnitlige biomasse af dafnier er derimod øget især p.g.a. stigning i de 25 % af søerne med størst forekomster. Dyreplanktons græsning Betragtet under et...

  11. er 2000

    DEFF Research Database (Denmark)

    Jensen, J. P.; Søndergaard, M.; Jeppesen, E.

    styrke det fagli-ge grundlag for de mil-jøpolitiske prioriteringer og beslut-ninger. En væsentlig del af denne opgave er overvågning af miljø og natur. Det er derfor et naturligt led i Danmarks Miljø-undersø-gelsers opgave at forestå den landsdækkende rapportering af overvågnings-program-met inden...

  12. Pre-emptive Quality Control Protects the ER from Protein Overload via the Proximity of ERAD Components and SRP

    Directory of Open Access Journals (Sweden)

    Hisae Kadowaki

    2015-11-01

    Full Text Available Cells possess ER quality control systems to adapt to ER stress and maintain their function. ER-stress-induced pre-emptive quality control (ER pQC selectively degrades ER proteins via translocational attenuation during ER stress. However, the molecular mechanism underlying this process remains unclear. Here, we find that most newly synthesized endogenous transthyretin proteins are rerouted to the cytosol without cleavage of the signal peptide, resulting in proteasomal degradation in hepatocytes during ER stress. Derlin family proteins (Derlins, which are ER-associated degradation components, reroute specific ER proteins, but not ER chaperones, from the translocon to the proteasome through interactions with the signal recognition particle (SRP. Moreover, the cytosolic chaperone Bag6 and the AAA-ATPase p97 contribute to the degradation of ER pQC substrates. These findings demonstrate that Derlins-mediated substrate-specific rerouting and Bag6- and p97-mediated effective degradation contribute to the maintenance of ER homeostasis without the need for translocation.

  13. Membrane Targeting of P-type ATPases in Plant Cells

    International Nuclear Information System (INIS)

    Harper, Jeffrey F.

    2004-01-01

    How membrane proteins are targeted to specific subcellular locations is a very complex and poorly understood area of research. Our long-term goal is to use P-type ATPases (ion pumps), in a model plant system Arabidopsis, as a paradigm to understand how members of a family of closely related membrane proteins can be targeted to different subcellular locations. The research is divided into two specific aims. The first aim is focused on determining the targeting destination of all 10 ACA-type calcium pumps (Arabidopsis Calcium ATPase) in Arabidopsis. ACAs represent a plant specific-subfamily of plasma membrane-type calcium pumps. In contrast to animals, the plant homologs have been found in multiple membrane systems, including the ER (ACA2), tonoplast (ACA4) and plasma membrane (ACA8). Their high degree of similarity provides a unique opportunity to use a comparative approach to delineate the membrane specific targeting information for each pump. One hypothesis to be tested is that an endomembrane located ACA can be re-directed to the plasma membrane by including targeting information from a plasma membrane isoform, ACA8. Our approach is to engineer domain swaps between pumps and monitor the targeting of chimeric proteins in plant cells using a Green Fluorescence Protein (GFP) as a tag. The second aim is to test the hypothesis that heterologous transporters can be engineered into plants and targeted to the plasma membrane by fusing them to a plasma membrane proton pump. As a test case we are evaluating the targeting properties of fusions made between a yeast sodium/proton exchanger (Sod2) and a proton pump (AHA2). This fusion may potentially lead to a new strategy for engineering salt resistant plants. Together these aims are designed to provide fundamental insights into the biogenesis and function of plant cell membrane systems

  14. Rab7a modulates ER stress and ER morphology.

    Science.gov (United States)

    Mateus, Duarte; Marini, Elettra Sara; Progida, Cinzia; Bakke, Oddmund

    2018-05-01

    The Endoplasmic Reticulum (ER) is a membranous organelle with diverse structural and functional domains. Peripheral ER includes interconnected tubules, and dense tubular arrays called "ER matrices" together with bona fide flat cisternae. Transitions between these states are regulated by membrane-associated proteins and cytosolic factors. Recently, the small GTPases Rab10 and Rab18 were reported to control ER shape by regulating ER dynamics and fusion. Here, we present evidence that another Rab protein, Rab7a, modulates the ER morphology by controlling the ER homeostasis and ER stress. Indeed, inhibition of Rab7a expression by siRNA or expression of the dominant negative mutant Rab7aT22 N, leads to enlargement of sheet-like ER structures and spreading towards the cell periphery. Notably, such alterations are ascribable neither to a direct modulation of the ER shaping proteins Reticulon-4b and CLIMP63, nor to interactions with Protrudin, a Rab7a-binding protein known to affect the ER organization. Conversely, depletion of Rab7a leads to basal ER stress, in turn causing ER membrane expansion. Both ER enlargement and basal ER stress are reverted in rescue experiments by Rab7a re-expression, as well as by the ER chemical chaperone tauroursodeoxycholic acid (TUDCA). Collectively, these findings reveal a new role of Rab7a in ER homeostasis, and indicate that genetic and pharmacological ER stress manipulation may restore ER morphology in Rab7a silenced cells. Copyright © 2018 Elsevier B.V. All rights reserved.

  15. Jeg Er blevet FRANKofil

    DEFF Research Database (Denmark)

    Andersson, Lasse

    2014-01-01

    afhængig af Frank Underwood fra serien House of Cards på den fremadstormende TV-streamingstjenesten Netflix. Jeg har opdaget et nyt internetbaseret datingforhold. Et surrealt, fedt miks af det kyniske og joviale personificeret i karakteren Frank Underwood, som er helt igennem ubehagelig, men fantastisk...... spillet af Kevin Spacey. Og tak til Spacey der for en tid har forladt teateret ’The Old Vic’ i London for at begejstre mig. Der er generelt to årsager til mine FRANKofile tilbøjeligheder. For det første er Netflix’s remake af den tyve år gamle BBC serie House of Cards efter min menig et stykke tv...... anden grund, til at jeg er blevet Frankofil, er, at jeg ikke skal sidde og vente på næste søndag efter søndag efter søndag for at få lov at se næste afsnit. Netflix lagde alle tretten timer af sæson 2 ud på nettet. Jeg afgør selv, hvornår jeg skal have mere Frank! Men på trods af mit narkomanlignende...

  16. A plasma membrane H + ATPase gene is germinationinduced in ...

    African Journals Online (AJOL)

    A plasma membrane H + ATPase gene is germinationinduced in wheat embryos. ... African Journal of Biotechnology ... of a germination specific plasma membrane H+-ATPase was analyzed by RTPCR and in situ RNA hybridization methods.

  17. Ondskaben er fortryllende

    DEFF Research Database (Denmark)

    Schubart, Rikke

    2013-01-01

    Indlæg om tv-serien Once Upon a Time (2011-), der hører til genren fairytale fantasy, der blander eventyr og fantasy. Her bliver alle eventyr brugt i en fortælling om alle beboere i en lille by, der ikke ved, at de i virkeligheden er eventyr-karakterer.......Indlæg om tv-serien Once Upon a Time (2011-), der hører til genren fairytale fantasy, der blander eventyr og fantasy. Her bliver alle eventyr brugt i en fortælling om alle beboere i en lille by, der ikke ved, at de i virkeligheden er eventyr-karakterer....

  18. Functional Analysis of P4-ATPases

    DEFF Research Database (Denmark)

    Theorin, Lisa

    and mammalian P4-ATPases have been studied extensively and the physiological function is mostly known, while the exact biochemistry and specific activity is mostly unknown. Even though the plant Arabidopsis thaliana has 12 P4-ATPases, not much is known about their function. In this study, the biochemical...... for purification of the complex by one-step purification. The ATPase activity of the ALA2/ALIS5 complex was stimulated in a highly specific manner by phosphatidylserine. Changes in the phosphatidylserine headgroup or alteration of the stereochemistry affected enzymatic activity. The results demonstrate that ALA2...... is specific for phosphatidylserine and that binding of the lipid to the substrate binding site requires a unique spatial configuration of the lipid head group. Detailed information on the substrate requirements lead the way towards the full function and transport pathway of lipid flippases in plants. Recent...

  19. Roles of transmembrane segment M1 of Na(+),K (+)-ATPase and Ca (2+)-ATPase, the gatekeeper and the pivot

    DEFF Research Database (Denmark)

    Einholm, Anja P.; Andersen, Jens Peter; Vilsen, Bente

    2007-01-01

    In this review we summarize mutagenesis work on the structure-function relationship of transmembrane segment M1 in the Na(+),K(+)-ATPase and the sarco(endo)plasmic reticulum Ca(2+)-ATPase. The original hypothesis that charged residues in the N-terminal part of M1 interact with the transported...... cations can be rejected. On the other hand hydrophobic residues in the middle part of M1 turned out to play crucial roles in Ca(2+) interaction/occlusion in Ca(2+)-ATPase and K(+) interaction/occlusion in Na(+),K(+)-ATPase. Leu(65) of the Ca(2+)-ATPase and Leu(99) of the Na(+),K(+)-ATPase, located...... of the extracytoplasmic gate in both the Ca(2+)-ATPase and the Na(+),K(+)-ATPase. Udgivelsesdato: 2007-Dec...

  20. P4 ATPases - lipid flippases and their role in disease

    NARCIS (Netherlands)

    Folmer, Dineke E.; Elferink, Ronald P. J. Oude; Paulusma, Coen C.

    2009-01-01

    P4 ATPases (type 4 P-type ATPases) are multispan transmembrane proteins that have been implicated in phospholipid translocation from the exoplasmic to the cytoplasmic leaflet of biological membranes. Studies in Saccharomyces cerevisiae have indicated that P4 ATPases are important in vesicle

  1. Na+/K+-ATPase: Activity and inhibition

    Science.gov (United States)

    Čolović, M.; Krstić, D.; Krinulović, K.; Momić, T.; Savić, J.; Vujačić, A.; Vasić, V.

    2009-09-01

    The aim of the study was to give an overview of the mechanism of inhibition of Na+/K+-ATPase activity induced by some specific and non specific inhibitors. For this purpose, the effects of some ouabain like compounds (digoxin, gitoxin), noble metals complexes ([PtCl2DMSO2], [AuCl4]-, [PdCl4]2-, [PdCl(dien)]+, [PdCl(Me4dien)]+), transition metal ions (Cu2+, Zn2+, Fe2+, Co2+), and heavy metal ions (Hg2+, Pb2+, Cd2+) on the activity of Na+/K+-ATPase from rat synaptic plasma membranes (SPM), porcine cerebral cortex and human erythrocytes were discussed.

  2. Myocardial Na,K-ATPase: Clinical aspects

    Science.gov (United States)

    Kjeldsen, Keld

    2003-01-01

    The specific binding of digitalis glycosides to Na,K-ATPase is used as a tool for Na,K-ATPase quantification with high accuracy and precision. In myocardial biopsies from patients with heart failure, total Na,K-ATPase concentration is decreased by around 40%; a correlation exists between a decrease in heart function and a decrease in Na,K-ATPase concentration. During digitalization, around 30% of remaining pumps are occupied by digoxin. Myocardial Na,K-ATPase is also influenced by other drugs used for the treatment of heart failure. Thus, potassium loss during diuretic therapy has been found to reduce myocardial Na,K-ATPase, whereas angiotensin-converting enzyme inhibitors may stimulate Na,K pump activity. Furthermore, hyperaldosteronism induced by heart failure has been found to decrease Na,K-ATPase activity. Accordingly, treatment with the aldosterone antagonist, spironolactone, may also influence Na,K-ATPase activity. The importance of Na,K pump modulation with heart disease, inhibition in digitalization and other effects of medication should be considered in the context of sodium, potassium and calcium regulation. It is recommended that digoxin be administered to heart failure patients who, after institution of mortality-reducing therapy, still have heart failure symptoms, and that the therapy be continued if symptoms are revealed or reduced. Digitalis glycosides are the only safe inotropic drugs for oral use that improve hemodynamics in heart failure. An important aspect of myocardial Na,K pump affection in heart disease is its influence on extracellular potassium (Ke) homeostasis. Two important aspects should be considered: potassium handling among myocytes, and effects of potassium entering the extracellular space of the heart via the bloodstream. It should be noted that both of these aspects of Ke homeostasis are affected by regulatory aspects, eg, regulation of the Na,K pump by physiological and pathophysiological conditions, as well as by medical

  3. Scenen er blevet din

    DEFF Research Database (Denmark)

    Helms, Niels Henrik

    2008-01-01

    Verden kan betragtes som en skueplads med mange forskellige scener. I bidrgaet vil læseren kunne stifte bekendtskab med forskellige refleksioner over netop denne metaforik som analyseoptik i forhold til begrebet innovation. Afsættet er ønsket om at finde en måde at analysere og begrebsliggøre inn...

  4. Er tang fremtidens superfood?

    DEFF Research Database (Denmark)

    Holdt, Susan Løvstad

    seneste år fået øjnene op for tang indenfor områderne: biokemikalier og bioaktive stoffer, ekstraktion, gastronomi, produktudvikling, foder, bioraffinaderier, dyrkning osv. Men hvad er det tang kan? Dette foredrag vil vise at vi med tangdyrkning kan rykke mere bæredygtig fødevareproduktion ud på havet...

  5. Ideologi er noget bras

    DEFF Research Database (Denmark)

    Hansen, Brian Benjamin; Bjerre, Henrik Jøker

    fra den slovenske filosof Slavoj Zizek stilles imidlertid det spørgsmål i bogen, om ideologien idag ikke blot har taget en anden form. Vi lever i dag ikke i post-ideologiske tider. Snarere er braset selv – X Factor, forbrugsobjekter, underholdning, følelser og balloner – blevet den næsten usynlige...

  6. Riget - alt er tilladt

    DEFF Research Database (Denmark)

    2017-01-01

    Riget indtager en nøglestilling, når overgangen mellem faser i Triers produktion skal karakteriseres. Riget danner bro mellem den billedfascination og de radikale stileksperimenter, der præger Triers tidlige produktion, og den genre- og plotbevidsthed, der er typisk for film som Breaking the Waves...

  7. Hvad er tidligt sprog

    DEFF Research Database (Denmark)

    Skriver Jensen, Anders

    2009-01-01

    Forfatteren argumenterer for at "early literacy" kan oversættes til "tidligt sprog"; særligt når der er tale om en helhedsorienteret tilgang med vægt på sprog som et alsidigt medie for kommunikation- og betydningsskabelse. Med inspiration fra Jerome Bruners kulturpsykologi diskuteres mulige...

  8. Er danskerne racister

    DEFF Research Database (Denmark)

    Bech, Henning; Necef, Mehmet Ümit

    Igennem de seneste årtier er det blevet almindeligt at tale om, at der er en udbredt racisme i Danmark. Påstande om danskernes racisme, fremmedhad og diskrimination optræder dagligt i offentligheden og i medierne, og der henvises ofte til, hvad ’forskerne’ og de ’videnskabelige undersøgelser’ siger...... om emnet. Der kan da næppe heller være tvivl om, at der forekommer racistiske holdninger hos nogle danskere. Men er problemet så stort, som det gøres til i den offentlige debat? Bogen ønsker at afklare, hvorvidt der er videnskabelig dokumentation for påstandene om danskernes racisme. Den går i dybden...... med en række forskeres og eksperters udtalelser på området og præsenterer en grundig analyse af deres fremstilling af dansk racisme i forhold til emner som kultur, seksualitet, kriminalitet og arbejdsmarked....

  9. Function of endoplasmic reticulum calcium ATPase in innate immunity-mediated programmed cell death

    Science.gov (United States)

    Zhu, Xiaohong; Caplan, Jeffrey; Mamillapalli, Padmavathi; Czymmek, Kirk; Dinesh-Kumar, Savithramma P

    2010-01-01

    Programmed cell death (PCD) initiated at the pathogen-infected sites during the plant innate immune response is thought to prevent the development of disease. Here, we describe the identification and characterization of an ER-localized type IIB Ca2+-ATPase (NbCA1) that function as a regulator of PCD. Silencing of NbCA1 accelerates viral immune receptor N- and fungal-immune receptor Cf9-mediated PCD, as well as non-host pathogen Pseudomonas syringae pv. tomato DC3000 and the general elicitor cryptogein-induced cell death. The accelerated PCD rescues loss-of-resistance phenotype of Rar1, HSP90-silenced plants, but not SGT1-silenced plants. Using a genetically encoded calcium sensor, we show that downregulation of NbCA1 results in the modulation of intracellular calcium signalling in response to cryptogein elicitor. We further show that NbCAM1 and NbrbohB function as downstream calcium decoders in N-immune receptor-mediated PCD. Our results indicate that ER-Ca2+-ATPase is a component of the calcium efflux pathway that controls PCD during an innate immune response. PMID:20075858

  10. The mechanism of Torsin ATPase activation.

    Science.gov (United States)

    Brown, Rebecca S H; Zhao, Chenguang; Chase, Anna R; Wang, Jimin; Schlieker, Christian

    2014-11-11

    Torsins are membrane-associated ATPases whose activity is dependent on two activating cofactors, lamina-associated polypeptide 1 (LAP1) and luminal domain-like LAP1 (LULL1). The mechanism by which these cofactors regulate Torsin activity has so far remained elusive. In this study, we identify a conserved domain in these activators that is predicted to adopt a fold resembling an AAA+ (ATPase associated with a variety of cellular activities) domain. Within these domains, a strictly conserved Arg residue present in both activating cofactors, but notably missing in Torsins, aligns with a key catalytic Arg found in AAA+ proteins. We demonstrate that cofactors and Torsins associate to form heterooligomeric assemblies with a defined Torsin-activator interface. In this arrangement, the highly conserved Arg residue present in either cofactor comes into close proximity with the nucleotide bound in the neighboring Torsin subunit. Because this invariant Arg is strictly required to stimulate Torsin ATPase activity but is dispensable for Torsin binding, we propose that LAP1 and LULL1 regulate Torsin ATPase activity through an active site complementation mechanism.

  11. Nucleotide binding to Na+/K+-ATPase

    Czech Academy of Sciences Publication Activity Database

    Kubala, Martin; Lánský, Zdeněk; Ettrich, R.; Plášek, J.; Teisinger, Jan; Amler, Evžen

    2005-01-01

    Roč. 272, č. S1 (2005), s. 191-191 E-ISSN 1742-4658. [FEBS Congress /30./ and IUBMB Conference /9./. 02.07.2005-07.07.2005, Budapest] Keywords : Na+/K+- ATPase * ATP binding * TNP-ATP Subject RIV: BO - Biophysics

  12. The plant plasma membrane H+-ATPase

    DEFF Research Database (Denmark)

    Ekberg, Kira

    of plants and fungi to generate electrochemical proton gradients. A recently published crystal structure of a plasma membrane H(+)-ATPase contributes to our knowledge about the mechanism of these essential enzymes. Together with biochemical and structural data presented in this thesis we are now able...

  13. Er HR ude i tovene?

    DEFF Research Database (Denmark)

    Poulfelt, Flemming

    2015-01-01

    HR: Er der behov for nytænkning i HR-land? Artikler i Harvard Business Review - bakket op af en dansk undersøgelse - konkluderer, at HR stadig mangler gennemslagskraft i virksomhederne. Er HR ude i tovene? ... For i undersøgelsen "Ny Dansk Ledelse" (maj 2015), som er baseret på danske lederes...

  14. Facebook er internettet nu

    DEFF Research Database (Denmark)

    Tække, Jesper

    2014-01-01

    Den store vision for world wide web var, at alle kunne få ubegrænset adgang til alverdens information. www var mangfoldigt og anarkistisk. I dag er der rigtig mange, der kun bruger nettet til at være på Facebook. Hvad bruger Facebook det til? Og hvad gør det ved vores udsyn?......Den store vision for world wide web var, at alle kunne få ubegrænset adgang til alverdens information. www var mangfoldigt og anarkistisk. I dag er der rigtig mange, der kun bruger nettet til at være på Facebook. Hvad bruger Facebook det til? Og hvad gør det ved vores udsyn?...

  15. Hvad er en by?

    DEFF Research Database (Denmark)

    Parby, Jakob; Thelle, Mikkel

    2011-01-01

    Nytænkning. Med de moderne megabyer er det slut med at forestille sig byen som noget, man planlægger og giver form. Vi skal lære at se dem som vildtvoksende urbane landskaber. Artikel om megabyer, byudvikling og forestillingen om byen gennem tiderne. Skrevet delvist i anledning af åbningen af...... udstillingen Citambulos på Københavns Museum, der handlede om Mexico City....

  16. The parietal cell gastric H, K-ATPase also functions as the Na, K-ATPase and Ca-ATPase in altered states.

    Science.gov (United States)

    Ray, Tushar

    2013-01-01

    This article offers an explanation for the apparent lack of Na, K-ATPase activity in parietal cells although ouabain has been known to inhibit gastric acid secretion since 1962. The gastric H, K-ATPase (proton-pump) seems to be acting in altered states, thus behaving like a Na, K-ATPase (Na-pump) and/or Ca-ATPase (Ca-pump) depending on cellular needs.  This conclusion is based on the following findings. First, parietal cell fractions do not exhibit Na, K-ATPase activity at pH 7.0 but do at pH 8.5. Second, the apical plasma membrane (APM) fraction exhibits a (Ca or Mg)-ATPase activity with negligible H, K-ATPase activity. However, when assayed with Mg alone in presence of the 80 k Da cytosolic proton-pump activator (HAF), the APM fraction reveals remarkably high H, K-ATPase activity, suggesting the observed low affinity of Ca (or Mg)-ATPase is an altered state of the latter. Third, calcium (between 1 and 4 µM) shows both stimulation and inhibition of the HAF-stimulated H, K-ATPase depending on its concentration, revealing a close interaction between the  proton-pump activator and local Ca concentration in gastric H, K-ATPase function. Such interactions suggest that Ca is acting as a terminal member of the intracellular signaling system for the HAF-regulated proton-pump. It appears that during resting state, the HAF-associated H, K-ATPase remains inhibited by Ca (>1 µM) and, prior to resumption of acid secretion the gastric H, K-ATPase acts temporarily as a Ca-pump for removing excess Ca from its immediate environment. This conclusion is consistent with the recent reports of immunochemical co-localization of the gastric H, K-ATPase and Ca-ATPase by superimposition in parietal cells, and a transitory efflux of Ca immediately preceding the onset of acid secretion. These new perspectives on proton-pump function would open new avenues for a fuller understanding of the intracellular regulation of the ubiquitous Na-pump.

  17. Amino Acid Availability Modulates Vacuolar H+-ATPase Assembly*

    Science.gov (United States)

    Stransky, Laura A.; Forgac, Michael

    2015-01-01

    The vacuolar H+-ATPase (V-ATPase) is an ATP-dependent proton pump composed of a peripheral ATPase domain (V1) and a membrane-integral proton-translocating domain (V0) and is involved in many normal and disease processes. An important mechanism of regulating V-ATPase activity is reversible assembly of the V1 and V0 domains. Increased assembly in mammalian cells occurs under various conditions and has been shown to involve PI3K. The V-ATPase is necessary for amino acid-induced activation of mechanistic target of rapamycin complex 1 (mTORC1), which is important in controlling cell growth in response to nutrient availability and growth signals. The V-ATPase undergoes amino acid-dependent interactions with the Ragulator complex, which is involved in recruitment of mTORC1 to the lysosomal membrane during amino acid sensing. We hypothesized that changes in the V-ATPase/Ragulator interaction might involve amino acid-dependent changes in V-ATPase assembly. To test this, we measured V-ATPase assembly by cell fractionation in HEK293T cells treated with and without amino acids. V-ATPase assembly increases upon amino acid starvation, and this effect is reversed upon readdition of amino acids. Lysosomes from amino acid-starved cells possess greater V-ATPase-dependent proton transport, indicating that assembled pumps are catalytically active. Amino acid-dependent changes in both V-ATPase assembly and activity are independent of PI3K and mTORC1 activity, indicating the involvement of signaling pathways distinct from those implicated previously in controlling assembly. By contrast, lysosomal neutralization blocks the amino acid-dependent change in assembly and reactivation of mTORC1 after amino acid starvation. These results identify an important new stimulus for controlling V-ATPase assembly. PMID:26378229

  18. Response of Ca2+-ATPase to clinorotaion of pea seedlings. O. M. Nedukha and E. L. Kordyum

    Science.gov (United States)

    Nedukha, Olena

    2016-07-01

    The present study was aimed to reveal of response of Ca2+-ATPase activity of cortex cells in distal elongation zone of Pisum sativum root to slow clinorotation. Pea seedlings were grown on a horizontal clinostat (2 rpm) and in the stationary control for 6 days. The electron-cytochemical method was used to examine the effects of imitated microgravity on the distribution of Ca2+-ATPase in outer layers of root cortex. The quantitative analysis of the density of cytochemical reaction products was measured using the Image J program. Electron microscopy showed the presence of electron-dense lead phosphate precipitated grains, the enzymatic activity reaction products on the plasma membrane, membranes of vesicular structures, endoplasmic reticulum (ER) and on organelles envelope in both of samples of the stationary control and clinorotated seedlings. We revealed the sensitivity of Ca2+-ATPase to clinorotation. The quantitative analysis of the area and density of enzymatic activity reaction products revealed that clinorotation led to the decrease of 3.4 times the density of reaction products on the plasma membrane and the increase of reaction products density on endomembranes and organelles membranes, in particular: in 2.2 times on mitochondria membranes; in 1.3 times - on membranes of ER; in 2.5 times - on tonoplast; by an order of magnitude greater - on contacting membranes of organelles with plasma membrane in comparison with such in cells of control samples. The data analysis can indicate an intensification of calcium pump on endomembranes, on envelopes of cytoplasmic organelles and nucleus. The obtained data suggest that the redistribution of Ca2+-ATPase activity in cells can be mediated by the activation of certain isoforms of enzyme or/and by an activation of Ca2+/H+ antiporter in plasma membrane that helps to maintain optimal calcium balance in plant cells under imitated microgravity.

  19. Hvad er tekstfiktion?

    DEFF Research Database (Denmark)

    Widell, Peter

    2012-01-01

    artiklen er et bidrag til belysningen af genteori og bygger på ideen om at karakterisere skønlitteratur som et restringeret sæt af universelle tekstparametre. Artiklen fokuserer især på parametren tekstfiktion, hvor jeg forsvarer den position, at en fiktiv tekst hverken kan karakteriseres som en ...... intentioner. Det betyder dog ikke, at sandhed og mening ikke spiller noget rolle ved identifikationen af fiktion, sådan som det hævdes af Walsh og andre....

  20. Sperm Na+, K+-ATPase α4 and plasma membrane Ca2+-ATPase (PMCA) 4 regulation in asthenozoospermia.

    Science.gov (United States)

    Lestari, Silvia W; Miati, Dessy Noor; Seoharso, P; Sugiyanto, R; Pujianto, Dwi A

    2017-10-01

    Asthenozoospermia, which is characterized by reduced motility, is one of the etiologies of male infertility. Its biochemical and functional consequences include altered ATPase activity. This study investigated the activities of Na + , K + -ATPase and Ca 2+ -ATPase and the expression of Na + , K + -ATPase α4 and PMCA4 isoforms in human sperm of asthenozoospermic infertile men. Nineteen samples from asthenozoospermic infertile couples were examined in this study. Computerized-assisted semen analysis (CASA) was performed, and the enzyme activity was measured based on the ability of ATPase to release organic phosphate from ATP as a substrate. The Na + , K + -ATPase α4 and PMCA4 isoform expression levels were measured by western immunoblotting, whereas the protein distribution was examined by immunocytochemistry. This showed that the Na + , K + -ATPase activity and the Na + , K + -ATPase α4 isoform expression were lower in the asthenozoospermia group than in the normozoospermia group (8.688±1.161 versus 13.851±1.884 µmol Pi/mg protein/h, respectively; p>0.05). In contrast, the Ca 2+ -ATPase activity was significantly higher in the asthenozoospermia group than in the normozoospermia group (11.154±1.186 versus 2.725±0.545 µmol Pi/mg protein/h, respectively; p0.05). The altered ATPase activity and isoform expression in asthenozoospermia may impair sperm structure and function.

  1. Lithium-induced inhibition of Na-K ATPase and Ca ATPase activities in rat brain synaptosome.

    Science.gov (United States)

    Cho, Y. W.

    1995-01-01

    To explore the action mechanism of lithium in the brain, the author investigated the effects of lithium on Na-K ATPase and Ca ATPase in rat brain synaptosomes prepared from forebrains by the method of Booth and Clark. The activities of Na-K ATPase and Ca ATPase were assayed by the level of inorganic phosphate liberated from the hydrolysis of ATP. Lithium at the optimum therapeutic concentration of 1 mM decreased the activity of Na-K ATPase from the control value of 19.08 +/- 0.29 to 18.27 +/- 0.10 micromoles Pi/mg protein/h and also reduced the activity of Ca ATPase from 6.38 +/- 0.12 to 5.64 +/- 0.12 micromoles Pi/mg protein/h. The decreased activity of Na-K ATPase will decrease the rate of Ca2+ efflux, probably via an Na-Ca exchange mechanism and will increase the rate of Ca2+ entry by the depolarization of nerve terminals. The reduced activity of Ca ATPase will result in the decreased efflux of Ca2+. As a Conclusion, it can be speculated that lithium elevates the intrasynaptosomal Ca2+ concentration via inhibition of the activities of Na-K ATPase and Ca ATPase, and this increased [Ca2+]i will cause the release of neurotransmitters and neurological effects of lithium. PMID:7598829

  2. Overproduction of PIB-Type ATPases

    DEFF Research Database (Denmark)

    Liu, Xiangyu; Sitsel, Oleg; Wang, Kaituo

    2016-01-01

    Understanding of the functions and mechanisms of fundamental processes in the cell requires structural information. Structural studies of membrane proteins typically necessitate large amounts of purified and preferably homogenous target protein. Here, we describe a rapid overproduction and purifi...... and purification strategy of a bacterial PIB-type ATPase for isolation of milligrams of target protein per liter Escherichia coli cell culture, with a final quality of the sample which is sufficient for generating high-resolution crystals....

  3. Transcriptional regulators of Na, K-ATPase subunits

    OpenAIRE

    Zhiqin eLi; Sigrid A Langhans

    2015-01-01

    The Na,K-ATPase classically serves as an ion pump creating an electrochemical gradient across the plasma membrane that is essential for transepithelial transport, nutrient uptake and membrane potential. In addition, Na,K-ATPase also functions as a receptor, a signal transducer and a cell adhesion molecule. With such diverse roles, it is understandable that the Na,K-ATPase subunits, the catalytic alpha-subunit, the beta-subunit and the FXYD proteins, are controlled extensively during developme...

  4. Transcriptional regulators of Na, K-ATPase subunits

    Directory of Open Access Journals (Sweden)

    Zhiqin eLi

    2015-10-01

    Full Text Available The Na,K-ATPase classically serves as an ion pump creating an electrochemical gradient across the plasma membrane that is essential for transepithelial transport, nutrient uptake and membrane potential. In addition, Na,K-ATPase also functions as a receptor, a signal transducer and a cell adhesion molecule. With such diverse roles, it is understandable that the Na,K-ATPase subunits, the catalytic alpha-subunit, the beta-subunit and the FXYD proteins, are controlled extensively during development and to accommodate physiological needs. The spatial and temporal expression of Na,K-ATPase is partially regulated at the transcriptional level. Numerous transcription factors, hormones, growth factors, lipids and extracellular stimuli modulate the transcription of the Na,K-ATPase subunits. Moreover, epigenetic mechanisms also contribute to the regulation of Na,K-ATPase expression. With the ever growing knowledge about diseases associated with the malfunction of Na,K-ATPase, this review aims at summarizing the best-characterized transcription regulators that modulate Na,K-ATPase subunit levels. As abnormal expression of Na,K-ATPase subunits have been observed in many carcinoma, we will also discuss transcription factors that are associated with epithelial-to-mesenchymal transition, a crucial step in the progression of many tumors to malignant disease.

  5. Conditions of activation of yeast plasma membrane ATPase.

    Science.gov (United States)

    Sychrová, H; Kotyk, A

    1985-04-08

    The in vivo activation of the H+-ATPase of baker's yeast plasma membrane found by Serrano in 1983 was demonstrated with D-glucose aerobically and anaerobically (as well as in a respiration-deficient mutant) and, after suitable induction, with maltose, trehalose, and galactose. The activated but not the control ATPase was sensitive to oligomycin. No activation was possible in a cell-free extract with added glucose. The ATPase was not activated in yeast protoplasts which may account for the absence of glucose-stimulated secondary active transports in these wall-less cells and provide support for a microscopic coupling between ATPase activity and these transports in yeast cells.

  6. Na(+),K (+)-ATPase as a docking station: protein-protein complexes of the Na(+),K (+)-ATPase.

    Science.gov (United States)

    Reinhard, Linda; Tidow, Henning; Clausen, Michael J; Nissen, Poul

    2013-01-01

    The Na(+),K(+)-ATPase, or sodium pump, is well known for its role in ion transport across the plasma membrane of animal cells. It carries out the transport of Na(+) ions out of the cell and of K(+) ions into the cell and thus maintains electrolyte and fluid balance. In addition to the fundamental ion-pumping function of the Na(+),K(+)-ATPase, recent work has suggested additional roles for Na(+),K(+)-ATPase in signal transduction and biomembrane structure. Several signaling pathways have been found to involve Na(+),K(+)-ATPase, which serves as a docking station for a fast-growing number of protein interaction partners. In this review, we focus on Na(+),K(+)-ATPase as a signal transducer, but also briefly discuss other Na(+),K(+)-ATPase protein-protein interactions, providing a comprehensive overview of the diverse signaling functions ascribed to this well-known enzyme.

  7. Understanding the mechanisms of ATPase beta family genes for cellular thermotolerance in crossbred bulls.

    Science.gov (United States)

    Deb, Rajib; Sajjanar, Basavaraj; Singh, Umesh; Alex, Rani; Raja, T V; Alyethodi, Rafeeque R; Kumar, Sushil; Sengar, Gyanendra; Sharma, Sheetal; Singh, Rani; Prakash, B

    2015-12-01

    Na+/K+-ATPase is an integral membrane protein composed of a large catalytic subunit (alpha), a smaller glycoprotein subunit (beta), and gamma subunit. The beta subunit is essential for ion recognition as well as maintenance of the membrane integrity. Present study was aimed to analyze the expression pattern of ATPase beta subunit genes (ATPase B1, ATPase B2, and ATPase B3) among the crossbred bulls under different ambient temperatures (20-44 °C). The present study was also aimed to look into the relationship of HSP70 with the ATPase beta family genes. Our results demonstrated that among beta family genes, transcript abundance of ATPase B1 and ATPase B2 is significantly (P ATPase Β1, ATPase B2, and ATPase B3 is highly correlated (P ATPase beta family genes for cellular thermotolerance in cattle.

  8. Rouw mag er zijn

    Directory of Open Access Journals (Sweden)

    Marlieke Moors

    2015-12-01

    Full Text Available ABSTRACTGrieving is allowedGrief is a human experience. Every form of loss shapes you into the human being you are today. Contrary to what earlier, unproven grief models postulate, grief does not have an end point. That is, bereavement does not have to be completely processed, but it should be integrated into someone’s life. The outdated grief models were often interpreted and used in a normative way, which led to a normative standard model. This portraits the belief that every mourner would experience similar symptoms and would go through a fixed pattern of phases. However, the updated vision emphasizes the individual and unique process of coping with loss: norms concerning grief should be banned. By means of literature research, interviews with professionals and personal experiences, it became clear that finding an equilibrium between restoration-orientated and loss-orientated coping styles is most beneficial. An important aspect in finding this balance is meaningfulness. Furthermore, the ability to bear a loss and to adapt accordingly are important components. Lastly, attaching significance to a loss is a constructive way of integrating the loss into one’s life. The death of a loved one should therefore not be forgotten or tucked away. After all, grief is the price we pay for love. SAMENVATTINGRouw mag er zijnRouw is een menselijke ervaring en elk verlies vormt je als mens. Rouw heeft, in tegenstelling tot wat de verouderde, niet bewezen rouwmodellen beweren, geen eindpunt. Verlies hoeft namelijk niet verwerkt te worden, maar moet juist geïntegreerd worden in iemands leven. De verouderde rouwmodellen zijn vaak normatief opgevat en toegepast, waaruit een normatief standaardmodel is ontstaan. Daarbij werd gedacht dat elke rouwende dezelfde symptomen zou vertonen en het rouwproces volgens vaste fasen zou verlopen. Binnen de vernieuwde visie wordt er juist van uitgegaan dat elk individu een unieke manier van reageren op rouw heeft. Er zou

  9. Han er her endnu

    DEFF Research Database (Denmark)

    Bonde, Lisbeth

    2012-01-01

    Interview med den kinesiske aktivist og billedkunstner Ai Weiwei, der sad fængslet i 81 dage i 2011. Hans pas er stadig (i februar 2014) inddraget af myndighederne, så han kan ikke forlade landet, selv om han har betalt en bøde på 13 mio. kr. for ”skatteunddragelse”. Både i sin kunst og i sine...... mange aktioner retter han en hård kritik af det korrupte kinesiske styre, som træder menneskerettighederne under fode. Men uretfærdighedsfølelsen har sin pris i et totalitært og topstyret samfund. Ligegyldigt, hvor mange vigtige udstillinger han har i udlandet, må han blive hjemme og styre aktiviteterne...... via cyberspace. I interviewet fortæller Ai Weiwei om de personlige omkostninger, som hans kamp mod systemet indebærer....

  10. Structural studies of the vacuolar membrane ATPase from Neurospora crassa and comparison with the tonoplast membrane ATPase and Zea mays

    International Nuclear Information System (INIS)

    Bowman, E.J.; Mandala, S.; Taiz, L.; Bowman, B.J.

    1986-01-01

    The H + translocating ATPase located on vacuolar membranes of Neurospora crassa was partially purified by solubilization in two detergents, Triton X-100 and N-hexadecyl-N,N-dimethyl-3-ammonio-1-propanesulfonate, followed by centrifugation on sucrose density gradients. Two polypeptides of M/sub r/ ≅ 70,000 and ≅ 62,000 consistently migrated with activity, along with several minor bands of lower molecular weight. Radioactively labeled inhibitors of ATPase activity, N-[ 14 C]ethylmaleimide and 7-chloro-4-nitro[ 14 C]benzo-2-oxa-1,3-diazole, labeled the M/sub r/ ≅ 70,000 polypeptide; this labeling was reduced in the presence of ATP. N,N'-[ 14 C]dicyclohexylcarbodiimide labeled a polypeptide of M/sub r/ ≅ 15,000. Estimation of the functional size of the vacuolar membrane ATPase by radiation inactivation gave a value of M/sub r/ 5.2 x 10 5 , 10-15% larger than the mitochondrial ATPase. The Neurospora vacuolar ATPase showed no crossreactivity with antiserum to plasma membrane or mitochrondrial ATPase but stongly crossreacted with antiserum against a polypeptide of M/sub r/ ≅ 70,000 associated with the tonoplast ATPase of corn coleoptiles. These results suggest that fungal and plant vacuolar ATPases may be large multisubunit complexes, somewhat similar to, but immunologically distinct from, known F 0 F 1 ATPases

  11. De 9 P’er

    DEFF Research Database (Denmark)

    Rennison, Betina Wolfgang

    2017-01-01

    Ledere skal i dag selv skabe deres eget rum til ledelse, men hvad er med til at sætte det, hvilke betingelser og udfordringer er der, og hvordan kan lederne skabe sig selv i et hav af forventninger? Dette katalog inviterer til refleksion herom....

  12. Fremtidens undervisningsmiljøer

    DEFF Research Database (Denmark)

    2013-01-01

    Som oplægget til dette temanummer af LOM også indikerede, så sætter vi fokus på fremtidens undervisningsmiljøer på universiteter og UCer. Fremtidens undervisningsmiljøer har mange facetter, hvilket samlingen af artikler også illustrerer. “Fremtidens Undervisningsmiljø” handler om eksisterende erf...

  13. Den rige personlighed er livsduelig

    DEFF Research Database (Denmark)

    Holm, Claus

    2015-01-01

    For omtrent 160 år siden formulerede Karl Marx forestillingen om en rig individualitet som det kommunistiske samfunds individideal. I dag får forestillingen relevans. Lyder det lidt besynderligt, er det ikke mærkeligt. For de fleste af os går næppe rundt og tror, at vi er lige på trapperne til...

  14. Humor er en alvorlig sag

    DEFF Research Database (Denmark)

    Søltoft, Pia

    2016-01-01

    I modsætning til ironi er humor for Kierkegaard fællesskabsgivende – ironikeren hævder sig selv, men humoristen har sympati med den, man ler med. Humor er hos Kierkegaard udtryk for, at humoristen forliger sig med tilværelsen og dens luner, og dermed grænser humoren hos Kierkegaard op til det...

  15. Brandulykker er et socialt problem

    DEFF Research Database (Denmark)

    Leth, Peter Mygind

    1999-01-01

    Det er de gamle, de syge, de handicappede og alkoholikerne, der brænder inde. Typisk har de tabt en cigaret eller tændstik på tøjet. En del af disse brandulykker opstår på plejehjem og andre institutioner, hvor det ofte er plejepersonalet, der opdager og slukker branden....

  16. Phosphorylation of the Na+,K+-ATPase and the H+,K+-ATPase

    DEFF Research Database (Denmark)

    Poulsen, Hanne; Morth, Jens Preben; Jensen, Jan Egebjerg

    2010-01-01

    pumps are very homologous, and at least one of the phosphorylation sites is conserved, namely a cAMP activated protein kinase (PKA) site, which is important for regulating pumping activity, either by changing the cellular distribution of the ATPases or by directly altering the kinetic properties...

  17. Na, K-ATPase as signaling transducer

    OpenAIRE

    Li, Juan

    2007-01-01

    It is now generally agreed that Na,K-ATPase (NKA), in addition to its role in the maintenance of Na+ and K+ gradients across the cell membrane, is a signal transducer. Our group has identified a novel signaling pathway where NKA interact with IP3R to form a signaling microdomain. Ouabain, a specific ligand of NKA, activates this pathway, triggers slow Ca2+ oscillations and activates NF-κB. In current study, the molecular mechanisms and some important downstream effects of NK...

  18. Substrate Specificity of Na+,Cl-(HCO3-)-ATPase.

    Science.gov (United States)

    Yurkiv, V A; Melikhov, V I; Shubin, V S

    2016-09-01

    We studied substrate specificity of Na + ,Cl - (HCO 3 - )-ATPase. In most cases, replacement of ATP for other phosphate-containing substances resulted in not only pronounced suppression of phosphohydrolase reactions, but also dramatic changes of their responsiveness to the stimulating effect of monovalent ions. The data showed that Na + ,Cl - (HCO 3 - )-ATPase is a highly specific enzyme for ATP.

  19. Regulation of the Plasma Membrane H+-ATPase

    DEFF Research Database (Denmark)

    Falhof, Janus

    The plasma membrane (PM) H+-ATPase is responsible for generating the electrochemical gradientthat drives the secondary transport of nutrients across the cellular membrane. It belongs to a familyof cation and lipid transporters that are vital to many organisms. PM H+-ATPases are Type P3AATPases...

  20. Kinase-Mediated Regulation of P4-ATPases

    DEFF Research Database (Denmark)

    Frøsig, Merethe Mørch

    designed a fast and efficient screening strategy to identify novel regulator proteins of P4-ATPases. The system is based on heterologous expression in a specially designed yeast strain, and regulatory proteins can be identified via change in activity of the P4-ATPase of interest. Hereby the first steps...

  1. Conformational changes in the AAA ATPase p97–p47 adaptor complex

    Science.gov (United States)

    Beuron, Fabienne; Dreveny, Ingrid; Yuan, Xuemei; Pye, Valerie E; Mckeown, Ciaran; Briggs, Louise C; Cliff, Matthew J; Kaneko, Yayoi; Wallis, Russell; Isaacson, Rivka L; Ladbury, John E; Matthews, Steve J; Kondo, Hisao; Zhang, Xiaodong; Freemont, Paul S

    2006-01-01

    The AAA+ATPase p97/VCP, helped by adaptor proteins, exerts its essential role in cellular events such as endoplasmic reticulum-associated protein degradation or the reassembly of Golgi, ER and the nuclear envelope after mitosis. Here, we report the three-dimensional cryo-electron microscopy structures at ∼20 Å resolution in two nucleotide states of the endogenous hexameric p97 in complex with a recombinant p47 trimer, one of the major p97 adaptor proteins involved in membrane fusion. Depending on the nucleotide state, we observe the p47 trimer to be in two distinct arrangements on top of the p97 hexamer. By combining the EM data with NMR and other biophysical measurements, we propose a model of ATP-dependent p97(N) domain motions that lead to a rearrangement of p47 domains, which could result in the disassembly of target protein complexes. PMID:16601695

  2. P4 ATPases: Flippases in Health and Disease

    Directory of Open Access Journals (Sweden)

    Coen C. Paulusma

    2013-04-01

    Full Text Available P4 ATPases catalyze the translocation of phospholipids from the exoplasmic to the cytosolic leaflet of biological membranes, a process termed “lipid flipping”. Accumulating evidence obtained in lower eukaryotes points to an important role for P4 ATPases in vesicular protein trafficking. The human genome encodes fourteen P4 ATPases (fifteen in mouse of which the cellular and physiological functions are slowly emerging. Thus far, deficiencies of at least two P4 ATPases, ATP8B1 and ATP8A2, are the cause of severe human disease. However, various mouse models and in vitro studies are contributing to our understanding of the cellular and physiological functions of P4-ATPases. This review summarizes current knowledge on the basic function of these phospholipid translocating proteins, their proposed action in intracellular vesicle transport and their physiological role.

  3. Min mentor er min voksenven

    DEFF Research Database (Denmark)

    Kaiser, Birte

    2009-01-01

    En artikel i bogen: 6 belysninger af vejledning, brydninger, forståelser & praksis. Artiklen er et sammendrag af en diskursanalyse fra 2008 fra projetrapporten: Ind under huden, www.vejledning.net...

  4. Demethoxycurcumin is a potent inhibitor of P-type ATPases from diverse kingdoms of life

    DEFF Research Database (Denmark)

    Dao, Trong Tuan; Sehgal, Pankaj; Thanh Tung, Truong

    2016-01-01

    the curcuminoids, demethoxycurcumin was the most potent inhibitor of all tested P-type ATPases from fungal (Pma1p; H+-ATPase), plant (AHA2; H+-ATPase) and animal (SERCA; Ca2+-ATPase) cells. All three curcuminoids acted as non-competitive antagonist to ATP and hence may bind to a highly conserved allosteric site...

  5. Thermophilic P-loop transport ATPases : Enzyme function and energetics at high temperature

    NARCIS (Netherlands)

    Pretz, Monika Gyöngyi

    2007-01-01

    Primary transport ATPases are divided into several superfamilies; amongst others including ATPases of the ABC transporter superfamily, the F-ATPase superfamily or the motor ATPases of the General Secretory (Sec) pathway. Motor proteins from these superfamilies show a low sequence similarity, except

  6. The amino-terminal 200 amino acids of the plasma membrane Na+,K+-ATPase alpha subunit confer ouabain sensitivity on the sarcoplasmic reticulum Ca(2+)-ATPase.

    OpenAIRE

    Ishii, T; Takeyasu, K

    1993-01-01

    Cardiac glycosides such as G-strophanthin (ouabain) bind to and inhibit the plasma membrane Na+,K(+)-ATPase but not the sarcoplasmic reticulum (SR) Ca(2+)-ATPase, whereas thapsigargin specifically blocks the SR Ca(2+)-ATPase. The chimera [n/c]CC, in which the amino-terminal amino acids Met1 to Asp162 of the SR Ca(2+)-ATPase (SERCA1) were replaced with the corresponding portion of the Na+,K(+)-ATPase alpha 1 subunit (Met1 to Asp200), retained thapsigargin- and Ca(2+)-sensitive ATPase activity,...

  7. Forenklingens fire F'er

    DEFF Research Database (Denmark)

    Bentzen, Tina Øllgaard

    2017-01-01

    At fjerne styring er det, man ofte forbinder med afbureaukratisering, men det er ikke tilstrækkeligt, når man vil gå fra flotte ambitioner til en styring, som reelt opleves enklere. For at forenkle må man også forandre, forankre og fastholde styring, og det må ske i et samspil mellem de aktører, ...

  8. Regulation of vacuolar H+-ATPase in microglia by RANKL

    International Nuclear Information System (INIS)

    Serrano, Eric M.; Ricofort, Ryan D.; Zuo, Jian; Ochotny, Noelle; Manolson, Morris F.; Holliday, L. Shannon

    2009-01-01

    Vacuolar H + -ATPases (V-ATPases) are large electrogenic proton pumps composed of numerous subunits that play vital housekeeping roles in the acidification of compartments of the endocytic pathway. Additionally, V-ATPases play specialized roles in certain cell types, a capacity that is linked to cell type selective expression of isoforms of some of the subunits. We detected low levels of the a3 isoform of the a-subunit in mouse brain extracts. Examination of various brain-derived cell types by immunoblotting showed a3 was expressed in the N9 microglia cell line and in primary microglia, but not in other cell types. The expression of a3 in osteoclasts requires stimulation by Receptor Activator of Nuclear Factor κB-ligand (RANKL). We found that Receptor Activator of Nuclear Factor κB (RANK) was expressed by microglia. Stimulation of microglia with RANKL triggered increased expression of a3. V-ATPases in microglia were shown to bind microfilaments, and stimulation with RANKL increased the proportion of V-ATPase associated with the detergent-insoluble cytoskeletal fraction and with actin. In summary, microglia express the a3-subunit of V-ATPase. The expression of a3 and the interaction between V-ATPases and microfilaments was modulated by RANKL. These data suggest a novel molecular pathway for regulating microglia.

  9. Intracellular alkalinization induces cytosolic Ca2+ increases by inhibiting sarco/endoplasmic reticulum Ca2+-ATPase (SERCA.

    Directory of Open Access Journals (Sweden)

    Sen Li

    Full Text Available Intracellular pH (pHi and Ca(2+ regulate essentially all aspects of cellular activities. Their inter-relationship has not been mechanistically explored. In this study, we used bases and acetic acid to manipulate the pHi. We found that transient pHi rise induced by both organic and inorganic bases, but not acidification induced by acid, produced elevation of cytosolic Ca(2+. The sources of the Ca(2+ increase are from the endoplasmic reticulum (ER Ca(2+ pools as well as from Ca(2+ influx. The store-mobilization component of the Ca(2+ increase induced by the pHi rise was not sensitive to antagonists for either IP(3-receptors or ryanodine receptors, but was due to inhibition of the sarco/endoplasmic reticulum Ca(2+-ATPase (SERCA, leading to depletion of the ER Ca(2+ store. We further showed that the physiological consequence of depletion of the ER Ca(2+ store by pHi rise is the activation of store-operated channels (SOCs of Orai1 and Stim1, leading to increased Ca(2+ influx. Taken together, our results indicate that intracellular alkalinization inhibits SERCA activity, similar to thapsigargin, thereby resulting in Ca(2+ leak from ER pools followed by Ca(2+ influx via SOCs.

  10. Different thermostabilities of sarcoplasmic reticulum (Ca2+ + Mg2+)-ATPases from rabbit and trout muscles.

    Science.gov (United States)

    de Toledo, F G; Albuquerque, M C; Goulart, B H; Chini, E N

    1995-05-01

    Trout and rabbit (Ca2+ + Mg2+)-ATPases from sarcoplasmic reticulum were compared for differences in thermal inactivation and susceptibility to trypsin digestion. The trout ATPase is more heat-sensitive than the rabbit ATPase and is stabilized by Ca2+, Na+, K+ and nucleotides. Solubilization of both ATPases shows that the two ATPases have different protein-intrinsic inactivation kinetics. When digested by trypsin, the two ATPases display different cleavage patterns. The present results indicate that the trout and rabbit ATPases have dissimilarities in protein structure that may explain the differences in thermal inactivation kinetics.

  11. Photosynthesis Activates Plasma Membrane H+-ATPase via Sugar Accumulation.

    Science.gov (United States)

    Okumura, Masaki; Inoue, Shin-Ichiro; Kuwata, Keiko; Kinoshita, Toshinori

    2016-05-01

    Plant plasma membrane H(+)-ATPase acts as a primary transporter via proton pumping and regulates diverse physiological responses by controlling secondary solute transport, pH homeostasis, and membrane potential. Phosphorylation of the penultimate threonine and the subsequent binding of 14-3-3 proteins in the carboxyl terminus of the enzyme are required for H(+)-ATPase activation. We showed previously that photosynthesis induces phosphorylation of the penultimate threonine in the nonvascular bryophyte Marchantia polymorpha However, (1) whether this response is conserved in vascular plants and (2) the process by which photosynthesis regulates H(+)-ATPase phosphorylation at the plasma membrane remain unresolved issues. Here, we report that photosynthesis induced the phosphorylation and activation of H(+)-ATPase in Arabidopsis (Arabidopsis thaliana) leaves via sugar accumulation. Light reversibly phosphorylated leaf H(+)-ATPase, and this process was inhibited by pharmacological and genetic suppression of photosynthesis. Immunohistochemical and biochemical analyses indicated that light-induced phosphorylation of H(+)-ATPase occurred autonomously in mesophyll cells. We also show that the phosphorylation status of H(+)-ATPase and photosynthetic sugar accumulation in leaves were positively correlated and that sugar treatment promoted phosphorylation. Furthermore, light-induced phosphorylation of H(+)-ATPase was strongly suppressed in a double mutant defective in ADP-glucose pyrophosphorylase and triose phosphate/phosphate translocator (adg1-1 tpt-2); these mutations strongly inhibited endogenous sugar accumulation. Overall, we show that photosynthesis activated H(+)-ATPase via sugar production in the mesophyll cells of vascular plants. Our work provides new insight into signaling from chloroplasts to the plasma membrane ion transport mechanism. © 2016 American Society of Plant Biologists. All Rights Reserved.

  12. Radioprotector modifying influence upon the ion transport ATPase activities

    International Nuclear Information System (INIS)

    Dvoretsky, A.I.; Egorova, E.G.; Ananieva, T.V.; Kulikova, I.A.

    1993-01-01

    The effects of aminothiol and biogenic amine radioprotectors (β-mercaptoethylamine, AET, serotonin, dopamine, histamine) on the basic ion transport enzymes, such as Na, K-ATP ase and Mg, Ca-ATPase activities were investigated in the tissues of numerous organs, with different radiosensitivity in the wistar rats. Experimental results showed that intraperitoneal injection of the used radioprotectors caused preliminary inhibition of the Na, K-ATPase activity in tissues from organs with different radioresistance, but had no influence on the Mg, Ca-ATPase activity in membranes of erythrocytes and rat brain cells. (2 tabs.)

  13. On Allosteric Modulation of P-Type Cu+-ATPases

    DEFF Research Database (Denmark)

    Mattle, Daniel; Sitsel, Oleg; Autzen, Henriette Elisabeth

    2013-01-01

    P-type ATPases perform active transport of various compounds across biological membranes and are crucial for ion homeostasis and the asymmetric composition of lipid bilayers. Although their functional cycle share principles of phosphoenzyme intermediates, P-type ATPases also show subclass...... of intramembranous Cu+ binding, and we suggest an alternative role for the proposed second site in copper translocation and proton exchange. The class-specific features demonstrate that topological diversity in P-type ATPases may tune a general energy coupling scheme to the translocation of compounds with remarkably...

  14. Insulin regulation of (Na+, K+)-ATPase

    International Nuclear Information System (INIS)

    Lytton, J.

    1985-01-01

    This thesis describes an investigation into the mechanism of insulin stimulation of (Na + ,K + )=ATPase in rat adipocytes. Two molecular forms of the catalytic subunit of the enzyme were identified and denoted α and α(+), due to their similarity to those isozymes previously described from rat brain. Insulin specifically stimulated the α(+) form of the enzyme. The two forms of the enzyme had quite different affinities for intracellular sodium ion; insulin affected only the lower affinity of α(+), shifting it toward a higher value. However, the sodium affinity of (Na + ,K + )-ATPase activity in isolated membranes was equally high for both forms of the enzyme. This suggests that the difference in sodium affinity between the two forms observed in the cell is not inherent within the structure of the sodium pump, but must depend upon a selective interaction with another molecule which has been lost upon membrane isolation. Immunoprecipitation of both the catalytic subunits either from extracts of whole cells which had been labelled with [ 32 P] orthophosphate, or from membranes which had been labelled with γ-[ 32 P]ATP demonstrated that less than 1 in 100 molecules had a covalently bound phosphate insulin had no influence on this value. The amino terminal sequences of the first 4 amino acids of the catalytic subunits of both α (isolated from rat kidney) and α(+) (from rat brainstem axolemma) were determined. The result shows two highly homologous but clearly different molecules. It can thus be concluded that the insulin sensitive version of the enzyme is not derived from the common α form by a post-translational modification

  15. Plasma membrane calcium ATPases and related disorders.

    Science.gov (United States)

    Giacomello, Marta; De Mario, Agnese; Scarlatti, Chiara; Primerano, Simona; Carafoli, Ernesto

    2013-03-01

    The plasma membrane Ca(2+) ATPases (PMCA pumps) cooperate with other transport systems in the plasma membrane and in the organelles in the regulation of cell Ca(2+). They have high Ca(2+) affinity and are thus the fine tuners of cytosolic Ca(2+). They belong to the superfamily of P-type ATPases: their four basic isoforms share the essential properties of the reaction cycle and the general membrane topography motif of 10 transmembrane domains and three large cytosolic units. However they also differ in other important properties, e.g., tissue distribution and regulatory mechanisms. Their chief regulator is calmodulin, that removes their C-terminal cytosolic tail from autoinhibitory binding sites next to the active site of the pump, restoring activity. The number of pump isoforms is increased to over 30 by alternative splicing of the transcripts at a N-terminal site (site A) and at site C within the C-terminal calmodulin binding domain: the splice variants are tissue specific and developmentally regulated. The importance of PMCAs in the maintenance of cellular Ca(2+) homeostasis is underlined by the disease phenotypes, genetic or acquired, caused by their malfunction. Non-genetic PMCA deficiencies have long been considered possible causative factors in disease conditions as important as cancer, hypertension, or neurodegeneration. Those of genetic origin are better characterized: some have now been discovered in humans as well. They concern all four PMCA isoforms, and range from cardiac dysfunctions, to deafness, to hypertension, to cerebellar ataxia. Copyright © 2012 Elsevier Ltd. All rights reserved.

  16. Advances in targeting the vacuolar proton-translocating ATPase (V-ATPase for anti-fungal therapy

    Directory of Open Access Journals (Sweden)

    Summer R. Hayek

    2014-01-01

    Full Text Available Vacuolar proton-translocating ATPase (V-ATPase is a membrane-bound, multi-subunit enzyme that uses the energy of ATP hydrolysis to pump protons across membranes. V-ATPase activity is critical for pH homeostasis and organelle acidification as well as for generation of the membrane potential that drives secondary transporters and cellular metabolism. V-ATPase is highly conserved across species and is best characterized in the model fungus Saccharomyces cerevisiae (S. cerevisiae. However, recent studies in mammals have identified significant alterations from fungi, particularly in the isoform composition of the 14 subunits and in the regulation of complex disassembly. These differences could be exploited for selectivity between fungi and humans and highlight the potential for V-ATPase as an anti-fungal drug target. Candida albicans (C. albicans is a major human fungal pathogen and causes fatality in 35% of systemic infections, even with anti-fungal treatment. The pathogenicity of C. albicans correlates with environmental, vacuolar, and cytoplasmic pH regulation, and V-ATPase appears to play a fundamental role in each of these processes. Genetic loss of V-ATPase in pathogenic fungi leads to defective virulence, and a comprehensive picture of the mechanisms involved is emerging. Recent studies have explored the practical utility of V-ATPase as an anti-fungal drug target in C. albicans, including pharmacological inhibition, azole therapy, and targeting of downstream pathways. This overview will discuss these studies as well as hypothetical ways to target V-ATPase and novel high-throughput methods for use in future drug discovery screens.

  17. OSMOTIC FRAGILITY AND Na + -K + + ATPase ACTIVITY OF ...

    African Journals Online (AJOL)

    + -K+ ATPase activity of the erythrocytes of HIV/AIDS patients. Whole blood was taken from subjects at the Human Virology Laboratory of the Nigerian Institute of Medical Research. Subjects were judged suitable for the various investigations by ...

  18. Stochastic Four-State Mechanochemical Model of F1-ATPase

    International Nuclear Information System (INIS)

    Wu Weixia; Zhan Yong; Zhao Tongjun; Han Yingrong; Chen Yafei

    2010-01-01

    F 1 -ATPase, a part of ATP synthase, can synthesize and hydrolyze ATP moleculars in which the central γ-subunit rotates inside the α 3 β 3 cylinder. A stochastic four-state mechanochemical coupling model of F 1 -ATPase is studied with the aid of the master equation. In this model, the ATP hydrolysis and synthesis are dependent on ATP, ADP, and Pi concentrations. The effects of ATP concentration, ADP concentration, and the external torque on the occupation probability of binding-state, the rotation rate and the diffusion coefficient of F 1 -ATPase are investigated. Moreover, the results from this model are compared with experiments. The mechanochemical mechanism F 1 -ATPase is qualitatively explained by the model. (general)

  19. V-ATPase-mediated granular acidification is regulated by the V-ATPase accessory subunit Ac45 in POMC-producing cells.

    NARCIS (Netherlands)

    Jansen, E.J.S.; Hafmans, T.G.M.; Martens, G.J.

    2010-01-01

    The vacuolar (H(+))-ATPase (V-ATPase) is an important proton pump, and multiple critical cell-biological processes depend on the proton gradient provided by the pump. Yet, the mechanism underlying the control of the V-ATPase is still elusive but has been hypothesized to involve an accessory subunit

  20. The non-gastric H,K-ATPase as a tool to study the ouabain-binding site in Na,K-ATPase.

    NARCIS (Netherlands)

    Pont, J.J.H.H.M. de; Swarts, H.G.P.; Karawajczyk, A.; Schaftenaar, G.; Willems, P.H.G.M.; Koenderink, J.B.

    2009-01-01

    Based on studies with chimeras between (non-)gastric H,K-ATPase and Na,K-ATPase, a model for the ouabain binding site has recently been presented (Qiu et al. J.Biol.Chem. 280 (2005) 32349). In this model, hydrogen bonds between specific amino acid residues of Na,K-ATPase and hydroxyl groups of

  1. Isolation and characterization of DNA-dependent ATPases from the Novikoff Hepatoma

    International Nuclear Information System (INIS)

    Thomas, D.C.

    1984-01-01

    Four DNA-dependent ATPases have been purified to apparent homogeneity from extracts of the Novikoff Hepatoma, and named ATPases II, III, IV, and V. The physical and enzymological properties of ATPases II, III, and V are nearly identical, and from tryptic peptide mapping these proteins were determined to be related, though they are still chromatographically distinct; all appear to be dimers. ATPaseIV is unique among the ATPases, and is probably a monomer. ATPase V appears much more stable to thermal inactivation than the similar curves generated by ATPases II, and III. ATPase IV, however, projects of a heat-inactivation curve intermediate to these two types. ATPase II is labelled to a much higher degree than the others when treated with a heterologous protein kinase using gamma-[ 32 P]-ATP. When ATPase II was treated with this kinase, and subsequently run over a DNA-cellulose column, the profile of ATPase II was found to contain small peaks of activity in the positions where ATPases III and V normally elute, suggesting that ATPase II may be a dephosphorylated form of the other two. The ATPases have been extensively characterized with respect to reaction products and requirements, substrate utilization, DNA effector requirements, and effects of ATP analogs

  2. Hvis en kartoffel er forkert, hvad er en Mars-bar så?

    DEFF Research Database (Denmark)

    Lichtenstein, Mia Beck; Thomsen, Freja; Hinze, Cecilie

    2016-01-01

    Danske unge er de slankeste af 41 europæiske unge, men de føler sig ofte tykke og går på slankekur. Hvad er årsagen, og hvorfor er det et problem?......Danske unge er de slankeste af 41 europæiske unge, men de føler sig ofte tykke og går på slankekur. Hvad er årsagen, og hvorfor er det et problem?...

  3. Characterization of the plasma membrane H+-ATPase in the liverwort Marchantia polymorpha.

    Science.gov (United States)

    Okumura, Masaki; Inoue, Shin-ichiro; Takahashi, Koji; Ishizaki, Kimitsune; Kohchi, Takayuki; Kinoshita, Toshinori

    2012-06-01

    The plasma membrane H(+)-ATPase generates an electrochemical gradient of H(+) across the plasma membrane that provides the driving force for solute transport and regulates pH homeostasis and membrane potential in plant cells. Recent studies have demonstrated that phosphorylation of the penultimate threonine in H(+)-ATPase and subsequent binding of a 14-3-3 protein is the major common activation mechanism for H(+)-ATPase in vascular plants. However, there is very little information on the plasma membrane H(+)-ATPase in nonvascular plant bryophytes. Here, we show that the liverwort Marchantia polymorpha, which is the most basal lineage of extant land plants, expresses both the penultimate threonine-containing H(+)-ATPase (pT H(+)-ATPase) and non-penultimate threonine-containing H(+)-ATPase (non-pT H(+)-ATPase) as in the green algae and that pT H(+)-ATPase is regulated by phosphorylation of its penultimate threonine. A search in the expressed sequence tag database of M. polymorpha revealed eight H(+)-ATPase genes, designated MpHA (for M. polymorpha H(+)-ATPase). Four isoforms are the pT H(+)-ATPase; the remaining isoforms are non-pT H(+)-ATPase. An apparent 95-kD protein was recognized by anti-H(+)-ATPase antibodies against an Arabidopsis (Arabidopsis thaliana) isoform and was phosphorylated on the penultimate threonine in response to the fungal toxin fusicoccin in thalli, indicating that the 95-kD protein contains pT H(+)-ATPase. Furthermore, we found that the pT H(+)-ATPase in thalli is phosphorylated in response to light, sucrose, and osmotic shock and that light-induced phosphorylation depends on photosynthesis. Our results define physiological signals for the regulation of pT H(+)-ATPase in the liverwort M. polymorpha, which is one of the earliest plants to acquire pT H(+)-ATPase.

  4. Den sproglige leg er super fly

    DEFF Research Database (Denmark)

    Just, Sine Nørholm

    2013-01-01

    Man kan rappe om alt. I hvert fald hvis man er Marvelous Mosell. I spændingsfeltet mellem fiktion og virkelighed skaber Mosell sin persona i et forjættende 80' er-univers der på en og samme tid er vildt overdrevet og helt autentisk.......Man kan rappe om alt. I hvert fald hvis man er Marvelous Mosell. I spændingsfeltet mellem fiktion og virkelighed skaber Mosell sin persona i et forjættende 80' er-univers der på en og samme tid er vildt overdrevet og helt autentisk....

  5. N-ov-emb-er

    Indian Academy of Sciences (India)

    " to. Andrew Z Fire - Stanford University School of Medicine Stanford, CA, USA. Craig C Mello - University of Massachusetts Medical School Worcester, MA, USA. -96----------------------------~~--------R-ES-O-N-A-N-C-E--1 -N-ov-e-m-b-er--2-oo-6.

  6. Hvad er ledelse af brugerinddragelse?

    DEFF Research Database (Denmark)

    Holm-Petersen, Christina; Navne, Laura Emdal

    2015-01-01

    Brugerinddragelse i det danske sundhedsvæsen står højt på den politiske dagsorden, men det er stadig en udfordring at implementere visionen i klinisk praksis. Ledelse af brugerinddragelse bliver aktuelt udpeget som en central nøgle til at føre visionen ud i livet. Samtidig er der kun relativt lidt......, at brugerinddragelse skal implementeres i en verden, hvor der allerede er en række andre mål tilstede. En central ledelsesudfordring er derfor, at nogle af målene med brugerinddragelse forudsætter nye måder at organisere ikke bare arbejdet og kompetencer på, men også relationer til patienter og pårørende. En væsentlig...... that organize relations Patient involvement in the health services in Denmark is high on the political agenda, though continues to be a challenge to implement. It is increasingly said that leadership is crucial to the implementation process. However, research into the role of leaders in patient involvement...

  7. Meiotic Clade AAA ATPases: Protein Polymer Disassembly Machines.

    Science.gov (United States)

    Monroe, Nicole; Hill, Christopher P

    2016-05-08

    Meiotic clade AAA ATPases (ATPases associated with diverse cellular activities), which were initially grouped on the basis of phylogenetic classification of their AAA ATPase cassette, include four relatively well characterized family members, Vps4, spastin, katanin and fidgetin. These enzymes all function to disassemble specific polymeric protein structures, with Vps4 disassembling the ESCRT-III polymers that are central to the many membrane-remodeling activities of the ESCRT (endosomal sorting complexes required for transport) pathway and spastin, katanin p60 and fidgetin affecting multiple aspects of cellular dynamics by severing microtubules. They share a common domain architecture that features an N-terminal MIT (microtubule interacting and trafficking) domain followed by a single AAA ATPase cassette. Meiotic clade AAA ATPases function as hexamers that can cycle between the active assembly and inactive monomers/dimers in a regulated process, and they appear to disassemble their polymeric substrates by translocating subunits through the central pore of their hexameric ring. Recent studies with Vps4 have shown that nucleotide-induced asymmetry is a requirement for substrate binding to the pore loops and that recruitment to the protein lattice via MIT domains also relieves autoinhibition and primes the AAA ATPase cassettes for substrate binding. The most striking, unifying feature of meiotic clade AAA ATPases may be their MIT domain, which is a module that is found in a wide variety of proteins that localize to ESCRT-III polymers. Spastin also displays an adjacent microtubule binding sequence, and the presence of both ESCRT-III and microtubule binding elements may underlie the recent findings that the ESCRT-III disassembly function of Vps4 and the microtubule-severing function of spastin, as well as potentially katanin and fidgetin, are highly coordinated. Copyright © 2015 Elsevier Ltd. All rights reserved.

  8. ER2OWL: Generating OWL Ontology from ER Diagram

    Science.gov (United States)

    Fahad, Muhammad

    Ontology is the fundamental part of Semantic Web. The goal of W3C is to bring the web into (its full potential) a semantic web with reusing previous systems and artifacts. Most legacy systems have been documented in structural analysis and structured design (SASD), especially in simple or Extended ER Diagram (ERD). Such systems need up-gradation to become the part of semantic web. In this paper, we present ERD to OWL-DL ontology transformation rules at concrete level. These rules facilitate an easy and understandable transformation from ERD to OWL. The set of rules for transformation is tested on a structured analysis and design example. The framework provides OWL ontology for semantic web fundamental. This framework helps software engineers in upgrading the structured analysis and design artifact ERD, to components of semantic web. Moreover our transformation tool, ER2OWL, reduces the cost and time for building OWL ontologies with the reuse of existing entity relationship models.

  9. Endoplasmic Reticulum (ER Stress and Endocrine Disorders

    Directory of Open Access Journals (Sweden)

    Daisuke Ariyasu

    2017-02-01

    Full Text Available The endoplasmic reticulum (ER is the organelle where secretory and membrane proteins are synthesized and folded. Unfolded proteins that are retained within the ER can cause ER stress. Eukaryotic cells have a defense system called the “unfolded protein response” (UPR, which protects cells from ER stress. Cells undergo apoptosis when ER stress exceeds the capacity of the UPR, which has been revealed to cause human diseases. Although neurodegenerative diseases are well-known ER stress-related diseases, it has been discovered that endocrine diseases are also related to ER stress. In this review, we focus on ER stress-related human endocrine disorders. In addition to diabetes mellitus, which is well characterized, several relatively rare genetic disorders such as familial neurohypophyseal diabetes insipidus (FNDI, Wolfram syndrome, and isolated growth hormone deficiency type II (IGHD2 are discussed in this article.

  10. Endoplasmic Reticulum (ER) Stress and Endocrine Disorders

    Science.gov (United States)

    Ariyasu, Daisuke; Yoshida, Hiderou; Hasegawa, Yukihiro

    2017-01-01

    The endoplasmic reticulum (ER) is the organelle where secretory and membrane proteins are synthesized and folded. Unfolded proteins that are retained within the ER can cause ER stress. Eukaryotic cells have a defense system called the “unfolded protein response” (UPR), which protects cells from ER stress. Cells undergo apoptosis when ER stress exceeds the capacity of the UPR, which has been revealed to cause human diseases. Although neurodegenerative diseases are well-known ER stress-related diseases, it has been discovered that endocrine diseases are also related to ER stress. In this review, we focus on ER stress-related human endocrine disorders. In addition to diabetes mellitus, which is well characterized, several relatively rare genetic disorders such as familial neurohypophyseal diabetes insipidus (FNDI), Wolfram syndrome, and isolated growth hormone deficiency type II (IGHD2) are discussed in this article. PMID:28208663

  11. Influence of hexavanadates on Na+/K+- ATPase activity

    Directory of Open Access Journals (Sweden)

    Zdravković Aleksandra

    2016-01-01

    Full Text Available Introduction: There is a great interest in use of polioximetalates in clinical medicine, primary as antibacterial, antiviral and antitumoral agents. Considering the key role of Na+/ K+- ATPase in normal functioning of most animal cells, as well as pivotal roles in cancer cell migration, the aim of this paper was to examine the influence of new synthesized hexavandates [V6-CH3][Na]2, [V6-NO2][TBA]2, [V6-C3][H]2, [V6-C5d][TBA]2 on Na+/K+- ATPase activity. Material and methods: The enzymatic activity of porcine cerebral cortex Na+/K+- ATPase was followed in both the absence and presence of increasing concentration of [V6-CH3] [Na]2, [V6-NO2][TBA]2, [V6-C3][H]2, [V6-C5d][TBA]2 (within the range 10-8 - 10-3 mol/L. The released Pi, liberated from the enzymatic hydrolysis of ATP, was determined by spectrophotometric method, using Perkin Elmer Lambda 35 UV-VIS spectrophotometer. Results: Investigated compounds inhibit the activity of Na+/K+ ATPase in dose-dependent manner within the investigated range. Obtained results indicate that all investigated compounds inhibit the Na+/K+ ATPase activity, but with different inhibiting power. [V6-NO2] [TBA]2 (IC50 = 1,87 × 10-5 mol/L was the most potent inhibitor of Na+/K+ ATPase, while [V6-C5d][TBA]2 showed the least potent inhibiting power (IC50 = 1,31 × 10-4 mol/L . The results are consistent with previously published concentration-dependent inhibitory effect of polyoxometalates (including polioxovandates on ATPase activity from different model syistems. Conclusion: Based on the results, we can conclude that the examined compounds inhibit Na+/K+- ATPase activity in a dose-dependent manner. Inhibiting power of tested hexavanadates are different, and weaker than inhibiting power of decavanadates (tested earlier on Na+/K+- ATPase activity, which is probably due to differences in charge, size and shape of these polioxometalates. Considering the role of this enzymes in the functioning of healthy cells and the

  12. The role of the N-domain in the ATPase activity of the mammalian AAA ATPase p97/VCP.

    Science.gov (United States)

    Niwa, Hajime; Ewens, Caroline A; Tsang, Chun; Yeung, Heidi O; Zhang, Xiaodong; Freemont, Paul S

    2012-03-09

    p97/valosin-containing protein (VCP) is a type II ATPase associated with various cellular activities that forms a homohexamer with each protomer containing an N-terminal domain (N-domain); two ATPase domains, D1 and D2; and a disordered C-terminal region. Little is known about the role of the N-domain or the C-terminal region in the p97 ATPase cycle. In the p97-associated human disease inclusion body myopathy associated with Paget disease of bone and frontotemporal dementia, the majority of missense mutations are located at the N-domain D1 interface. Structure-based predictions suggest that such mutations affect the interaction of the N-domain with D1. Here we have tested ten major inclusion body myopathy associated with Paget disease of bone and frontotemporal dementia-linked mutants for ATPase activity and found that all have increased activity over the wild type, with one mutant, p97(A232E), having three times higher activity. Further mutagenesis of p97(A232E) shows that the increase in ATPase activity is mediated through D2 and requires both the N-domain and a flexible ND1 linker. A disulfide mutation that locks the N-domain to D1 in a coplanar position reversibly abrogates ATPase activity. A cryo-EM reconstruction of p97(A232E) suggests that the N-domains are flexible. Removal of the C-terminal region also reduces ATPase activity. Taken together, our data suggest that the conformation of the N-domain in relation to the D1-D2 hexamer is directly linked to ATP hydrolysis and that the C-terminal region is required for hexamer stability. This leads us to propose a model where the N-domain adopts either of two conformations: a flexible conformation compatible with ATP hydrolysis or a coplanar conformation that is inactive.

  13. The Role of the N-Domain in the ATPase Activity of the Mammalian AAA ATPase p97/VCP*

    Science.gov (United States)

    Niwa, Hajime; Ewens, Caroline A.; Tsang, Chun; Yeung, Heidi O.; Zhang, Xiaodong; Freemont, Paul S.

    2012-01-01

    p97/valosin-containing protein (VCP) is a type II ATPase associated with various cellular activities that forms a homohexamer with each protomer containing an N-terminal domain (N-domain); two ATPase domains, D1 and D2; and a disordered C-terminal region. Little is known about the role of the N-domain or the C-terminal region in the p97 ATPase cycle. In the p97-associated human disease inclusion body myopathy associated with Paget disease of bone and frontotemporal dementia, the majority of missense mutations are located at the N-domain D1 interface. Structure-based predictions suggest that such mutations affect the interaction of the N-domain with D1. Here we have tested ten major inclusion body myopathy associated with Paget disease of bone and frontotemporal dementia-linked mutants for ATPase activity and found that all have increased activity over the wild type, with one mutant, p97A232E, having three times higher activity. Further mutagenesis of p97A232E shows that the increase in ATPase activity is mediated through D2 and requires both the N-domain and a flexible ND1 linker. A disulfide mutation that locks the N-domain to D1 in a coplanar position reversibly abrogates ATPase activity. A cryo-EM reconstruction of p97A232E suggests that the N-domains are flexible. Removal of the C-terminal region also reduces ATPase activity. Taken together, our data suggest that the conformation of the N-domain in relation to the D1-D2 hexamer is directly linked to ATP hydrolysis and that the C-terminal region is required for hexamer stability. This leads us to propose a model where the N-domain adopts either of two conformations: a flexible conformation compatible with ATP hydrolysis or a coplanar conformation that is inactive. PMID:22270372

  14. Towards the structure of yeast and mammalian P4-ATPases

    DEFF Research Database (Denmark)

    Lyons, Joseph; Laban, Milena; Mikkelsen, Stine

    2017-01-01

    P4-ATPases are members of the P-type ATPase superfamily that drive the inward translocation (flipping) of lipids within the membrane. These lipid flippase largely function as binary complexes with an auxiliary protein from the CDC50 family. The bulk of our knowledge has derived genetic and bioche......P4-ATPases are members of the P-type ATPase superfamily that drive the inward translocation (flipping) of lipids within the membrane. These lipid flippase largely function as binary complexes with an auxiliary protein from the CDC50 family. The bulk of our knowledge has derived genetic...... a basis for the analysis of reported mutagenesis data, we aim to solve the first molecular structures of the PS transporting P4-ATPases using electron microscopy. To date, negative stain EM analysis, on detergent, amphipol and saposin-lipoprotein nanoparticle (Salipro) reconstituted of both Drs2p/CDC50p...... and bATP8A2/CDC50A, has yielded comparable low-resolution envelopes of these two transporters, highlighting the bulk architecture of the complex. Current efforts and progress on the functional characterization and cryo-EM analysis of both lipid transporters reconstituted in Salipro are described...

  15. Characterization of the vacuolar H sup + -ATPase of higher plants

    Energy Technology Data Exchange (ETDEWEB)

    Manolson, M F

    1988-01-01

    The tonoplast H{sup +}-ATPase of Beta vulgaris L. was partially purified by Triton X-100 solubilization and Sepharose 4B chromatography resulting in the enrichment of two polypeptides. Kinetic analysis of ({alpha}-{sup 32}P) BzATP labeling identified the 57 kDa polypeptide as a nucleotide-binding subunit with a possible regulatory function. In addition, ({sup 14}C) DCCD-labeling identified a 16 kDa polypeptide as a putative transmembrane proton channel. It is concluded that the tonoplast H{sup +}-ATPase is a multimer composed of at least three polypeptides. Anti-57 and anti-67 kDa sera reacted with polypeptides of the corresponding size in bovine chromaffin granules, bovine clathrin-coated vesicles, and yeast vacuolar membranes, suggesting common structural features and common ancestry for endomembrane H{sup +}-ATPase of different organelles and different phyla. Anti-57 serum was used to isolate a cDNA encoding the corresponding subunit from Arabidopsis. Protein sequence analysis revealed homologies between endomembrane, F{sub 0}F{sub 1} and archaebacterial ATPases, suggesting that these different classes of ATPases have evolved from a common ancestor.

  16. A method to measure hydrolytic activity of adenosinetriphosphatases (ATPases.

    Directory of Open Access Journals (Sweden)

    Gianluca Bartolommei

    Full Text Available The detection of small amounts (nanomoles of inorganic phosphate has a great interest in biochemistry. In particular, phosphate detection is useful to evaluate the rate of hydrolysis of phosphatases, that are enzymes able to remove phosphate from their substrate by hydrolytic cleavage. The hydrolysis rate is correlated to enzyme activity, an extremely important functional parameter. Among phosphatases there are the cation transporting adenosinetriphosphatases (ATPases, that produce inorganic phosphate by cleavage of the γ-phosphate of ATP. These membrane transporters have many fundamental physiological roles and are emerging as potential drug targets. ATPase hydrolytic activity is measured to test enzyme functionality, but it also provides useful information on possible inhibitory effects of molecules that interfere with the hydrolytic process. We have optimized a molybdenum-based protocol that makes use of potassium antimony (III oxide tartrate (originally employed for phosphate detection in environmental analysis to allow its use with phosphatase enzymes. In particular, the method was successfully applied to native and recombinant ATPases to demonstrate its reliability, validity, sensitivity and versatility. Our method introduces significant improvements to well-established experimental assays, which are currently employed for ATPase activity measurements. Therefore, it may be valuable in biochemical and biomedical investigations of ATPase enzymes, in combination with more specific tests, as well as in high throughput drug screening.

  17. Hormonal regulation of Na -K -ATPase in cultured epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Johnson, J.P.; Jones, D.; Wiesmann, W.P.

    1986-08-01

    Aldosterone and insulin stimulate Na transport through mechanisms involving protein synthesis. Na -K -ATPase has been implicated in the action of both hormones. The authors examined the effect of aldosterone and insulin on Na -K -ATPase in epithelial cells in culture derived from toad urinary bladder (TB6C) and toad kidney (A6). Aldosterone, but not insulin, increases short-circuit current (I/sub sc/) in TB6C cells. Aldosterone increases Na -K -(TSP)ATPase activity after 18 h of incubation, but no effect can be seen at 3 and 6 h. Amiloride, which inhibits aldosterone-induced increases in I/sub sc/, has no effect on either basal or aldosterone stimulated enzyme activity. Both aldosterone and insulin increase I/sub sc/ in A6 cells and when added together are synergistic. Aldosterone stimulates enzyme activity in A6 cells, but insulin alone has no effect. However, aldosterone and insulin together stimulate enzyme activity more than aldosterone alone. It appears that stimulation of Na -K -ATPase activity is involved in aldosterone action in both cell lines but does not appear to be due to increased Na entry, since enhanced enzyme activity is not inhibited by amiloride. In contrast, insulin alone has no direct effect on Na -K -ATPase, although the increased enzyme activity following both agents in combination may explain their synergism on I/sub sc/.

  18. Hvornår er man ung?

    DEFF Research Database (Denmark)

    Gundelach, Peter; Nørregård-Nielsen, Esther C.

    2002-01-01

    Hvornår er man ung, og hvornår er man voksen? Er der forskelle i befolkningens værdier i forhold til arbejde og politik, når det undersøges ud fra henholdsvis et alders- eller generationsperspektiv? Baseret på data fra den danske del af den internationale værdiundersøgelse vises at der er så store...

  19. Hvorfor er sygeplejersker usynlige i offentlige medier?

    DEFF Research Database (Denmark)

    Joensen, Annemi Lund; Hall, Elisabeth

    2015-01-01

    Når der er sygeplejerelevante emner til debat i de offentlige medier på Færøerne, bærer debatten præg af sygeplejerskers manglende deltagelse. Sygeplejerskerne er usynlige. Et eksempel på dette er en debat om besparelser inden for ældreomsorgen. Til trods for at besparelsen fik omfattende konsekv...

  20. Jordkvalitetskriterier for PAH'er

    DEFF Research Database (Denmark)

    Carlsen, Lars

    2009-01-01

    For to år siden forhøjede Miljøstyrelsen den administrativt fastsatte grænseværdi for polycycliske hydrocarboner (PAH’er) i jord med en faktor 3, hvilket i praksis betød at en række grunde, der tidligere var klassifi ceret som forurenede, nu ikke klassifi ceres sådan. Den hidtidige og den nye met...

  1. Structural and functional studies of heavy metal ATPases

    DEFF Research Database (Denmark)

    Sitsel, Oleg

    2015-01-01

    to handle heavy metal ions. LpCopA is then compared to its two human homologues ATP7A and ATP7B, which cause the severe Menkes and Wilson diseases when malfunctioning. The differences between the three proteins are described and disease-causing mutations in the human proteins are analyzed. The crystal......Copper and zinc are trace elements that are crucial for the well-being of all cells and are an indispensable part of many proteins. At the same time, the intracellular levels of these metals require careful regulation, as an excess or deficiency may be lethal. P1B-ATPases are key players in Cu......+ and Zn2+ homeostasis that belong to the superfamily of P-type ATPases, transmembrane proteins which are present in virtually all lifeforms, with functions ranging from membrane potential generation to muscle relaxation. The goal of this thesis is to improve our understanding of P1B-ATPases by focusing...

  2. Protein import into chloroplasts requires a chloroplast ATPase

    International Nuclear Information System (INIS)

    Pain, D.; Blobel, G.

    1987-01-01

    The authors have transcribed mRNA from a cDNA clone coding for pea ribulose-1,5-bisphosphate carboxylase, translated the mRNA in a wheat germ cell-free system, and studied the energy requirement for posttranslational import of the [ 35 S]methionine-labeled protein into the stroma of pea chloroplasts. They found that import depends on ATP hydrolysis within the stroma. Import is not inhibited when H + , K + , Na + , or divalent cation gradients across the chloroplast membranes are dissipated by ionophores, as long as exogenously added ATP is also present during the import reaction. The data suggest that protein import into the chloroplast stroma requires a chloroplast ATPase that does not function to generate a membrane potential for driving the import reaction but that exerts its effect in another, yet-to-be-determined, mode. They have carried out a preliminary characterization of this ATPase regarding its nucleotide specificity and the effects of various ATPase inhibitors

  3. Review: The HSP90 molecular chaperone-an enigmatic ATPase.

    Science.gov (United States)

    Pearl, Laurence H

    2016-08-01

    The HSP90 molecular chaperone is involved in the activation and cellular stabilization of a range of 'client' proteins, of which oncogenic protein kinases and nuclear steroid hormone receptors are of particular biomedical significance. Work over the last two decades has revealed a conformational cycle critical to the biological function of HSP90, coupled to an inherent ATPase activity that is regulated and manipulated by many of the co-chaperones proteins with which it collaborates. Pharmacological inhibition of HSP90 ATPase activity results in degradation of client proteins in vivo, and is a promising target for development of new cancer therapeutics. Despite this, the actual function that HSP90s conformationally-coupled ATPase activity provides in its biological role as a molecular chaperone remains obscure. © 2016 Wiley Periodicals, Inc. Biopolymers 105: 594-607, 2016. © 2016 The Authors. Biopolymers Published by Wiley Periodicals, Inc.

  4. Autoinhibitory Regulation of Plasma Membrane H+-ATPases

    DEFF Research Database (Denmark)

    Pedersen, Jesper Torbøl

    Electrochemical gradients across cell membranes are essential for nutrient uptake. In plant and fungal cells the electrochemical gradient across the plasma membrane (PM) can build much higher than in mammalian cells. The protein responsible for this gradient is the essential PM H+-ATPase that uses...... resolution 3D structure the mechanism behind is only poorly understood. This thesis aimed at illuminating the autoinhibitory mechanism in plant and yeast PM H+-ATPases and below some of our main findings will be highlighted. The two terminal domains of the PM H+-ATPases have several amino acid residues...... that can be phosphorylated, and it has been demonstrated that these phosphorylation sites in both plant and yeast are highly involved in the regulation of terminal autoinhibition. In this study we used a phylogenetic analysis to investigate the evolutionary development of these phosphorylation sites...

  5. Na+,K+-ATPase Na+ affinity in rat skeletal muscle fiber types

    DEFF Research Database (Denmark)

    Kristensen, Michael; Juel, Carsten

    2010-01-01

    Previous studies in expression systems have found different ion activation of the Na(+)/K(+)-ATPase isozymes, which suggest that different muscles have different ion affinities. The rate of ATP hydrolysis was used to quantify Na(+),K(+)-ATPase activity, and the Na(+) affinity of Na(+),K(+)-ATPase...

  6. Structural and functional studies of a Cu+-ATPase from Legionella pneumophila

    DEFF Research Database (Denmark)

    Mattle, Daniel

    During his studies, Daniel Mattle explored the copper(I) export mechanism of a P-type Cu+ ATPase from Legionella pneumophila – a homologue to the human Cu+ ATPases. Cu+ ATPases are responsible for the homeostatic control of the physiological relevant – but toxic – copper(I) cations. To assess...

  7. Regulatory Mechanisms in the P4-ATPase Complex

    DEFF Research Database (Denmark)

    Costa, Sara

    . The functionality on the P4-ATPase complex is essential for several cellular processes, such as vesicle-mediated transport. However, the specific role of flippase activity in vesicle biogenesis and the regulatory mechanism behind this process is still poorly understood. In these studies, we identified...... affordable alternative using a microscope-based cytometer. This system can simultaneously provide information on flippase activity and expression levels. Taken together, the findings described in this thesis provide new tools for P4-ATPase characterization and valuable insights into the regulation...

  8. F F1-ATPase as biosensor to detect single virus

    International Nuclear Information System (INIS)

    Liu, XiaoLong; Zhang, Yun; Yue, JiaChang; Jiang, PeiDong; Zhang, ZhenXi

    2006-01-01

    F F 1 -ATPase within chromatophore was constructed as a biosensor (immuno-rotary biosensor) for the purpose of capturing single virus. Capture of virus was based on antibody-antigen reaction. The detection of virus based on proton flux change driven by ATP-synthesis of F F 1 -ATPase, which was indicated by F1300, was directly observed by a fluorescence microscope. The results demonstrate that the biosensor loading of virus particles has remarkable signal-to-noise ratio (3.8:1) compared to its control at single molecular level, and will be convenient, quick, and even super-sensitive for detecting virus particles

  9. A structural overview of the plasma membrane Na+,K+-ATPase and H+-ATPase ion pumps

    DEFF Research Database (Denmark)

    Morth, Jens Preben; Pedersen, Bjørn Panella; Buch-Pedersen, Morten Jeppe

    2011-01-01

    transport systems that are responsible for uptake and extrusion of metabolites and other ions. The ion gradients are also both directly and indirectly used to control pH homeostasis and to regulate cell volume. The plasma membrane H(+)-ATPase maintains a proton gradient in plants and fungi and the Na(+),K(+)-ATPase...... maintains a Na(+) and K(+) gradient in animal cells. Structural information provides insight into the function of these two distinct but related P-type pumps.......Plasma membrane ATPases are primary active transporters of cations that maintain steep concentration gradients. The ion gradients and membrane potentials derived from them form the basis for a range of essential cellular processes, in particular Na(+)-dependent and proton-dependent secondary...

  10. A structural overview of the plasma membrane Na+,K+-ATPase and H+-ATPase ion pumps

    DEFF Research Database (Denmark)

    Morth, Jens Preben; Pedersen, Bjørn Panella; Buch-Pedersen, Morten Jeppe

    2011-01-01

    transport systems that are responsible for uptake and extrusion of metabolites and other ions. The ion gradients are also both directly and indirectly used to control pH homeostasis and to regulate cell volume. The plasma membrane H(+)-ATPase maintains a proton gradient in plants and fungi and the Na......Plasma membrane ATPases are primary active transporters of cations that maintain steep concentration gradients. The ion gradients and membrane potentials derived from them form the basis for a range of essential cellular processes, in particular Na(+)-dependent and proton-dependent secondary......(+),K(+)-ATPase maintains a Na(+) and K(+) gradient in animal cells. Structural information provides insight into the function of these two distinct but related P-type pumps....

  11. Hormonal regulation of Na+/K+-dependent ATPase activity and pump function in corneal endothelial cells.

    Science.gov (United States)

    Hatou, Shin

    2011-10-01

    Na- and K-dependent ATPase (Na,K-ATPase) in the basolateral membrane of corneal endothelial cells plays an important role in the pump function of the corneal endothelium. We investigated the role of dexamethasone in the regulation of Na,K-ATPase activity and pump function in these cells. Mouse corneal endothelial cells were exposed to dexamethasone or insulin. ATPase activity was evaluated by spectrophotometric measurement, and pump function was measured using an Ussing chamber. Western blotting and immunocytochemistry were performed to measure the expression of the Na,K-ATPase α1-subunit. Dexamethasone increased Na,K-ATPase activity and the pump function of endothelial cells. Western blot analysis indicated that dexamethasone increased the expression of the Na,K-ATPase α1-subunit but decreased the ratio of active to inactive Na,K-ATPase α1-subunit. Insulin increased Na,K-ATPase activity and pump function of cultured corneal endothelial cells. These effects were transient and blocked by protein kinase C inhibitors and inhibitors of protein phosphatases 1 (PP1) and 2A (PP2A). Western blot analysis indicated that insulin decreased the amount of inactive Na,K-ATPase α1-subunit, but the expression of total Na,K-ATPase α1-subunit was unchanged. Immunocytochemistry showed that insulin increased cell surface expression of the Na,K-ATPase α1-subunit. Our results suggest that dexamethasone and insulin stimulate Na,K-ATPase activity in mouse corneal endothelial cells. The effect of dexamethasone activation in these cells was mediated by Na,K-ATPase synthesis and an increased enzymatic activity because of dephosphorylation of Na,K-ATPase α1-subunits. The effect of insulin is mediated by the protein kinase C, PP1, and/or PP2A pathways.

  12. Effects of aqueous extract of Hibiscus sabdariffa on renal Na(+)-K(+)-ATPase and Ca(2+)-Mg(2+)-ATPase activities in Wistar rats.

    Science.gov (United States)

    Olatunji, Lawrence A; Usman, Taofeek O; Adebayo, Joseph O; Olatunji, Victoria A

    2012-09-01

    To investigate the effects of oral administration of aqueous extract of Hibiscus sabdariffa on renal Na(+)-K(+)-ATPase and Ca(2+)-Mg(2+)-ATPase activities in rats. The 25 and 50 mg/(kg·d) of aqueous extracts of H. sabdariffa were respectively given to rats in the experimental groups for 28 d, and rats in the control group received an appropriate volume of distilled water as vehicle. Na(+)-K(+)-ATPase and Ca(2+)-Mg(2+)-ATPase activities in the kidney were assayed by spectrophotometric method. Administrations of 25 and 50 mg/(kg·d) of aqueous extract of H. sabdariffa significantly decreased the Ca(2+)-Mg(2+)-ATPase activity in the kidney of rats (Psabdariffa may preserve the renal function despite a decreased renal Ca(2+)-Mg(2+)-ATPase activity.

  13. In and out of the cation pumps: P-type ATPase structure revisited

    DEFF Research Database (Denmark)

    Bublitz, Maike; Poulsen, Hanne; Morth, Jens Preben

    2010-01-01

    Active transport across membranes is a crucial requirement for life. P-type ATPases build up electrochemical gradients at the expense of ATP by forming and splitting a covalent phosphoenzyme intermediate, coupled to conformational changes in the transmembrane section where the ions are translocated....... The marked increment during the last three years in the number of crystal structures of P-type ATPases has greatly improved our understanding of the similarities and differences of pumps with different ion specificities, since the structures of the Ca2+-ATPase, the Na+,K+-ATPase and the H+-ATPase can now...

  14. Crystal structure of a copper-transporting PIB-type ATPase

    DEFF Research Database (Denmark)

    Gourdon, Pontus Emanuel; Liu, Xiang-Yu; Skjørringe, Tina

    2011-01-01

    Heavy-metal homeostasis and detoxification is crucial for cell viability. P-type ATPases of the class IB (PIB) are essential in these processes, actively extruding heavy metals from the cytoplasm of cells. Here we present the structure of a PIB-ATPase, a Legionella pneumophila CopA Cu(+)-ATPase, ......Heavy-metal homeostasis and detoxification is crucial for cell viability. P-type ATPases of the class IB (PIB) are essential in these processes, actively extruding heavy metals from the cytoplasm of cells. Here we present the structure of a PIB-ATPase, a Legionella pneumophila CopA Cu...

  15. Review: P4-ATPases as Phospholipid Flippases-Structure, Function, and Enigmas

    DEFF Research Database (Denmark)

    Andersen, Jens P; Vestergaard, Anna L; Mikkelsen, Stine A

    2016-01-01

    group is propelled along against its concentration gradient with the hydrocarbon chains projecting out into the lipid phase by movement of an isoleucine located at the position corresponding to an ion binding glutamate in the Ca2+- and Na+/K+-ATPases. Hence, the P4-ATPase mechanism is quite similar...... on properties of mammalian and yeast P4-ATPases for which most mechanistic insight is available. However, the structure, function and enigmas associated with mammalian and yeast P4-ATPases most likely extend to P4-ATPases of plants and other organisms....

  16. The parietal cell gastric H, K-ATPase also functions as the Na, K-ATPase and Ca-ATPase in altered states [v2; ref status: indexed, http://f1000r.es/1tc

    Directory of Open Access Journals (Sweden)

    Tushar Ray

    2013-09-01

    Full Text Available This article offers an explanation for the apparent lack of Na, K-ATPase activity in parietal cells although ouabain has been known to inhibit gastric acid secretion since 1962. The gastric H, K-ATPase (proton-pump seems to be acting in altered states, thus behaving like a Na, K-ATPase (Na-pump and/or Ca-ATPase (Ca-pump depending on cellular needs.  This conclusion is based on the following findings. First, parietal cell fractions do not exhibit Na, K-ATPase activity at pH 7.0 but do at pH 8.5. Second, the apical plasma membrane (APM fraction exhibits a (Ca or Mg-ATPase activity with negligible H, K-ATPase activity. However, when assayed with Mg alone in presence of the 80 k Da cytosolic proton-pump activator (HAF, the APM fraction reveals remarkably high H, K-ATPase activity, suggesting the observed low affinity of Ca (or Mg-ATPase is an altered state of the latter. Third, calcium (between 1 and 4 µM shows both stimulation and inhibition of the HAF-stimulated H, K-ATPase depending on its concentration, revealing a close interaction between the  proton-pump activator and local Ca concentration in gastric H, K-ATPase function. Such interactions suggest that Ca is acting as a terminal member of the intracellular signaling system for the HAF-regulated proton-pump. It appears that during resting state, the HAF-associated H, K-ATPase remains inhibited by Ca (>1 µM and, prior to resumption of acid secretion the gastric H, K-ATPase acts temporarily as a Ca-pump for removing excess Ca from its immediate environment. This conclusion is consistent with the recent reports of immunochemical co-localization of the gastric H, K-ATPase and Ca-ATPase by superimposition in parietal cells, and a transitory efflux of Ca immediately preceding the onset of acid secretion. These new perspectives on proton-pump function would open new avenues for a fuller understanding of the intracellular regulation of the ubiquitous Na-pump.

  17. The gastric H, K-ATPase system also functions as the Na, K-ATPase and Ca-ATPase in altered states [v1; ref status: indexed, http://f1000r.es/1eo

    Directory of Open Access Journals (Sweden)

    Tushar Ray

    2013-07-01

    Full Text Available This article offers an explanation for the apparent lack of Na, K-ATPase activity in parietal cells although ouabain has been known to inhibit gastric acid secretion since 1962. The gastric H, K-ATPase (proton-pump seems to be acting in altered states, thus behaving like a Na, K-ATPase (Na-pump and/or Ca-ATPase (Ca-pump depending on cellular needs.  This conclusion is based on the following findings. First, parietal cell fractions do not exhibit Na, K-ATPase activity at pH 7.0 but do at pH 8.5. Second, the apical plasma membrane (APM fraction exhibits a (Ca or Mg-ATPase activity with negligible H, K-ATPase activity. However, when assayed with Mg alone in presence of the 80 k Da cytosolic proton-pump activator (HAF, the APM fraction reveals remarkably high H, K-ATPase activity, suggesting the observed low affinity of Ca (or Mg-ATPase is an altered state of the latter. Third, calcium (between 1 and 4 µM shows both stimulation and inhibition of the HAF-stimulated H, K-ATPase depending on its concentration, revealing a close interaction between the  proton-pump activator and local Ca concentration in gastric H, K-ATPase function. Such interactions suggest that Ca is acting as a terminal member of the intracellular signaling system for the HAF-regulated proton-pump. It appears that during resting state, the HAF-associated H, K-ATPase remains inhibited by Ca (>1 µM and, prior to resumption of acid secretion the gastric H, K-ATPase acts temporarily as a Ca-pump for removing excess Ca from its immediate environment. This conclusion is consistent with the recent reports of immunochemical co-localization of the gastric H, K-ATPase and Ca-ATPase by superimposition in parietal cells, and a transitory efflux of Ca immediately preceding the onset of acid secretion. These new perspectives on proton-pump function would open new avenues for a fuller understanding of the intracellular regulation of the ubiquitous Na-pump.

  18. New ATPase regulators-p97 goes to the PUB

    DEFF Research Database (Denmark)

    Madsen, Louise; Seeger, Michael; Semple, Colin A

    2009-01-01

    The conserved eukaryotic AAA-type ATPase complex, known as p97 or VCP in mammals and Cdc48 in yeast, is involved in a number of cellular pathways, including fusion of homotypic membranes, protein degradation, and activation of membrane-bound transcription factors. Most likely, p97 is directed...

  19. Probing the functional subunits of the tonoplast H+-ATPase

    International Nuclear Information System (INIS)

    Randall, S.K.; Lai, S.; Sze, H.

    1986-01-01

    The tonoplast ATPase of oat roots is composed of at least three polypeptides of 72, 60, and 16 kDa. The 16 kDA polypeptide covalently binds N,N'-dicyclohexylcarbodiimide and is postulated to be a component of the proton channel. Initial studies to identify other subunits indicate that both the 72 and 60 kDa subunits covalently bind 14 C]-7-chloro-4-nitrobenzo-2-oxa-1,3-diazole and [ 14 C]N-ethylamleimide, inhibitors of the tonoplast ATPase. ATP prevents binding of these inhibitors suggesting that both the 72 and 60 kDa subunits are involved in substrate binding. Polyclonal antibody has been made to the 72 kDa subunit. Western blot analysis of tonoplast vesicles reveals single reactive polypeptide (72 kDa). The antibody shows no cross-reactivity towards either the mitochondrial F 1 -ATPase or the plasma membrane ATPase. This antibody specifically inhibits ATP hydrolysis and ATP-dependent H + pumping in native tonoplast vesicles. The authors conclude that the 72 kDa subunit is intimately associated with the catalytic (or ATP-binding) site

  20. RNAi-based silencing of genes encoding the vacuolar- ATPase ...

    African Journals Online (AJOL)

    RNAi-based silencing of genes encoding the vacuolar- ATPase subunits a and c in pink bollworm (Pectinophora gossypiella). Ahmed M. A. Mohammed. Abstract. RNA interference is a post- transcriptional gene regulation mechanism that is predominantly found in eukaryotic organisms. RNAi demonstrated a successful ...

  1. Changes in erythrocyte ATPase activity under different pathological ...

    African Journals Online (AJOL)

    Changes in erythrocyte ATPase activity under different pathological conditions. Ali A Kherd, Nawal Helmi, Khadijah Saeed Balamash, Taha A Kumosani, Shareefa A AL-Ghamdi, Qari M, Etimad A Huwait, Soonham S Yaghmoor, Alaama Nabil, Maryam A AL-Ghamdi, Said S Moselhy ...

  2. The Kdp-ATPase system and its regulation

    Indian Academy of Sciences (India)

    2007-03-15

    Mar 15, 2007 ... K+, the dominant intracellular cation, is required for various physiological processes like turgor homeostasis, pH regulation etc. Bacterial cells have evolved many diverse K+ transporters to maintain the desired concentration of internal K+. In E. coli, the KdpATPase (comprising of the KdpFABC complex), ...

  3. Asymmetric incorporation of Na+, K+-ATPase into phospholipid vesicles

    NARCIS (Netherlands)

    Jackson, R.L.; Verkleij, A.J.; Zoelen, E.J.J. van; Lane, L.K.; Schwartz, A.; Deenen, L.L.M. van

    Purified lamb kidney Na+, K+-ATPase, consisting solely of the Mτ = 95,000 catalytic subunit and the Mτ- 44,000 glycoprotein, was solubilized with Triton X-100 and incorporated into unilamellar phospholipid vesicles. Freeze-fracture electron microscopy of the vesicles showed intramembranous particles

  4. RNAi-based silencing of genes encoding the vacuolar- ATPase ...

    African Journals Online (AJOL)

    2016-11-09

    Nov 9, 2016 ... Spodoptera exigua larval development by silencing chitin synthase gene with RNA interference. Bull. Entomol. Res. 98:613-619. Dow JAT (1999). The Multifunctional Drosophila melanogaster V-. ATPase is encoded by a multigene family. J. Bioenerg. Biomembr. 31:75-83. Fire A, Xu SQ, Montgomery MK, ...

  5. Models for the a subunits of the Thermus thermophilus V/A-ATPase and Saccharomyces cerevisiae V-ATPase enzymes by cryo-EM and evolutionary covariance

    Science.gov (United States)

    Schep, Daniel G.; Rubinstein, John L.

    2016-01-01

    Rotary ATPases couple ATP synthesis or hydrolysis to proton translocation across a membrane. However, understanding proton translocation has been hampered by a lack of structural information for the membrane-embedded a subunit. The V/A-ATPase from the eubacterium Thermus thermophilus is similar in structure to the eukaryotic V-ATPase but has a simpler subunit composition and functions in vivo to synthesize ATP rather than pump protons. We determined the T. thermophilus V/A-ATPase structure by cryo-EM at 6.4 Å resolution. Evolutionary covariance analysis allowed tracing of the a subunit sequence within the map, providing a complete model of the rotary ATPase. Comparing the membrane-embedded regions of the T. thermophilus V/A-ATPase and eukaryotic V-ATPase from Saccharomyces cerevisiae allowed identification of the α-helices that belong to the a subunit and revealed the existence of previously unknown subunits in the eukaryotic enzyme. Subsequent evolutionary covariance analysis enabled construction of a model of the a subunit in the S. cerevisae V-ATPase that explains numerous biochemical studies of that enzyme. Comparing the two a subunit structures determined here with a structure of the distantly related a subunit from the bovine F-type ATP synthase revealed a conserved pattern of residues, suggesting a common mechanism for proton transport in all rotary ATPases. PMID:26951669

  6. Vacuolar ATPases, like F1,F0-ATPases, show a strong dependence of the reaction velocity on the binding of more than one ATP per enzyme

    International Nuclear Information System (INIS)

    Kasho, V.N.; Boyer, P.D.

    1989-01-01

    Recent studies with vacuolar ATPases have shown that multiple copies catalytic subunits are present and that these have definite sequence homology with catalytic subunits of the F 1 , F 0 -ATPases. Experiments are reported that assess whether the vacuolar ATPases may have the unusual catalytic cooperativity with sequential catalytic site participation as in the binding change mechanism for the F 1 ,F 0 -ATPases. The extent of reversal of bound ATP hydrolysis to bound ADP and P i as medium ATP concentration was lowered was determined by 18 O-exchange measurements for yeast and neurospora vacuolar ATPases. The results show a pronounced increase in the extent of water oxygen incorporation into the P i formed as ATP concentration is decreased to the micromolar range. The F 1 ,F 0 -ATPase from neurospora mitochondria showed an event more pronounced modulation, similar to that of other F 1 -type ATPases. The vacuolar ATPases thus appear to have a catalytic mechanism quite analogous to that of the F 1 ,F 0 -ATPases

  7. Epigallocatechin-3-Gallate Protects Erythrocyte Ca2+-ATPase and Na+/K+-ATPase Against Oxidative Induced Damage During Aging in Humans

    Directory of Open Access Journals (Sweden)

    Prabhanshu Kumar

    2014-10-01

    Full Text Available Purpose: The main purpose of this study was to investigate the protective role of epigallocatechin-3-gallate on tertiary butyl hydroperoxide induced oxidative damage in erythrocyte during aging in humans. Methods: Human erythrocyte membrane bound Ca2+-ATPase and Na+/K+-ATPase activities were determined as a function of human age. Protective role of epigallocatechin-3-gallate was evaluated by in vitro experiments by adding epigallocatechin-3-gallate in concentration dependent manner (final concentration range 10-7M to 10-4M to the enzyme assay medium. Oxidative stress was induced in vitro by incubating washed erythrocyte ghosts with tertiary butyl hydroperoxide (10-5 M final concentration. Results: We have reported concentration dependent effect of epigallocatechin-3-gallate on tertiary butyl hydroperoxide induced damage on activities of Ca2+-ATPase and Na+/K+-ATPase during aging in humans. We have detected a significant (p < 0.001 decreased activity of Ca2+-ATPase and Na+/K+ -ATPase as a function of human age. Epigallocatechin-3-gallate protected ATPases against tertiary butyl hydroperoxide induced damage in concentration dependent manner during aging in humans. Conclusion: Epigallocatechin-3-gallate is a powerful antioxidant that is capable of protecting erythrocyte Ca2+-ATPase and Na+/K+ -ATPase against oxidative stress during aging in humans. We may propose hypothesis that a high intake of catechin rich diet may provide some protection against development of aging and age related diseases.

  8. Sodium ions as substitutes for protons in the gastric H,K-ATPase

    International Nuclear Information System (INIS)

    Polvani, C.; Sachs, G.; Blostein, R.

    1989-01-01

    In view of the striking homology among various ion-translocating ATPases including Na,K-ATPase, Ca-ATPase, and H,K-ATPase, and the recent evidence that protons can replace cytoplasmic sodium as well as potassium in the reaction mechanism of the Na,K-ATPase (Polvani, C., and Blostein, R. (1988) J. Biol. Chem. 263, 16757-16763), we studied the role of sodium as a substitute for protons in the H,K-ATPase reaction. Using hog gastric H,K-ATPase-rich inside-out membrane vesicles we observed 22Na+ influx which was stimulated by intravesicular potassium ions (K+i) at pH 8.5 but not at pH 7.1. This sodium influx was observed in medium containing ATP and was inhibited by vanadate and SCH28080, a selective inhibitor of the gastric H,K-ATPase. At least 2-fold accumulation of sodium was observed at pH 8.5. Experiments aimed to determine the sidedness of the alkaline pH requirement for K+i-dependent sodium influx showed that K+i-activated sodium influx depends on pHout and is unaffected by changes in pHin. These results support the conclusion that sodium ions substitute for protons in the H,K-ATPase reaction mechanism and provide evidence for a similarity in ion selectivity and/or binding domains of the Na,K-ATPase and the gastric H,K-ATPase enzymes

  9. Characterization of Na+K+-ATPase in bovine sperm.

    Science.gov (United States)

    Hickey, Katie D; Buhr, Mary M

    2012-04-15

    Existing as a ubiquitous transmembrane protein, Na(+)K(+)-ATPase affects sperm fertility and capacitation through ion transport and a recently identified signaling function. Functional Na(+)K(+)-ATPase is a dimer of α and β subunits, each with isoforms (four and three, respectively). Since specific isoform pairings and locations may influence or indicate function, the objective of this study was to identify and localize subunits of Na(+)K(+)-ATPase in fresh bull sperm by immunoblotting and immunocytochemistry using antibodies against α1 and 3, and all β isoforms. Relative quantity of Na(+)K(+)-ATPase in head plasma membranes (HPM's) from sperm of different bulls was determined by densitometry of immunoblot bands, and compared to bovine kidney. Sperm and kidney specifically bound all antibodies at kDa equivalent to commercial controls, and to additional lower kDa bands in HPM. Immunofluorescence of intact sperm confirmed that all isoforms were present in the head region of sperm and that α3 was also uniformly distributed post-equatorially. Permeabilization exposing internal membranes typically resulted in an increase in fluorescence, indicating that some antibody binding sites were present on the inner surface of the HPM or the acrosomal membrane. Deglycosylation of β1 reduced the kDa of bands in sperm, rat brain and kidney, with the kDa of the deglycosylated bands differing among tissues. Two-dimensional blots of β1 revealed three distinct spots. Based on the unique quantity, location and structure Na(+)K(+)-ATPase subunits in sperm, we inferred that this protein has unique functions in sperm. Copyright © 2012 Elsevier Inc. All rights reserved.

  10. ER Stress and Lipid Metabolism in Adipocytes

    Directory of Open Access Journals (Sweden)

    Beth S. Zha

    2012-01-01

    Full Text Available The role of endoplasmic reticulum (ER stress is a rapidly emerging field of interest in the pathogenesis of metabolic diseases. Recent studies have shown that chronic activation of ER stress is closely linked to dysregulation of lipid metabolism in several metabolically important cells including hepatocytes, macrophages, β-cells, and adipocytes. Adipocytes are one of the major cell types involved in the pathogenesis of the metabolic syndrome. Recent advances in dissecting the cellular and molecular mechanisms involved in the regulation of adipogenesis and lipid metabolism indicate that activation of ER stress plays a central role in regulating adipocyte function. In this paper, we discuss the current understanding of the potential role of ER stress in lipid metabolism in adipocytes. In addition, we touch upon the interaction of ER stress and autophagy as well as inflammation. Inhibition of ER stress has the potential of decreasing the pathology in adipose tissue that is seen with energy overbalance.

  11. Endoplasmic reticulum: ER stress regulates mitochondrial bioenergetics

    Science.gov (United States)

    Bravo, Roberto; Gutierrez, Tomás; Paredes, Felipe; Gatica, Damián; Rodriguez, Andrea E.; Pedrozo, Zully; Chiong, Mario; Parra, Valentina; Quest, Andrew F.G.; Rothermel, Beverly A.; Lavandero, Sergio

    2014-01-01

    Endoplasmic reticulum (ER) stress activates an adaptive unfolded protein response (UPR) that facilitates cellular repair, however, under prolonged ER stress, the UPR can ultimately trigger apoptosis thereby terminating damaged cells. The molecular mechanisms responsible for execution of the cell death program are relatively well characterized, but the metabolic events taking place during the adaptive phase of ER stress remain largely undefined. Here we discuss emerging evidence regarding the metabolic changes that occur during the onset of ER stress and how ER influences mitochondrial function through mechanisms involving calcium transfer, thereby facilitating cellular adaptation. Finally, we highlight how dysregulation of ER–mitochondrial calcium homeostasis during prolonged ER stress is emerging as a novel mechanism implicated in the onset of metabolic disorders. PMID:22064245

  12. Expression of Na,K-ATPase and H,K-ATPase Isoforms with the Baculovirus Expression System

    NARCIS (Netherlands)

    Koenderink, J.B.; Swarts, H.G.

    2016-01-01

    P-type ATPases can be expressed in several cell systems. The baculovirus expressions system uses an insect virus to enter and express proteins in Sf9 insect cells. This expression system is a lytic system in which the cells will die a few days after viral infection. Subsequently, the expressed

  13. Regulation of proximal tubule vacuolar H+-ATPase by PKA and AMP-activated protein kinase

    Science.gov (United States)

    Al-bataineh, Mohammad M.; Gong, Fan; Marciszyn, Allison L.; Myerburg, Michael M.

    2014-01-01

    The vacuolar H+-ATPase (V-ATPase) mediates ATP-driven H+ transport across membranes. This pump is present at the apical membrane of kidney proximal tubule cells and intercalated cells. Defects in the V-ATPase and in proximal tubule function can cause renal tubular acidosis. We examined the role of protein kinase A (PKA) and AMP-activated protein kinase (AMPK) in the regulation of the V-ATPase in the proximal tubule as these two kinases coregulate the V-ATPase in the collecting duct. As the proximal tubule V-ATPases have different subunit compositions from other nephron segments, we postulated that V-ATPase regulation in the proximal tubule could differ from other kidney tubule segments. Immunofluorescence labeling of rat ex vivo kidney slices revealed that the V-ATPase was present in the proximal tubule both at the apical pole, colocalizing with the brush-border marker wheat germ agglutinin, and in the cytosol when slices were incubated in buffer alone. When slices were incubated with a cAMP analog and a phosphodiesterase inhibitor, the V-ATPase accumulated at the apical pole of S3 segment cells. These PKA activators also increased V-ATPase apical membrane expression as well as the rate of V-ATPase-dependent extracellular acidification in S3 cell monolayers relative to untreated cells. However, the AMPK activator AICAR decreased PKA-induced V-ATPase apical accumulation in proximal tubules of kidney slices and decreased V-ATPase activity in S3 cell monolayers. Our results suggest that in proximal tubule the V-ATPase subcellular localization and activity are acutely coregulated via PKA downstream of hormonal signals and via AMPK downstream of metabolic stress. PMID:24553431

  14. Optimal fodring af goldkøer

    DEFF Research Database (Denmark)

    Bjerre-Harpøth, Vibeke; Damgaard, Birthe Marie

    2013-01-01

    Et forsøg har vist, at køer på lavt energiniveau i goldperioden var fysiologisk sundere og havde mindre risiko for at udvikle stofskiftesygdomme end køer på et højt energiniveau. Forsøget viste også, at køer på et normalt energiniveau i senlaktationen gav mere mælk i den efterfølgende laktation e...

  15. Er moral bare noget vi leger?

    DEFF Research Database (Denmark)

    Thomsen, Frej Klem

    2014-01-01

    Hvis man er skeptiker, så er moral nemlig altid kun et udtryk for psykologi og kultur, også når det handler om for eksempel misbrug af børn i Tønder-sagen eller terror-angreb i Madrid, London og Mumbai .......Hvis man er skeptiker, så er moral nemlig altid kun et udtryk for psykologi og kultur, også når det handler om for eksempel misbrug af børn i Tønder-sagen eller terror-angreb i Madrid, London og Mumbai ....

  16. Thermodynamic assessments of the Ag-Er and Er-Y systems

    International Nuclear Information System (INIS)

    Wang, S.L.; Wang, C.P.; Liu, X.J.; Tang, A.T.; Pan, F.S.; Ishida, K.

    2010-01-01

    The phase diagrams and thermodynamic properties in the Ag-Er and Er-Y binary systems have been assessed by using the CALPHAD (Calculation of Phase Diagrams) method on the basis of the experimental data including the thermodynamic properties and phase equilibria. The Gibbs free energies of the liquid, bcc, fcc, and hcp phases were described by the subregular solution model with the Redlich-Kister equation, and those of intermetallic compounds (Ag 2 Er and AgEr phases) were treated as stoichiometric compounds, and Ag 51 Er 14 phase was modeled by the sublattice model in the Ag-Er binary system. The thermodynamic parameters of the Ag-Er and Er-Y binary systems were obtained, and an agreement between the calculated results and experimental data was obtained for each binary system.

  17. A tomato ER-type Ca2+-ATPase, LCA1, has a low thapsigargin-sensitivity and can transport manganese

    DEFF Research Database (Denmark)

    Johnson, Neil A.; Liu, F; Weeks, P. D.

    2008-01-01

    and directly via in vitro ATP hydrolysis. We found LCA1 to be approximately 300-fold less sensitive to thapsigargin than animal SERCAs, whereas ECA1 was thapsigargin-resistant. LCA1 showed typical pharmacological sensitivities to cyclopiazonic acid, vanadate, and eosin, consistent with it being a P...

  18. Neoclassical transport in ERS regime

    International Nuclear Information System (INIS)

    Lin, Z.; Tang, W.; Lee, W.W.

    1996-01-01

    The core ion thermal conductivity has been observed to fall below the standard neoclassical level in recent TFTR reversed magnetic shear discharges. Due to the combination of high central q and small local inverse aspect ratio, the ion poloidal gyroradius can be larger than the minor radius and comparable to the pressure gradient scale length in this ERS regime. It is then expected that finite orbit width effects play a key role in modifying the standard picture of neoclassical transport. Specifically, both the trapped particle fraction and the banana orbit width axe reduced by the finite minor radius and possibly by the pressure-gradient-driven radial electric field. In addition, the steep pressure gradient can generate neoclassical poloidal flows which, in turn, could reduce the particle and heat transports. Results from analytic estimates as well as those from full toroidal gyrokinetic neoclassical simulations will be presented

  19. Ungdomslitteratur former(er) sig

    DEFF Research Database (Denmark)

    Henkel, Ayoe Qvist

    2016-01-01

    Igennem en 'mediesensitiv' analyse af romanen "Akavet" af Ronnie Andersen (2014) og perspektivering til andre aktuelle romaner for og med unge undersøger artiklen, hvordan ungdomslitteratur udvikler sig i dialog med digitale og mediebaserede impulser, og hvilke konsekvenser for udsigelserne om...... ungdomsliv og ungdomslitteraturens æstetik og mulige egenart, denne udvikling har. Artiklen baserer sig på en materialitetstilgang særligt inspireret af N. Katherine Hayles, som ikke tidligere har fået opmærksomhed i læsninger af ungdomslitteratur eller i diskussioner af ungdomslitteraturens mulige egenart......, at ungdomslitteratur drejer sig om unges udviklingsproces fra barndom og til voksenhed og dermed skildrer en overgang præget af linearitet, modning og vækst. Artiklen konkluderer, at der er ungdomslitteratur, der realiseres på andre præmisser, og Akavet kan ses som eksponent for en bevægelse fra ungdomslitteratur som...

  20. Low Estrogen Receptor (ER)-Positive Breast Cancer and Neoadjuvant Systemic Chemotherapy: Is Response Similar to Typical ER-Positive or ER-Negative Disease?

    Science.gov (United States)

    Landmann, Alessandra; Farrugia, Daniel J; Zhu, Li; Diego, Emilia J; Johnson, Ronald R; Soran, Atilla; Dabbs, David J; Clark, Beth Z; Puhalla, Shannon L; Jankowitz, Rachel C; Brufsky, Adam M; Ahrendt, Gretchen M; McAuliffe, Priscilla F; Bhargava, Rohit

    2018-05-08

    Pathologic complete response (pCR) rate after neoadjuvant chemotherapy was compared between 141 estrogen receptor (ER)-negative (43%), 41 low ER+ (13%), 47 moderate ER+ (14%), and 98 high ER+ (30%) tumors. Human epidermal growth factor receptor 2-positive cases, cases without semiquantitative ER score, and patients treated with neoadjuvant endocrine therapy alone were excluded. The pCR rate of low ER+ tumors was similar to the pCR rate of ER- tumors (37% and 26% for low ER and ER- respectively, P = .1722) but significantly different from the pCR rate of moderately ER+ (11%, P = .0049) and high ER+ tumors (4%, P < .0001). Patients with pCR had an excellent prognosis regardless of the ER status. In patients with residual disease (no pCR), the recurrence and death rate were higher in ER- and low ER+ cases compared with moderate and high ER+ cases. Low ER+ breast cancers are biologically similar to ER- tumors. Semiquantitative ER H-score is an important determinant of response to neoadjuvant chemotherapy.

  1. Regulation of cardiac remodeling by cardiac Na/K-ATPase isoforms

    Directory of Open Access Journals (Sweden)

    Lijun Catherine Liu

    2016-09-01

    Full Text Available Cardiac remodeling occurs after cardiac pressure/volume overload or myocardial injury during the development of heart failure and is a determinant of heart failure. Preventing or reversing remodeling is a goal of heart failure therapy. Human cardiomyocyte Na+/K+-ATPase has multiple α isoforms (1-3. The expression of the α subunit of the Na+/K+-ATPase is often altered in hypertrophic and failing hearts. The mechanisms are unclear. There are limited data from human cardiomyocytes. Abundant evidences from rodents show that Na+/K+-ATPase regulates cardiac contractility, cell signaling, hypertrophy and fibrosis. The α1 isoform of the Na+/K+-ATPase is the ubiquitous isoform and possesses both pumping and signaling functions. The α2 isoform of the Na+/K+-ATPase regulates intracellular Ca2+ signaling, contractility and pathological hypertrophy. The α3 isoform of the Na+/K+-ATPase may also be a target for cardiac hypertrophy. Restoration of cardiac Na+/K+-ATPase expression may be an effective approach for prevention of cardiac remodeling. In this article, we will overview: (1 the distribution and function of isoform specific Na+/K+-ATPase in the cardiomyocytes. (2 the role of cardiac Na+/K+-ATPase in the regulation of cell signaling, contractility, cardiac hypertrophy and fibrosis in vitro and in vivo. Selective targeting of cardiac Na+/K+-ATPase isoform may offer a new target for the prevention of cardiac remodeling.

  2. Differential effects of inhibitors and detergents on the Ca2+-ATPase and Mg2+-ATPase activities of the plasma membrane of a human oat cell carcinoma

    International Nuclear Information System (INIS)

    Knowles, A.F.; Lawrence, C.M.

    1986-01-01

    Plasma membranes of human oat cell carcinoma possess Mg 2+ - and Ca 2+ -dependent ATPase activities of similar magnitude. These activities exhibit the unusual characteristic of being inactiviated by prolonged incubation of the membrane with 1-2 mM dithiothreitol (DTT). Inactivation by DTT was prevented by lowering the incubation temperature, elevation of the membrane protein concentration, and addition of ATP. Fluorosulfonylbenzoyl adenosine (FSBA), an affinity ATP analog, also inactivates these activities. The Ca 2+ -ATPase activity appears to be more sensitive to both DTT and FSBA. The Ca 2+ -ATPase activity is more easily inactivated by Triton X-100, while the Mg 2+ -ATPase is preferentially activated by digitonin. These differential effects of inhibitors and detergents suggest that the Ca 2+ -ATPase and Mg 2+ -ATPase are separate enzymes. Incubation of oat cell carcinoma plasma membrane with [ 3 H]FSBA resulted in the labeling of several proteins. A labelled 35,000 dalton protein corresponds to the molecular weight of the oat cell carcinoma plasma membrane Ca 2+ -ATPase previously purified in this laboratory. The identity of one or more of the other labelled proteins with the Mg 2+ -ATPase has not been demonstrated, but is presently under investigation

  3. The non-gastric H,K-ATPase is oligomycin-sensitive and can function as an H+,NH4(+)-ATPase.

    NARCIS (Netherlands)

    Swarts, H.G.P.; Koenderink, J.B.; Willems, P.H.G.M.; Pont, J.J.H.H.M. de

    2005-01-01

    We used the baculovirus/Sf9 expression system to gain new information on the mechanistic properties of the rat non-gastric H,K-ATPase, an enzyme that is implicated in potassium homeostasis. The alpha2-subunit of this enzyme (HKalpha2) required a beta-subunit for ATPase activity thereby showing a

  4. Sperm Na+, K+-ATPase and Ca2+-ATPase activity: A preliminary study of comparison of swim up and density gradient centrifugation methods for sperm preparation

    Science.gov (United States)

    Lestari, Silvia W.; Larasati, Manggiasih D.; Asmarinah, Mansur, Indra G.

    2018-02-01

    As one of the treatment for infertility, the success rate of Intrauterine Insemination (IUI) is still relatively low. Several sperm preparation methods, swim-up (SU) and the density-gradient centrifugation (DGC) are frequently used to select for better sperm quality which also contribute to IUI failure. Sperm selection methods mainly separate the motile from the immotile sperm, eliminating the seminal plasma. The sperm motility involves the structure and function of sperm membrane in maintaining the balance of ion transport system which is regulated by the Na+, K+-ATPase, and Ca2+-ATPase enzymes. This study aims to re-evaluate the efficiency of these methods in selecting for sperm before being used for IUI and based the evaluation on sperm Na+,K+-ATPase and Ca2+-ATPase activities. Fourteen infertile men from couples who underwent IUI were involved in this study. The SU and DGC methods were used for the sperm preparation. Semen analysis was performed based on the reference value of World Health Organization (WHO) 2010. After isolating the membrane fraction of sperms, the Na+, K+-ATPase activity was defined as the difference in the released inorganic phosphate (Pi) with and without the existence of 10 mM ouabain in the reaction, while the Ca2+-ATPase was determined as the difference in Pi contents with and without the existence of 55 µm CaCl2. The prepared sperm demonstrated a higher percentage of motile sperm compared to sperm from the whole semen. Additionally, the percentage of motile sperm of post-DGC showed higher result than the sperm from post-SU. The velocity of sperm showed similar pattern with the percentage of motile sperm, in which the velocity of prepared sperm was higher than the sperm from whole semen. Furthermore, the sperm velocity of post-DGC was higher compared to the sperm from post-SU. The Na+, K+-ATPase activity of prepared sperm was higher compared to whole semen, whereas Na+, K+-ATPase activity in the post DGC was higher than post SU. The Ca2

  5. Multiple band structure in 156Er

    International Nuclear Information System (INIS)

    Sunyar, A.W.; Der Mateosian, E.; Kistner, O.C.; Johnson, A.; Lumpkin, A.H.; Thieberger, P.

    1976-01-01

    The 142 Nd( 18 O,4n) 156 Er reaction at 90-95 MeV was used to study 156 Er high-spin states to spin 24. In addition to the background ground-state band, two well developed off-spin side bands, one of each parity, were observed. (Auth.)

  6. Er der en underviser til stede?

    DEFF Research Database (Denmark)

    Christiansen, René Boyer; Nortvig, Anne-Mette; Rosenlund, Lea Tilde

    2015-01-01

    I de senere år er MOOC (Massive Open Online Courses) blevet en faktor på uddannelsesområdet. MOOC-forskning er et forholdsvist nyt felt, men har inden for de seneste 2-3 år udviklet sig med hastige skridt (Liyanagunawardena et al. 2013, Bayne & Ross 2014). En stor del af denne forskning har haft...

  7. Coronavirus infection, ER stress and Apoptosis

    Directory of Open Access Journals (Sweden)

    TO SING eFUNG

    2014-06-01

    Full Text Available The replication of coronavirus, a family of important animal and human pathogens, is closely associated with the cellular membrane compartments, especially the endoplasmic reticulum (ER. Coronavirus infection of cultured cells was previously shown to cause ER stress and induce the unfolded protein response (UPR, a process that aims to restore the ER homeostasis by global translation shutdown and increasing the ER folding capacity. However under prolonged ER stress, UPR can also induce apoptotic cell death. Accumulating evidence from recent studies has shown that induction of ER stress and UPR may constitute a major aspect of coronavirus-host interaction. Activation of the three branches of UPR modulates a wide variety of signaling pathways, such as mitogen-activated protein (MAP kinases activation, autophagy, apoptosis and innate immune response. ER stress and UPR activation may therefore contribute significantly to the viral replication and pathogenesis during coronavirus infection. In this review, we summarize current knowledge on coronavirus-induced ER stress and UPR activation, with emphasis on their cross-talking to apoptotic signaling.

  8. Led strategisk med 3 F'er

    DEFF Research Database (Denmark)

    Klausen, Kurt Klaudi

    2017-01-01

    Artiklen gennemgår kort de tre F'er: at fremskrive, forudse og forestille som strategiske værktøjer til strategic forecast......Artiklen gennemgår kort de tre F'er: at fremskrive, forudse og forestille som strategiske værktøjer til strategic forecast...

  9. Et kors er ikke bare et kors

    DEFF Research Database (Denmark)

    Bach-Nielsen, Carsten

    2015-01-01

    Om et kors, der er rejst uden for den danske kirke i Bruxelles; formodentlig en afledning af det såkaldte Brabanterkors.......Om et kors, der er rejst uden for den danske kirke i Bruxelles; formodentlig en afledning af det såkaldte Brabanterkors....

  10. Er formen blevet vigtigst i politik?

    DEFF Research Database (Denmark)

    Etemadi, Maziar Haghpaidar

    2005-01-01

    RETORIK: Retorikken er kunsten at tage ordet - skriftligt eller mundtligt - og udtrykke sig smukt og velformuleret. Gorgias (ca. 490-ca. 390 f. Kr.) fra Leontinoi på Sicilien betragtes som ophavsmanden til den første endelige formulering af retorikkens virke og muligheder: "Talen er en mægtig...

  11. En isoleret verden er en bedre verden

    DEFF Research Database (Denmark)

    Enrum, Nils

    2014-01-01

    Man skal ikke kaste med sten, hvis man bor i et hus med energimærke D ... Men problemet med varmetab fra danske boliger er til at løse......Man skal ikke kaste med sten, hvis man bor i et hus med energimærke D ... Men problemet med varmetab fra danske boliger er til at løse...

  12. "Det man siger er man selv..."

    DEFF Research Database (Denmark)

    Næsby, Torben; Nørgaard, Britta; Uddholm, Mats

    forhold, der er hermeneutisk, strukturelt og relationelt bestemt. Praksisviden kan ikke være objektiv i gængs forstand, men det behøver ikke at diskvalificere denne viden. Forståelse er altid knyttet til den sag og bundet til den situation man står overfor og i som professionel og som menneske....

  13. Mechanism of photoinactivation of plant plasma membrane ATPases

    International Nuclear Information System (INIS)

    Imbrie, C.W.; Murphy, T.M.

    1984-01-01

    UV radiation at 290 and 365 nm inactivates two forms of the K + -stimulated ATPase associated with the plasma membrane of suspension-cultured cells of Rosa damascena. One form is 15 and 36 times more sensitive than the other to 290 and 365 nm, respectively. For both forms, the inactivation requires oxygen, is inhibited by azide and diazobicyclo(2.2.2.2)octane, but not glycerol, and is enhanced up to 7.5 times in deuterium oxide solvent. Inactivation occurs concomitantly with loss of absorbance at 290 nm. Cs + and NO 3 - , quenchers of tryptophan fluorescence, inhibit inactivation. The results suggest that inactivation involves singlet-oxygen mediated destruction of tryptophans in the ATPases. (author)

  14. Structural and functional characterization of P4-ATPase lipid flippases

    DEFF Research Database (Denmark)

    Ulstrup, Jakob

    2018-01-01

    to its much larger substrate and how the mechanism allowing the transport unfolds. This is one of the central questions in the field known as the “giant substrate problem”. To this date, no structural information of P4-ATPases is available. The focus of this thesis is divided into two projects, both...... focusing on P4-ATPases from the yeast organism Saccharomyces cerevisiae: I. The structural characterization of the flippase Drs2p in complex with its auxiliary subunit Cdc50p. II. Establishing a protocol for obtaining a homogenous sample of the flippase Neo1p suitable for biochemical characterization...... and substrate identification. Part I was performed using X-ray crystallography and single-particle electron microscopy as the main methods. A 3D envelope was obtained by cryo-EM extending to a resolution of 4.4 Å. This envelope reveals the first structural insight of the conformational organization of the Drs2p...

  15. Arginine substitution of a cysteine in transmembrane helix M8 converts Na+,K+-ATPase to an electroneutral pump similar to H+,K+-ATPase.

    Science.gov (United States)

    Holm, Rikke; Khandelwal, Jaanki; Einholm, Anja P; Andersen, Jens P; Artigas, Pablo; Vilsen, Bente

    2017-01-10

    Na + ,K + -ATPase and H + ,K + -ATPase are electrogenic and nonelectrogenic ion pumps, respectively. The underlying structural basis for this difference has not been established, and it has not been revealed how the H + ,K + -ATPase avoids binding of Na + at the site corresponding to the Na + -specific site of the Na + ,K + -ATPase (site III). In this study, we addressed these questions by using site-directed mutagenesis in combination with enzymatic, transport, and electrophysiological functional measurements. Replacement of the cysteine C932 in transmembrane helix M8 of Na + ,K + -ATPase with arginine, present in the H + ,K + -ATPase at the corresponding position, converted the normal 3Na + :2K + :1ATP stoichiometry of the Na + ,K + -ATPase to electroneutral 2Na + :2K + :1ATP stoichiometry similar to the electroneutral transport mode of the H + ,K + -ATPase. The electroneutral C932R mutant of the Na + ,K + -ATPase retained a wild-type-like enzyme turnover rate for ATP hydrolysis and rate of cellular K + uptake. Only a relatively minor reduction of apparent Na + affinity for activation of phosphorylation from ATP was observed for C932R, whereas replacement of C932 with leucine or phenylalanine, the latter of a size comparable to arginine, led to spectacular reductions of apparent Na + affinity without changing the electrogenicity. From these results, in combination with structural considerations, it appears that the guanidine + group of the M8 arginine replaces Na + at the third site, thus preventing Na + binding there, although allowing Na + to bind at the two other sites and become transported. Hence, in the H + ,K + -ATPase, the ability of the M8 arginine to donate an internal cation binding at the third site is decisive for the electroneutral transport mode of this pump.

  16. Protein import into chloroplasts requires a chloroplast ATPase

    Energy Technology Data Exchange (ETDEWEB)

    Pain, D.; Blobel, G.

    1987-05-01

    The authors have transcribed mRNA from a cDNA clone coding for pea ribulose-1,5-bisphosphate carboxylase, translated the mRNA in a wheat germ cell-free system, and studied the energy requirement for posttranslational import of the (/sup 35/S)methionine-labeled protein into the stroma of pea chloroplasts. They found that import depends on ATP hydrolysis within the stroma. Import is not inhibited when H/sup +/, K/sup +/, Na/sup +/, or divalent cation gradients across the chloroplast membranes are dissipated by ionophores, as long as exogenously added ATP is also present during the import reaction. The data suggest that protein import into the chloroplast stroma requires a chloroplast ATPase that does not function to generate a membrane potential for driving the import reaction but that exerts its effect in another, yet-to-be-determined, mode. They have carried out a preliminary characterization of this ATPase regarding its nucleotide specificity and the effects of various ATPase inhibitors.

  17. V-ATPase as an effective therapeutic target for sarcomas

    Energy Technology Data Exchange (ETDEWEB)

    Perut, Francesca, E-mail: francesca.perut@ior.it [Laboratory for Orthopaedic Pathophysiology and Regenerative Medicine, Istituto Ortopedico Rizzoli, Bologna (Italy); Avnet, Sofia; Fotia, Caterina; Baglìo, Serena Rubina; Salerno, Manuela [Laboratory for Orthopaedic Pathophysiology and Regenerative Medicine, Istituto Ortopedico Rizzoli, Bologna (Italy); Hosogi, Shigekuni [Laboratory for Orthopaedic Pathophysiology and Regenerative Medicine, Istituto Ortopedico Rizzoli, Bologna (Italy); Department of Molecular Cell Physiology, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, Kyoto (Japan); Kusuzaki, Katsuyuki [Department of Molecular Cell Physiology, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, Kyoto (Japan); Baldini, Nicola [Laboratory for Orthopaedic Pathophysiology and Regenerative Medicine, Istituto Ortopedico Rizzoli, Bologna (Italy); Department of Biomedical and Neuromotor Sciences, University of Bologna, Bologna (Italy)

    2014-01-01

    Malignant tumors show intense glycolysis and, as a consequence, high lactate production and proton efflux activity. We investigated proton dynamics in osteosarcoma, rhabdomyosarcoma, and chondrosarcoma, and evaluated the effects of esomeprazole as a therapeutic agent interfering with tumor acidic microenvironment. All sarcomas were able to survive in an acidic microenvironment (up to 5.9–6.0 pH) and abundant acidic lysosomes were found in all sarcoma subtypes. V-ATPase, a proton pump that acidifies intracellular compartments and transports protons across the plasma membrane, was detected in all cell types with a histotype-specific expression pattern. Esomeprazole administration interfered with proton compartmentalization in acidic organelles and induced a significant dose-dependent toxicity. Among the different histotypes, rhabdomyosarcoma, expressing the highest levels of V-ATPase and whose lysosomes are most acidic, was mostly susceptible to ESOM treatment. - Highlights: • Osteosarcoma, rhabdomyosarcoma, and chondrosarcoma survive in acidic microenvironment. • At acidic extracellular pH, sarcoma survival is dependent on V-ATPase expression. • Esomeprazole administration induce a significant dose-dependent toxicity.

  18. Radiation inactivation analysis of chloroplast CF0-CF1 ATPase

    International Nuclear Information System (INIS)

    Wang, M.Y.; Chien, L.F.; Pan, R.L.

    1988-01-01

    Radiation inactivation technique was employed to measure the functional size of adenosine triphosphatase of spinach chloroplasts. The functional size for acid-base-induced ATP synthesis was 450 +/- 24 kilodaltons; for phenazine methosulfate-mediated ATP synthesis, 613 +/- 33 kilodaltons; and for methanol-activated ATP hydrolysis, 280 +/- 14 kilodaltons. The difference (170 +/- 57 kilodaltons) between 450 +/- 24 and 280 +/- 14 kilodaltons is explained to be the molecular mass of proton channel (coupling factor 0) across the thylakoid membrane. Our data suggest that the stoichiometry of subunits I, II, and III of coupling factor 0 is 1:2:15. Ca2+- and Mg2+-ATPase activated by methanol, heat, and trypsin digestion have a similar functional size. However, anions such as SO 3 (2-) and CO 3 (2-) increased the molecular mass for both ATPase's (except trypsin-activated Mg2+-ATPase) by 12-30%. Soluble coupling factor 1 has a larger target size than that of membrane-bound. This is interpreted as the cold effect during irradiation

  19. Formation of oriented membrane multilayers of Na/K-ATPase

    International Nuclear Information System (INIS)

    Pachence, J.M.; Knott, R.; Edelman, I.S.; Schoenborn, B.P.; Wallace, B.A.

    1982-01-01

    The isolated membrane-bound enzyme retains its ouabain-sensitive ATP hydrolysis activity, and produces ATP-dependent Na + and K + fluxes when incorporated into phospholipid vesicles. The ultimate goal of this work is to determine its low resolution structure using both X-ray and neutron diffraction. A number of methods were used to impart lamellar stacking order to highly purified pig Na/K-ATPase membranes. Upon partial dehydration, x-ray diffraction from Na/K-ATPase membrane multilayers at 98% relative humidity yielded discrete reflections of 118 A periodicity, diffracting to 1/14.8 A -1 , additionally, continuous diffraction to 1/10 A -1 was obtained. Subjecting the membrane multilayers to high magnetic fields improved the quality of the lamellar diffraction dramatically. Neutron diffraction studies of the partially dehydrated Na/K-ATPase membrane multilayers detected a mosaic spread of 2 0 when the samples were subjected to a magnetic field of 5 Tesla perpendicular to the membrane surface; the reflections were narrower than the camera line width; hence, the lattice disorder has also decreased significantly, although only four orders were measured

  20. Porphyromonas gingivalis is highly sensitive to inhibitors of a proton-pumping ATPase.

    Science.gov (United States)

    Sekiya, Mizuki; Shimoyama, Yu; Ishikawa, Taichi; Sasaki, Minoru; Futai, Masamitsu; Nakanishi-Matsui, Mayumi

    2018-04-15

    Porphyromonas gingivalis is a well-known Gram-negative bacterium that causes periodontal disease. The bacterium metabolizes amino acids and peptides to obtain energy. An ion gradient across its plasma membrane is thought to be essential for nutrient import. However, it is unclear whether an ion-pumping ATPase responsible for the gradient is required for bacterial growth. Here, we report the inhibitory effect of protonophores and inhibitors of a proton-pumping ATPase on the growth of P. gingivalis. Among the compounds examined, curcumin and citreoviridin appreciably reduced the bacterial growth. Furthermore, these compounds inhibited the ATPase activity in the bacterial membrane, where the A-type proton-pumping ATPase (A-ATPase) is located. This study suggests that curcumin and citreoviridin inhibit the bacterial growth by inhibiting the A-ATPase in the P. gingivalis membrane. Copyright © 2018 Elsevier Inc. All rights reserved.

  1. Human and rodent muscle Na(+)-K(+)-ATPase in diabetes related to insulin, starvation, and training

    DEFF Research Database (Denmark)

    Schmidt, T A; Hasselbalch, S; Farrell, P A

    1994-01-01

    cerebral cortex Na(+)-K(+)-ATPase concentration as a result of diabetes, semistarvation, or insulin treatment. In human subjects, Na(+)-K(+)-ATPase concentration in vastus lateralis muscle biopsies was 17 and 22% greater (P dependent diabetes...... mellitus (n = 24) and insulin-dependent diabetes mellitus (n = 7) than in control subjects (n = 8). A positive linear correlation between muscle Na(+)-K(+)-ATPase and plasma insulin concentrations was observed (r = 0.50, P = 0.006; n = 29). Thus, insulin seems a regulator of muscle Na......(+)-K(+)-ATPase concentration, reduction of muscle Na(+)-K(+)-ATPase concentration with untreated diabetes bears similarities with undernourishment, and physical conditioning may ameliorate the muscle Na(+)-K(+)-ATPase concentration decrease induced by diabetes....

  2. The role of the plasma membrane H+-ATPase in plant-microbe interactions.

    Science.gov (United States)

    Elmore, James Mitch; Coaker, Gitta

    2011-05-01

    Plasma membrane (PM) H+-ATPases are the primary pumps responsible for the establishment of cellular membrane potential in plants. In addition to regulating basic aspects of plant cell function, these enzymes contribute to signaling events in response to diverse environmental stimuli. Here, we focus on the roles of the PM H+-ATPase during plant-pathogen interactions. PM H+-ATPases are dynamically regulated during plant immune responses and recent quantitative proteomics studies suggest complex spatial and temporal modulation of PM H+-ATPase activity during early pathogen recognition events. Additional data indicate that PM H+-ATPases cooperate with the plant immune signaling protein RIN4 to regulate stomatal apertures during bacterial invasion of leaf tissue. Furthermore, pathogens have evolved mechanisms to manipulate PM H+-ATPase activity during infection. Thus, these ubiquitous plant enzymes contribute to plant immune responses and are targeted by pathogens to increase plant susceptibility.

  3. Subcellular localization of H(+)-ATPase from pumpkin hypocotyls (Cucurbita maxima L.) by membrane fractionation.

    Science.gov (United States)

    Scherer, G F

    1984-03-01

    A new method of preparing sealed vesicles from membrane fractions of pumpkin hypocotyls in ethanolamine-containing buffers was used to investigate the subcellular localization of H(+)-ATPase measured as nigericin-stimulated ATPase. In a fluorescence-quench assay, the H(+) pump was directly demonstrated. The H(+) pump was substrate-specific for Mg·ATP and 0.1 mM diethylstilbestrol completely prevented the development of a Δ pH. The presence of unsupecific phosphatase hampered the detection of nigericin-stimulated ATPase. Unspecific phosphatases could be demonstrated by comparing the broad substrate specificity of the hydrolytic activities of the fractions with the clear preference for Mg·ATP as the substrate for the proton pump. Inhibitor studies showed that neither orthovanadate nor molybdate are absolutely specific for ATPase or acid phosphatase, respectively. Diethylstilbestrol seemed to be a specific inhibitor of ATPase activity in fractions containing nigericin-stimulated ATPase, but it stimulated acid phosphatase which tended to obscure its effect on ATPase activity. Nigericin-stimulated ATPase had its optimum at pH 6.0 and the nigericin effect was K(+)-dependent. The combination of valinomycin and carbonylcyanide m-chlorophenylhydrazone had a similar effect to nigericin, but singly these ionophores were much less stimulatory. After prolonged centrifugation on linear sucrose gradients, nigericin-stimulated ATPase correlated in dense fractions with plasma membrane markers but a part of it remained at the interphase. This lessdense part of the nigericin-stimulated ATPase could be derived from tonoplast vesicles because α-mannosidase, an enzyme of the vacuolar sap, remained in the upper part of the gradient. Nigericinstimulated ATPase did not correlate with the mitochondrial marker, cytochrome c oxidase, whereas azide inhibition of ATPase activity did.

  4. [Properties and localization of Mg- and Ca-ATpase activities in wheat embryo cell nuclei].

    Science.gov (United States)

    Vasil'eva, N A; Belkina, G G; Stepanenko, S Y; Atalykova, F I; Oparin, A I

    1978-05-01

    The isolated nuclei of wheat embryo possess the ATPase activity. The addition of Mg2+ and Ca2+ significantly increases the activities of nuclear ATPases, whereas Hg2+, Cu2+ and Mn2+ inhibit the activity. The activating effect of Mg2+ is enhanced by an addition of Na and K ions. The activity of wheat embryo nuclear Mg-ATPase is higher than its Ca-ATPase activity; both ATPases also differ in their pH optima. Separation of total nuclear protein according to the solubility of its individual protein components in wheat and strong salt solutions, using the detergents, as well as ammonium sulfate precipitation and dialysis do not result in separation of Mg-activated and Ca-activated ATPases, although their levels of activities and ratios change in the course of fractionation. The Mg- and Ca-ATPase activities of the wheat embryo nuclei were found in the nuclear fraction of albumin, in nonhistone proteins and nuclear membranes. In the albumin nuclear fraction and subfractions of non-histone proteins the higher level of activity is observed in Ca-ATPase, whereas in the nuclei and soluble fractions of residual proteins in Mg-ATPase.

  5. LOCALIZATION OF Na+, K+-ATPASE AND OTHER ENZYMES IN TELEOST PSEUDOBRANCH

    Science.gov (United States)

    Dendy, Leslie A.; Deter, Russell L.; Philpott, Charles W.

    1973-01-01

    In an effort to determine the subcellular localization of sodium- and potassium-activated adenosine triphosphatase (Na+, K+-ATPase) in the pseudobranch of the pinfish Lagodon rhomboides, this tissue was fractionated by differential centrifugation and the activities of several marker enzymes in the fractions were measured. Cytochrome c oxidase was found primarily in the mitochondrial-light mitochondrial (M+L) fraction. Phosphoglucomutase appeared almost exclusively in the soluble (S) fraction. Monoamine oxidase was concentrated in the nuclear (N) fraction, with a significant amount also in the microsomal (P) fraction but little in M+L or S. Na+, K+-ATPase and ouabain insensitive Mg2+-ATPase were distributed in N, M+L, and P, the former having its highest specific activity in P and the latter in M+L. Rate sedimentation analysis of the M+L fraction indicated that cytochrome c oxidase and Mg2+-ATPase were associated with a rapidly sedimenting particle population (presumably mitochondria), while Na+, K+-ATPase was found primarily in a slowly sedimenting component. At least 75% of the Na+, K+-ATPase in M+L appeared to be associated with structures containing no Mg2+-ATPase. Kinetic properties of the two ATPases were studied in the P fraction and were typical of these enzymes in other tissues. Na+, K+-ATPase activity was highly dependent on the ratio of Na+ and K+ concentrations but independent of absolute concentrations over at least a fourfold range. PMID:4349221

  6. Regulation of glycolytic oscillations by mitochondrial and plasma membrane H+-ATPases

    DEFF Research Database (Denmark)

    Olsen, Lars Folke; Andersen, Ann Zahle; Lunding, Anita

    2009-01-01

    ,3'-diethyloxacarbocyanine iodide. The responses of glycolytic and membrane potential oscillations to a number of inhibitors of glycolysis, mitochondrial electron flow, and mitochondrial and plasma membrane H(+)-ATPase were investigated. Furthermore, the glycolytic flux was determined as the rate of production of ethanol....../ATP antiporter and the mitochondrial F(0)F(1)-ATPase. The results further suggest that ATP hydrolysis, through the action of the mitochondrial F(0)F(1)-ATPase and plasma membrane H(+)-ATPase, are important in regulating these oscillations. We conclude that it is glycolysis that drives the oscillations...

  7. Some assembly required: Contributions of Tom Stevens' lab to the V-ATPase field.

    Science.gov (United States)

    Graham, Laurie A; Finnigan, Gregory C; Kane, Patricia M

    2018-06-01

    Tom Stevens' lab has explored the subunit composition and assembly of the yeast V-ATPase for more than 30 years. Early studies helped establish yeast as the predominant model system for study of V-ATPase proton pumps and led to the discovery of protein splicing of the V-ATPase catalytic subunit. The Vma - phenotype, characteristic of loss-of-V-ATPase activity in yeast was key in determining the enzyme's subunit composition via yeast genetics. V-ATPase subunit composition proved to be highly conserved among eukaryotes. Genetic screens for new vma mutants led to identification of a set of dedicated V-ATPase assembly factors and helped unravel the complex pathways for V-ATPase assembly. In later years, exploration of the evolutionary history of several V-ATPase subunits provided new information about the enzyme's structure and function. This review highlights V-ATPase work in the Stevens' lab between 1987 and 2017. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  8. Plasma Membrane ATPase Activity following Reversible and Irreversible Freezing Injury 1

    Science.gov (United States)

    Iswari, S.; Palta, Jiwan P.

    1989-01-01

    Plasma membrane ATPase has been proposed as a site of functional alteration during early stages of freezing injury. To test this, plasma membrane was purified from Solanum leaflets by a single step partitioning of microsomes in a dextran-polyethylene glycol two phase system. Addition of lysolecithin in the ATPase assay produced up to 10-fold increase in ATPase activity. ATPase activity was specific for ATP with a Km around 0.4 millimolar. Presence of the ATPase enzyme was identified by immunoblotting with oat ATPase antibodies. Using the phase partitioning method, plasma membrane was isolated from Solanum commersonii leaflets which had four different degrees of freezing damage, namely, slight (reversible), partial (partially reversible), substantial and total (irreversible). With slight (reversible) damage the plasma membrane ATPase specific activity increased 1.5- to 2-fold and its Km was decreased by about 3-fold, whereas the specific activity of cytochrome c reductase and cytochrome c oxidase in the microsomes were not different from the control. However, with substantial (lethal, irreversible) damage, there was a loss of membrane protein, decrease in plasma membrane ATPase specific activity and decrease in Km, while cytochrome c oxidase and cytochrome c reductase were unaffected. These results support the hypothesis that plasma membrane ATPase is altered by slight freeze-thaw stress. Images Figure 1 Figure 2 PMID:16666856

  9. Abscisic acid induction of vacuolar H+-ATPase activity in mesembryanthemum crystallinum is developmentally regulated

    Science.gov (United States)

    Barkla; Vera-Estrella; Maldonado-Gama; Pantoja

    1999-07-01

    Abscisic acid (ABA) has been implicated as a key component in water-deficit-induced responses, including those triggered by drought, NaCl, and low- temperature stress. In this study a role for ABA in mediating the NaCl-stress-induced increases in tonoplast H+-translocating ATPase (V-ATPase) and Na+/H+ antiport activity in Mesembryanthemum crystallinum, leading to vacuolar Na+ sequestration, were investigated. NaCl or ABA treatment of adult M. crystallinum plants induced V-ATPase H+ transport activity, and when applied in combination, an additive effect on V-ATPase stimulation was observed. In contrast, treatment of juvenile plants with ABA did not induce V-ATPase activity, whereas NaCl treatment resulted in a similar response to that observed in adult plants. Na+/H+ antiport activity was induced in both juvenile and adult plants by NaCl, but ABA had no effect at either developmental stage. Results indicate that ABA-induced changes in V-ATPase activity are dependent on the plant reaching its adult phase, whereas NaCl-induced increases in V-ATPase and Na+/H+ antiport activity are independent of plant age. This suggests that ABA-induced V-ATPase activity may be linked to the stress-induced, developmentally programmed switch from C3 metabolism to Crassulacean acid metabolism in adult plants, whereas, vacuolar Na+ sequestration, mediated by the V-ATPase and Na+/H+ antiport, is regulated through ABA-independent pathways.

  10. Supervision og de tre k´er

    DEFF Research Database (Denmark)

    Schilling, Benedicte; Jacobsen, Claus Haugaard; Nielsen, Jan

    2010-01-01

    Kontrol, kontrakt og kontekst er supervisionens tre k'er. Men hvad er supervision i det hele taget for en størrelse, der spillerså central en rolle for den psykologfaglige profession?......Kontrol, kontrakt og kontekst er supervisionens tre k'er. Men hvad er supervision i det hele taget for en størrelse, der spillerså central en rolle for den psykologfaglige profession?...

  11. En hat er vel en hat ... eller?

    DEFF Research Database (Denmark)

    Bøilerehauge, Dorrit

    2007-01-01

    Mens vi kører i høj fart på "the information highway" - i den helt store bus midt på vejen - glemmer vi ofte noget meget vigtigt i vores glæde over, at verden tilsyneladende åbner sig for os, og vi kan kommunikere med hele verden med ét klik. Vi glemmer, at selvom trafikskilte som oftest er ens -...... - så er selve trafikken og forventningerne blandt medtrafikanterne helt forskellige fra det, vi er vant til. Udgivelsesdato: Februar...

  12. Ytringsfriheden er ikke nødvendig

    DEFF Research Database (Denmark)

    Lauritzen, Pia

    2008-01-01

    nemlig godt konstatere, at ytringsfriheden ikke er nødvendig uden dermed at give afkald på frihed og demokrati.Vi kan oven i købet mene, at ytringsfriheden ophører med at være frihed, når vi betragter den som nødvendig. Og det er lige nøjagtig det, jeg efterlyser: at vi bruger filosoffer som Immanuel...... Kant og Luigi Pareyson til at forstå, at friheden er betydelig mere kompleks, end vi gør den til i øjeblikket....

  13. Hvordan er det å leve med CRPS?

    OpenAIRE

    Hovind, Thomas

    2014-01-01

    Hensikten med denne studien er å se på hvordan pasienter med komplekst regionalt smertesyndrom (CRPS) opplever å leve med tilstanden og hvordan disse pasientene har opplevd å bli møtt i helsevesenet. CRPS er en langvarig tilstand med sterke smerter som kan medføre økt smerterespons, redusert funksjon og nedsatt livskvalitet hos pasientene. Studien er en kvalitativ undersøkelse basert på dybdeintervju av fire pasienter med CRPS. Pasientene ble rekruttert fra en smerteavdeling i Nord-Norge og u...

  14. Branding er ikke til at komme udenom

    DEFF Research Database (Denmark)

    Hermansen, Judy

    2008-01-01

    Søren Askegaard definerer branding som "et iscenesat og afkodet symbolunivers". Det vil sige, at vi på den ene side har virksomhedens strategiske profilering, hvor den forsøger at skabe sig en særlig differentieret og positioneret profil, som adskiller den fra andre virksomheder. "Og det er denne...... strategiske kommunikation, der gør at branding er et ledelsesbegreb og ikke kun et markedsføringsbegreb. Det er ikke staffage, men værdier, som skal gennemsyre alle organisationens aktiviteter." Udgivelsesdato: maj...

  15. Regulation of branchial V-H(+)-ATPase, Na(+)/K(+)-ATPase and NHE2 in response to acid and base infusions in the Pacific spiny dogfish (Squalus acanthias).

    Science.gov (United States)

    Tresguerres, Martin; Katoh, Fumi; Fenton, Heather; Jasinska, Edyta; Goss, Greg G

    2005-01-01

    To study the mechanisms of branchial acid-base regulation, Pacific spiny dogfish were infused intravenously for 24 h with either HCl (495+/- 79 micromol kg(-1) h(-1)) or NaHCO(3) (981+/-235 micromol kg(-1) h(-1)). Infusion of HCl produced a transient reduction in blood pH. Despite continued infusion of acid, pH returned to normal by 12 h. Infusion of NaHCO(3) resulted in a new steady-state acid-base status at approximately 0.3 pH units higher than the controls. Immunostained serial sections of gill revealed the presence of separate vacuolar proton ATPase (V-H(+)-ATPase)-rich or sodium-potassium ATPase (Na(+)/K(+)-ATPase)-rich cells in all fish examined. A minority of the cells also labeled positive for both transporters. Gill cell membranes prepared from NaHCO(3)-infused fish showed significant increases in both V-H(+)-ATPase abundance (300+/-81%) and activity. In addition, we found that V-H(+)-ATPase subcellular localization was mainly cytoplasmic in control and HCl-infused fish, while NaHCO(3)-infused fish demonstrated a distinctly basolateral staining pattern. Western analysis in gill membranes from HCl-infused fish also revealed increased abundance of Na(+)/H(+) exchanger 2 (213+/-5%) and Na(+)/K(+)-ATPase (315+/-88%) compared to the control.

  16. Size of the plasma membrane H+-ATPase from Neurospora crassa determined by radiation inactivation and comparison with the sarcoplasmic reticulum Ca2+-ATPase from skeletal muscle

    International Nuclear Information System (INIS)

    Bowman, B.J.; Berenski, C.J.; Jung, C.Y.

    1985-01-01

    Using radiation inactivation, the authors have measured the size of the H + -ATPase in Neurospora crassa plasma membranes. Membranes were exposed to either high energy electrons from a Van de Graaff generator or to gamma irradiation from 60 Co. Both forms of radiation caused an exponential loss of ATPase activity in parallel with the physical destruction of the Mr = 104,000 polypeptide of which this enzyme is composed. By applying target theory, the size of the H + -ATPase in situ was found to be approximately 2.3 X 10(5) daltons. They also used radiation inactivation to measure the size of the Ca 2+ -ATPase of sarcoplasmic reticulum and got a value of approximately 2.4 X 10(5) daltons, in agreement with previous reports. By irradiating a mixture of Neurospora plasma membranes and rabbit sarcoplasmic reticulum, they directly compared the sizes of these two ATPases and found them to be essentially the same. The authors conclude that both H + -ATPase and Ca 2+ -ATPase are oligomeric enzymes, most likely composed of two approximately 100,000-dalton polypeptides

  17. Alzheimer's Toxic Amyloid Beta Oligomers: Unwelcome Visitors to the Na/K ATPase alpha3 Docking Station.

    Science.gov (United States)

    DiChiara, Thomas; DiNunno, Nadia; Clark, Jeffrey; Bu, Riana Lo; Cline, Erika N; Rollins, Madeline G; Gong, Yuesong; Brody, David L; Sligar, Stephen G; Velasco, Pauline T; Viola, Kirsten L; Klein, William L

    2017-03-01

    Toxic amyloid beta oligomers (AβOs) are known to accumulate in Alzheimer's disease (AD) and in animal models of AD. Their structure is heterogeneous, and they are found in both intracellular and extracellular milieu. When given to CNS cultures or injected ICV into non-human primates and other non-transgenic animals, AβOs have been found to cause impaired synaptic plasticity, loss of memory function, tau hyperphosphorylation and tangle formation, synapse elimination, oxidative and ER stress, inflammatory microglial activation, and selective nerve cell death. Memory loss and pathology in transgenic models are prevented by AβO antibodies, while Aducanumab, an antibody that targets AβOs as well as fibrillar Aβ, has provided cognitive benefit to humans in early clinical trials. AβOs have now been investigated in more than 3000 studies and are widely thought to be the major toxic form of Aβ. Although much has been learned about the downstream mechanisms of AβO action, a major gap concerns the earliest steps: How do AβOs initially interact with surface membranes to generate neuron-damaging transmembrane events? Findings from Ohnishi et al (PNAS 2005) combined with new results presented here are consistent with the hypothesis that AβOs act as neurotoxins because they attach to particular membrane protein docks containing Na/K ATPase-α3, where they inhibit ATPase activity and pathologically restructure dock composition and topology in a manner leading to excessive Ca++ build-up. Better understanding of the mechanism that makes attachment of AβOs to vulnerable neurons a neurotoxic phenomenon should open the door to therapeutics and diagnostics targeting the first step of a complex pathway that leads to neural damage and dementia.

  18. ER Consolidated Quarterly Report October 2014

    Energy Technology Data Exchange (ETDEWEB)

    Cochran, John R. [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States)

    2014-10-01

    This Environmental Restoration Operations (ER) Consolidated Quarterly Report (ER Quarterly Report) provides the status of ongoing corrective actions and related Long- Term Stewardship (LTS) activities being implemented by Sandia National Laboratories, New Mexico (SNL/NM) ER for the April, May, and June 2014 quarterly reporting period. Section 2.0 provides the status of ER Operations activities including closure activities for the Mixed Waste Landfill (MWL), project management and site closure, and hydrogeologic characterizations. Section 3.0 provides the status of LTS activities that relate to the Chemical Waste Landfill (CWL) and the associated Corrective Action Management Unit (CAMU). Section 4.0 provides the references noted in Section I of this report.

  19. Effects of phenol on ATPase activities in crude gill homogenates of rainbow trout (Salmo gairdneri Richardson)

    Energy Technology Data Exchange (ETDEWEB)

    Poston, T.M.

    1979-01-01

    The ATPase specific activities from crude gill homogenates of rainbow trout were lower than those from microsomal preparations reported in the literature. Sodium pump activity (ouabain sensitive NaK-ATPase) was demonstrable at 37/sup 0/C. An ouabain insensitive NaK-ATPase was demonstrable at temperatures below 30/sup 0/C and may represent a Na-ATPase activity reported by others. Energy of activation at 25/sup 0/C for total NaK-ATPase ws 10,500 cal.mole/sup -1/. Mg-baseline activity had an energy of activation at 25/sup 0/C of 15,600 cal.mole/sup -1/. Mg-baseline activity was thermally labile at temperatures in excess of 30/sup 0/C. Concentrations of Mg/sup +2/ in excess of 5 mM appeared to inhibit total NaK-ATPase activity. At 37/sup 0/C, Na/sup +/ and K/sup +/ exerted little, if any, stimulatory effect on ATPase activities, in spite of the fact that 37/sup 0/C was the only temperature at which sodium pump activity was demonstrable. MS-222 failed to produce any discernible changes in any of the demonstrable ATPase activities in crude gill homogenates. Total NaK-ATPase activities were more sensitive than Mg-baseline activities to in vitro inhibition by phenol. Concentrations of phenol which produce 50% inhibition in total NaK-ATPase produced only 35% inhibition in Mg-baseline activity. The nature of in vitro inhibition was uncompetitive. Sodium pump activity was unaffected by phenol at concentrations as high as 25 mM. An effort was made to demonstrate an in vivo effects of phenol on rainbow trout gill ATPase activites. An infestation of a parasite (Gyrodactilus) on the experimental fish precludes any definative assessment of in vivo effects.

  20. Proton accumulation and ATPase activity in Golgi apparatus-enriched vesicles from rat liver

    International Nuclear Information System (INIS)

    Yeh, H.I.; van Rossum, G.D.

    1991-01-01

    We have studied the mechanism by which liver Golgi apparatus maintains the acidity of its contents, using a subcellular fraction from rat liver highly enriched in Golgi marker enzymes. Proton accumulation (measured by quenching of acridine-orange fluorescence) and anion-dependent ATPase were characterized and compared. Maximal ATPase and proton accumulation required ATP; GTP and other nucleotides gave 10% to 30% of maximal activity. Among anions, Cl- and Br- approximately doubled the activities; others were much less effective. Half-maximal increase of ATPase and H+ uptake required 55 mmol/L and 27 mmol/L Cl-, respectively. In predominantly chloride media, SCN- and NO3- markedly inhibited H+ uptake. Nitrate competitively inhibited both the chloride-dependent ATPase (apparent Ki 6 mmol/L) and proton uptake (apparent Ki 2 mmol/L). Nitrate and SCN- also inhibited uptake of 36Cl. Replacing K+ with Na+ had no effect on the initial rate of proton uptake but somewhat reduced the steady state attained. Replacement of K+ with NH4+ and choline reduced proton uptake without affecting ATPase. The ATPase and H+ uptake were supported equally well by Mg2+ or Mn2+. The ATPase was competitively inhibited by 4-acetamido-4'-isothiocyano-stilbene-2,2'-disulfonic acid (apparent Ki 39 mumol/L). Other agents inhibiting both H+ uptake and ATPase were N-ethylmaleimide, N,N'-dicyclohexylcarbodiimide, chlorpromazine, diethylstilbestrol, Zn2+, Co2+ and Cu2+. In the Cl- medium, accumulated protons were released by ionophores at the relative rates, monensin = nigericin greater than valinomycin greater than carbonyl cyanide mchlorophenylhydrazone; the last of these also reduced ATPase activity. In the absence of Cl-, monensin and valinomycin both stimulated the ATPase. These results show a close association between ATPase activity and acidification of liver Golgi vesicles

  1. Vacuolar ATPase regulates surfactant secretion in rat alveolar type II cells by modulating lamellar body calcium.

    Directory of Open Access Journals (Sweden)

    Narendranath Reddy Chintagari

    2010-02-01

    Full Text Available Lung surfactant reduces surface tension and maintains the stability of alveoli. How surfactant is released from alveolar epithelial type II cells is not fully understood. Vacuolar ATPase (V-ATPase is the enzyme responsible for pumping H(+ into lamellar bodies and is required for the processing of surfactant proteins and the packaging of surfactant lipids. However, its role in lung surfactant secretion is unknown. Proteomic analysis revealed that vacuolar ATPase (V-ATPase dominated the alveolar type II cell lipid raft proteome. Western blotting confirmed the association of V-ATPase a1 and B1/2 subunits with lipid rafts and their enrichment in lamellar bodies. The dissipation of lamellar body pH gradient by Bafilomycin A1 (Baf A1, an inhibitor of V-ATPase, increased surfactant secretion. Baf A1-stimulated secretion was blocked by the intracellular Ca(2+ chelator, BAPTA-AM, the protein kinase C (PKC inhibitor, staurosporine, and the Ca(2+/calmodulin-dependent protein kinase II (CaMKII, KN-62. Baf A1 induced Ca(2+ release from isolated lamellar bodies. Thapsigargin reduced the Baf A1-induced secretion, indicating cross-talk between lamellar body and endoplasmic reticulum Ca(2+ pools. Stimulation of type II cells with surfactant secretagogues dissipated the pH gradient across lamellar bodies and disassembled the V-ATPase complex, indicating the physiological relevance of the V-ATPase-mediated surfactant secretion. Finally, silencing of V-ATPase a1 and B2 subunits decreased stimulated surfactant secretion, indicating that these subunits were crucial for surfactant secretion. We conclude that V-ATPase regulates surfactant secretion via an increased Ca(2+ mobilization from lamellar bodies and endoplasmic reticulum, and the activation of PKC and CaMKII. Our finding revealed a previously unrealized role of V-ATPase in surfactant secretion.

  2. Er iPad digital udeskole?

    DEFF Research Database (Denmark)

    Schilhab, Theresa

    2014-01-01

    Hvor ’konkret’ skal en oplevelse være, for at intuitionen om, at det selvoplevede styrker læringen, holder? Er brug af tablets (e.g. iPads) et ligebyrdigt alternativ til den konkrete oplevelse?......Hvor ’konkret’ skal en oplevelse være, for at intuitionen om, at det selvoplevede styrker læringen, holder? Er brug af tablets (e.g. iPads) et ligebyrdigt alternativ til den konkrete oplevelse?...

  3. Snip snap snude - dobbelthagerne er ude

    DEFF Research Database (Denmark)

    Larsen, Malene Charlotte; Kofoed, Jette

    2015-01-01

    Det handler om at være grim. Det står i skarp kontrast til det pæne og polerede på Instagram. Det er flygtigt, useriøst og ligegyldigt. Og så forsvinder det heldigvis igen, så man behøver kun at forholde sig til det i de få sekunder, det foregår. Snapchat vokser i popularitet og er nu det...

  4. Håndbold er en praksis!

    DEFF Research Database (Denmark)

    Olsen, Poul Bitsch

    2008-01-01

    Dansk håndbold er en kæmpemæssig succeshistorie. Ser man snævert på den udvikling, elitedelen af sporten har gennemlevet de seneste 15 år, viser stort set alle tænkelige parametre fremgang. De internationale resultater for lands og klubhold er med få udfald blevet stadigt bedre, omsætningen i klu...

  5. Arctigenin alleviates ER stress via activating AMPK

    Science.gov (United States)

    Gu, Yuan; Sun, Xiao-xiao; Ye, Ji-ming; He, Li; Yan, Shou-sheng; Zhang, Hao-hao; Hu, Li-hong; Yuan, Jun-ying; Yu, Qiang

    2012-01-01

    Aim: To investigate the protective effects of arctigenin (ATG), a phenylpropanoid dibenzylbutyrolactone lignan from Arctium lappa L (Compositae), against ER stress in vitro and the underlying mechanisms. Methods: A cell-based screening assay for ER stress regulators was established. Cell viability was measured using MTT assay. PCR and Western blotting were used to analyze gene and protein expression. Silencing of the CaMKKβ, LKB1, and AMPKα1 genes was achieved by RNA interference (RNAi). An ATP bioluminescent assay kit was employed to measure the intracellular ATP levels. Results: ATG (2.5, 5 and 10 μmol/L) inhibited cell death and unfolded protein response (UPR) in a concentration-dependent manner in cells treated with the ER stress inducer brefeldin A (100 nmol/L). ATG (1, 5 and 10 μmol/L) significantly attenuated protein synthesis in cells through inhibiting mTOR-p70S6K signaling and eEF2 activity, which were partially reversed by silencing AMPKα1 with RNAi. ATG (1-50 μmol/L) reduced intracellular ATP level and activated AMPK through inhibiting complex I-mediated respiration. Pretreatment of cells with the AMPK inhibitor compound C (25 μmol/L) rescued the inhibitory effects of ATG on ER stress. Furthermore, ATG (2.5 and 5 μmol/L) efficiently activated AMPK and reduced the ER stress and cell death induced by palmitate (2 mmol/L) in INS-1 β cells. Conclusion: ATG is an effective ER stress alleviator, which protects cells against ER stress through activating AMPK, thus attenuating protein translation and reducing ER load. PMID:22705729

  6. Bæredygtighed hos SMV'er

    DEFF Research Database (Denmark)

    2016-01-01

    Valg af materiale/medie/form: YouTube Valg af arbejdsform: Videoerne kan bruges som caseundervisning Begrundelse for valg af materiale/medie/form/arbejdsform: Caseundervisning er velegnet til at lære om praksis......Valg af materiale/medie/form: YouTube Valg af arbejdsform: Videoerne kan bruges som caseundervisning Begrundelse for valg af materiale/medie/form/arbejdsform: Caseundervisning er velegnet til at lære om praksis...

  7. Hvad litteraturen kan, er ofte det rigtige ..

    DEFF Research Database (Denmark)

    Bundsgaard, Jeppe; Kühn, Lisbet

    2010-01-01

    Læsning i dansk handler i første omgang om at blive god til at læse, men på mellemtrinnet er det ved at være tiden til også at bruge skønlitteraturen til det, den er god til, nemlig til at få indsigt i menneskelivets kompleksitet. I dette kapitel anbefales det at gøre noget med litteraturen, fx f...

  8. Hvilke Dødehavstekster er "sekteriske"?

    DEFF Research Database (Denmark)

    Holst, Søren

    2003-01-01

    Skønt Dødehavsrullerne antageligt tilhørte en gruppering, der boede i Qumran, tyder alt på, at mange af teksterne er forfattet andre steder. Artiklen undersøger den særlige sprogbrug, som antageligt er typisk for Qumran-samfundet, og som præger forbavsende få af tekserne. En meget stor del af...

  9. Trump er godt nyt for verdensfreden

    DEFF Research Database (Denmark)

    Jakobsen, Peter Viggo

    2017-01-01

    Trumps uberegnelighed øger ikke risikoen for krig. Den mindsker den ved at få de allierede til at gøre mere og modstanderne til at forhandle. Det er godt for verdensfreden......Trumps uberegnelighed øger ikke risikoen for krig. Den mindsker den ved at få de allierede til at gøre mere og modstanderne til at forhandle. Det er godt for verdensfreden...

  10. Characterization of the ER-Targeted Low Affinity Ca2+ Probe D4ER

    Directory of Open Access Journals (Sweden)

    Elisa Greotti

    2016-09-01

    Full Text Available Calcium ion (Ca2+ is a ubiquitous intracellular messenger and changes in its concentration impact on nearly every aspect of cell life. Endoplasmic reticulum (ER represents the major intracellular Ca2+ store and the free Ca2+ concentration ([Ca2+] within its lumen ([Ca2+]ER can reach levels higher than 1 mM. Several genetically-encoded ER-targeted Ca2+ sensors have been developed over the last years. However, most of them are non-ratiometric and, thus, their signal is difficult to calibrate in live cells and is affected by shifts in the focal plane and artifactual movements of the sample. On the other hand, existing ratiometric Ca2+ probes are plagued by different drawbacks, such as a double dissociation constant (Kd for Ca2+, low dynamic range, and an affinity for the cation that is too high for the levels of [Ca2+] in the ER lumen. Here, we report the characterization of a recently generated ER-targeted, Förster resonance energy transfer (FRET-based, Cameleon probe, named D4ER, characterized by suitable Ca2+ affinity and dynamic range for monitoring [Ca2+] variations within the ER. As an example, resting [Ca2+]ER have been evaluated in a known paradigm of altered ER Ca2+ homeostasis, i.e., in cells expressing a mutated form of the familial Alzheimer’s Disease-linked protein Presenilin 2 (PS2. The lower Ca2+ affinity of the D4ER probe, compared to that of the previously generated D1ER, allowed the detection of a conspicuous, more clear-cut, reduction in ER Ca2+ content in cells expressing mutated PS2, compared to controls.

  11. Expression of a prokaryotic P-type ATPase in E. coli Plasma Membranes and Purification by Ni2+-affinity chromatography

    Directory of Open Access Journals (Sweden)

    Geisler Markus

    1998-01-01

    Full Text Available In order to characterize the P-type ATPase from Synechocystis 6803 [Geisler (1993 et al. J. Mol. Biol. 234, 1284] and to facilitate its purification, we expressed an N-terminal 6xHis-tagged version of the ATPase in an ATPase deficient E. coli strain. The expressed ATPase was immunodetected as a dominant band of about 97 kDa localized to the E. coli plasma membranes representing about 20-25% of the membrane protein. The purification of the Synecho-cystis 6xHis-ATPase by single-step Ni-affinity chromatography under native and denaturating conditions is described. ATPase activity and the formation of phosphointermediates verify the full function of the enzyme: the ATPase is inhibited by vanadate (IC50= 119 &mgr;M and the formation of phosphorylated enzyme intermediates shown by acidic PAGE depends on calcium, indicating that the Synechocystis P-ATPase functions as a calcium pump.

  12. Relationship between serum adiponectin level and ATPase activity of erythrocyte membrance in patients with 2-type diabetes

    International Nuclear Information System (INIS)

    Song Jiejin

    2008-01-01

    Objective: To explore the possible mechanism of development nephrosis as related to changes of serum adiponectin levels and alteration of activities of Na + ·K + -ATPase and Ca +2 ·Mg +2 -ATPase of erythrocyte membrance in patients with 2-type diabetes. Methods: Serum adiponectin levels (with RIA) and erythrocyte membrance (prepared with Reilnila method) Na + ·K + - ATPase and Ca +2 ·Mg +2 -ATPase activity were determined in 45 DM2 patients without nephropathy, 31 DM2 patients with nephropathy and 30 controls. Results: Serum adiponectin levels in the diabetic patients were significantly lower than those in controls (P + ·K + -ATPase and Ca +2 ·Mg +2 -ATPase activities were also significantly lower than those in controls (P + ·K + -ATPase and Ca +2 ·Mg +2 -ATPase activities of erythrocyte membrance. (authors)

  13. ER-to-plasma membrane tethering proteins regulate cell signaling and ER morphology.

    Science.gov (United States)

    Manford, Andrew G; Stefan, Christopher J; Yuan, Helen L; Macgurn, Jason A; Emr, Scott D

    2012-12-11

    Endoplasmic reticulum-plasma membrane (ER-PM) junctions are conserved structures defined as regions of the ER that tightly associate with the plasma membrane. However, little is known about the mechanisms that tether these organelles together and why such connections are maintained. Using a quantitative proteomic approach, we identified three families of ER-PM tethering proteins in yeast: Ist2 (related to mammalian TMEM16 ion channels), the tricalbins (Tcb1/2/3, orthologs of the extended synaptotagmins), and Scs2 and Scs22 (vesicle-associated membrane protein-associated proteins). Loss of all six tethering proteins results in the separation of the ER from the PM and the accumulation of cytoplasmic ER. Importantly, we find that phosphoinositide signaling is misregulated at the PM, and the unfolded protein response is constitutively activated in the ER in cells lacking ER-PM tether proteins. These results reveal critical roles for ER-PM contacts in cell signaling, organelle morphology, and ER function. Copyright © 2012 Elsevier Inc. All rights reserved.

  14. Cyclophilin B interacts with sodium-potassium ATPase and is required for pump activity in proximal tubule cells of the kidney.

    Directory of Open Access Journals (Sweden)

    Guillermo Suñé

    Full Text Available Cyclophilins (Cyps, the intracellular receptors for Cyclosporine A (CsA, are responsible for peptidyl-prolyl cis-trans isomerisation and for chaperoning several membrane proteins. Those functions are inhibited upon CsA binding. Albeit its great benefits as immunosuppressant, the use of CsA has been limited by undesirable nephrotoxic effects, including sodium retention, hypertension, hyperkalemia, interstial fibrosis and progressive renal failure in transplant recipients. In this report, we focused on the identification of novel CypB-interacting proteins to understand the role of CypB in kidney function and, in turn, to gain further insight into the molecular mechanisms of CsA-induced toxicity. By means of yeast two-hybrid screens with human kidney cDNA, we discovered a novel interaction between CypB and the membrane Na/K-ATPase β1 subunit protein (Na/K-β1 that was confirmed by pull-down, co-immunoprecipitation and confocal microscopy, in proximal tubule-derived HK-2 cells. The Na/K-ATPase pump, a key plasma membrane transporter, is responsible for maintenance of electrical Na+ and K+ gradients across the membrane. We showed that CypB silencing produced similar effects on Na/K-ATPase activity than CsA treatment in HK-2 cells. It was also observed an enrichment of both alpha and beta subunits in the ER, what suggested a possible failure on the maturation and routing of the pump from this compartment towards the plasma membrane. These data indicate that CypB through its interaction with Na/K-β1 might regulate maturation and trafficking of the pump through the secretory pathway, offering new insights into the relationship between cyclophilins and the nephrotoxic effects of CsA.

  15. Cyclophilin B interacts with sodium-potassium ATPase and is required for pump activity in proximal tubule cells of the kidney.

    Science.gov (United States)

    Suñé, Guillermo; Sarró, Eduard; Puigmulé, Marta; López-Hellín, Joan; Zufferey, Madeleine; Pertel, Thomas; Luban, Jeremy; Meseguer, Anna

    2010-11-10

    Cyclophilins (Cyps), the intracellular receptors for Cyclosporine A (CsA), are responsible for peptidyl-prolyl cis-trans isomerisation and for chaperoning several membrane proteins. Those functions are inhibited upon CsA binding. Albeit its great benefits as immunosuppressant, the use of CsA has been limited by undesirable nephrotoxic effects, including sodium retention, hypertension, hyperkalemia, interstial fibrosis and progressive renal failure in transplant recipients. In this report, we focused on the identification of novel CypB-interacting proteins to understand the role of CypB in kidney function and, in turn, to gain further insight into the molecular mechanisms of CsA-induced toxicity. By means of yeast two-hybrid screens with human kidney cDNA, we discovered a novel interaction between CypB and the membrane Na/K-ATPase β1 subunit protein (Na/K-β1) that was confirmed by pull-down, co-immunoprecipitation and confocal microscopy, in proximal tubule-derived HK-2 cells. The Na/K-ATPase pump, a key plasma membrane transporter, is responsible for maintenance of electrical Na+ and K+ gradients across the membrane. We showed that CypB silencing produced similar effects on Na/K-ATPase activity than CsA treatment in HK-2 cells. It was also observed an enrichment of both alpha and beta subunits in the ER, what suggested a possible failure on the maturation and routing of the pump from this compartment towards the plasma membrane. These data indicate that CypB through its interaction with Na/K-β1 might regulate maturation and trafficking of the pump through the secretory pathway, offering new insights into the relationship between cyclophilins and the nephrotoxic effects of CsA.

  16. Asn792 participates in the hydrogen bond network around the K+-binding pocket of gastric H,K-ATPase.

    NARCIS (Netherlands)

    Swarts, H.G.P.; Koenderink, J.B.; Willems, P.H.G.M.; Krieger, E.; Pont, J.J.H.H.M. de

    2005-01-01

    Asn792 present in M5 of gastric H,K-ATPase is highly conserved within the P-type ATPase family. A direct role in K+ binding was postulated for Na,K-ATPase but was not found in a recent model for gastric H,K-ATPase (Koenderink, J. B., Swarts, H. G. P., Willems, P. H. G. M., Krieger, E., and De Pont,

  17. Carbonylation Modification Regulates Na/K-ATPase Signaling and Salt Sensitivity: A Review and a Hypothesis.

    Science.gov (United States)

    Shah, Preeya T; Martin, Rebecca; Yan, Yanling; Shapiro, Joseph I; Liu, Jiang

    2016-01-01

    Na/K-ATPase signaling has been implicated in different physiological and pathophysiological conditions. Accumulating evidence indicates that oxidative stress not only regulates the Na/K-ATPase enzymatic activity, but also regulates its signaling and other functions. While cardiotonic steroids (CTS)-induced increase in reactive oxygen species (ROS) generation is an intermediate step in CTS-mediated Na/K-ATPase signaling, increase in ROS alone also stimulates Na/K-ATPase signaling. Based on literature and our observations, we hypothesize that ROS have biphasic effects on Na/K-ATPase signaling, transcellular sodium transport, and urinary sodium excretion. Oxidative modulation, in particular site specific carbonylation of the Na/K-ATPase α1 subunit, is a critical step in proximal tubular Na/K-ATPase signaling and decreased transcellular sodium transport leading to increases in urinary sodium excretion. However, once this system is overstimulated, the signaling, and associated changes in sodium excretion are blunted. This review aims to evaluate ROS-mediated carbonylation of the Na/K-ATPase, and its potential role in the regulation of pump signaling and sodium reabsorption in the renal proximal tubule (RPT).

  18. Nitric oxide and Na,K-ATPase activity in rat skeletal muscle.

    Science.gov (United States)

    Juel, C

    2016-04-01

    It has been suggested that nitric oxide (NO) stimulates the Na,K-ATPase in cardiac myocytes. Therefore, the aims of this study were to investigate whether NO increases Na,K-ATPase activity in skeletal muscle and, if that is the case, to identify the underlying mechanism. The study used isolated rat muscle, muscle homogenates and purified membranes as model systems. Na,K-ATPase activity was quantified from phosphate release due to ATP hydrolysis. Exposure to the NO donor spermine NONOate (10 μm) increased the maximal Na,K-ATPase activity by 27% in isolated glycolytic muscles, but had no effect in oxidative muscles. Spermine NONOate increased the maximal Na,K-ATPase activity by 58% (P Na,K-ATPase α-isoform. Incubation with cGMP (1 mm) increased the maximal Na,K-ATPase activity in homogenates from glycolytic muscle by 16% (P Na,K-ATPase in glycolytic skeletal muscle. Direct S-nitrosylation and interference with S-glutathionylation seem to be excluded. In addition, phosphorylation of phospholemman at serine 68 is not involved. Most likely, the NO/cGMP/protein kinase G signalling pathway is involved. © 2015 Scandinavian Physiological Society. Published by John Wiley & Sons Ltd.

  19. Electrophysiological analysis of the mutated Na,K-ATPase cation binding pocket.

    NARCIS (Netherlands)

    Koenderink, J.B.; Geibel, S.; Grabsch, E.; Pont, J.J.H.H.M. de; Bamberg, E.; Friedrich, T.

    2003-01-01

    Na,K-ATPase mediates net electrogenic transport by extruding three Na+ ions and importing two K+ ions across the plasma membrane during each reaction cycle. We mutated putative cation coordinating amino acids in transmembrane hairpin M5-M6 of rat Na,K-ATPase: Asp776 (Gln, Asp, Ala), Glu779 (Asp,

  20. Excess capacity of H+ ATPase and inverse respiratory control in Escherichia coli

    DEFF Research Database (Denmark)

    Jensen, Peter Ruhdal; Westerhoff, Hans V.; Michelsen, Ole

    1993-01-01

    With succinate as free-energy source, Escherichia coli generating virtually all ATP by oxidative phosphorylation might be expected heavily to tax its ATP generating capacity. To examine this the H+-ATPase (ATP synthase) was modulated over a 30-fold range. Decreasing the amount of H+-ATPase reduce...

  1. Membrane-bound ATPase contributes to hop resistance of Lactobacillus brevis

    NARCIS (Netherlands)

    Sakamoto, K; van Veen, HW; Saito, H; Kobayashi, H; Konings, WN

    2002-01-01

    The activity of the membrane-bound H+-ATPase of the beer spoilage bacterium Lactobacillus brevis ABBC45 increased upon adaptation to bacteriostatic hop compounds. The ATPase activity was optimal around pH 5.6 and increased up to fourfold when L. brevis was exposed to 666 muM hop compounds. The

  2. Study on the changes in the levels of membrane-bound ATPases ...

    African Journals Online (AJOL)

    An attempt has been made to determine the deleterious effects of λ cyhalothrin- induced in fresh water tilapia (Oreochromis mossambicus) with respect to changes in the activities of membrane-bound ATPases (Na+/K+, Mg+ and Ca2+ ATPase) and mineral status ...

  3. Purification and functional motifs of the recombinant ATPase of orf virus.

    Science.gov (United States)

    Lin, Fong-Yuan; Chan, Kun-Wei; Wang, Chi-Young; Wong, Min-Liang; Hsu, Wei-Li

    2011-10-01

    Our previous study showed that the recombinant ATPase encoded by the A32L gene of orf virus displayed ATP hydrolysis activity as predicted from its amino acids sequence. This viral ATPase contains four known functional motifs (motifs I-IV) and a novel AYDG motif; they are essential for ATP hydrolysis reaction by binding ATP and magnesium ions. The motifs I and II correspond with the Walker A and B motifs of the typical ATPase, respectively. To examine the biochemical roles of these five conserved motifs, recombinant ATPases of five deletion mutants derived from the Taiping strain were expressed and purified. Their ATPase functions were assayed and compared with those of two wild type strains, Taiping and Nantou isolated in Taiwan. Our results showed that deletions at motifs I-III or IV exhibited lower activity than that of the wild type. Interestingly, deletion of AYDG motif decreased the ATPase activity more significantly than those of motifs I-IV deletions. Divalent ions such as magnesium and calcium were essential for ATPase activity. Moreover, our recombinant proteins of orf virus also demonstrated GTPase activity, though weaker than the original ATPase activity. Copyright © 2011 Elsevier Inc. All rights reserved.

  4. Further investigations on the inorganic phosphate binding site of beef heart mitochondrial F1-ATPase

    International Nuclear Information System (INIS)

    Pougeois, R.; Lauquin, G.J.

    1985-01-01

    The possibility that 4-azido-2-nitrophenyl phosphate (ANPP), a photoreactive derivative of inorganic phosphate (P /sub i/ ), could mimic ATP was investigated. ANPP was hydrolyzed in the dark by sarcoplasmic reticulum Ca 2+ -ATPase in the presence of Ca 2+ but not in the presence of ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid. ANPP was not hydrolyzed by purified mitochondrial F1-ATPase; however, ADP and ATP protected F1-ATPase against ANPP photoinactivation. On the other hand, the trinitrophenyl nucleotide analogues (TNP-ADP, TNP-ATP, and TNP-AMP-PNP), which bind specifically at the two catalytic sites of F1-ATPase, abolished P /sub i/ binding on F1-ATPase; they do not protect F1-ATPase against ANPP photoinactivation. Furthermore, ANPP-photoinactivated F1-ATPase binds the TNP analogues in the same way as the native enzyme. The Pi binding site of F1-ATPase, which is shown to be photolabeled by ANPP, does not appear to be at the gamma-phosphate position of the catalytic sites

  5. Mammary gland involution is associated with rapid down regulation of major mammary Ca**2+-ATPases

    Science.gov (United States)

    Sixty percent of calcium in milk is transported across the mammary cells apical membrane by the plasma membrane Ca**2+-ATPase 2 (PMCA2). The effect of abrupt cessation of milk production on the Ca**2+-ATPases and mammary calcium transport is unknown. We found that 24 hours after stopping milk prod...

  6. Two types of essential carboxyl groups in Rhodospirillum rubrum proton ATPase

    International Nuclear Information System (INIS)

    Ceccarelli, E.; Vallejos, R.H.

    1983-01-01

    Two different types of essential carboxyl groups were detected in the extrinsic component of the proton ATPase of Rhodospirillum rubrum. Chemical modification of R. rubrum chromatophores or its solubilized ATPase by Woodward's reagent K resulted in inactivation of photophosphorylating and ATPase activities. The apparent order of reaction was nearly 1 with respect to reagent concentration and similar K1 were obtained for the soluble and membrane-bound ATPases suggesting that inactivation was associated with modification of one essential carboxyl group located in the soluble component of the proton ATPase. Inactivation was prevented by adenine nucleotides but not by divalent cations. Dicyclohexylcarbodiimide completely inhibited the solubilized ATPase with a K1 of 5.2 mM and a K2 of 0.81 min-1. Mg2+ afforded nearly complete protection with a Kd of 2.8 mM. Two moles of [14C]dicyclohexylcarbodiimide were incorporated per mole of enzyme for complete inactivation but in the presence of 30 mM MgCl2 only one mole was incorporated and there was no inhibition. The labeling was recovered mostly from the beta subunit. The incorporation of the labeled reagent into the ATPase was not prevented by previous modification with Woodward's reagent K. It is concluded that both reagents modified two different essential carboxyl groups in the soluble ATPase from R. rubrum

  7. Modulation of FXYD interaction with Na,K-ATPase by anionic phospholipids and protein kinase phosphorylation

    DEFF Research Database (Denmark)

    Cornelius, Flemming; Mahmmoud, Yasser Ahmed

    2007-01-01

    acids of FXYD10 had been cleaved by mild, controlled trypsin treatment. Several kinetic properties of the Na,K-ATPase reaction cycle as well as the FXYD-regulation of Na,K-ATPase activity were found to be affected by acidic phospholipids like PI, PS, and PG. This takes into consideration the Na+ and K...

  8. Cation Transport Coupled to ATP Hydrolysis by the (Na, K)-ATPase: An Integrated, Animated Model

    Science.gov (United States)

    Leone, Francisco A.; Furriel, Rosa P. M.; McNamara, John C.; Horisberger, Jean D.; Borin, Ivana A.

    2010-01-01

    An Adobe[R] animation is presented for use in undergraduate Biochemistry courses, illustrating the mechanism of Na[superscript +] and K[superscript +] translocation coupled to ATP hydrolysis by the (Na, K)-ATPase, a P[subscript 2c]-type ATPase, or ATP-powered ion pump that actively translocates cations across plasma membranes. The enzyme is also…

  9. Disease mutations reveal residues critical to the interaction of P4-ATPases with lipid substrates

    DEFF Research Database (Denmark)

    Gantzel, Rasmus H; Mogensen, Louise S; Mikkelsen, Stine A

    2017-01-01

    Phospholipid flippases (P4-ATPases) translocate specific phospholipids from the exoplasmic to the cytoplasmic leaflet of membranes. While there is good evidence that the overall molecular structure of flippases is similar to that of P-type ATPase ion-pumps, the transport pathway for the "giant...

  10. Hemin reconstitutes proton extrusion in an H+-ATPase-negative mutant of Lactococcus lactis

    DEFF Research Database (Denmark)

    Blank, L.M.; Købmann, Brian Jensen; Michelsen, Ole

    2001-01-01

    H+-ATPase is considered essential for growth of Lactococcus lactis. However, media containing hemin restored the aerobic growth of an H+-ATPase-negative mutant, suggesting that hemin complements proton extrusion. We show that inverted membrane vesicles prepared from hemin-grown L. lactis cells...

  11. Na,K-ATPase activity modulates Src activation: A role for ATP/ADP ratio.

    NARCIS (Netherlands)

    Weigand, K.M.; Swarts, H.G.P.; Fedosova, N.U.; Russel, F.G.M.; Koenderink, J.B.

    2012-01-01

    Digitalis-like compounds (DLCs), specific inhibitors of Na,K-ATPase, are implicated in cellular signaling. Exposure of cell cultures to ouabain, a well-known DLC, leads to up- or down regulation of various processes and involves activation of Src kinase. Since Na,K-ATPase is the only known target

  12. Hierarchy of mechanisms involved in generating Na/K-ATPase polarity in MDCK epithelial cells

    NARCIS (Netherlands)

    Mays, R.W.; Siemers, K.A.; Fritz, B.A.; Lowe, A.W.; van Meer, G.; Nelson, W.J.

    1995-01-01

    We have studied mechanisms involved in generating a polarized distribution of Na/K-ATPase in the basal-lateral membrane of two clones of MDCK II cells. Both clones exhibit polarized distributions of marker proteins of the apical and basal-lateral membranes, including Na/K-ATPase, at steady state.

  13. MgATP hydrolysis destabilizes the interaction between subunit H and yeast V1-ATPase, highlighting H's role in V-ATPase regulation by reversible disassembly.

    Science.gov (United States)

    Sharma, Stuti; Oot, Rebecca A; Wilkens, Stephan

    2018-05-12

    Vacuolar H+-ATPases (V-ATPases; V1Vo-ATPases) are rotary motor proton pumps that acidify intracellular compartments and in some tissues, the extracellular space. V-ATPase is regulated by reversible disassembly into autoinhibited V1-ATPase and Vo proton channel sectors. An important player in V-ATPase regulation is subunit H, which binds at the interface of V1 and Vo. H is required for MgATPase activity in holo V-ATPase, but also for stabilizing the MgADP inhibited state in membrane detached V1. However, how H fulfills these two functions is poorly understood. To characterize the H-V1 interaction and its role in reversible disassembly, we determined binding affinities of full length H and its N-terminal domain (HNT) for an isolated heterodimer of subunits E and G (EG), the N-terminal domain of subunit a (aNT), and V1 lacking subunit H (V1ΔH). Using isothermal titration calorimetry (ITC) and biolayer interferometry (BLI), we show that HNT binds EG with moderate affinity, that full length H binds aNT weakly, and that both H and HNT bind V1ΔH with high affinity. We also found that only one molecule of HNT binds V1ΔH with high affinity, suggesting conformational asymmetry of the three EG heterodimers in V1ΔH. Moreover, MgATP hydrolysis-driven conformational changes in V1 destabilized the interaction of H, or HNT, with V1ΔH, suggesting an interplay between MgADP inhibition and subunit H. Our observation that H binding is affected by MgATP hydrolysis in V1 points to H's role in the mechanism of reversible disassembly. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.

  14. Direct interaction of the Golgi V-ATPase a-subunit isoform with PI(4)P drives localization of Golgi V-ATPases in yeast.

    Science.gov (United States)

    Banerjee, Subhrajit; Kane, Patricia M

    2017-09-15

    Luminal pH and phosphoinositide content are fundamental features of organelle identity. Vacuolar H + -ATPases (V-ATPases) drive organelle acidification in all eukaryotes, and membrane-bound a-subunit isoforms of the V-ATPase are implicated in organelle-specific targeting and regulation. Earlier work demonstrated that the endolysosomal lipid PI(3,5)P 2 activates V-ATPases containing the vacuolar a-subunit isoform in Saccharomyces cerevisiae Here we demonstrate that PI(4)P, the predominant Golgi phosphatidylinositol (PI) species, directly interacts with the cytosolic amino terminal (NT) domain of the yeast Golgi V-ATPase a-isoform Stv1. Lysine-84 of Stv1NT is essential for interaction with PI(4)P in vitro and in vivo, and interaction with PI(4)P is required for efficient localization of Stv1-containing V-ATPases. The cytosolic NT domain of the human V-ATPase a2 isoform specifically interacts with PI(4)P in vitro, consistent with its Golgi localization and function. We propose that NT domains of V o a-subunit isoforms interact specifically with PI lipids in their organelles of residence. These interactions can transmit organelle-specific targeting or regulation information to V-ATPases. © 2017 Banerjee and Kane. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

  15. The influence of blood plasma of irradiated animals on activity of Ca2+ - ATPase and Mg2+ - ATPase in plasma membrane of thymocytes

    International Nuclear Information System (INIS)

    Dreval', V.I.

    1994-01-01

    Rats were irradiated at doses 1.5, 4.0, 7.0 and 10 Gy. After 1, 8, 15, 22 and 30 days the effect of blood plasma on activity of Ca 2+ -ATPase and Mg 2+ -ATPase in plasma membrane of thymocytes was investigated. It was found that the raise of irradiation dose leads to increasing of blood plasma effect on membrane-bound enzymes

  16. In and out of the cation pumps: P-type ATPase structure revisited

    DEFF Research Database (Denmark)

    Bublitz, Maike; Poulsen, Hanne; Morth, Jens Preben

    2010-01-01

    . The marked increment during the last three years in the number of crystal structures of P-type ATPases has greatly improved our understanding of the similarities and differences of pumps with different ion specificities, since the structures of the Ca2+-ATPase, the Na+,K+-ATPase and the H+-ATPase can now......Active transport across membranes is a crucial requirement for life. P-type ATPases build up electrochemical gradients at the expense of ATP by forming and splitting a covalent phosphoenzyme intermediate, coupled to conformational changes in the transmembrane section where the ions are translocated...... be compared directly. Mechanisms for ion gating, charge neutralization and backflow prevention are starting to emerge from comparative structural analysis; and in combination with functional studies of mutated pumps this provides a framework for speculating on how the ions are bound and released as well...

  17. The Structure and Function of the Na,K-ATPase Isoforms in Health and Disease

    Directory of Open Access Journals (Sweden)

    Michael V. Clausen

    2017-06-01

    Full Text Available The sodium and potassium gradients across the plasma membrane are used by animal cells for numerous processes, and the range of demands requires that the responsible ion pump, the Na,K-ATPase, can be fine-tuned to the different cellular needs. Therefore, several isoforms are expressed of each of the three subunits that make a Na,K-ATPase, the alpha, beta and FXYD subunits. This review summarizes the various roles and expression patterns of the Na,K-ATPase subunit isoforms and maps the sequence variations to compare the differences structurally. Mutations in the Na,K-ATPase genes encoding alpha subunit isoforms have severe physiological consequences, causing very distinct, often neurological diseases. The differences in the pathophysiological effects of mutations further underline how the kinetic parameters, regulation and proteomic interactions of the Na,K-ATPase isoforms are optimized for the individual cellular needs.

  18. The Structure and Function of the Na,K-ATPase Isoforms in Health and Disease.

    Science.gov (United States)

    Clausen, Michael V; Hilbers, Florian; Poulsen, Hanne

    2017-01-01

    The sodium and potassium gradients across the plasma membrane are used by animal cells for numerous processes, and the range of demands requires that the responsible ion pump, the Na,K-ATPase, can be fine-tuned to the different cellular needs. Therefore, several isoforms are expressed of each of the three subunits that make a Na,K-ATPase, the alpha, beta and FXYD subunits. This review summarizes the various roles and expression patterns of the Na,K-ATPase subunit isoforms and maps the sequence variations to compare the differences structurally. Mutations in the Na,K-ATPase genes encoding alpha subunit isoforms have severe physiological consequences, causing very distinct, often neurological diseases. The differences in the pathophysiological effects of mutations further underline how the kinetic parameters, regulation and proteomic interactions of the Na,K-ATPase isoforms are optimized for the individual cellular needs.

  19. Functional analysis of a potential regulatory K+-binding site in the Na+, K+-ATPase

    DEFF Research Database (Denmark)

    Schack, Vivien Rodacker; Vilsen, Bente

    The Na+, K+-ATPase functions by actively transporting 3 Na+ ions out of and 2 K+ ions into the cell, thereby creating ion gradients crucial for many physiological processes. Recently, a combined structural and functional study of the closely related Ca2+-ATPase indicated the presence...... of a regulatory K+-binding site in the P-domain of the enzyme, identifying E732 as being of particular importance (Sorensen, Clausen et al. 2004). In addition, P709 is thought to play a significant role in the structural organization of this site. Both E732 and P709 are highly conserved among P-type ATPases (E732...... is present as either glutamic acid or aspartic acid), which supports their importance and additionally raises the question whether this site may play a general role among P-type ATPases. In Na+, K+-ATPase, K+ functions directly as a substrate for membrane binding sites, however, an additional regulatory...

  20. Phosphorylation of plant plasma membrane H+-ATPase by the heterologous host S. cerevisiae

    DEFF Research Database (Denmark)

    Rudashevskaya, Elena; Ye, Juanying; Young, Clifford

     It is known, that phosphorylation of both plant and yeast plasma membrane H+-ATPase results in enzyme activation or inhibition. Several sites at the regulatory C-terminus of the enzyme have been found to undergo phosphorylation in vivo in both plant and yeast. The C-termini of plant H...... of heterologous system of yeast cells, expressing plant proton pump. Therefore identification of possible regulatory effects by phosphorylation events in plant H+-ATPase in the system is significant. A number of putative phosphorylation sites at regulatory C-domain of H+-ATPase (AHA2) have been point...... functioning of the residues and suggests, that plant H+-ATPase could be regulated by phosphorylation at several sites being in yeast cells. Plant H+-ATPase purified from yeast cells by his-tag affinity chromatography was subjected to IMAC and TiO2 for enrichment of phosphopeptides. The phosphopeptides were...

  1. Nitric oxide and Na,K-ATPase activity in rat skeletal muscle

    DEFF Research Database (Denmark)

    Juel, Carsten

    2016-01-01

    Aim: It has been suggested that nitric oxide (NO) stimulates the Na,K-ATPase in cardiac myocytes. Therefore, the aims of this study were to investigate whether NO increases Na,K-ATPase activity in skeletal muscle and, if that is the case, to identify the underlying mechanism. Method: The study used...... isolated rat muscle, muscle homogenates and purified membranes as model systems. Na,K-ATPase activity was quantified from phosphate release due to ATP hydrolysis. Results: Exposure to the NO donor spermine NONOate (10 μm) increased the maximal Na,K-ATPase activity by 27% in isolated glycolytic muscles...... activity was depressed by oxidized glutathione. Conclusion: NO and cGMP stimulate the Na,K-ATPase in glycolytic skeletal muscle. Direct S-nitrosylation and interference with S-glutathionylation seem to be excluded. In addition, phosphorylation of phospholemman at serine 68 is not involved. Most likely...

  2. Increasing acidification of nonreplicating Lactococcus lactis Delta thyA mutants by incorporating ATPase activity

    DEFF Research Database (Denmark)

    Pedersen, Martin Bastian; Købmann, Brian Jensen; Jensen, Peter Ruhdal

    2002-01-01

    % of that of exponentially growing MBP71. However, when nonspecific ATPase activity was incorporated into MBP71, the lactic acid flux was restored to 100% but not above that point, indicating that control over the flux switched from ATP demand to ATP supply (i.e., to sugar transport and glycolysis). As determined by growing...... nonreplicating cells with high ATPase activity on various sugar sources, it appeared that glycolysis exerted the majority of the control. ATPase activity also stimulated the rate of acidification by noureplicating MBP71 growing in milk, and pH 5.2 was reached 40% faster than it was without ATPase activity. We...... concluded that ATPase activity is a functional means of increasing acidification by nonreplicating L. lactis....

  3. In vitro effects of toxaphene on mitochondrial calcium ATPase and calcium uptake in selected rat tissues

    International Nuclear Information System (INIS)

    Trottman, C.H.; Rao, K.S.P.; Morrow, W.; Uzodinma, J.E.; Desaiah, D.

    1985-01-01

    In vitro effects of toxaphene on Ca 2+ -ATPase activity and 45 Ca 2+ -uptake were studied in mitochondrial fractions of heart, kidney and liver tissues of rat. Mitochondrial fractions were prepared by the conventional centrifugation method. Ca 2+ -ATPase activity was determined by measuring the inorganic phosphate liberated during ATP hydrolysis. Toxaphene inhibited Ca 2+ -ATPase in a concentration dependent manner in all the three tissues. Substrate activation kinetics, with heart, kidney and liver tissue fractions, revealed that toxaphene inhibited Ca 2+ -ATPase activity non-competetively by decreasing the maximum velocity of the enzyme without affecting the enzyme-substrate affinity. Toxaphene also inhibited mitochondrial 45 Ca 2+ -uptake in the three selected tissues in a concentration dependent manner. These results indicate that toxaphene is an inhibitor of mitochondrial Ca 2+ -ATPase and calcium transport in heart, kidney and liver tissues of rat. 19 references, 5 figures

  4. The role of Na,K-ATPase/Src-kinase signaling pathway in the vascular wall contaction

    DEFF Research Database (Denmark)

    Bouzinova, Elena

    Aim: Na,K-ATPase is essential for maintaining the transmembrane ion gradient and might initiate various intracellular signaling. These signals possibly act through a modification of the local ion concentrations or via Src-kinase activation. It is known that inhibition of the α-2 isoform of Na......,K-ATPase by ouabain elevates blood pressure. Consequently, ouabain was shown to potentiate arterial contraction in vitro. In contrast, we have demonstrated that siRNA-induced down-regulation of the α-2 isoform Na,K-ATPase expression reduced arterial sensitivity to agonist stimulation and prevented the effect......) phosphorylation assay. Down-regulation of the α-2 isoform Na,K-ATPase prevented the inhibitory effect of Src inhibitors on arterial contraction. Conclusions: The pro-contractile action of ouabain-sensitive Na,K-ATPase inhibition is associated with Src-kinase inhibition suggesting the role of this signaling...

  5. Effect of hindlimb unweighting on single soleus fiber maximal shortening velocity and ATPase activity

    Science.gov (United States)

    Mcdonald, K. S.; Fitts, R. H.

    1993-01-01

    The effect of hindlimb unweighting (HU) for 1 to 3 wks on the shortening velocity of a soleus fiber, its ATPase content, and the relative contents of the slow and fast myosin was investigated by measuring fiber force, V(0), ATPase activity, and myosin content in SDS protein profiles of a single rat soleus fiber suspended between a motor arm and a transducer. It was found that HU induces a progressive increase in fiber V(0) that is likely caused, at least in part, by an increase in the fiber's myofibrillar ATPase activity. The HU-induced increases in V(0) and ATPase were associated with the presence of a greater percentage of fast type IIa fibers. However, a large population of fibers after 1, 2, and 3 wks of HU showed increases in V(0) and ATPase but displayed the same myosin protein profile on SDS gels as control fibers.

  6. Glutamate transporter activity promotes enhanced Na+/K+-ATPase -mediated extracellular K+ management during neuronal activity

    DEFF Research Database (Denmark)

    Larsen, Brian R; Holm, Rikke; Vilsen, Bente

    2016-01-01

    , in addition, Na+ /K+ -ATPase-mediated K+ clearance could be governed by astrocytic [Na+ ]i . During most neuronal activity, glutamate is released in the synaptic cleft and is re-absorbed by astrocytic Na+ -coupled glutamate transporters, thereby elevating [Na+ ]i . It thus remains unresolved whether...... the different Na+ /K+ -ATPase isoforms are controlled by [K+ ]o or [Na+ ]i during neuronal activity. Hippocampal slice recordings of stimulus-induced [K+ ]o transients with ion-sensitive microelectrodes revealed reduced Na+ /K+ -ATPase-mediated K+ management upon parallel inhibition of the glutamate transporter......+ affinity to the α1 and α2 isoforms than the β2 isoform. In summary, enhanced astrocytic Na+ /K+ -ATPase-dependent K+ clearance was obtained with parallel glutamate transport activity. The astrocytic Na+ /K+ -ATPase isoform constellation α2β1 appeared to be specifically geared to respond to the [Na+ ]i...

  7. TRANSPARENT TESTA 13 is a tonoplast P3A -ATPase required for vacuolar deposition of proanthocyanidins in Arabidopsis thaliana seeds

    NARCIS (Netherlands)

    Appelhagen, I.; Nordholt, N.; Seidel, T.; Spelt, K.; Koes, R.; Quattrochio, F.; Sagasser, M.; Weisshaar, B.

    2015-01-01

    Intracellular pH homeostasis is essential for all living cells. In plants, pH is usually maintained by three structurally distinct and differentially localized types of proton pump: P-type H(+) -ATPases in the plasma membrane, and multimeric vacuolar-type H(+) -ATPases (V-ATPases) and vacuolar H(+)

  8. Specific inhibition of p97/VCP ATPase and kinetic analysis demonstrate interaction between D1 and D2 ATPase domains.

    Science.gov (United States)

    Chou, Tsui-Fen; Bulfer, Stacie L; Weihl, Conrad C; Li, Kelin; Lis, Lev G; Walters, Michael A; Schoenen, Frank J; Lin, Henry J; Deshaies, Raymond J; Arkin, Michelle R

    2014-07-29

    The p97 AAA (ATPase associated with diverse cellular activities), also called VCP (valosin-containing protein), is an important therapeutic target for cancer and neurodegenerative diseases. p97 forms a hexamer composed of two AAA domains (D1 and D2) that form two stacked rings and an N-terminal domain that binds numerous cofactor proteins. The interplay between the three domains in p97 is complex, and a deeper biochemical understanding is needed in order to design selective p97 inhibitors as therapeutic agents. It is clear that the D2 ATPase domain hydrolyzes ATP in vitro, but whether D1 contributes to ATPase activity is controversial. Here, we use Walker A and B mutants to demonstrate that D1 is capable of hydrolyzing ATP and show for the first time that nucleotide binding in the D2 domain increases the catalytic efficiency (kcat/Km) of D1 ATP hydrolysis 280-fold, by increasing kcat 7-fold and decreasing Km about 40-fold. We further show that an ND1 construct lacking D2 but including the linker between D1 and D2 is catalytically active, resolving a conflict in the literature. Applying enzymatic observations to small-molecule inhibitors, we show that four p97 inhibitors (DBeQ, ML240, ML241, and NMS-873) have differential responses to Walker A and B mutations, to disease-causing IBMPFD mutations, and to the presence of the N domain binding cofactor protein p47. These differential effects provide the first evidence that p97 cofactors and disease mutations can alter p97 inhibitor potency and suggest the possibility of developing context-dependent inhibitors of p97. Copyright © 2014 Elsevier Ltd. All rights reserved.

  9. H+-ATPase activity from storage tissue of Beta vulgaris. IV. N,N'-dicyclohexylcarbodiimide binding and inhibition of the plasma membrane H+-ATPase

    International Nuclear Information System (INIS)

    Oleski, N.A.; Bennett, A.B.

    1987-01-01

    The molecular weight and isoelectric point of the plasma membrane H + -ATPase from red beet storage tissue were determined using N,N'-dicyclohexylcarbodiimide (DCCD) and a H + -ATPase antibody. When plasma membrane vesicles were incubated with 20 micromolar [ 14 C]-DCCD at 0 0 C, a single 97,000 dalton protein was visualized on a fluorography of a sodium dodecyl sulfate polyacrylamide gel. A close correlation between [ 14 C]DCCD labeling of the 97,000 dalton protein and the extent of ATPase inhibition over a range of DCCD concentration suggests that this 97,000 dalton protein is a component of the plasma membrane H + -ATPase. An antibody raised against the plasma membrane H + -ATPase of Neurospora crassa cross-reacted with the 97,000 dalton DCCD-binding protein, further supporting the identity of this protein. Immunoblots of two-dimensional gels of red beet plasma membrane vesicles indicated the isoelectric point of the H + -ATPase to be 6.5

  10. Association with β-COP Regulates the Trafficking of the Newly Synthesized Na,K-ATPase*

    Science.gov (United States)

    Morton, Michael J.; Farr, Glen A.; Hull, Michael; Capendeguy, Oihana; Horisberger, Jean-Daniel; Caplan, Michael J.

    2010-01-01

    Plasma membrane expression of the Na,K-ATPase requires assembly of its α- and β-subunits. Using a novel labeling technique to identify Na,K-ATPase partner proteins, we detected an interaction between the Na,K-ATPase α-subunit and the coat protein, β-COP, a component of the COP-I complex. When expressed in the absence of the Na,K-ATPase β-subunit, the Na,K-ATPase α-subunit interacts with β-COP, is retained in the endoplasmic reticulum, and is targeted for degradation. In the presence of the Na,K-ATPase β-subunit, the α-subunit does not interact with β-COP and traffics to the plasma membrane. Pulse-chase experiments demonstrate that in cells expressing both the Na,K-ATPase α- and β-subunits, newly synthesized α-subunit associates with β-COP immediately after its synthesis but that this interaction does not constitute an obligate intermediate in the assembly of the α- and β-subunits to form the pump holoenzyme. The interaction with β-COP was reduced by mutating a dibasic motif at Lys54 in the Na,K-ATPase α-subunit. This mutant α-subunit is not retained in the endoplasmic reticulum and reaches the plasma membrane, even in the absence of Na,K-ATPase β-subunit expression. Although the Lys54 α-subunit reaches the cell surface without need for β-subunit assembly, it is only functional as an ion-transporting ATPase in the presence of the β-subunit. PMID:20801885

  11. Combined effects of EGFR tyrosine kinase inhibitors and vATPase inhibitors in NSCLC cells

    Energy Technology Data Exchange (ETDEWEB)

    Jin, Hyeon-Ok [KIRAMS Radiation Biobank, Korea Institute of Radiological and Medical Sciences, 75 Nowon-ro, Nowon-gu, Seoul, 139–706 (Korea, Republic of); Hong, Sung-Eun [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, 75 Nowon-ro, Nowon-gu, Seoul, 139–706 (Korea, Republic of); Kim, Chang Soon [Department of Microbiological Engineering, Kon-Kuk University, 120 Neungdong-ro, Gwangjin-gu, Seoul, 143–701 (Korea, Republic of); Park, Jin-Ah; Kim, Jin-Hee; Kim, Ji-Young; Kim, Bora [KIRAMS Radiation Biobank, Korea Institute of Radiological and Medical Sciences, 75 Nowon-ro, Nowon-gu, Seoul, 139–706 (Korea, Republic of); Chang, Yoon Hwan; Hong, Seok-Il; Hong, Young Jun [Department of Laboratory Medicine, Korea Cancer Center Hospital, Korea Institute of Radiological and Medical Sciences, 75 Nowon-ro, Nowon-gu, Seoul, 139–706 (Korea, Republic of); Park, In-Chul, E-mail: parkic@kirams.re.kr [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, 75 Nowon-ro, Nowon-gu, Seoul, 139–706 (Korea, Republic of); Lee, Jin Kyung, E-mail: jklee@kirams.re.kr [KIRAMS Radiation Biobank, Korea Institute of Radiological and Medical Sciences, 75 Nowon-ro, Nowon-gu, Seoul, 139–706 (Korea, Republic of); Department of Laboratory Medicine, Korea Cancer Center Hospital, Korea Institute of Radiological and Medical Sciences, 75 Nowon-ro, Nowon-gu, Seoul, 139–706 (Korea, Republic of)

    2015-08-15

    Despite excellent initial clinical responses of non-small cell lung cancer (NSCLC) patients to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs), many patients eventually develop resistance. According to a recent report, vacuolar H + ATPase (vATPase) is overexpressed and is associated with chemotherapy drug resistance in NSCLC. We investigated the combined effects of EGFR TKIs and vATPase inhibitors and their underlying mechanisms in the regulation of NSCLC cell death. We found that combined treatment with EGFR TKIs (erlotinib, gefitinib, or lapatinib) and vATPase inhibitors (bafilomycin A1 or concanamycin A) enhanced synergistic cell death compared to treatments with each drug alone. Treatment with bafilomycin A1 or concanamycin A led to the induction of Bnip3 expression in an Hif-1α dependent manner. Knock-down of Hif-1α or Bnip3 by siRNA further enhanced cell death induced by bafilomycin A1, suggesting that Hif-1α/Bnip3 induction promoted resistance to cell death induced by the vATPase inhibitors. EGFR TKIs suppressed Hif-1α and Bnip3 expression induced by the vATPase inhibitors, suggesting that they enhanced the sensitivity of the cells to these inhibitors by decreasing Hif-1α/Bnip3 expression. Taken together, we conclude that EGFR TKIs enhance the sensitivity of NSCLC cells to vATPase inhibitors by decreasing Hif-1α/Bnip3 expression. We suggest that combined treatment with EGFR TKIs and vATPase inhibitors is potentially effective for the treatment of NSCLC. - Highlights: • Co-treatment with EGFR TKIs and vATPase inhibitors induces synergistic cell death • EGFR TKIs enhance cell sensitivity to vATPase inhibitors via Hif-1α downregulation • Co-treatment of these inhibitors is potentially effective for the treatment of NSCLC.

  12. Oxidative stress (glutathionylation and Na,K-ATPase activity in rat skeletal muscle.

    Directory of Open Access Journals (Sweden)

    Carsten Juel

    Full Text Available Changes in ion distribution across skeletal muscle membranes during muscle activity affect excitability and may impair force development. These changes are counteracted by the Na,K-ATPase. Regulation of the Na,K-ATPase is therefore important for skeletal muscle function. The present study investigated the presence of oxidative stress (glutathionylation on the Na,K-ATPase in rat skeletal muscle membranes.Immunoprecipitation with an anti-glutathione antibody and subsequent immunodetection of Na,K-ATPase protein subunits demonstrated 9.0±1.3% and 4.1±1.0% glutathionylation of the α isoforms in oxidative and glycolytic skeletal muscle, respectively. In oxidative muscle, 20.0±6.1% of the β1 units were glutathionylated, whereas 14.8±2.8% of the β2-subunits appear to be glutathionylated in glycolytic muscle. Treatment with the reducing agent dithiothreitol (DTT, 1 mM increased the in vitro maximal Na,K-ATPase activity by 19% (P<0.05 in membranes from glycolytic muscle. Oxidized glutathione (GSSG, 0-10 mM increased the in vitro glutathionylation level detected with antibodies, and decreased the in vitro maximal Na,K-ATPase activity in a dose-dependent manner, and with a larger effect in oxidative compared to glycolytic skeletal muscle.This study demonstrates the existence of basal glutathionylation of both the α and the β units of rat skeletal muscle Na,K-ATPase. In addition, the study suggests a negative correlation between glutathionylation levels and maximal Na,K-ATPase activity.Glutathionylation likely contributes to the complex regulation of Na,K-ATPase function in skeletal muscle. Especially, glutathionylation induced by oxidative stress may have a role in Na,K-ATPase regulation during prolonged muscle activity.

  13. The AAA+ ATPase p97, a cellular multitool.

    Science.gov (United States)

    Stach, Lasse; Freemont, Paul S

    2017-08-17

    The AAA+ (ATPases associated with diverse cellular activities) ATPase p97 is essential to a wide range of cellular functions, including endoplasmic reticulum-associated degradation, membrane fusion, NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) activation and chromatin-associated processes, which are regulated by ubiquitination. p97 acts downstream from ubiquitin signaling events and utilizes the energy from ATP hydrolysis to extract its substrate proteins from cellular structures or multiprotein complexes. A multitude of p97 cofactors have evolved which are essential to p97 function. Ubiquitin-interacting domains and p97-binding domains combine to form bi-functional cofactors, whose complexes with p97 enable the enzyme to interact with a wide range of ubiquitinated substrates. A set of mutations in p97 have been shown to cause the multisystem proteinopathy inclusion body myopathy associated with Paget's disease of bone and frontotemporal dementia. In addition, p97 inhibition has been identified as a promising approach to provoke proteotoxic stress in tumors. In this review, we will describe the cellular processes governed by p97, how the cofactors interact with both p97 and its ubiquitinated substrates, p97 enzymology and the current status in developing p97 inhibitors for cancer therapy. © 2017 The Author(s).

  14. Activation of the Na+,K+-ATPase in Narcine brasiliensis

    International Nuclear Information System (INIS)

    Blum, H.; Nioka, Shoko; Johnson, R.G. Jr.

    1990-01-01

    The in vivo activation and turnover rates of the sodium pump (Na + ,K + -ATPase) were investigated in the electrocytes of the electric organ of the elasmobranch Narcine brasiliensis. The Narcine electric organ appears to be an excellent model for the study of sodium pump activation in an excitable tissue. The sodium transmembrane gradient and high-energy phosphagens were concurrently measured by 23 Na and 31 P NMR spectroscopy. The resting electric organ, which depends primarily on anaerobic metabolism displays a high concentration of phosphocreatin (PCr). It has an intracellular sodium concentration ([Na + ] i ) of 20±10 milliequivalents/liter as estimated by NMR. Electrical stimulation of the nerves innervating the electric organ results in an increase in [Na + ] i in the electrolyte and rapid depletion of PCr. Ouabain causes an 85% decrease in utilization of high-energy phosphagens, indicating that rapid PCr turnover in this tissue is mainly due to Na + ,K + -ATPase activity. From these data the authors can determine that the rate of sodium pump turnover increases by >3 orders of magnitude within several hundred milliseconds. The authors conclude that cholinergic stimulation of the electric organ causes a rapid and extremely large increase in sodium pump turnover, which is regulated predominantly by factors other than [Na + ] i

  15. Synthesis of Er and Er : Yb doped sol–gel derived silica glass and ...

    Indian Academy of Sciences (India)

    Unknown

    Materials Science Centre, †Central Research Facility, Optical Fibre Unit, Indian Institute of Technology,. Kharagpur 721 302, India. MS received 1 March 2004; revised 4 July 2004. Abstract. Er3+ and Er3+ : Yb3+ doped optical quality, crack and bubble free glasses for possible use in mak- ing laser material have been ...

  16. Excessive signal transduction of gain-of-function variants of the calcium-sensing receptor (CaSR are associated with increased ER to cytosol calcium gradient.

    Directory of Open Access Journals (Sweden)

    Marianna Ranieri

    Full Text Available In humans, gain-of-function mutations of the calcium-sensing receptor (CASR gene are the cause of autosomal dominant hypocalcemia or type 5 Bartter syndrome characterized by an abnormality of calcium metabolism with low parathyroid hormone levels and excessive renal calcium excretion. Functional characterization of CaSR activating variants has been so far limited at demonstrating an increased sensitivity to external calcium leading to lower Ca-EC50. Here we combine high resolution fluorescence based techniques and provide evidence that for the efficiency of calcium signaling system, cells expressing gain-of-function variants of CaSR monitor cytosolic and ER calcium levels increasing the expression of the Sarco-Endoplasmic Reticulum Calcium-ATPase (SERCA and reducing expression of Plasma Membrane Calcium-ATPase (PMCA. Wild-type CaSR (hCaSR-wt and its gain-of-function (hCaSR-R990G; hCaSR-N124K variants were transiently transfected in HEK-293 cells. Basal intracellular calcium concentration was significantly lower in cells expressing hCaSR-wt and its gain of function variants compared to mock. In line, FRET studies using the D1ER probe, which detects [Ca2+]ER directly, demonstrated significantly higher calcium accumulation in cells expressing the gain of function CaSR variants compared to hCaSR-wt. Consistently, cells expressing activating CaSR variants showed a significant increase in SERCA activity and expression and a reduced PMCA expression. This combined parallel regulation in protein expression increases the ER to cytosol calcium gradient explaining the higher sensitivity of CaSR gain-of-function variants to external calcium. This control principle provides a general explanation of how cells reliably connect (and exacerbate receptor inputs to cell function.

  17. Site of Er ions in silica layers codoped with Si nanoclusters and Er

    International Nuclear Information System (INIS)

    Pellegrino, P.; Garrido, B.; Arbiol, J.; Garcia, C.; Lebour, Y.; Morante, J.R.

    2006-01-01

    Silica layers implanted with Si and Er ions to various doses and annealed at 950 deg. C have been investigated by means of energy-filtered transmission electron microscopy (EFTEM) and high annular angle dark field (HAADF). EFTEM analysis reveals Si nanoclusters (Si-nc) with an average size around 3 nm for high Si content (15 at. %) whereas no clusters can be imaged for the lowest Si excess (5 at. %). Raman scattering supports that amorphous Si precipitates are present in all the samples. Moreover, the filtered images show that Er ions appear preferentially located outside the Si-nc. HAADF analysis confirms that the Er atoms form agglomerations of 5-10 nm size when the Er concentration exceeds 1x10 20 cm -3 . This observation correlates well with the reduction of the Er population excitable by Si nanoclusters, in the best case corresponding to 10% of the total. A suitable tuning of the annealing drastically reduces this deleterious effect

  18. DNA as a component of ER materials

    International Nuclear Information System (INIS)

    Minagawa, K; Aoki, Y; Berber, M R; Mori, T; Tanaka, M

    2009-01-01

    Deoxyribonucleic acid (DNA), which is known as a typical biopolymer, has been utilized for a few types of ER materials. Suspensions were prepared with the particles of DNA, DNA/lipid complexes, and LDH (layered double hydroxide)/DNA composites. The purified DNA showed larger ER effect than the others, but this particle tended to absorb water, which caused less stability. Preliminary experiments of preparing composite with LDH indicated that this inorganic material would be useful for hydrophobic modification of DNA particles, although further optimization of composite preparation is needed. In addition, the LDH/DNA suspensions showed interesting behaviours under some conditions, which indicated possibility for controlling ER property in a wide range.

  19. DNA as a component of ER materials

    Energy Technology Data Exchange (ETDEWEB)

    Minagawa, K; Aoki, Y; Berber, M R [Institute of Technology and Science, University of Tokushima, Tokushima 770-8506 (Japan); Mori, T [Department of Applied Chemistry, Faculty of Engineering, Kyushu University, Fukuoka 819-0395 (Japan); Tanaka, M [Faculty of Pharmaceutical Science, Tokushima Bunri University, Tokushima 770-8514 (Japan)], E-mail: minagawa@chem.tokushima-u.ac.jp

    2009-02-01

    Deoxyribonucleic acid (DNA), which is known as a typical biopolymer, has been utilized for a few types of ER materials. Suspensions were prepared with the particles of DNA, DNA/lipid complexes, and LDH (layered double hydroxide)/DNA composites. The purified DNA showed larger ER effect than the others, but this particle tended to absorb water, which caused less stability. Preliminary experiments of preparing composite with LDH indicated that this inorganic material would be useful for hydrophobic modification of DNA particles, although further optimization of composite preparation is needed. In addition, the LDH/DNA suspensions showed interesting behaviours under some conditions, which indicated possibility for controlling ER property in a wide range.

  20. ARM CLASIC ER2 CRS/EDOP

    Energy Technology Data Exchange (ETDEWEB)

    Gerald Heymsfield

    2010-12-20

    Data was taken with the NASA ER-2 aircraft with the Cloud Radar System and other instruments in conjunction with the DOE ARM CLASIC field campaign. The flights were near the SGP site in north Central Oklahoma and targeted small developing convection. The CRS is a 94 GHz nadir pointing Doppler radar. Also on board the ER-2 was the Cloud Physics Lidar (CPL). Seven science flights were conducted but the weather conditions did not cooperate in that there was neither developing convection, or there was heavy rain.

  1. Psyken er en grænseflade

    DEFF Research Database (Denmark)

    Hoffmeyer, Jesper

    2011-01-01

    En computer er i sig selv blot et stykke mekanik. Det afgørende sker i mødet mellem maskinen, mennesker og fællesskaber: på grænsefladen. Præcis som hos levende væsner......En computer er i sig selv blot et stykke mekanik. Det afgørende sker i mødet mellem maskinen, mennesker og fællesskaber: på grænsefladen. Præcis som hos levende væsner...

  2. Structure and function of the latent F0-F1-ATPase complex of Micrococcus lysodeikticus

    International Nuclear Information System (INIS)

    Chung, Y.S.

    1988-01-01

    The latent F 0 F 1 -ATPase from Micrococcus luteus (lysodeikticus) has been purified to homogeneity, and nine distinct subunit bands were observed on SDS-PAGE. Five of nine bands corresponded to the F 1 subunits and the other four bands are likely to be subunits a, a', b, and c of the F 0 segment of the complex. The subunit designated as a' probably arises from proteolytic cleavage of the 25,5000 Mr subunit a. The F 0 F 1 -ATPase complex has a molecular weight of approximately 1,060,000, as determined by Fast Protein Liquid Chromatography (FPLC). It is assumed that the F 0 F 1 -ATPase peak obtained by FPLC was a dimer and that molecular weight of the F 0 F 1 -ATPase monomer was accordingly 530,000. The stoichiometry of the subunits was determined with 14 C-labeled F 0 F 1 -ATPase prepared from cells grown on medium containing 14 C-amino acids. Antibodies to the native and SDS-denatured F 1 and F 0 F 1 -ATPase as well as to individual SDS-dissociated subunits have been generated for immunochemical analysis. The arrangement of the subunits in F 1 and F 0 F 1 -ATPase have been investigated using bifunctional chemical cross-linking agents

  3. Congruence between PM H+-ATPase and NADPH oxidase during root growth: a necessary probability.

    Science.gov (United States)

    Majumdar, Arkajo; Kar, Rup Kumar

    2018-07-01

    Plasma membrane (PM) H + -ATPase and NADPH oxidase (NOX) are two key enzymes responsible for cell wall relaxation during elongation growth through apoplastic acidification and production of ˙OH radical via O 2 ˙ - , respectively. Our experiments revealed a putative feed-forward loop between these enzymes in growing roots of Vigna radiata (L.) Wilczek seedlings. Thus, NOX activity was found to be dependent on proton gradient generated across PM by H + -ATPase as evident from pharmacological experiments using carbonyl cyanide m-chlorophenylhydrazone (CCCP; protonophore) and sodium ortho-vanadate (PM H + -ATPase inhibitor). Conversely, H + -ATPase activity retarded in response to different ROS scavengers [CuCl 2 , N, N' -dimethylthiourea (DMTU) and catalase] and NOX inhibitors [ZnCl 2 and diphenyleneiodonium (DPI)], while H 2 O 2 promoted PM H + -ATPase activity at lower concentrations. Repressing effects of Ca +2 antagonists (La +3 and EGTA) on the activity of both the enzymes indicate its possible mediation. Since, unlike animal NOX, the plant versions do not possess proton channel activity, harmonized functioning of PM H + -ATPase and NOX appears to be justified. Plasma membrane NADPH oxidase and H + -ATPase are functionally synchronized and they work cooperatively to maintain the membrane electrical balance while mediating plant cell growth through wall relaxation.

  4. Regulation of alpha1 Na/K-ATPase expression by cholesterol.

    Science.gov (United States)

    Chen, Yiliang; Li, Xin; Ye, Qiqi; Tian, Jiang; Jing, Runming; Xie, Zijian

    2011-04-29

    We have reported that α1 Na/K-ATPase regulates the trafficking of caveolin-1 and consequently alters cholesterol distribution in the plasma membrane. Here, we report the reciprocal regulation of α1 Na/K-ATPase by cholesterol. Acute exposure of LLC-PK1 cells to methyl β-cyclodextrin led to parallel decreases in cellular cholesterol and the expression of α1 Na/K-ATPase. Cholesterol repletion fully reversed the effect of methyl β-cyclodextrin. Moreover, inhibition of intracellular cholesterol trafficking to the plasma membrane by compound U18666A had the same effect on α1 Na/K-ATPase. Similarly, the expression of α1, but not α2 and α3, Na/K-ATPase was significantly reduced in the target organs of Niemann-Pick type C mice where the intracellular cholesterol trafficking is blocked. Mechanistically, decreases in the plasma membrane cholesterol activated Src kinase and stimulated the endocytosis and degradation of α1 Na/K-ATPase through Src- and ubiquitination-dependent pathways. Thus, the new findings, taken together with what we have already reported, revealed a previously unrecognized feed-forward mechanism by which cells can utilize the Src-dependent interplay among Na/K-ATPase, caveolin-1, and cholesterol to effectively alter the structure and function of the plasma membrane.

  5. Regulation of vacuolar H{sup +}-ATPase in microglia by RANKL

    Energy Technology Data Exchange (ETDEWEB)

    Serrano, Eric M.; Ricofort, Ryan D.; Zuo, Jian [Department of Orthodontics, University of Florida College of Dentistry, Gainesville, FL 32610 (United States); Ochotny, Noelle [Department of Pharmacology, University of Toronto, Toronto, Ont., Canada M5G 1G6 (Canada); Manolson, Morris F. [Faculty of Dentistry, University of Toronto, Toronto, Ont., Canada M5G 1G6 (Canada); Holliday, L. Shannon, E-mail: sholliday@dental.ufl.edu [Department of Orthodontics, University of Florida College of Dentistry, Gainesville, FL 32610 (United States); Department of Anatomy and Cell Biology, University of Florida College of Medicine, Gainesville, FL 32610 (United States)

    2009-11-06

    Vacuolar H{sup +}-ATPases (V-ATPases) are large electrogenic proton pumps composed of numerous subunits that play vital housekeeping roles in the acidification of compartments of the endocytic pathway. Additionally, V-ATPases play specialized roles in certain cell types, a capacity that is linked to cell type selective expression of isoforms of some of the subunits. We detected low levels of the a3 isoform of the a-subunit in mouse brain extracts. Examination of various brain-derived cell types by immunoblotting showed a3 was expressed in the N9 microglia cell line and in primary microglia, but not in other cell types. The expression of a3 in osteoclasts requires stimulation by Receptor Activator of Nuclear Factor {kappa}B-ligand (RANKL). We found that Receptor Activator of Nuclear Factor {kappa}B (RANK) was expressed by microglia. Stimulation of microglia with RANKL triggered increased expression of a3. V-ATPases in microglia were shown to bind microfilaments, and stimulation with RANKL increased the proportion of V-ATPase associated with the detergent-insoluble cytoskeletal fraction and with actin. In summary, microglia express the a3-subunit of V-ATPase. The expression of a3 and the interaction between V-ATPases and microfilaments was modulated by RANKL. These data suggest a novel molecular pathway for regulating microglia.

  6. Inositol phosphates influence the membrane bound Ca2+/Mg2+ stimulated ATPase from human erythrocyte membranes

    International Nuclear Information System (INIS)

    Kester, M.; Ekholm, J.; Kumar, R.; Hanahan, D.J.

    1986-01-01

    The modulation by exogenous inositol phosphates of the membrane Ca 2+ /Mg 2+ ATPase from saponin/EGTA lysed human erythrocytes was determined in a buffer (pH 7.6) containing histidine, 80 mM, MgCl 2 , 3.3 mM, NaCl, 74 mM, KCl, 30 mM, Na 2 ATP, 2.3 mM, ouabain, 0.83 mM, with variable amounts of CaCl 2 and EGTA. The ATPase assay was linear with time at 44 0 C. The inositol phosphates were commercially obtained and were also prepared from 32 P labeled rabbit platelet inositol phospholipids. Inositol triphosphate (IP 3 ) elevated the Ca 2+ /Mg 2+ ATPase activity over basal levels in a dose, time, and calcium dependent manner and were increased up to 85% of control values. Activities for the Na + /K + -ATPase and a Mg 2+ ATPase were not effected by IP 3 . Ca 2+ /Mg 2+ APTase activity with IP 2 or IP 3 could be synergistically elevated with calmodulin addition. The activation of the ATPase with IP 3 was calcium dependent in a range from .001 to .02 mM. The apparent Km and Vmax values were determined for IP 3 stimulated Ca 2+ /Mg 2+ ATPase

  7. Single-molecule analysis of inhibitory pausing states of V1-ATPase.

    Science.gov (United States)

    Uner, Naciye Esma; Nishikawa, Yoshihiro; Okuno, Daichi; Nakano, Masahiro; Yokoyama, Ken; Noji, Hiroyuki

    2012-08-17

    V(1)-ATPase, the hydrophilic V-ATPase domain, is a rotary motor fueled by ATP hydrolysis. Here, we found that Thermus thermophilus V(1)-ATPase shows two types of inhibitory pauses interrupting continuous rotation: a short pause (SP, 4.2 s) that occurred frequently during rotation, and a long inhibitory pause (LP, >30 min) that terminated all active rotations. Both pauses occurred at the same angle for ATP binding and hydrolysis. Kinetic analysis revealed that the time constants of inactivation into and activation from the SP were too short to represent biochemically predicted ADP inhibition, suggesting that SP is a newly identified inhibitory state of V(1)-ATPase. The time constant of inactivation into LP was 17 min, consistent with one of the two time constants governing the inactivation process observed in bulk ATPase assay. When forcibly rotated in the forward direction, V(1) in LP resumed active rotation. Solution ADP suppressed the probability of mechanical activation, suggesting that mechanical rotation enhanced inhibitory ADP release. These features were highly consistent with mechanical activation of ADP-inhibited F(1), suggesting that LP represents the ADP-inhibited state of V(1)-ATPase. Mechanical activation largely depended on the direction and angular displacement of forced rotation, implying that V(1)-ATPase rotation modulates the off rate of ADP.

  8. Response of plasma membrane H+-ATPase in rice (Oryza sativa) seedlings to simulated acid rain.

    Science.gov (United States)

    Liang, Chanjuan; Ge, Yuqing; Su, Lei; Bu, Jinjin

    2015-01-01

    Understanding the adaptation of plants to acid rain is important to find feasible approaches to alleviate such damage to plants. We studied effects of acid rain on plasma membrane H(+)-ATPase activity and transcription, intracellular H(+), membrane permeability, photosynthetic efficiency, and relative growth rate during stress and recovery periods. Simulated acid rain at pH 5.5 did not affect plasma membrane H(+)-ATPase activity, intracellular H(+), membrane permeability, photosynthetic efficiency, and relative growth rate. Plasma membrane H(+)-ATPase activity and transcription in leaves treated with acid rain at pH 3.5 was increased to maintain ion homeostasis by transporting excessive H(+) out of cells. Then intracellular H(+) was close to the control after a 5-day recovery, alleviating damage on membrane and sustaining photosynthetic efficiency and growth. Simulated acid rain at pH 2.5 inhibited plasma membrane H(+)-ATPase activity by decreasing the expression of H(+)-ATPase at transcription level, resulting in membrane damage and abnormal intracellular H(+), and reduction in photosynthetic efficiency and relative growth rate. After a 5-day recovery, all parameters in leaves treated with pH 2.5 acid rain show alleviated damage, implying that the increased plasma membrane H(+)-ATPase activity and its high expression were involved in repairing process in acid rain-stressed plants. Our study suggests that plasma membrane H(+)-ATPase can play a role in adaptation to acid rain for rice seedlings.

  9. Quantitative measurement of membrane Na+-K+ ATPase activity using thallium-201: comparison with rubidium-86

    International Nuclear Information System (INIS)

    Lee, Jae Tae; Shon, Sang Kyun; Lee, Kyu Bo; Lee, In Kyu

    1998-01-01

    Na + -K + ATPase activity has been estimated by the degree of inhibition of cation transport by cardiac glycosides (ouabain) using Rb-86 as a substrate. The biological characteristics of Tl-201 is known to be similar to those of potassium as a transport substrate in the presence of glucose, insulin or phobol myristate acetate (PMA). The purpose of this study was to measure ouabain sensitive Na + -K + ATPase activity using Tl-201 and compare with that using Rb-86. Smooth muscle cells isolated from rat aorta or human placental umbilical artery were cultured, and used to measure cellular Na + -K + ATPase activity. Na + -K + ATPase activity was measured as a percentage decrease in cellular uptake of Tl-201 or Rb-86 by ouabain under the presence of glucose, insulin or PMA in media. Na + -K + ATPase activity measured with Tl-201, as a transport substrate, was not different from those measured with Rb-86 in rat or human smooth muscle cell preparation. Incubation with high concentration glucose resulted in about 30% decrease in enzyme activity. In contrast, insulin or PMA resulted in 50-70% or 28% increase from baseline activity, respectively. These results suggests that Tl-201 could replace Rb-86 in measurement of ouabain sensitive Na + -K + ATPase activity in vitro. High level of glucose concentration decreased cellular Na + -K + ATPase activity, but insulin or PMA increased it

  10. Characterization and effect of light on the plasma membrane H(+) -ATPase of bean leaves

    Science.gov (United States)

    Linnemeyer, P. A.; Van Volkenburgh, E.; Cleland, R. E.

    1990-01-01

    Proton excretion from bean (Phaseolus vulgaris L.) leaf cells is increased by bright white light. To test whether this could be due, at least in part, to an increase in plasma membrane (PM) ATPase activity, PM vesicles were isolated from primary leaves by phase partitioning and used to characterize PM ATPase activity and changes in response to light. ATPase activity was characterized as magnesium ion dependent, vanadate sensitive, and slightly stimulated by potassium chloride. The pH optimum was 6.5, the Km was approximately 0.30 millimolar ATP, and the activity was about 60% latent. PM vesicles were prepared from leaves of plants grown for 11 days in dim red light (growing slowly) or grown for 10 days in dim red light and then transferred to bright white-light for 1 day (growing rapidly). For both light treatments, ATPase specific activity was approximately 600 to 700 nanomoles per milligram protein per minute, and the latency, Km, and sensitivity to potassium chloride were also similar. PM vesicles from plants grown in complete darkness, however, exhibited a twofold greater specific activity. We conclude that the promotion of leaf growth and proton excretion by bright white light is not due to an increase in ATPase specific activity. Light does influence ATPase activity, however; both dim red light and bright white light decreased the ATPase specific activity by nearly 50% as compared with dark-grown leaves.

  11. Specialized functional diversity and interactions of the Na,K-ATPase

    Directory of Open Access Journals (Sweden)

    Igor I. Krivoi

    2016-05-01

    Full Text Available Na,K-ATPase is a protein ubiquitously expressed in the plasma membrane of all animal cells and vitally essential for their functions. A specialized functional diversity of the Na,K-ATPase isozymes is provided by molecular heterogeneity, distinct subcellular localizations and functional interactions with molecular environment. Studies over the last decades clearly demonstrated complex and isoform-specific reciprocal functional interactions between the Na,K-ATPase and neighboring proteins and lipids. These interactions are enabled by a spatially restricted ion homeostasis, direct protein-protein/lipid interactions and protein kinase signaling pathways. In addition to its ‘classical’ function in ion translocation, the Na,K-ATPase is now considered as one of the most important signaling molecules in neuronal, epithelial, skeletal, cardiac and vascular tissues. Accordingly, the Na,K-ATPase forms specialized sub-cellular multimolecular microdomains which act as receptors to circulating endogenous cardiotonic steroids triggering a number of signaling pathways. Changes in these endogenous cardiotonic steroid levels and initiated signaling responses have significant adaptive values for tissues and whole organisms under numerous physiological and pathophysiological conditions. This review discusses recent progress in the studies of functional interactions between the Na,K-ATPase and molecular microenvironment, the Na,K-ATPase-dependent signaling pathways and their significance for diversity of cell function.

  12. ER Stress: A Therapeutic Target in Rheumatoid Arthritis?

    Science.gov (United States)

    Rahmati, Marveh; Moosavi, Mohammad Amin; McDermott, Michael F

    2018-04-22

    Diverse physiological and pathological conditions that impact on protein folding of the endoplasmic reticulum (ER) cause ER stress. The unfolded protein response (UPR) and the ER-associated degradation (ERAD) pathway are activated to cope with ER stress. In rheumatoid arthritis (RA), inflammation and ER stress work in parallel by driving inflammatory cells to release cytokines that induce chronic ER stress pathways. This chronic ER stress may contribute to the pathogenesis of RA through synoviocyte proliferation and proinflammatory cytokine production. Therefore, ER stress pathways and their constituent elements are attractive targets for RA drug development. In this review, we integrate current knowledge of the contribution of ER stress to the overall pathogenesis of RA, and suggest some therapeutic implications of these discoveries. Copyright © 2018 Elsevier Ltd. All rights reserved.

  13. Comparative properties of caveolar and noncaveolar preparations of kidney Na+/K+-ATPase.

    Science.gov (United States)

    Liu, Lijun; Ivanov, Alexander V; Gable, Marjorie E; Jolivel, Florent; Morrill, Gene A; Askari, Amir

    2011-10-11

    To evaluate previously proposed functions of renal caveolar Na(+)/K(+)-ATPase, we modified the standard procedures for the preparation of the purified membrane-bound kidney enzyme, separated the caveolar and noncaveolar pools, and compared their properties. While the subunits of Na(+)/K(+)-ATPase (α,β,γ) constituted most of the protein content of the noncaveolar pool, the caveolar pool also contained caveolins and major caveolar proteins annexin-2 tetramer and E-cadherin. Ouabain-sensitive Na(+)/K(+)-ATPase activities of the two pools had similar properties and equal molar activities, indicating that the caveolar enzyme retains its ion transport function and does not contain nonpumping enzyme. As minor constituents, both caveolar and noncaveolar pools also contained Src, EGFR, PI3K, and several other proteins known to be involved in stimulous-induced signaling by Na(+)/K(+)-ATPase, indicating that signaling function is not limited to the caveolar pool. Endogenous Src was active in both pools but was not further activated by ouabain, calling into question direct interaction of Src with native Na(+)/K(+)-ATPase. Chemical cross-linking, co-immunoprecipitation, and immunodetection studies showed that in the caveolar pool, caveolin-1 oligomers, annexin-2 tetramers, and oligomers of the α,β,γ-protomers of Na(+)/K(+)-ATPase form a large multiprotein complex. In conjunction with known roles of E-cadherin and the β-subunit of Na(+)/K(+)-ATPase in cell adhesion and noted intercellular β,β-contacts within the structure of Na(+)/K(+)-ATPase, our findings suggest that interacting caveolar Na(+)/K(+)-ATPases located at renal adherens junctions maintain contact of two adjacent cells, conduct essential ion pumping, and are capable of locus-specific signaling in junctional cells.

  14. Src-independent ERK signaling through the rat α3 isoform of Na/K-ATPase.

    Science.gov (United States)

    Madan, Namrata; Xu, Yunhui; Duan, Qiming; Banerjee, Moumita; Larre, Isabel; Pierre, Sandrine V; Xie, Zijian

    2017-03-01

    The Na/K-ATPase α1 polypeptide supports both ion-pumping and signaling functions. The Na/K-ATPase α3 polypeptide differs from α1 in both its primary structure and its tissue distribution. The expression of α3 seems particularly important in neurons, and recent clinical evidence supports a unique role of this isoform in normal brain function. The nature of this specific role of α3 has remained elusive, because the ubiquitous presence of α1 has hindered efforts to characterize α3-specific functions in mammalian cell systems. Using Na/K-ATPase α1 knockdown pig kidney cells (PY-17), we generated the first stable mammalian cell line expressing a ouabain-resistant form of rat Na/K-ATPase α3 in the absence of endogenous pig α1 detectable by Western blotting. In these cells, Na/K-ATPase α3 formed a functional ion-pumping enzyme and rescued the expression of Na/K-ATPase β1 and caveolin-1 to levels comparable with those observed in PY-17 cells rescued with a rat Na/K-ATPase α1 (AAC-19). The α3-containing enzymes had lower Na + affinity and lower ouabain-sensitive transport activity than their α1-containing counterparts under basal conditions, but showed a greater capacity to be activated when intracellular Na + was increased. In contrast to Na/K-ATPase α1, α3 could not regulate Src. Upon exposure to ouabain, Src activation did not occur, yet ERK was activated through Src-independent pathways involving PI3K and PKC. Hence, α3 expression confers signaling and pumping properties that are clearly distinct from that of cells expressing Na/K-ATPase α1. Copyright © 2017 the American Physiological Society.

  15. Long-term regulation of Na,K-ATPase pump during T-cell proliferation.

    Science.gov (United States)

    Karitskaya, Inna; Aksenov, Nikolay; Vassilieva, Irina; Zenin, Valerii; Marakhova, Irina

    2010-09-01

    The aim of the study was to elucidate the mechanism responsible for the proliferation-related regulation of Na,K-ATPase pump. Our data demonstrate that in mitogen-stimulated human blood lymphocytes, enhanced ouabain-sensitive Rb(K) fluxes in the middle/late stage of G(0)/G(1)/S transit are associated with the increased number of Na,K-ATPase pumps expressed at the cell surface (as determined by the [(3)H]ouabain binding). Analysis of total RNA (reverse transcription-polymerase chain reaction) and protein (Western blotting) showed a threefold increase in the level of Na,K-ATPase alpha1-subunit and beta1-subunit mRNAs and significant increase in the Na,K-ATPase alpha1-subunit protein during the first day of mitogen-induced proliferation. The elevated K transport as well as the increased expression of Na,K-ATPase is closely associated with the IL-2-dependent stage of T-cell response. The pharmacological inhibition of IL-2-induced MEK/ERK or JAK/STAT cascades suppressed the IL-2-induced proliferation and reduced the functional and protein expressions of Na,K-ATPase. It is concluded that during the lymphocyte transition from resting stage to proliferation, (1) long-term activation of Na,K-ATPase pump is due to the enhanced expression of Na,K-ATPase protein and mRNA, and (2) the cytokine signaling via the IL-2 receptor is necessary for the cell cycle-associated upregulation of Na,K-ATPase.

  16. Abscisic Acid Induction of Vacuolar H+-ATPase Activity in Mesembryanthemum crystallinum Is Developmentally Regulated1

    Science.gov (United States)

    Barkla, Bronwyn J.; Vera-Estrella, Rosario; Maldonado-Gama, Minerva; Pantoja, Omar

    1999-01-01

    Abscisic acid (ABA) has been implicated as a key component in water-deficit-induced responses, including those triggered by drought, NaCl, and low- temperature stress. In this study a role for ABA in mediating the NaCl-stress-induced increases in tonoplast H+-translocating ATPase (V-ATPase) and Na+/H+ antiport activity in Mesembryanthemum crystallinum, leading to vacuolar Na+ sequestration, were investigated. NaCl or ABA treatment of adult M. crystallinum plants induced V-ATPase H+ transport activity, and when applied in combination, an additive effect on V-ATPase stimulation was observed. In contrast, treatment of juvenile plants with ABA did not induce V-ATPase activity, whereas NaCl treatment resulted in a similar response to that observed in adult plants. Na+/H+ antiport activity was induced in both juvenile and adult plants by NaCl, but ABA had no effect at either developmental stage. Results indicate that ABA-induced changes in V-ATPase activity are dependent on the plant reaching its adult phase, whereas NaCl-induced increases in V-ATPase and Na+/H+ antiport activity are independent of plant age. This suggests that ABA-induced V-ATPase activity may be linked to the stress-induced, developmentally programmed switch from C3 metabolism to Crassulacean acid metabolism in adult plants, whereas, vacuolar Na+ sequestration, mediated by the V-ATPase and Na+/H+ antiport, is regulated through ABA-independent pathways. PMID:10398716

  17. Alteration of aluminium inhibition of synaptosomal (Na(+)/K(+))ATPase by colestipol administration.

    Science.gov (United States)

    Silva, V S; Oliveira, L; Gonçalves, P P

    2013-11-01

    The ability of aluminium to inhibit the (Na(+)/K(+))ATPase activity has been observed by several authors. During chronic dietary exposure to AlCl3, brain (Na(+)/K(+))ATPase activity drops, even if no alterations of catalytic subunit protein expression and of energy charge potential are observed. The aluminium effect on (Na(+)/K(+))ATPase activity seems to implicate the reduction of interacting protomers within the oligomeric ensemble of the membrane-bound (Na(+)/K(+))ATPase. The activity of (Na(+)/K(+))ATPase is altered by the microviscosity of lipid environment. We studied if aluminium inhibitory effect on (Na(+)/K(+))ATPase is modified by alterations in synaptosomal membrane cholesterol content. Adult male Wistar rats were submitted to chronic dietary AlCl3 exposure (0.03 g/day of AlCl3) and/or to colestipol, a hypolidaemic drug (0.31 g/day) during 4 months. The activity of (Na(+)/K(+))ATPase was studied in brain cortex synaptosomes with different cholesterol contents. Additionally, we incubate synaptosomes with methyl-β-cyclodextrin for both enrichment and depletion of membrane cholesterol content, with or without 300 μM AlCl3. This enzyme activity was significantly reduced by micromolar AlCl3 added in vitro and when aluminium was orally administered to rats. The oral administration of colestipol reduced the cholesterol content and concomitantly inhibited the (Na(+)/K(+))ATPase. The aluminium inhibitory effect on synaptosomal (Na(+)/K(+))ATPase was reduced by cholesterol depletion both in vitro and in vivo. Copyright © 2013 Elsevier Inc. All rights reserved.

  18. The effects of dexamethasone on the Na,K-ATPase activity and pump function of corneal endothelial cells.

    Science.gov (United States)

    Hatou, Shin; Yamada, Masakazu; Mochizuki, Hiroshi; Shiraishi, Atsushi; Joko, Takeshi; Nishida, Teruo

    2009-05-01

    The Na(+)- and K(+)-dependent ATPase (Na,K-ATPase) expressed in the basolateral membrane of corneal endothelial cells plays an important role in the pump function of the corneal endothelium. We investigated the possible role of dexamethasone in the regulation of Na,K-ATPase activity and pump function in corneal endothelial cells. Confluent monolayers of mouse corneal endothelial cells were exposed to dexamethasone. ATPase activity of the cells was evaluated by spectrophotometric measurement of phosphate released from ATP with the use of ammonium molybdate, with Na,K-ATPase activity being defined as the portion of total ATPase activity sensitive to ouabain. Pump function of the cells was measured with the use of an Ussing chamber, with the pump function attributable to Na,K-ATPase activity being defined as the portion of the total short-circuit current sensitive to ouabain. Western blot analysis was examined to measure the expression of the Na,K-ATPase alpha(1)-subunit. Dexamethasone (1 or 10 microM) increased the Na,K-ATPase activity and pump function of the cultured cells. These effects of dexamethasone were blocked by cycloheximide, a protein synthesis inhibitor. Western blot analysis also indicated that dexamethasone increased the expression of the Na,K-ATPase alpha(1)-subunit, whereas it decreased the expression of the phospho-Na,K-ATPase alpha(1)-subunit. Our results suggest that dexamethasone stimulates Na,K-ATPase activity in mouse corneal endothelial cells. The effect of dexamethasone activation in these cells is mediated by Na,K-ATPase synthesis and increase in an enzymatic activity by dephosphorylation of Na,K-ATPase alpha(1)-subunits.

  19. A novel mechanism of P-type ATPase autoinhibition involving both termini of the protein

    DEFF Research Database (Denmark)

    Ekberg, Kira; Palmgren, Michael; Veierskov, Bjarke

    2010-01-01

    The activity of many P-type ATPases is found to be regulated by interacting proteins or autoinhibitory elements located in N- or C-terminal extensions. An extended C terminus of fungal and plant P-type plasma membrane H+-ATPases has long been recognized to be part of a regulatory apparatus....... This identifies the first group of P-type ATPases for which both ends of the polypeptide chain constitute regulatory domains, which together contribute to the autoinhibitory apparatus. This suggests an intricate mechanism of cis-regulation with both termini of the protein communicating to obtain the necessary...

  20. Regulation of α1 Na/K-ATPase Expression by Cholesterol*

    OpenAIRE

    Chen, Yiliang; Li, Xin; Ye, Qiqi; Tian, Jiang; Jing, Runming; Xie, Zijian

    2011-01-01

    We have reported that α1 Na/K-ATPase regulates the trafficking of caveolin-1 and consequently alters cholesterol distribution in the plasma membrane. Here, we report the reciprocal regulation of α1 Na/K-ATPase by cholesterol. Acute exposure of LLC-PK1 cells to methyl β-cyclodextrin led to parallel decreases in cellular cholesterol and the expression of α1 Na/K-ATPase. Cholesterol repletion fully reversed the effect of methyl β-cyclodextrin. Moreover, inhibition of intracellular cholesterol tr...

  1. Human Hsp70 molecular chaperone binds two calcium ions within the ATPase domain.

    Science.gov (United States)

    Sriram, M; Osipiuk, J; Freeman, B; Morimoto, R; Joachimiak, A

    1997-03-15

    The 70 kDa heat shock proteins (Hsp70) are a family of molecular chaperones, which promote protein folding and participate in many cellular functions. The Hsp70 chaperones are composed of two major domains. The N-terminal ATPase domain binds to and hydrolyzes ATP, whereas the C-terminal domain is required for polypeptide binding. Cooperation of both domains is needed for protein folding. The crystal structure of bovine Hsc70 ATPase domain (bATPase) has been determined and, more recently, the crystal structure of the peptide-binding domain of a related chaperone, DnaK, in complex with peptide substrate has been obtained. The molecular chaperone activity and conformational switch are functionally linked with ATP hydrolysis. A high-resolution structure of the ATPase domain is required to provide an understanding of the mechanism of ATP hydrolysis and how it affects communication between C- and N-terminal domains. The crystal structure of the human Hsp70 ATPase domain (hATPase) has been determined and refined at 1. 84 A, using synchrotron radiation at 120K. Two calcium sites were identified: the first calcium binds within the catalytic pocket, bridging ADP and inorganic phosphate, and the second calcium is tightly coordinated on the protein surface by Glu231, Asp232 and the carbonyl of His227. Overall, the structure of hATPase is similar to bATPase. Differences between them are found in the loops, the sites of amino acid substitution and the calcium-binding sites. Human Hsp70 chaperone is phosphorylated in vitro in the presence of divalent ions, calcium being the most effective. The structural similarity of hATPase and bATPase and the sequence similarity within the Hsp70 chaperone family suggest a universal mechanism of ATP hydrolysis among all Hsp70 molecular chaperones. Two calcium ions have been found in the hATPase structure. One corresponds to the magnesium site in bATPase and appears to be important for ATP hydrolysis and in vitro phosphorylation. Local changes

  2. Na+,K+-ATPase concentration in rodent and human heart and skeletal muscle

    DEFF Research Database (Denmark)

    Kjeldsen, K; Bjerregaard, P; Richter, Erik

    1988-01-01

    rats, cardiomyopathic hamsters, and human subjects. These methods have earlier been shown to quantify the Na+,K+-ATPase concentration in muscle tissue with high accuracy. When rats were swim trained for six weeks the heart ventricular muscle Na+,K+-ATPase concentration was increased by 20% (p less than...... was increased by up to 46% (p less than 0.001) and decreased by up to 30% (p less than 0.005) after training and immobilisation respectively. Cardiomyopathic hamsters showed a reduction of 33% (p less than 0.005) in the heart ventricular Na+,K+-ATPase concentration compared with normal hamsters. This decrease...

  3. The effect of near-UV light on Na-K-ATPase of the rat lens

    International Nuclear Information System (INIS)

    Torriglia, A.; Zigman, S.

    1988-01-01

    The influence of in vitro near-UV radiation exposure on the physical state of the rat lens and on its membrane-bound Na-K-ATPase activity was investigated. Lens swelling was correlated to the appearance of opacities and the inactivation of the enzyme. The results show a significant decrease in the Na-K-ATPase activity which may be an early change leading to osmotic type cataracts. The dose-effect curves obtained for cortical and epithelial enzymes were different. Since the data do not follow a mono-exponential function, the existence of two forms of Na-K-ATPase in the lens is discussed. (author)

  4. Na+,K+-ATPase as the Target Enzyme for Organic and Inorganic Compounds

    Directory of Open Access Journals (Sweden)

    Tatjana Momić

    2008-12-01

    Full Text Available This paper gives an overview of the literature data concerning specific and non specific inhibitors of Na+,K+-ATPase receptor. The immobilization approaches developed to improve the rather low time and temperature stability of Na+,K+-ATPase, as well to preserve the enzyme properties were overviewed. The functional immobilization of Na+,K+-ATPase receptor as the target, with preservation of the full functional protein activity and access of various substances to an optimum number of binding sites under controlled conditions in the combination with high sensitive technology for the detection of enzyme activity is the basis for application of this enzyme in medical, pharmaceutical and environmental research.

  5. Effect of near-UV light on Na-K-ATPase of the rat lens

    Energy Technology Data Exchange (ETDEWEB)

    Torriglia, A.; Zigman, S.

    1988-06-01

    The influence of in vitro near-UV radiation exposure on the physical state of the rat lens and on its membrane-bound Na-K-ATPase activity was investigated. Lens swelling was correlated to the appearance of opacities and the inactivation of the enzyme. The results show a significant decrease in the Na-K-ATPase activity which may be an early change leading to osmotic type cataracts. The dose-effect curves obtained for cortical and epithelial enzymes were different. Since the data do not follow a mono-exponential function, the existence of two forms of Na-K-ATPase in the lens is discussed.

  6. Na,K-ATPase: a molecular target for Leptospira interrogans endotoxin

    Directory of Open Access Journals (Sweden)

    Younes-Ibrahim M.

    1997-01-01

    Full Text Available On the basis of our report that a glycolipoprotein fraction (GLP extracted from Leptospira interrogans contains a potent inhibitor of renal Na,K-ATPase, we proposed that GLP-induced inhibition of Na,K-ATPase might be the primary cellular defect in the physiopathology of leptospirosis. The present study was designed to test this hypothesis by determining whether or not 1 GLP inhibits all the isoforms of Na,K-ATPase which are expressed in the tissues affected by leptospirosis, 2 Na,K-ATPase from leptospirosis-resistant species, such as the rat, is sensitive to GLP, 3 GLP inhibits Na,K-ATPase from intact cells, and 4 GLP inhibits ouabain-sensitive H,K-ATPase. The results indicate that in the rabbit, a leptospirosis-sensitive species, GLP inhibits with similar efficiency (apparent IC50: 120-220 µg protein GLP/ml all isoforms of Na,K-ATPase known to be expressed in target tissues for the disease. Na,K-ATPase from rat kidney displays a sensitivity to GLP similar to that of the rabbit kidney enzyme (apparent IC50: 25-80 and 50-150 µg protein GLP/ml for rat and rabbit, respectively, indicating that resistance to the disease does not result from the resistance of Na,K-ATPase to GLP. GLP also reduces ouabain-sensitive rubidium uptake in rat thick ascending limbs (pmol mm-1 min-1 ± SEM; control: 23.8 ± 1.8; GLP, 88 µg protein/ml: 8.2 ± 0.9, demonstrating that it is active in intact cells. Finally, GLP had no demonstrable effect on renal H,K-ATPase activity, even on the ouabain-sensitive form, indicating that the active principle of GLP is more specific for Na,K-ATPase than ouabain itself. Although the hypothesis remains to be demonstrated in vivo, the present findings are compatible with the putative role of GLP-induced inhibition of Na,K-ATPase as an initial mechanism in the physiopathology of leptospirosis

  7. The stochastic chemomechanics of the F(1)-ATPase molecular motor.

    Science.gov (United States)

    Gaspard, P; Gerritsma, E

    2007-08-21

    We report a theoretical study of the F(1)-ATPase molecular rotary motor experimentally studied by R. Yasuda, H. Noji, M. Yoshida, K. Kinosita Jr., H. Itoh [Nature 410 (2001) 898]. The motor is modeled as a stochastic process for the angle of its shaft and the chemical state of its catalytic sites. The stochastic process is ruled by six coupled Fokker-Planck equations for the biased diffusion of the angle and the random jumps between the chemical states. The model reproduces the experimental observations that the motor proceeds by substeps and the rotation rate saturates at high concentrations of adenosine triphosphate or at low values of the friction coefficient. Moreover, predictions are made about the dependence of the rotation rate on temperature, and about the behavior of the F(1) motor under the effect of an external torque, especially, in the regime of synthesis of adenosine triphosphate.

  8. Crystals of Na(+)/K(+)-ATPase with bound cisplatin.

    Science.gov (United States)

    Huliciak, Miroslav; Reinhard, Linda; Laursen, Mette; Fedosova, Natalya; Nissen, Poul; Kubala, Martin

    2014-12-01

    Cisplatin is the most widely used chemotherapeutics for cancer treatment, however, its administration is connected to inevitable adverse effects. Previous studies suggested that cisplatin is able to inhibit Na(+)/K(+)-ATPase (NKA), the enzyme responsible for maintaining electrochemical potential and sodium gradient across the plasma membrane. Here we report a crystallographic analysis of cisplatin bound to NKA in the ouabain bound E2P form. Despite a moderate resolution (7.4 Å and 7.9 Å), the anomalous scattering from platinum and a model representation from a recently published structure enabled localization of seven cisplatin binding sites by anomalous difference Fourier maps. Comparison with NKA structures in the E1P conformation suggested two possible inhibitory mechanisms for cisplatin. Binding to Met151 can block the N-terminal pathway for transported cations, while binding to Met171 can hinder the interaction of cytoplasmic domains during the catalytic cycle. Copyright © 2014 Elsevier Inc. All rights reserved.

  9. Is the ATPase from halobacterium saccharovorum an evolutionary relic?

    Science.gov (United States)

    Hochstein, L. I.; Altekar, W.; Kristjansson, H.

    1986-01-01

    The ATP Synthase Complex present in the membranes of mitochondria, chloroplasts or bacteria is composed of 2 sectors: FO, an integral membrane protein consisting of 3 subunits mediating proton translocation across the membrane and F1, the catalytic component composed of 5 non-identical subunits. The apparent early origin of the ATP Synthase Complex, as implied by its ubiquitous distribution, seems inconsistent with its structural and functional complexity and raises the question if simpler versions of the ATP Synthase exist. Such an ATP Synthase has been searched for in various Archaebacteria. A purified halobacterial ATPase activity which possesses certain properties consistent with those of an ATP Synthase but which has a different subunit structure is described.

  10. ATPase and GTPase Tangos Drive Intracellular Protein Transport.

    Science.gov (United States)

    Shan, Shu-Ou

    2016-12-01

    The GTPase superfamily of proteins provides molecular switches to regulate numerous cellular processes. The 'GTPase switch' paradigm, in which external regulatory factors control the switch of a GTPase between 'on' and 'off' states, has been used to interpret the regulatory mechanism of many GTPases. However, recent work unveiled a class of nucleotide hydrolases that do not adhere to this classical paradigm. Instead, they use nucleotide-dependent dimerization cycles to regulate key cellular processes. In this review article, recent studies of dimeric GTPases and ATPases involved in intracellular protein targeting are summarized. It is suggested that these proteins can use the conformational plasticity at their dimer interface to generate multiple points of regulation, thereby providing the driving force and spatiotemporal coordination of complex cellular pathways. Copyright © 2016 Elsevier Ltd. All rights reserved.

  11. ATPase Activity Measurements by an Enzyme-Coupled Spectrophotometric Assay.

    Science.gov (United States)

    Sehgal, Pankaj; Olesen, Claus; Møller, Jesper V

    2016-01-01

    Enzymatic coupled assays are usually based on the spectrophotometric registration of changes in NADH/NAD(+) or NADPH/NADP(+) absorption at 340 nm accompanying the oxidation/reduction of reactants that by dehydrogenases and other helper enzymes are linked to the activity of the enzymatic reaction under study. The present NADH-ATP-coupled assay for ATPase activity is a seemingly somewhat complicated procedure, but in practice adaptation to performance is easily acquired. It is a more safe and elegant method than colorimetric methods, but not suitable for handling large number of samples, and also presupposes that the activity of the helper enzymes is not severely affected by the chemical environment of the sample in which it is tested.

  12. Structural studies of Ca2+-ATPase ligand and regulatory complexes

    DEFF Research Database (Denmark)

    Drachmann, Nikolaj Düring

    2015-01-01

    , the surrounding membrane itself has a huge influence on SERCA structure and function. Changes in the membrane thickness can alter the activity of the ATPase significantly, and even cause changes in the stoichiometry of ion transport. Structural studies on SERCA in the presence of four different phosphatidyl...... choline lipids with different aliphatic chain length and saturation show three specific lipid binding sites. The four different lipids analysed bind to the same binding sites with varying degrees of disorder. The study contributes to understanding the complex interplay between the surrounding membrane...... to explore the possibilities for an efficient screening of ligand-bound SERCA structures, serial femtosecond crystallography experiments of microcrystals of SERCA1a in the Ca2+ bound state and in a vanadate stabilised E2 state was conducted. A structure obtained at 2.8 Å maximum resolution of the proof...

  13. Elucidating Functional Aspects of P-type ATPases

    DEFF Research Database (Denmark)

    Autzen, Henriette Elisabeth

    2015-01-01

    and helped enlighten how thapsigargin, a potent inhibitor of SERCA1a, depends on a water mediated hydrogen bond network when bound to SERCA1a. Furthermore, molecular dynamics (MD) simulations of the same P-type ATPase were used to assess a long-standing question whether cholesterol affects SERCA1a through...... similar to that of the wild type (WT) protein. The discrepancy between the newly determined crystal structure of LpCopA and the functional manifestations of the missense mutation in human CopA, could indicate that LpCopA is insufficient in structurally elucidating the effect of disease-causing mutations...... in the human CopA proteins. MD simulations, which combine coarse-grained (CG) and atomistic procedures, were set up in order to elucidate mechanistic implications exerted by the lipid bilayer on LpCopA. The MD simulations of LpCopA corroborated previous and new in vivo activity data and showed...

  14. Ledelse af samskabelse er en ny ledelsesdisciplin

    DEFF Research Database (Denmark)

    Ulrich, Jens; Martin Nielsen, Kim; Bartram, Jan

    2018-01-01

    Samskabelse er blevet det nye orienteringspunkt for rigtig mange offentlige organisationer. Det skyldes en udbredt konsensus om, at velfærdsopgaverne ikke kan løses inden for de vante organisatoriske rammer, men må løses i tæt kontakt med de parter, som opgaverne vedrører. Denne nye tilgang til...

  15. Der er fuld religionsfrihed i Danmark, men..

    DEFF Research Database (Denmark)

    Christoffersen, Lisbet

    2018-01-01

    Artiklen analyserer en rapport, udgivet af FN's special rapporteur on Freedom of Religion and Belief, Heiner Bielefeldt. Rapporten vedrører Danmark. Analysen tilslutter sig rapporten på de fleste punkter, men kritiserer begrebet 'blød diskrimination'; kritiserer at rapporten ikke er gået tættere på...

  16. Hvad er kildekritik?

    DEFF Research Database (Denmark)

    Olden-Jørgensen, Sebastian

    2009-01-01

    Hvad er Kildekritik? I stedet for at forstå kildekritik som et stringent system, grundlagt af Leopold von Ranke og i Danmark kanoniseret af Kristian Erslev, kunne man med større historiografisk ret og faglig frugtbarhed opfatte kildekritikken som en faglig tradition med tre lag afsat (sedimentere...

  17. Hvad er PET og fMRI?

    DEFF Research Database (Denmark)

    B. Paulson, Olaf; Balslev, Daniela; Gerlach, Christian

    2007-01-01

    Hvilken rolle spiller følelserne for vores tankeliv, og hvorfor har vi overhovedet følelser? I kraft af avancerede hjerneskanningsteknikker er den moderne hjernevidenskab blevet i stand til at besvare disse spørgsmål. Gennem artiklerne i denne antologi, skrevet af ledende danske forskere, bliver ...

  18. Creating Smart-er Cities: An Overview

    Science.gov (United States)

    Allwinkle, Sam; Cruickshank, Peter

    2011-01-01

    The following offers an overview of what it means for cities to be "smart." It draws the supporting definitions and critical insights into smart cities from a series of papers presented at the 2009 Trans-national Conference on Creating Smart(er) Cities. What the papers all have in common is their desire to overcome the all too often…

  19. Geometric calibration of ERS satellite SAR images

    DEFF Research Database (Denmark)

    Mohr, Johan Jacob; Madsen, Søren Nørvang

    2001-01-01

    Geometric calibration of the European Remote Sensing (ERS) Satellite synthetic aperture radar (SAR) slant range images is important in relation to mapping areas without ground reference points and also in relation to automated processing. The relevant SAR system parameters are discussed...

  20. Er nogen mere lige end andre?

    DEFF Research Database (Denmark)

    Møberg, Rasmus Juul

    normativ fordring. Kohlis (2007) formulering af normalbiografien er en tredeling af livsforløbet (uddannelse – arbejdsliv - pension). En tredeling der bliver kritiseret for at være køns- og familieblind, hvorfor linked lives-begrebet bliver introduceret. Begrebet har den teoretiske implikation...... analyser tager højde for afhængighed over tid i den afhængige variabel. Overordnet set er det tre aspekter af datas karakteristika, der fordrer specielle metoder. Det er censurering af data, tidsmæssig variation i de uafhængige variable samt potentiel korrelation mellem målingerne af den afhængige variabel...... for disse ændringer, hvorved risikoen for en given transition beregnes mere præcist. Afhandlingens empiriske grundlag er registerdata for alle personer i Danmark, der tilhører fødselskohorterne 1968, 1972 og 1976. Data dækker over personlige, familiære forhold og oplysninger på arbejdspladsniveau, hvor den...

  1. Hvad nu hvis sommerfuglen er ekshibitionist?

    DEFF Research Database (Denmark)

    Rösing, Lilian Munk

    2008-01-01

    Groft sagt vil denne artikel, ved at aflæse hvad hhv. Inger Christensen, Sigmund Freud og Jacques Lacan har at sige om sommerfugle, argumentere for at der er et afgørende skel mellem naturvidenskab og litteraturvidenskab; mellem naturen, dyret, sommerfuglen på den ene side og bevidstheden...

  2. Hvad er hjemkundskabslæreren?

    DEFF Research Database (Denmark)

    Benn, Jette

    2009-01-01

        En appetizer: De praktisk-musiske fag på dagsordenen Temaet for denne artikel er hjemkundskabsfaget, -professionen og dens praktisering belyst gennem et interviewstudie fra 2008 af lærerstuderende og deres lærere. Dette studie relateres til forfatterens tidligere studie af fag og profession i...

  3. Er Storm P. en hardcore vagabond?

    DEFF Research Database (Denmark)

    Sortkær, Allan

    2002-01-01

    Den vagabond, som vi kender som Storm P.s, er ikke en figur, der kom fra en guddommelig inspiration eller deslige. Den var en allerede velkendt figur, før Storm P. tog den til sig, og figuren gennemgik radikale forandringer gennem Storm P.s liv: Krads social satire, hypervoldelig eller hyggelig...

  4. Er Frank rationel demokrat eller naiv fetichist?

    DEFF Research Database (Denmark)

    Rösing, Lilian Munk

    2014-01-01

    Artiklen er en diskussion af demokrati-begrebet i Rune Lykkebergs Alle har ret. Over for Lykkebregs demokrati-begreb sætter den Rancières og Zizeks forståelse af demokratiet som en åbning for det, den sociale orden ikke kan rumme. Det fører til en kritik af Lykkebergs anvendelse af begreberne om...

  5. Hvad er militær ledelse?

    DEFF Research Database (Denmark)

    Barfod, Jakob Rømer

    militær ledelse unik og derved muliggøre en opdeling, således at Forsvarets skoler kun underviser i den del af ledelse, som andre civile uddannelsesinstitutioner ikke menes at kunne undervise i. Men er det muligt at adskille begreber og praksis på den måde? Dette brief undersøger nogle betydninger af...

  6. Finanskrisen - er den markedsorienterede regulering skurken?

    DEFF Research Database (Denmark)

    Grosen, Anders

    2009-01-01

    markedsorienterede opgørelse tilsyneladende skyld i bankernes dårlige resultater. I hvert fald er standardsætteren inden for international regnskabsregulering, IASB, i efteråret 2008 på rekordtid kommet de betrængte virksomheder - herunder bankerne - til undsætning med en regnskabsmæssig hjælpepakke som supplement...

  7. Reconstruction of the complete ouabain-binding pocket of Na,K-ATPase in gastric H,K-ATPase by substitution of only seven amino acids.

    Science.gov (United States)

    Qiu, Li Yan; Krieger, Elmar; Schaftenaar, Gijs; Swarts, Herman G P; Willems, Peter H G M; De Pont, Jan Joep H H M; Koenderink, Jan B

    2005-09-16

    Although cardiac glycosides have been used as drugs for more than 2 centuries and their primary target, the sodium pump (Na,K-ATPase), has already been known for 4 decades, their exact binding site is still elusive. In our efforts to define the molecular basis of digitalis glycosides binding we started from the fact that a closely related enzyme, the gastric H,K-ATPase, does not bind glycosides like ouabain. Previously, we showed that a chimera of these two enzymes, in which only the M3-M4 and M5-M6 hairpins were of Na,K-ATPase, bound ouabain with high affinity (Koenderink, J. B., Hermsen, H. P. H., Swarts, H. G. P., Willems, P. H. G. M., and De Pont, J. J. H. H. M. (2000) Proc. Natl. Acad. Sci. U. S. A. 97, 11209-11214). We also demonstrated that only three amino acids (Phe(783), Thr(797), and Asp(804)) present in the M5-M6 hairpin of Na,K-ATPase were sufficient to confer high affinity ouabain binding to a chimera which contained in addition the M3-M4 hairpin of Na,K-ATPase (Qiu, L. Y., Koenderink, J. B., Swarts, H. G., Willems, P. H., and De Pont, J. J. H. H. M. (2003) J. Biol. Chem. 278, 47240-47244). To further pinpoint the ouabain-binding site here we used a chimera-based loss-of-function strategy and identified four amino acids (Glu(312), Val(314), Ile(315), Gly(319)), all present in M4, as being important for ouabain binding. In a final gain-of-function study we showed that a gastric H,K-ATPase that contained Glu(312), Val(314), Ile(315), Gly(319), Phe(783), Thr(797), and Asp(804) of Na,K-ATPase bound ouabain with the same affinity as the native enzyme. Based on the E(2)P crystal structure of Ca(2+)-ATPase we constructed a homology model for the ouabain-binding site of Na,K-ATPase involving all seven amino acids as well as several earlier postulated amino acids.

  8. Effects of material growth technique and Mg doping on Er3+ photoluminescence in Er-implanted GaN

    International Nuclear Information System (INIS)

    Kim, S.; Henry, R. L.; Wickenden, A. E.; Koleske, D. D.; Rhee, S. J.; White, J. O.; Myoung, J. M.; Kim, K.; Li, X.; Coleman, J. J.

    2001-01-01

    Photoluminescence (PL) and photoluminescence excitation (PLE) spectroscopies have been carried out at 6 K on the ∼1540 nm 4 I 13/2 - 4 I 15/2 emissions of Er 3+ in Er-implanted and annealed GaN. These studies revealed the existence of multiple Er 3+ centers and associated PL spectra in Er-implanted GaN films grown by metalorganic chemical vapor deposition, hydride vapor phase epitaxy, and molecular beam epitaxy. The results demonstrate that the multiple Er 3+ PL centers and below-gap defect-related absorption bands by which they are selectively excited are universal features of Er-implanted GaN grown by different techniques. It is suggested that implantation-induced defects common to all the GaN samples are responsible for the Er site distortions that give rise to the distinctive, selectively excited Er 3+ PL spectra. The investigations of selectively excited Er 3+ PL and PLE spectra have also been extended to Er-implanted samples of Mg-doped GaN grown by various techniques. In each of these samples, the so-called violet-pumped Er 3+ PL band and its associated broad violet PLE band are significantly enhanced relative to the PL and PLE of the other selectively excited Er 3+ PL centers. More importantly, the violet-pumped Er 3+ PL spectrum dominates the above-gap excited Er 3+ PL spectrum of Er-implanted Mg-doped GaN, whereas it was unobservable under above-gap excitation in Er-implanted undoped GaN. These results confirm the hypothesis that appropriate codopants can increase the efficiency of trap-mediated above-gap excitation of Er 3+ emission in Er-implanted GaN. [copyright] 2001 American Institute of Physics

  9. Endoplasmic reticulum protein targeting of phospholamban: a common role for an N-terminal di-arginine motif in ER retention?

    Directory of Open Access Journals (Sweden)

    Parveen Sharma

    2010-07-01

    Full Text Available Phospholamban (PLN is an effective inhibitor of the sarco(endoplasmic reticulum Ca(2+-ATPase, which transports Ca(2+ into the SR lumen, leading to muscle relaxation. A mutation of PLN in which one of the di-arginine residues at positions 13 and 14 was deleted led to a severe, early onset dilated cardiomyopathy. Here we were interested in determining the cellular mechanisms involved in this disease-causing mutation.Mutations deleting codons for either or both Arg13 or Arg14 resulted in the mislocalization of PLN from the ER. Our data show that PLN is recycled via the retrograde Golgi to ER membrane traffic pathway involving COP-I vesicles, since co-immunoprecipitation assays determined that COP I interactions are dependent on an intact di-arginine motif as PLN RDelta14 did not co-precipitate with COP I containing vesicles. Bioinformatic analysis determined that the di-arginine motif is present in the first 25 residues in a large number of all ER/SR Gene Ontology (GO annotated proteins. Mutations in the di-arginine motif of the Sigma 1-type opioid receptor, the beta-subunit of the signal recognition particle receptor, and Sterol-O-acyltransferase, three proteins identified in our bioinformatic screen also caused mislocalization of these known ER-resident proteins.We conclude that PLN is enriched in the ER due to COP I-mediated transport that is dependent on its intact di-arginine motif and that the N-terminal di-arginine motif may act as a general ER retrieval sequence.

  10. Altered expression and insulin-induced trafficking of Na+-K+-ATPase in rat skeletal muscle

    DEFF Research Database (Denmark)

    Galuska, Dana; Kotova, Olga; Barres, Romain

    2009-01-01

    Skeletal muscle Na(+)-K(+)-ATPase plays a central role in the clearance of K(+) from the extracellular fluid, therefore maintaining blood [K(+)]. Na(+)-K(+)-ATPase activity in peripheral tissue is impaired in insulin resistant states. We determined effects of high-fat diet (HFD) and exercise......(+)-K(+)-ATPase activity after 4 wk of HFD. Exercise training restored alpha(1)-, alpha(2)-, and beta(1)-subunit expression and Na(+)-K(+)-ATPase activity to control levels and reduced beta(2)-subunit expression 2.2-fold (P ... phospholemman. Phospholemman mRNA and protein expression were increased after HFD and restored to control levels after ET. Insulin-stimulated translocation of the alpha(2)-subunit to plasma membrane was impaired by HFD, whereas alpha(1)-subunit translocation remained unchanged. Alterations in sodium pump...

  11. Copper-transporting P-type ATPases use a unique ion-release pathway

    DEFF Research Database (Denmark)

    Andersson, Magnus; Mattle, Daniel; Sitsel, Oleg

    2014-01-01

    Heavy metals in cells are typically regulated by PIB-type ATPases. The first structure of the class, a Cu(+)-ATPase from Legionella pneumophila (LpCopA), outlined a copper transport pathway across the membrane, which was inferred to be occluded. Here we show by molecular dynamics simulations...... that extracellular water solvated the transmembrane (TM) domain, results indicative of a Cu(+)-release pathway. Furthermore, a new LpCopA crystal structure determined at 2.8-Å resolution, trapped in the preceding E2P state, delineated the same passage, and site-directed-mutagenesis activity assays support...... a functional role for the conduit. The structural similarities between the TM domains of the two conformations suggest that Cu(+)-ATPases couple dephosphorylation and ion extrusion differently than do the well-characterized PII-type ATPases. The ion pathway explains why certain Menkes' and Wilson's disease...

  12. Spatial distribution of Na+-K+-ATPase in dendritic spines dissected by nanoscale superresolution STED microscopy

    Directory of Open Access Journals (Sweden)

    Bondar Alexander

    2011-01-01

    Full Text Available Abstract Background The Na+,K+-ATPase plays an important role for ion homeostasis in virtually all mammalian cells, including neurons. Despite this, there is as yet little known about the isoform specific distribution in neurons. Results With help of superresolving stimulated emission depletion microscopy the spatial distribution of Na+,K+-ATPase in dendritic spines of cultured striatum neurons have been dissected. The found compartmentalized distribution provides a strong evidence for the confinement of neuronal Na+,K+-ATPase (α3 isoform in the postsynaptic region of the spine. Conclusions A compartmentalized distribution may have implications for the generation of local sodium gradients within the spine and for the structural and functional interaction between the sodium pump and other synaptic proteins. Superresolution microscopy has thus opened up a new perspective to elucidate the nature of the physiological function, regulation and signaling role of Na+,K+-ATPase from its topological distribution in dendritic spines.

  13. Plant P4-ATPases: lipid translocators with a role in membrane traficking

    DEFF Research Database (Denmark)

    Lopez Marques, Rosa Laura

    a large family of membrane proteins involved in pumping different physiologically-relevant substrates across biological membranes [4]. The members of the P4 subfamily (also known as flippases) catalyze the energy-driven translocation of lipids necessary for establishing transbilayer lipid asymmetry [5......], a feature necessary for correct functioning of the cells [6,7]. Deletion of one or more P4-ATPase genes causes defects in vesicle budding in various organisms [8-10] and some members of the yeast family have been shown to interact with the vesiculation machinery [11,12]. Thus, unraveling the key features...... of P4-ATPase functioning is crucial to understand the mechanisms underlying the whole secretory and endocytic pathways. In the model plant Arabidopsis, 12 members of the P4-ATPase family have been described (ALA1-ALA12, for Aminophospholipid ATPase) [4]. In the past years, we have characterized several...

  14. RIN4 functions with plasma membrane H+-ATPases to regulate stomatal apertures during pathogen attack

    DEFF Research Database (Denmark)

    Liu, Jun; Elmore, James M.; Fuglsang, Anja Thoe

    2009-01-01

    Abstract Pathogen perception by the plant innate immune system is of central importance to plant survival and productivity. The Arabidopsis protein RIN4 is a negative regulator of plant immunity. In order to identify additional proteins involved in RIN4- mediated immune signal transduction, we...... purified components of the RIN4 protein complex. We identified six novel proteins that had not previously been implicated in RIN4 signaling, including the plasma membrane (PM) H+-ATPases AHA1 and/or AHA2. RIN4 interacts with AHA1 and AHA2 both in vitro and in vivo. RIN4 overexpression and knockout lines...... exhibit differential PM H+-ATPase activity. PM H+-ATPase activation induces stomatal opening, enabling bacteria to gain entry into the plant leaf; inactivation induces stomatal closure thus restricting bacterial invasion. The rin4 knockout line exhibited reduced PM H+-ATPase activity and, importantly, its...

  15. The MLH1 ATPase domain is needed for suppressing aberrant formation of interstitial telomeric sequences.

    Science.gov (United States)

    Jia, Pingping; Chai, Weihang

    2018-05-01

    Genome instability gives rise to cancer. MLH1, commonly known for its important role in mismatch repair (MMR), DNA damage signaling and double-strand break (DSB) repair, safeguards genome stability. Recently we have reported a novel role of MLH1 in preventing aberrant formation of interstitial telomeric sequences (ITSs) at intra-chromosomal regions. Deficiency in MLH1, in particular its N-terminus, leads to an increase of ITSs. Here, we identify that the ATPase activity in the MLH1 N-terminal domain is important for suppressing the formation of ITSs. The ATPase activity is also needed for recruiting MLH1 to DSBs. Moreover, defective ATPase activity of MLH1 causes an increase in micronuclei formation. Our results highlight the crucial role of MLH1's ATPase domain in preventing the aberrant formation of telomeric sequences at the intra-chromosomal regions and preserving genome stability. Copyright © 2018 Elsevier B.V. All rights reserved.

  16. Targeting vacuolar H+-ATPases as a new strategy against cancer.

    Science.gov (United States)

    Fais, Stefano; De Milito, Angelo; You, Haiyan; Qin, Wenxin

    2007-11-15

    Growing evidence suggests a key role of tumor acidic microenvironment in cancer development, progression, and metastasis. As a consequence, the need for compounds that specifically target the mechanism(s) responsible for the low pH of tumors is increasing. Among the key regulators of the tumor acidic microenvironment, vacuolar H(+)-ATPases (V-ATPases) play an important role. These proteins cover a number of functions in a variety of normal as well as tumor cells, in which they pump ions across the membranes. We discuss here some recent results showing that a molecular inhibition of V-ATPases by small interfering RNA in vivo as well as a pharmacologic inhibition through proton pump inhibitors led to tumor cytotoxicity and marked inhibition of human tumor growth in xenograft models. These results propose V-ATPases as a key target for new strategies in cancer treatment.

  17. Preparation of a highly concentrated, completely monomeric, active sarcoplasmic reticulum Ca2+-ATPase.

    Science.gov (United States)

    Lüdi, H; Hasselbach, W

    1985-11-21

    Sarcoplasmic reticulum vesicles from fast skeletal muscle were partially delipidated with sodium cholate at high ionic strength and sedimented in a discontinuous sucrose gradient. Phospholipid content was reduced from 0.777 mumol/mg protein to 0.242 mumol/mg protein. As judged from gel electrophoresis and high pressure liquid gel chromatography, accessory proteins were removed during centrifugation and the Ca2+-ATPase was obtained in an almost pure form. Addition of myristoylglycerophosphocholine (1 mg/mg protein) reactivates ATPase and dinitrophenylphosphatase activity to the same degree obtained with native vesicles. Using the analytical ultracentrifuge it could be demonstrated that the reactivated Ca2+-ATPase was present exclusively in a monomeric state. These results were obtained at high and low ionic strength and up to a protein concentration of 10 mg/ml. Therefore this preparation should be very useful to investigate differences between oligomeric and monomeric Ca2+-ATPase.

  18. Plasma Membrane H(+)-ATPase Regulation in the Center of Plant Physiology.

    Science.gov (United States)

    Falhof, Janus; Pedersen, Jesper Torbøl; Fuglsang, Anja Thoe; Palmgren, Michael

    2016-03-07

    The plasma membrane (PM) H(+)-ATPase is an important ion pump in the plant cell membrane. By extruding protons from the cell and generating a membrane potential, this pump energizes the PM, which is a prerequisite for growth. Modification of the autoinhibitory terminal domains activates PM H(+)-ATPase activity, and on this basis it has been hypothesized that these regulatory termini are targets for physiological factors that activate or inhibit proton pumping. In this review, we focus on the posttranslational regulation of the PM H(+)-ATPase and place regulation of the pump in an evolutionary and physiological context. The emerging picture is that multiple signals regulating plant growth interfere with the posttranslational regulation of the PM H(+)-ATPase. Copyright © 2016 The Author. Published by Elsevier Inc. All rights reserved.

  19. The AAA protein spastin possesses two levels of basal ATPase activity.

    Science.gov (United States)

    Fan, Xiangyu; Lin, Zhijie; Fan, Guanghui; Lu, Jing; Hou, Yongfei; Habai, Gulijiazi; Sun, Linyue; Yu, Pengpeng; Shen, Yuequan; Wen, Maorong; Wang, Chunguang

    2018-04-30

    The AAA ATPase spastin is a microtubule-severing enzyme that plays important roles in various cellular events including axon regeneration. Herein, we found that the basal ATPase activity of spastin is negatively regulated by spastin concentration. By determining a spastin crystal structure, we demonstrate the necessity of intersubunit interactions between spastin AAA domains. Neutralization of the positive charges in the microtubule-binding domain (MTBD) of spastin dramatically decreases the ATPase activity at low concentration, although the ATP-hydrolyzing potential is not affected. These results demonstrate that, in addition to the AAA domain, the MTBD region of spastin is also involved in regulating ATPase activity, making interactions between spastin protomers more complicated than expected. © 2018 Federation of European Biochemical Societies.

  20. Radionuclide assay of membrane Na+, K+-ATPase activity of peserved red blood cells

    International Nuclear Information System (INIS)

    Trusov, V.V.; Zelenin, A.A.; Marizin, S.A.

    1986-01-01

    The radionuclide tests were used to investigate the influence of varying blood preservatives on erythrocylic membrane Na + , K + -ATPase activity in samples of whole blood and packed red blood cells from normal donors prepared by standard methods. The tests were performed before and after seven days of preservation under standard conditions. It was found that blood preservations lowered membrane Na + , K + -ATPase activity: its minimum reduction was recorded with citroglucopnosphate, while glugicir induced a significant drop in Na + , K + -ATPase activity of preserved red blood cells regardless of the type of the blood transfusion solution. The assay of membrane Na + , K + -ATPase activity of preserved red blood cells with the use of 86 Rb could be recommended as an evaluation test for preserved blood and its components

  1. CAMEX-4 ER-2 HIGH ALTITUDE DROPSONDE V1

    Data.gov (United States)

    National Aeronautics and Space Administration — The CAMEX-4 ER-2 High Altitude Dropsonde dataset was collected by the ER-2 High Altitude Dropsonde System (EHAD), which used dropwinsondes fitted with Global...

  2. CAMEX-4 ER-2 LIGHTNING INSTRUMENT PACKAGE (LIP) V1

    Data.gov (United States)

    National Aeronautics and Space Administration — The CAMEX-4 ER-2 Lightning Instrument Package (LIP) dataset was collected by the ER-2 LIP, which allows the vector components of the electric field (i.e, Ex, Ey, Ez...

  3. Facets of operational performance in an emergency room (ER)

    NARCIS (Netherlands)

    van der Vaart, Taco; Vastag, Gyula; Wijngaard, Jacob

    This paper, using detailed time measurements of patients complemented by interviews with hospital management and staff, examines three facets of an emergency room's (ER) operational performance: (1) effectiveness of the triage system in rationing patient treatment; (2) factors influencing ER's

  4. ER Consolidated Qtrly Rpt_April-June 2016_October 2016

    Energy Technology Data Exchange (ETDEWEB)

    Cochran, John R. [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States)

    2016-10-01

    This Environmental Restoration Operations (ER) Consolidated Quarterly Report (ER Quarterly Report) provides the status of ongoing corrective action activities being implemented by Sandia National Laboratories, New Mexico (SNL/NM) for the April, May, and June 2016 quarterly reporting period.

  5. GPM Ground Validation Navigation Data ER-2 OLYMPEX V1

    Data.gov (United States)

    National Aeronautics and Space Administration — The GPM Ground Validation NASA ER-2 Navigation Data OLYMPEX dataset supplies navigation data collected by the NASA ER-2 aircraft for flights that occurred during...

  6. Senior Sport er læring i virkeligheden

    DEFF Research Database (Denmark)

    Meyer, Erna Rosenlund

    2014-01-01

    Samarbejde mellem fysioterapeutudddannelsen på UCN og Aalborg senior Sport er givende for alle parter, inklusiv studerende.......Samarbejde mellem fysioterapeutudddannelsen på UCN og Aalborg senior Sport er givende for alle parter, inklusiv studerende....

  7. Role of insulin in regulation of Na+-/K+-dependent ATPase activity and pump function in corneal endothelial cells.

    Science.gov (United States)

    Hatou, Shin; Yamada, Masakazu; Akune, Yoko; Mochizuki, Hiroshi; Shiraishi, Atsushi; Joko, Takeshi; Nishida, Teruo; Tsubota, Kazuo

    2010-08-01

    The Na(+)-/K(+)-dependent ATPase (Na,K-ATPase) expressed in the basolateral membrane of corneal endothelial cells plays an important role in the pump function of the corneal endothelium. The role of insulin in the regulation of Na,K-ATPase activity and pump function in corneal endothelial cells was investigated. Confluent monolayers of mouse corneal endothelial cells were exposed to insulin. ATPase activity was evaluated by spectrophotometric measurement of phosphate released from ATP with the use of ammonium molybdate; Na,K-ATPase activity was defined as the portion of total ATPase activity sensitive to ouabain. Pump function was measured with the use of a Ussing chamber; pump function attributable to Na,K-ATPase activity was defined as the portion of the total short-circuit current sensitive to ouabain. Western blot analysis and immunocytochemistry were performed to measure the expression of the Na,K-ATPase alpha(1)-subunit. Insulin increased the Na,K-ATPase activity and pump function of cultured corneal endothelial cells. These effects were blocked by protein kinase C (PKC) inhibitors and protein phosphatases 1 and 2A inhibitor. Western blot analysis indicated that insulin decreased the ratio of the inactive Na,K-ATPase alpha(1)-subunit. Immunocytochemistry indicated that insulin increased the cell surface expression of the Na,K-ATPase alpha(1)-subunit. These results suggest that insulin increases the Na,K-ATPase activity and pump function of cultured corneal endothelial cells. The effect of insulin is mediated by PKC and presumably results in the activation of PP1, 2A, or both, which are essential for activating Na,K-ATPase by alpha(1)-subunit dephosphorylation.

  8. Response of membrane-bound ATPase of Micrococcus luteus to heat and ultraviolet light

    International Nuclear Information System (INIS)

    Volotovskij, J.; Risi, S.; Dose, K.

    1976-01-01

    It is shown that the properties of ATPase (EC 3.6.1.3) of Micrococcus luteus depend only to some extent on the state of the membrane to which it is attached. Its interaction with the membrane appears to be largely controlled by polar forces. It is shown, however, that the UV-sensitivity of the membrane-bound ATPase is also significantly influenced by the state of membrane lipids. (orig.) [de

  9. Response of membrane-bound ATPase of Micrococcus luteus to heat and ultraviolet light

    Energy Technology Data Exchange (ETDEWEB)

    Volotovskii, J; Risi, S; Dose, K [Mainz Univ. (F.R. Germany). Inst. fuer Biochemie

    1976-03-01

    It is shown that the properties of ATPase (EC 3.6.1.3) of Micrococcus luteus depend only to some extent on the state of the membrane to which it is attached. Its interaction with the membrane appears to be largely controlled by polar forces. It is shown, however, that the UV-sensitivity of the membrane-bound ATPase is also significantly influenced by the state of membrane lipids.

  10. The V-ATPase a2-subunit as a putative endosomal pH-sensor.

    Science.gov (United States)

    Marshansky, V

    2007-11-01

    V-ATPase (vesicular H(+)-ATPase)-driven intravesicular acidification is crucial for vesicular trafficking. Defects in vesicular acidification and trafficking have recently been recognized as essential determinants of various human diseases. An important role of endosomal acidification in receptor-ligand dissociation and in activation of lysosomal hydrolytic enzymes is well established. However, the molecular mechanisms by which luminal pH information is transmitted to the cytosolic small GTPases that control trafficking events such as budding, coat formation and fusion are unknown. Here, we discuss our recent discovery that endosomal V-ATPase is a pH-sensor regulating the degradative pathway. According to our model, V-ATPase is responsible for: (i) the generation of a pH gradient between vesicular membranes; (ii) sensing of intravesicular pH; and (iii) transmitting this information to the cytosolic side of the membrane. We also propose the hypothetical molecular mechanism involved in function of the V-ATPase a2-subunit as a putative pH-sensor. Based on extensive experimental evidence on the crucial role of histidine residues in the function of PSPs (pH-sensing proteins) in eukaryotic cells, we hypothesize that pH-sensitive histidine residues within the intra-endosomal loops and/or C-terminal luminal tail of the a2-subunit could also be involved in the pH-sensing function of V-ATPase. However, in order to identify putative pH-sensitive histidine residues and to test this hypothesis, it is absolutely essential that we increase our understanding of the folding and transmembrane topology of the a-subunit isoforms of V-ATPase. Thus the crucial role of intra-endosomal histidine residues in pH-dependent conformational changes of the V-ATPase a2-isoform, its interaction with cytosolic small GTPases and ultimately in its acidification-dependent regulation of the endosomal/lysosomal protein degradative pathway remain to be determined.

  11. Na/K-ATPase Signaling and Salt Sensitivity: The Role of Oxidative Stress

    Directory of Open Access Journals (Sweden)

    Jiang Liu

    2017-03-01

    Full Text Available Other than genetic regulation of salt sensitivity of blood pressure, many factors have been shown to regulate renal sodium handling which contributes to long-term blood pressure regulation and have been extensively reviewed. Here we present our progress on the Na/K-ATPase signaling mediated sodium reabsorption in renal proximal tubules, from cardiotonic steroids-mediated to reactive oxygen species (ROS-mediated Na/K-ATPase signaling that contributes to experimental salt sensitivity.

  12. Photoaffinity labelling of a small protein component of a purified (Na+-K+)ATPase

    International Nuclear Information System (INIS)

    Rogers, T.B.; Lazdunski, M.

    1979-01-01

    Studies have been carried out on the photoaffinity labelling of the (Na + -K + )ATPase from the electric organ of Electrophorus electricus. The aims were to see if different photoaffinity labels of the ouabain binding site, are capable of labelling a small protein component and to know if there is a small protein component, in addition to the major protein chains with molecular weights in the regions of 100 000 and 50 000, which is present in other purified (Na + -K + )ATPase preparations. (Auth.)

  13. Er frygten for Kina vores eneste håb?

    DEFF Research Database (Denmark)

    Mehlsen, Camilla

    2011-01-01

    Man siger, at håbet er lysegrønt, men i det 21. århundrede er håbet rødt som riget i midten. Mens optimismen vinder frem i Asien med Kina som bannerfører, er Vesten ramt af frygt, og det forplanter sig negativt til uddannelsessystemet. Er positive følelser hemmeligheden bag, at asiatiske lande st...... stormer frem i internationale undersøgelser som PISA?...

  14. Functional interaction of nicotinic acetylcholine receptors and Na+/K+ ATPase from Locusta migratoria manilensis (Meyen).

    Science.gov (United States)

    Bao, Haibo; Sun, Huahua; Xiao, Youxin; Zhang, Yixi; Wang, Xin; Xu, Xiaoyong; Liu, Zewen; Fang, Jichao; Li, Zhong

    2015-03-06

    Associated proteins are important for the correct functioning of nicotinic acetylcholine receptors (nAChRs). In the present study, a neonicotinoid-agarose affinity column was used to isolate related proteins from a solubilized membrane preparation from the nervous system of Locusta migratoria manilensis (Meyen). 1530 peptides were identified and most of them were involved in the membranous structure, molecular interaction and cellular communication. Among these peptides, Na(+)/K(+) ATPase had the highest MASCOT score and were involved in the molecular interaction, which suggested that Na(+)/K(+) ATPase and nAChRs might have strong and stable interactions in insect central nervous system. In the present study, functional interactions between nAChRs and Na(+)/K(+) ATPase were examined by heterologous expression in Xenopus oocytes. The results showed that the activated nAChRs increased pump currents of Na(+)/K(+) ATPase, which did not require current flow through open nAChRs. In turn, Na(+)/K(+) ATPase significantly increased agonist sensitivities of nAChRs in a pump activity-independent manner and reduced the maximum current (Imax) of nAChRs. These findings provide novel insights concerning the functional interactions between insect nAChRs and Na(+)/K(+) ATPase.

  15. Milrinone and thyroid hormone stimulate myocardial membrane Ca2+-ATPase activity and share structural homologies.

    Science.gov (United States)

    Mylotte, K M; Cody, V; Davis, P J; Davis, F B; Blas, S D; Schoenl, M

    1985-01-01

    We have recently shown that thyroid hormone in physiological concentrations stimulates sarcolemma-enriched rabbit-myocardial-membrane Ca2+-ATPase in vitro. In this study, milrinone [2-methyl-5-cyano-(3,4'-bipyridin)-6(1H)-one], a cardiac inotropic agent, was thyromimetic in the same system. At clinically achievable concentrations (50-500 nM), milrinone significantly stimulated membrane Ca2+-ATPase in vitro. This action was antagonized by W-7 [N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide], an agent that also blocks thyroid hormone action on the Ca2+-ATPase, at concentrations as low as 5 microM. Progressive additions of milrinone to membranes incubated with a fixed concentration of thyroxine (0.10 nM) or triiodothyronine resulted in a progressive obliteration of the thyroid hormone effect on Ca2+-ATPase. Amrinone [5-amino-(3,4'-bipyridin)-6(1H)-one], the parent bipyridine of milrinone, had no effect on myocardial Ca2+-ATPase activity. X-ray crystallographic analysis of milrinone and amrinone revealed structural homologies between the phenolic ring of thyroxine and the substituted ring of milrinone, whereas amrinone did not share these homologies. The mechanism(s) of the inotropic actions of thyroxine and of milrinone is not clearly understood, but these observations implicate Ca2+-ATPase, a calcium pump-associated enzyme, as one mediator of the effects on the heart of these two compounds. PMID:2933747

  16. The Role of the Plasma Membrane H+-ATPase in Plant Responses to Aluminum Toxicity

    Directory of Open Access Journals (Sweden)

    Jiarong Zhang

    2017-10-01

    Full Text Available Aluminum (Al toxicity is a key factor limiting plant growth and crop production on acid soils. Increasing the plant Al-detoxification capacity and/or breeding Al-resistant cultivars are a cost-effective strategy to support crop growth on acidic soils. The plasma membrane H+-ATPase plays a central role in all plant physiological processes. Changes in the activity of the plasma membrane H+-ATPase through regulating the expression and phosphorylation of this enzyme are also involved in many plant responses to Al toxicity. The plasma membrane H+-ATPase mediated H+ influx may be associated with the maintenance of cytosolic pH and the plasma membrane gradients as well as Al-induced citrate efflux mediated by a H+-ATPase-coupled MATE co-transport system. In particular, modulating the activity of plasma membrane H+-ATPase through application of its activators (e.g., magnesium or IAA or using transgenics has effectively enhanced plant resistance to Al stress in several species. In this review, we critically assess the available knowledge on the role of the plasma membrane H+-ATPase in plant responses to Al stress, incorporating physiological and molecular aspects.

  17. Mechanisms of Rose Bengal inhibition on SecA ATPase and ion channel activities.

    Science.gov (United States)

    Hsieh, Ying-Hsin; Huang, Ying-Ju; Jin, Jin-Shan; Yu, Liyan; Yang, Hsiuchin; Jiang, Chun; Wang, Binghe; Tai, Phang C

    2014-11-14

    SecA is an essential protein possessing ATPase activity in bacterial protein translocation for which Rose Bengal (RB) is the first reported sub-micromolar inhibitor in ATPase activity and protein translocation. Here, we examined the mechanisms of inhibition on various forms of SecA ATPase by conventional enzymatic assays, and by monitoring the SecA-dependent channel activity in the semi-physiological system in cells. We build on the previous observation that SecA with liposomes form active protein-conducting channels in the oocytes. Such ion channel activity is enhanced by purified Escherichia coli SecYEG-SecDF·YajC liposome complexes. Inhibition by RB could be monitored, providing correlation of in vitro activity and intact cell functionality. In this work, we found the intrinsic SecA ATPase is inhibited by RB competitively at low ATP concentration, and non-competitively at high ATP concentrations while the translocation ATPase with precursors and SecYEG is inhibited non-competitively by RB. The Inhibition by RB on SecA channel activity in the oocytes with exogenous ATP-Mg(2+), mimicking translocation ATPase activity, is also non-competitive. The non-competitive inhibition on channel activity has also been observed with SecA from other bacteria which otherwise would be difficult to examine without the cognate precursors and membranes. Copyright © 2014 Elsevier Inc. All rights reserved.

  18. Plasmalemmal V-H+-ATPases regulate intracellular pH in human lung microvascular endothelial cells

    International Nuclear Information System (INIS)

    Rojas, Jose D.; Sennoune, Souad R.; Maiti, Debasish; Martinez, Gloria M.; Bakunts, Karina; Wesson, Donald E.; Martinez-Zaguilan, Raul

    2004-01-01

    The lung endothelium layer is exposed to continuous CO 2 transit which exposes the endothelium to a substantial acid load that could be detrimental to cell function. The Na + /H + exchanger and HCO 3 - -dependent H + -transporting mechanisms regulate intracellular pH (pH cyt ) in most cells. Cells that cope with high acid loads might require additional primary energy-dependent mechanisms. V-H + -ATPases localized at the plasma membranes (pmV-ATPases) have emerged as a novel pH regulatory system. We hypothesized that human lung microvascular endothelial (HLMVE) cells use pmV-ATPases, in addition to Na + /H + exchanger and HCO 3 - -based H + -transporting mechanisms, to maintain pH cyt homeostasis. Immunocytochemical studies revealed V-H + -ATPase at the plasma membrane, in addition to the predicted distribution in vacuolar compartments. Acid-loaded HLMVE cells exhibited proton fluxes in the absence of Na + and HCO 3 - that were similar to those observed in the presence of either Na + , or Na + and HCO 3 - . The Na + - and HCO 3 - -independent pH cyt recovery was inhibited by bafilomycin A 1 , a V-H + -ATPase inhibitor. These studies show a Na + - and HCO 3 - -independent pH cyt regulatory mechanism in HLMVE cells that is mediated by pmV-ATPases

  19. Production and characterization of a monoclonal antibody to H+ATPase

    International Nuclear Information System (INIS)

    Yurko, M.; Fitch, F; Gluck, S.

    1986-01-01

    Acidification of endocytic vesicles is carried out by an ATP-dependent proton pump, H+ATPase, an FOF1 type enzyme comprised of at least 5 major subunits of 70, 56, 45, 35, and 17 kDa. A monoclonal antibody, H6.1, to H+ATPase from bovine kidney medulla, was raised to enable the structural characterization and localization of the pump. Several criteria were used to show that H6.1 recognized H+ATPase. 1.) H6.1 immunoprecipitated N-ethylmaleimide-sensitive and vanadate- and azide-insensitive solubilized ATPase activity (and GTPase activity) from both crude and purified enzyme preparations. 2.) H6.1 immunoprecipitated oligomycin-insensitive ATP-dependent proton transporting vesicles made from bovine kidney medulla, rat kidney, and CHO cells. 3.) H6.1 specifically immuno-precipitated the 5 subunits of H+ATPase from a partially purified preparation of the enzyme that had been labelled with I-125. H6.1 was then used as an immunocytochemical probe for the localization of H+ATPase. In bovine kidney medullary collecting duct, there was an intense apical staining of selected cells. In proximal tubule and in cultured CHO cells there was a granular pattern of staining characteristic of endocytic vesicles and lysosomes, suggesting that the kidney and CHO cell proton pumps are structurally related

  20. Molecular architecture of the N-type ATPase rotor ring from Burkholderia pseudomallei.

    Science.gov (United States)

    Schulz, Sarah; Wilkes, Martin; Mills, Deryck J; Kühlbrandt, Werner; Meier, Thomas

    2017-04-01

    The genome of the highly infectious bacterium Burkholderia pseudomallei harbors an atp operon that encodes an N-type rotary ATPase, in addition to an operon for a regular F-type rotary ATPase. The molecular architecture of N-type ATPases is unknown and their biochemical properties and cellular functions are largely unexplored. We studied the B. pseudomallei N 1 N o -type ATPase and investigated the structure and ion specificity of its membrane-embedded c-ring rotor by single-particle electron cryo-microscopy. Of several amphiphilic compounds tested for solubilizing the complex, the choice of the low-density, low-CMC detergent LDAO was optimal in terms of map quality and resolution. The cryoEM map of the c-ring at 6.1 Å resolution reveals a heptadecameric oligomer with a molecular mass of ~141 kDa. Biochemical measurements indicate that the c 17 ring is H + specific, demonstrating that the ATPase is proton-coupled. The c 17 ring stoichiometry results in a very high ion-to-ATP ratio of 5.7. We propose that this N-ATPase is a highly efficient proton pump that helps these melioidosis-causing bacteria to survive in the hostile, acidic environment of phagosomes. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

  1. Stabilisation of Na,K-ATPase structure by the cardiotonic steroid ouabain

    International Nuclear Information System (INIS)

    Miles, Andrew J.; Fedosova, Natalya U.; Hoffmann, Søren V.; Wallace, B.A.; Esmann, Mikael

    2013-01-01

    Highlights: •Ouabain binding to pig and shark Na,K-ATPase enhances thermal stability. •Ouabain stabilises both membrane-bound and solubilised Na,K-ATPase. •Synchrotron radiation circular dichroism is used for structure determination. •Secondary structure in general is not affected by ouabain binding. •Stabilisation is due to re-arrangement of tertiary structure. -- Abstract: Cardiotonic steroids such as ouabain bind with high affinity to the membrane-bound cation-transporting P-type Na,K-ATPase, leading to complete inhibition of the enzyme. Using synchrotron radiation circular dichroism spectroscopy we show that the enzyme-ouabain complex is less susceptible to thermal denaturation (unfolding) than the ouabain-free enzyme, and this protection is observed with Na,K-ATPase purified from pig kidney as well as from shark rectal glands. It is also shown that detergent-solubilised preparations of Na,K-ATPase are stabilised by ouabain, which could account for the successful crystallisation of Na,K-ATPase in the ouabain-bound form. The secondary structure is not significantly affected by the binding of ouabain. Ouabain appears however, to induce a reorganization of the tertiary structure towards a more compact protein structure which is less prone to unfolding; recent crystal structures of the two enzymes are consistent with this interpretation. These circular dichroism spectroscopic studies in solution therefore provide complementary information to that provided by crystallography

  2. Stabilisation of Na,K-ATPase structure by the cardiotonic steroid ouabain

    Energy Technology Data Exchange (ETDEWEB)

    Miles, Andrew J. [Institute of Structural and Molecular Biology, Birkbeck College, University of London, London WC1E 7HX (United Kingdom); Fedosova, Natalya U. [Department of Biomedicine, Aarhus University, DK-8000 Aarhus (Denmark); Hoffmann, Søren V. [ISA, Department of Physics and Astronomy, Aarhus University, DK-8000 Aarhus (Denmark); Wallace, B.A. [Institute of Structural and Molecular Biology, Birkbeck College, University of London, London WC1E 7HX (United Kingdom); Esmann, Mikael, E-mail: me@biophys.au.dk [Department of Biomedicine, Aarhus University, DK-8000 Aarhus (Denmark)

    2013-05-31

    Highlights: •Ouabain binding to pig and shark Na,K-ATPase enhances thermal stability. •Ouabain stabilises both membrane-bound and solubilised Na,K-ATPase. •Synchrotron radiation circular dichroism is used for structure determination. •Secondary structure in general is not affected by ouabain binding. •Stabilisation is due to re-arrangement of tertiary structure. -- Abstract: Cardiotonic steroids such as ouabain bind with high affinity to the membrane-bound cation-transporting P-type Na,K-ATPase, leading to complete inhibition of the enzyme. Using synchrotron radiation circular dichroism spectroscopy we show that the enzyme-ouabain complex is less susceptible to thermal denaturation (unfolding) than the ouabain-free enzyme, and this protection is observed with Na,K-ATPase purified from pig kidney as well as from shark rectal glands. It is also shown that detergent-solubilised preparations of Na,K-ATPase are stabilised by ouabain, which could account for the successful crystallisation of Na,K-ATPase in the ouabain-bound form. The secondary structure is not significantly affected by the binding of ouabain. Ouabain appears however, to induce a reorganization of the tertiary structure towards a more compact protein structure which is less prone to unfolding; recent crystal structures of the two enzymes are consistent with this interpretation. These circular dichroism spectroscopic studies in solution therefore provide complementary information to that provided by crystallography.

  3. Effect of endurance swimming on rat cardiac myofibrillar ATPase with experimental diabetes.

    Science.gov (United States)

    Belcastro, A N; Maybank, P; Rossiter, M; Secord, D

    1985-09-01

    Diabetes is characterized by depressed cardiac functional properties attributed to Ca2+-activated ATPase activity. In contrast, endurance swimming enhances the cardiac functional properties and Ca2+-activated myofibril ATPase. Thus, the purpose of this study was to observe if the changes associated with experimental diabetes can be ameliorated with training. Diabetes was induced with a single i.v. injection of streptozotocin (60 mg/kg). Blood and urine glucose concentrations were 802 +/- 44 and 6965 +/- 617 mg/dL, respectively. The training control and training diabetic animals were made to swim (+/- 2% body weight) 4 days/week for 8 weeks. Cardiac myofibril, at 10 microM free Ca2+ concentration was reduced by 54% in the sedentary diabetics compared with sedentary control animals (p less than 0.05). Swim training enhanced the Ca2+-activated myofibril ATPase activities for the normal animals. The diabetic animals, which swam for 8 weeks, had further reduced their Ca2+-activated myofibril ATPase activity when compared with sedentary diabetics (p less than 0.05). Similarly, the Mg2+-stimulated myofibril ATPase activity was depressed by 31% in diabetics following endurance swimming. It is concluded that the depressed Ca2+-activated myofibril ATPase activity of diabetic hearts is not reversible with endurance swimming.

  4. Glycolytic control of vacuolar-type ATPase activity: A mechanism to regulate influenza viral infection

    Energy Technology Data Exchange (ETDEWEB)

    Kohio, Hinissan P.; Adamson, Amy L., E-mail: aladamso@uncg.edu

    2013-09-15

    As new influenza virus strains emerge, finding new mechanisms to control infection is imperative. In this study, we found that we could control influenza infection of mammalian cells by altering the level of glucose given to cells. Higher glucose concentrations induced a dose-specific increase in influenza infection. Linking influenza virus infection with glycolysis, we found that viral replication was significantly reduced after cells were treated with glycolytic inhibitors. Addition of extracellular ATP after glycolytic inhibition restored influenza infection. We also determined that higher levels of glucose promoted the assembly of the vacuolar-type ATPase within cells, and increased vacuolar-type ATPase proton-transport activity. The increase of viral infection via high glucose levels could be reversed by inhibition of the proton pump, linking glucose metabolism, vacuolar-type ATPase activity, and influenza viral infection. Taken together, we propose that altering glucose metabolism may be a potential new approach to inhibit influenza viral infection. - Highlights: • Increased glucose levels increase Influenza A viral infection of MDCK cells. • Inhibition of the glycolytic enzyme hexokinase inhibited Influenza A viral infection. • Inhibition of hexokinase induced disassembly the V-ATPase. • Disassembly of the V-ATPase and Influenza A infection was bypassed with ATP. • The state of V-ATPase assembly correlated with Influenza A infection of cells.

  5. Purification, characterization and crystallization of the F-ATPase from Paracoccus denitrificans.

    Science.gov (United States)

    Morales-Rios, Edgar; Watt, Ian N; Zhang, Qifeng; Ding, Shujing; Fearnley, Ian M; Montgomery, Martin G; Wakelam, Michael J O; Walker, John E

    2015-09-01

    The structures of F-ATPases have been determined predominantly with mitochondrial enzymes, but hitherto no F-ATPase has been crystallized intact. A high-resolution model of the bovine enzyme built up from separate sub-structures determined by X-ray crystallography contains about 85% of the entire complex, but it lacks a crucial region that provides a transmembrane proton pathway involved in the generation of the rotary mechanism that drives the synthesis of ATP. Here the isolation, characterization and crystallization of an integral F-ATPase complex from the α-proteobacterium Paracoccus denitrificans are described. Unlike many eubacterial F-ATPases, which can both synthesize and hydrolyse ATP, the P. denitrificans enzyme can only carry out the synthetic reaction. The mechanism of inhibition of its ATP hydrolytic activity involves a ζ inhibitor protein, which binds to the catalytic F₁-domain of the enzyme. The complex that has been crystallized, and the crystals themselves, contain the nine core proteins of the complete F-ATPase complex plus the ζ inhibitor protein. The formation of crystals depends upon the presence of bound bacterial cardiolipin and phospholipid molecules; when they were removed, the complex failed to crystallize. The experiments open the way to an atomic structure of an F-ATPase complex. © 2015 The Authors.

  6. Glycolytic control of vacuolar-type ATPase activity: A mechanism to regulate influenza viral infection

    International Nuclear Information System (INIS)

    Kohio, Hinissan P.; Adamson, Amy L.

    2013-01-01

    As new influenza virus strains emerge, finding new mechanisms to control infection is imperative. In this study, we found that we could control influenza infection of mammalian cells by altering the level of glucose given to cells. Higher glucose concentrations induced a dose-specific increase in influenza infection. Linking influenza virus infection with glycolysis, we found that viral replication was significantly reduced after cells were treated with glycolytic inhibitors. Addition of extracellular ATP after glycolytic inhibition restored influenza infection. We also determined that higher levels of glucose promoted the assembly of the vacuolar-type ATPase within cells, and increased vacuolar-type ATPase proton-transport activity. The increase of viral infection via high glucose levels could be reversed by inhibition of the proton pump, linking glucose metabolism, vacuolar-type ATPase activity, and influenza viral infection. Taken together, we propose that altering glucose metabolism may be a potential new approach to inhibit influenza viral infection. - Highlights: • Increased glucose levels increase Influenza A viral infection of MDCK cells. • Inhibition of the glycolytic enzyme hexokinase inhibited Influenza A viral infection. • Inhibition of hexokinase induced disassembly the V-ATPase. • Disassembly of the V-ATPase and Influenza A infection was bypassed with ATP. • The state of V-ATPase assembly correlated with Influenza A infection of cells

  7. Biochemical characterization of the plasma membrane H+ - ATPase from red beet (Beta vulgaris) hypocotyl tissue

    International Nuclear Information System (INIS)

    Oleski, N.A.

    1986-01-01

    Several biochemical techniques including selective solubilization followed by gel filtration or various types of affinity chromatography, and antibody production were employed in an attempt to purify the plasma membrane H + - ATPase from red beet hypocotyl tissue. While the enzyme could not be purified using any of these methods, it was possible to successfully conduct a more detailed biochemical analysis of the H + - ATPase. The molecular weight and isoelectric point of the enzyme were determined using N,N'dicyclohexylcarbodiimide (DCCD) and a H + - ATPase antibody. When plasma membrane vesicles were incubated with 20 μM [ 14 C]-DCCD at 0 C, a single 97,000 dalton protein was apparent on a fluorograph. A close correlation between [ 14 C]-DCCD labelling of the 97,000 dalton protein and the extent of ATPase inhibition over a range of DCCD concentrations suggests that this 97,000 dalton protein is a component of the plasma membrane H + - ATPase. An antibody raised against the plasma membrane H + - ATPase of Neurospora crassa cross-reacted with the 97,000 dalton DCCD-binding protein, further supporting the identity of this protein. Immunoblots of two dimensional gels of red beet plasma membrane vesicles indicated the isoelectric point of the enzyme to be pH 6.5

  8. Erythrocyte membrane ATPase and calcium pumping activities in porcine malignant hyperthermia

    International Nuclear Information System (INIS)

    Thatte, H.S.; Mickelson, J.R.; Addis, P.B.; Louis, C.F.

    1987-01-01

    To investigate possible abnormalities in erythrocyte membrane enzyme activities in the pharmacogenetic disorder MH, membrane ATPase activities have been examined in erythrocyte ghosts prepared from red blood cells of MHS and normal swine. While no differences were noted in Mg2+-ATPase activities, the (Na+, K+)-ATPase activity of MHS erythrocyte ghosts was less than that of normal ghosts. Ca2+-ATPase activity exhibited low- and high-affinity Ca2+-binding sites in both types of erythrocyte ghost. While the Km for Ca2+ was greater for normal than for MHS erythrocyte ghosts at the high-affinity Ca2+-binding site, the reverse was true at the low-affinity Ca2+-binding site. Irrespective of the type of calcium binding site occupied, the Vmax for normal erythrocyte ghost Ca2+-ATPase activity was greater than that for MHS ghosts. In the presence of calmodulin, there was now no difference between MHS and normal erythrocyte ghosts in either the Km for Ca2+ or the Vmax of the Ca2+-ATPase activity. To determine if the calcium pumping activity of intact MHS and normal pig erythrocytes differed, calcium efflux from the 45 Ca-loaded erythrocytes was determined; this activity was significantly greater for MHS than for normal erythrocytes. Thus, the present study confirms that there are abnormalities in the membranes of MHS pig red blood cells. However, we conclude that these abnormalities are unlikely to result in an impaired ability of MHS erythrocytes to regulate their cytosolic Ca2+ concentration

  9. TCSP ER-2 DOPPLER RADAR (EDOP) V1

    Data.gov (United States)

    National Aeronautics and Space Administration — The TCSP ER-2 DOPPLER RADAR (EDOP) dataset was collected by the ER-2 Doppler radar (EDOP), which is an X-band (9.6 GHz) Doppler radar mounted in the nose of the ER-2...

  10. CAMEX-4 ER-2 DOPPLER RADAR V1

    Data.gov (United States)

    National Aeronautics and Space Administration — The CAMEX-4 ER-2 Doppler Radar dataset was collected by the ER-2 Doppler radar (EDOP), which is an X-band (9.6 GHz) Doppler radar mounted in the nose of ER-2. The...

  11. SEC16 in COPII coat dynamics at ER exit sites

    NARCIS (Netherlands)

    Sprangers, Joep; Rabouille, Catherine

    Protein export from the endoplasmic reticulum (ER), the first step in protein transport through the secretory pathway, is mediated by coatomer protein II (COPII)-coated vesicles at ER exit sites. COPII coat assembly on the ER is well understood and the conserved large hydrophilic protein Sec16

  12. Magnetic properties of ErGa3

    International Nuclear Information System (INIS)

    Murasik, A.; Czopnik, A.; Keller, L.; Fischer, P.

    1999-01-01

    Bulk magnetization measurements and magnetic phase diagram for ErGa 3 show that zero-external magnetic field it undergoes two successive transitions at T 1 = 2.6 K and T 2 = 2.8 K, respectively. Its magnetic ordering examined by neutron diffraction, can be derived from the so-called [1/2, 1/2, 0] structure, i.e. one in which the successive antiparallel (110) sheets of spins have additionally superimposed on them a sinusoidal modulation parallel to the [100] axis. The temperature dependence of neutron diffraction diagrams studied with powder and single crystal samples revealed, that in the range of (2.6 - 2.78) k there occurs an abrupt reorientation of the Er 3+ spins from the [110]-type direction, towards the [100] axis. This rotation can be attributed to the T 1 transition found in the magnetic phase diagram. (author)

  13. SYP73 Anchors the ER to the Actin Cytoskeleton for Maintenance of ER Integrity and Streaming in Arabidopsis.

    Science.gov (United States)

    Cao, Pengfei; Renna, Luciana; Stefano, Giovanni; Brandizzi, Federica

    2016-12-05

    The endoplasmic reticulum (ER) is an essential organelle that spreads throughout the cytoplasm as one interconnected network of narrow tubules and dilated cisternae that enclose a single lumen. The ER network undergoes extensive remodeling, which critically depends on membrane-cytoskeleton interactions [1]. In plants, the ER is also highly mobile, and its streaming contributes significantly to the movement of other organelles [2, 3]. The remodeling and motility of the plant ER rely mainly on actin [4] and to a minor extent on microtubules [5]. Although a three-way interaction between the ER, cytosolic myosin-XI, and F-actin mediates the plant ER streaming [6], the mechanisms underlying stable interaction of the ER membrane with actin are unknown. Early electron microscopy studies suggested a direct attachment of the plant ER with actin filaments [7, 8], but it is plausible that yet-unknown proteins facilitate anchoring of the ER membrane with the cytoskeleton. We demonstrate here that SYP73, a member of the plant Syp7 subgroup of SNARE proteins [9] containing actin-binding domains, is a novel ER membrane-associated actin-binding protein. We show that overexpression of SYP73 causes a striking rearrangement of the ER over actin and that, similar to mutations of myosin-XI [4, 10, 11], loss of SYP73 reduces ER streaming and affects overall ER network morphology and plant growth. We propose a model for plant ER remodeling whereby the dynamic rearrangement and streaming of the ER network depend on the propelling action of myosin-XI over actin coupled with a SYP73-mediated bridging, which dynamically anchors the ER membrane with actin filaments. Copyright © 2016 Elsevier Ltd. All rights reserved.

  14. Virtual Reality er dit nye Skype

    DEFF Research Database (Denmark)

    Bødker, Mads

    2016-01-01

    VR er mediet, der kan gøre den flade skærm og det begrænsede blik i for eksempel Skype til et lidt gammeldags digitalt samarbejde. Ved at sætte folk i stand til at kommunikere, dele virtuelle genstande og give dem en øget oplevelse af at være i et rum sammen, kan man mediere nye aspekter af samar...

  15. Her er din motion i 2012

    DEFF Research Database (Denmark)

    Suhr, Lasse Kristian

    2014-01-01

    Motion i naturen, en indendørs fitnessform inspireret af junglens lianer og høj intensitet med crossfit. Det er ifølge en ekspert nogle af de træningstyper, der vil være mest populære i 2012. Året hvor fleksibilitet og frihed vil blive endnu vigtigere, for den måde vi holder os i form på....

  16. Selskaberne er en del af staten

    DEFF Research Database (Denmark)

    Hein Jessen, Mathias

    2016-01-01

    Lækket af Panama Papers fremhæver endnu engang den paradoksale rolle, ikke mindst multinationale selskaber spiller i politik og økonomi: De er både skabt af staten, og fremstår stadig som uden for og i opposition til staten. Hvis vi vil gøre noget ved problemet, må vi forstå, hvordan stat og...

  17. Hamlet er manden i mit liv

    DEFF Research Database (Denmark)

    Rösing, Lilian Munk

    2009-01-01

    Hamlet er manden i mit liv. Min elskede. Jeg har en drøm om at frelse ham. At redde ham ud af sin mors soveværelse og ind i mit. Væk fra det ødipale Helvede han befinder sig i, fastlåst mellem et svin af en faderskikkelse (Claudius) og en alt for idealiseret far (gamle Hamlet), vågende over sin m...

  18. Electrorheological (ER) Fluids: A Research Needs Assessment

    Science.gov (United States)

    1993-05-01

    is given as line (g). Experimental results a correspond to polyaniline in silicone oil, e.c•,=36 (Gow 1990); * hollow silica spheres in corn oil ye...Malishauskas, 1981) (Fig. 18b). In Minsk, we have designed a new model of a screw valve (Shulman, 1978b). Its electrodes are fabricated in the form of...Takimoto, both of whom were very gracious. Dr. Takimoto is primarily involved in ER research. I was not shown any facilities while there, primarily

  19. Musikkens Hus i Aalborg er demensvenligt

    DEFF Research Database (Denmark)

    Ridder, Hanne Mette Ochsner

    2017-01-01

    . Der blev afholdt frokostmøder hvor medarbejdere og brugere af Musikkens Hus fik undervisning om demens. Der blev også udvekslet gode idéer til hvad man i Musikkens Hus kan gøre for at byde mennesker med demens velkomne. at vi skal læse en masse, gå på kurser eller forpligte os til at være besøgsven...

  20. ER-12-1 completion report

    International Nuclear Information System (INIS)

    Russell, C.E.; Gillespie, D.; Cole, J.C.; Drellack, S.L.

    1996-12-01

    The objective of drillhole ER-12-1 was to determine the hydrogeology of paleozoic carbonate rocks and of the Eleana Formation, a regional aquitard, in an area potentially downgradient from underground nuclear testing conducted in nearby Rainier Mesa. This objective was addressed through the drilling of well ER-12-1 at N886,640.26 E640,538.85 Nevada Central Coordinates. Drilling of the 1094 m (3588 ft) well began on July 19, 1991 and was completed on October 17, 1991. Drilling problems included hole deviation and hole instability that prevented the timely completion of this borehole. Drilling methods used include rotary tri-cone and rotary hammer drilling with conventional and reverse circulation using air/water, air/foam (Davis mix), and bentonite mud. Geologic cuttings and geophysical logs were obtained from the well. The rocks penetrated by the ER-12-1 drillhole are a complex assemblage of Silurian, Devonian, and Mississippian sedimentary rocks that are bounded by numerous faults that show substantial stratigraphic offset. The final 7.3 m (24 ft) of this hole penetrated an unusual intrusive rock of Cretaceous age. The geology of this borehole was substantially different from that expected, with the Tongue Wash Fault encountered at a much shallower depth, paleozoic rocks shuffled out of stratigraphic sequence, and the presence of an altered biotite-rich microporphyritic igneous rock at the bottom of the borehole. Conodont CAI analyses and rock pyrolysis analyses indicate that the carbonate rocks in ER-12-1, as well as the intervening sheets of Eleana siltstone, have been thermally overprinted following movement on the faults that separate them. The probable source of heat for this thermal disturbance is the microporphyritic intrusion encountered at the bottom of the hole, and its age establishes that the major fault activity must have occurred prior to 102.3+0.5 Ma (middle Cretaceous)

  1. Friluftsinstallationer i træer

    DEFF Research Database (Denmark)

    Skov, Simon; Thomsen, Iben Margrete

    2015-01-01

    Abebaner, treetop walking, high rope adventure, slackline, træhuse og parkour-baner er alle installationer, som bruger træerne som støtte, enten midlertidigt eller permanent. Den nye sport, turistattraktionen eller team building-redskabet sætter i alle tilfælde træerne på prøve. Der følger positiv...

  2. The role of the AR/ER ratio in ER-positive breast cancer patients.

    Science.gov (United States)

    Rangel, Nelson; Rondon-Lagos, Milena; Annaratone, Laura; Osella-Abate, Simona; Metovic, Jasna; Mano, Maria Piera; Bertero, Luca; Cassoni, Paola; Sapino, Anna; Castellano, Isabella

    2018-03-01

    The significance of androgen receptor (AR) in breast cancer (BC) management is not fully defined, and it is still ambiguous how the level of AR expression influences oestrogen receptor-positive (ER+) tumours. The aim of the present study was to analyse the prognostic impact of AR/ER ratio, evaluated by immunohistochemistry (IHC), correlating this value with clinical, pathological and molecular characteristics. We retrospectively selected a cohort of 402 ER+BC patients. On each tumour, IHC analyses for AR, ER, PgR, HER2 and Ki67 were performed and AR+ cases were used to calculate the AR/ER value. A cut-off of ≥2 was selected using receiver-operating characteristic (ROC) curve analyses. RNA from 19 cases with AR/ER≥2 was extracted and used for Prosigna-PAM50 assays. Tumours with AR/ER≥2 (6%) showed more frequent metastatic lymph nodes, larger size, higher histological grade and lower PgR levels than cases with AR/ERAR/ER≥2 had worse disease-free interval (DFI) and disease-specific survival (DSS) (hazard ratios (HR) = 4.96 for DFI and HR = 8.69 for DSS, both P  ≤ 0.004). According to the Prosigna-PAM50 assay, 63% (12/19) of these cases resulted in intermediate or high risk of recurrence categories. Additionally, although all samples were positive for ER assessed by IHC, the molecular test assigned 47.4% (9/19) of BCs to intrinsic non-luminal subtypes. In conclusion, the AR/ER ratio ≥2 identifies a subgroup of patients with aggressive biological features and may represent an additional independent marker of worse BC prognosis. Moreover, the Prosigna-PAM50 results indicate that a significant number of cases with AR/ER≥2 could be non-luminal tumours. © 2018 Society for Endocrinology.

  3. Torque Generation of Enterococcus hirae V-ATPase*

    Science.gov (United States)

    Ueno, Hiroshi; Minagawa, Yoshihiro; Hara, Mayu; Rahman, Suhaila; Yamato, Ichiro; Muneyuki, Eiro; Noji, Hiroyuki; Murata, Takeshi; Iino, Ryota

    2014-01-01

    V-ATPase (VoV1) converts the chemical free energy of ATP into an ion-motive force across the cell membrane via mechanical rotation. This energy conversion requires proper interactions between the rotor and stator in VoV1 for tight coupling among chemical reaction, torque generation, and ion transport. We developed an Escherichia coli expression system for Enterococcus hirae VoV1 (EhVoV1) and established a single-molecule rotation assay to measure the torque generated. Recombinant and native EhVoV1 exhibited almost identical dependence of ATP hydrolysis activity on sodium ion and ATP concentrations, indicating their functional equivalence. In a single-molecule rotation assay with a low load probe at high ATP concentration, EhVoV1 only showed the “clear” state without apparent backward steps, whereas EhV1 showed two states, “clear” and “unclear.” Furthermore, EhVoV1 showed slower rotation than EhV1 without the three distinct pauses separated by 120° that were observed in EhV1. When using a large probe, EhVoV1 showed faster rotation than EhV1, and the torque of EhVoV1 estimated from the continuous rotation was nearly double that of EhV1. On the other hand, stepping torque of EhV1 in the clear state was comparable with that of EhVoV1. These results indicate that rotor-stator interactions of the Vo moiety and/or sodium ion transport limit the rotation driven by the V1 moiety, and the rotor-stator interactions in EhVoV1 are stabilized by two peripheral stalks to generate a larger torque than that of isolated EhV1. However, the torque value was substantially lower than that of other rotary ATPases, implying the low energy conversion efficiency of EhVoV1. PMID:25258315

  4. Kinesin-2 KIF3AB exhibits novel ATPase characteristics.

    Science.gov (United States)

    Albracht, Clayton D; Rank, Katherine C; Obrzut, Steven; Rayment, Ivan; Gilbert, Susan P

    2014-10-03

    KIF3AB is an N-terminal processive kinesin-2 family member best known for its role in intraflagellar transport. There has been significant interest in KIF3AB in defining the key principles that underlie the processivity of KIF3AB in comparison with homodimeric processive kinesins. To define the ATPase mechanism and coordination of KIF3A and KIF3B stepping, a presteady-state kinetic analysis was pursued. For these studies, a truncated murine KIF3AB was generated. The results presented show that microtubule association was fast at 5.7 μm(-1) s(-1), followed by rate-limiting ADP release at 12.8 s(-1). ATP binding at 7.5 μm(-1) s(-1) was followed by an ATP-promoted isomerization at 84 s(-1) to form the intermediate poised for ATP hydrolysis, which then occurred at 33 s(-1). ATP hydrolysis was required for dissociation of the microtubule·KIF3AB complex, which was observed at 22 s(-1). The dissociation step showed an apparent affinity for ATP that was very weak (K½,ATP at 133 μm). Moreover, the linear fit of the initial ATP concentration dependence of the dissociation kinetics revealed an apparent second-order rate constant at 0.09 μm(-1) s(-1), which is inconsistent with fast ATP binding at 7.5 μm(-1) s(-1) and a Kd ,ATP at 6.1 μm. These results suggest that ATP binding per se cannot account for the apparent weak K½,ATP at 133 μm. The steady-state ATPase Km ,ATP, as well as the dissociation kinetics, reveal an unusual property of KIF3AB that is not yet well understood and also suggests that the mechanochemistry of KIF3AB is tuned somewhat differently from homodimeric processive kinesins. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  5. ER stress, autophagy, and RNA viruses

    Directory of Open Access Journals (Sweden)

    Jia-Rong eJheng

    2014-08-01

    Full Text Available Endoplasmic reticulum (ER stress is a general term for representing the pathway by which various stimuli affect ER functions. ER stress induces the evolutionarily conserved signaling pathways, called the unfolded protein response (UPR, which compromises the stimulus and then determines whether the cell survives or dies. In recent years, ongoing research has suggested that these pathways may be linked to the autophagic response, which plays a key role in the cell’s response to various stressors. Autophagy performs a self-digestion function, and its activation protects cells against certain pathogens. However, the link between the UPR and autophagy may be more complicated. These two systems may act dependently, or the induction of one system may interfere with the other. Experimental studies have found that different viruses modulate these mechanisms to allow them to escape the host immune response or, worse, to exploit the host’s defense to their advantage; thus, this topic is a critical area in antiviral research. In this review, we summarize the current knowledge about how RNA viruses, including influenza virus, poliovirus, coxsackievirus, enterovirus 71, Japanese encephalitis virus, hepatitis C virus, and dengue virus, regulate these processes. We also discuss recent discoveries and how these will produce novel strategies for antiviral treatment.

  6. Sund livsstil er mere end fysik

    DEFF Research Database (Denmark)

    Nybo, Lars; Thing, Lone Friis; Astrup, Arne

    2015-01-01

    . I virkeligheden hænger tingene sammen, og i et holistisk sundhedsbillede er de psykiske og sociale dimensioner mindst lige så vigtige som de fysiske. Fællesskabet under og efter fodboldtræningen kan for eksempel være en lige så vigtig del af at være aktiv som selve træningen. Og selv om nogle...... begynder på en aktiv livsstil ud fra en ydre motivation om det fysiske aspekt af ’mere sundhed’, så er der meget, som tyder på, at den indre motivation overtager, og at de i stedet bliver drevet af glæden ved den øgede livskvalitet. Nogle mennesker fascineres af at følge en fast træningsplan og have et...... behandling af inaktivitetsrelaterede lidelser bør dog ikke være det vigtigste argument for at oplyse om og opfordre til en sund livsstil. Livskvalitetsaspektet skal være i højsædet, og det er for så vidt en legitim bekymring, at der kommer et overdrevet fokus på samfundsøkonomi og forebyggelse af somatisk...

  7. Na/K Pump and Beyond: Na/K-ATPase as a Modulator of Apoptosis and Autophagy.

    Science.gov (United States)

    Felippe Gonçalves-de-Albuquerque, Cassiano; Ribeiro Silva, Adriana; Ignácio da Silva, Camila; Caire Castro-Faria-Neto, Hugo; Burth, Patrícia

    2017-04-21

    Lung cancer is a leading cause of global cancer deaths. Na/K-ATPase has been studied as a target for cancer treatment. Cardiotonic steroids (CS) trigger intracellular signalling upon binding to Na/K-ATPase. Normal lung and tumour cells frequently express different pump isoforms. Thus, Na/K-ATPase is a powerful target for lung cancer treatment. Drugs targeting Na/K-ATPase may induce apoptosis and autophagy in transformed cells. We argue that Na/K-ATPase has a role as a potential target in chemotherapy in lung cancer treatment. We discuss the effects of Na/K-ATPase ligands and molecular pathways inducing deleterious effects on lung cancer cells, especially those leading to apoptosis and autophagy.

  8. Curcumin modulation of Na,K-ATPase: phosphoenzyme accumulation, decreased K+ occlusion, and inhibition of hydrolytic activity

    DEFF Research Database (Denmark)

    Mahmmoud, Yasser Ahmed

    2005-01-01

    Curcumin, the major constitute of tumeric, is an important nutraceutical that has been shown to be useful in the treatment of many diseases. As an inhibitor of the sarcoplasmic reticulum Ca2+-ATPase, curcumin was shown to correct cystic fibrosis (CF) defects in some model systems, whereas others...... have reported no or little effects on CF after curcumin treatment, suggesting that curcumin effect is not due to simple inhibition of the Ca2+-ATPase. We tested the hypothesis that curcumin may modulate other members of the P2-type ATPase superfamily by studying the effects of curcumin on the activity...... and kinetic properties of the Na,K-ATPase. Curcumin treatment inhibited Na,K-ATPase activity in a dose-dependent manner (K0.514.6 M). Curcumin decreased the apparent affinity of Na,K-ATPase for K+ and increased it for Na+ and ATP. Kinetic analyses indicated that curcumin induces a three-fold reduction...

  9. Na/K Pump and Beyond: Na/K-ATPase as a Modulator of Apoptosis and Autophagy

    Directory of Open Access Journals (Sweden)

    Cassiano Felippe Gonçalves-de-Albuquerque

    2017-04-01

    Full Text Available Lung cancer is a leading cause of global cancer deaths. Na/K-ATPase has been studied as a target for cancer treatment. Cardiotonic steroids (CS trigger intracellular signalling upon binding to Na/K-ATPase. Normal lung and tumour cells frequently express different pump isoforms. Thus, Na/K-ATPase is a powerful target for lung cancer treatment. Drugs targeting Na/K-ATPase may induce apoptosis and autophagy in transformed cells. We argue that Na/K-ATPase has a role as a potential target in chemotherapy in lung cancer treatment. We discuss the effects of Na/K-ATPase ligands and molecular pathways inducing deleterious effects on lung cancer cells, especially those leading to apoptosis and autophagy.

  10. Quaternary structure of the ATPase complex of human 26S proteasomes determined by chemical cross-linking

    DEFF Research Database (Denmark)

    Hartmann-Petersen, R; Tanaka, K; Hendil, K B

    2001-01-01

    and substrate specificity. Among the approximately 18 subunits of PA700 regulator, six are ATPases. The ATPases presumably recognize, unfold, and translocate substrates into the interior of the 26S proteasome. It is generally believed that the ATPases form a hexameric ring. By means of chemical cross......-linking, immunoprecipitation, and blotting, we have determined that the ATPases are organized in the order S6-S6'-S10b-S8-S4-S7. Additionally, we found cross-links between the ATPase S10b and the 20S proteasome subunit alpha6. Together with the previously known interaction between S8 and alpha1 and between S4 and alpha7......, these data establish the relative orientations of ATPases with respect to the 20S proteasome....

  11. The ATPase of the phi29 DNA packaging motor is a member of the hexameric AAA+ superfamily.

    Science.gov (United States)

    Schwartz, Chad; De Donatis, Gian Marco; Fang, Huaming; Guo, Peixuan

    2013-08-15

    The AAA+ superfamily of proteins is a class of motor ATPases performing a wide range of functions that typically exist as hexamers. The ATPase of phi29 DNA packaging motor has long been a subject of debate in terms of stoichiometry and mechanism of action. Here, we confirmed the stoichiometry of phi29 motor ATPase to be a hexamer and provide data suggesting that the phi29 motor ATPase is a member of the classical hexameric AAA+ superfamily. Native PAGE, EMSA, capillary electrophoresis, ATP titration, and binomial distribution assay show that the ATPase is a hexamer. Mutations in the known Walker motifs of the ATPase validated our previous assumptions that the protein exists as another member of this AAA+ superfamily. Our data also supports the finding that the phi29 DNA packaging motor uses a revolution mechanism without rotation or coiling (Schwartz et al., this issue). Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

  12. The structure of the 168Er nucleus and the 166Er(t,p) 168 Er reaction in terms of the sdg interacting boson model

    Science.gov (United States)

    Akiyama, Y.; Heyde, K.; Arima, A.; Yoshinaga, N.

    1986-05-01

    Extending the interacting boson model by incorporating besides s and d, also the g-boson, we can describe the population of positive parity states of 168Er in the 166Er(t,P) 168Er reaction rather well. In particular, the excitation of I,Kπi = 4,3 +1; 2,2 +2; 0,0 +3 and 0,0 +4 states is much improved over the sd-IBM approach.

  13. MicroRNAs meet calcium: joint venture in ER proteostasis.

    Science.gov (United States)

    Finger, Fabian; Hoppe, Thorsten

    2014-11-04

    The endoplasmic reticulum (ER) is a cellular compartment that has a key function in protein translation and folding. Maintaining its integrity is of fundamental importance for organism's physiology and viability. The dynamic regulation of intraluminal ER Ca(2+) concentration directly influences the activity of ER-resident chaperones and stress response pathways that balance protein load and folding capacity. We review the emerging evidence that microRNAs play important roles in adjusting these processes to frequently changing intracellular and environmental conditions to modify ER Ca(2+) handling and storage and maintain ER homeostasis. Copyright © 2014, American Association for the Advancement of Science.

  14. Vidensledelse er også en social praksis

    DEFF Research Database (Denmark)

    Lauring, Jakob; Waldstrøm, Christian

    2006-01-01

    lys er det utroligt vigtigt at personalefunktionen i virksomheden er klar over de processer der udspiller sig, og aktivt tager fat om problemerne der ligger i disse barrierer. Vores egne undersøgelser viser, at på trods af opstillingen af teknologiske systemer til vidensdeling, er der stadig...... væsentlige ledelsesmæssige opgaver i forhold til sociale aspekter ved arbejdet i en organisation, som skal varetages hvis vidensdelingen skal fremmes succesfuldt. Det er væsentligt at forstå, at vidensledelse ikke blot handler om cirkulering af information, men også er forbundet mere generelt til ledelse som...

  15. H,K-ATPase and carbonic anhydrase response to chronic systemic rat gastric hypoxia

    Directory of Open Access Journals (Sweden)

    Ulfah Lutfiah

    2015-11-01

    Full Text Available Background: Hypoxia may induce gastric ulcer associated with excessive hidrogen chloride (HCl secretion. Synthesis of HCl involves 2 enzymes, H,K-ATPase and carbonic anhydrase (CA. This study aimed to clarify the underlying cause of gastric ulcer in chronic hypoxic condition, by investigating the H,K-ATPase and CA9 response in rats.Methods: This study was an in vivo experiment, to know the relationship between hypoxia to expression of H,K-ATPase and CA9 mRNA, and H,K-ATPase and total CA specific activity of chronic systemic rat gastric hypoxia. The result was compared to control. Data was analyzed by SPSS. If the data distribution was normal and homogeneous, ANOVA and LSD post-hoc test were used. However, if the distribution was not normal and not homogeneous, and still as such after transformation, data was treated in non-parametric using Kruskal-Wallis and Mann Whitney test. Twenty five male Sprague-Dawley rats were divided into 5 groups: rats undergoing hypoxia for 1, 3, 5, and 7 days placed in hypoxia chamber (10% O2, 90% N2, and one control group. Following this treatment, stomach of the rats was extracted and homogenized. Expression of H,K-ATPase and CA9 mRNA was measured using real time RT-PCR. Specific activity of H,K-ATPase was measured using phosphate standard solution, and specific activity of total CA was measured using p-nitrophenol solution.Results: The expression of H,K-ATPase mRNA was higher in the first day (2.159, and drastically lowered from the third to seventh day (0.289; 0.108; 0.062. Specific activities of H,K-ATPase was slightly higher in the first day (0.765, then was lowered in the third (0.685 and fifth day (0.655, and was higher in the seventh day (0.884. The expression of CA9 mRNA was lowered progressively from the first to seventh day (0.84; 0.766; 0.736; 0.343. Specific activities of total CA was low in the first day (0.083, and was higher from the third to seventh day (0.111; 0.136; 0.144.Conclusion: In hypoxia

  16. Crystal structure of the sodium-potassium pump (Na+,K+-ATPase) with bound potassium and ouabain

    OpenAIRE

    Ogawa, Haruo; Shinoda, Takehiro; Cornelius, Flemming; Toyoshima, Chikashi

    2009-01-01

    The sodium-potassium pump (Na+,K+-ATPase) is responsible for establishing Na+ and K+ concentration gradients across the plasma membrane and therefore plays an essential role in, for instance, generating action potentials. Cardiac glycosides, prescribed for congestive heart failure for more than 2 centuries, are efficient inhibitors of this ATPase. Here we describe a crystal structure of Na+,K+-ATPase with bound ouabain, a representative cardiac glycoside, at 2.8 Å resolution in a state analog...

  17. Evaluation of Na+, K+-ATPase activity in the brain of young rats after acute administration of fenproporex

    OpenAIRE

    Rezin, Gislaine T.; Scaini, Giselli; Gonçalves, Cinara L.; Ferreira, Gabriela K.; Cardoso, Mariane R.; Ferreira, Andréa G.K.; Cunha, Maira J.; Schmitz, Felipe; Varela, Roger B.; Quevedo, João; Wyse, Angela T.S.; Streck, Emilio L.

    2013-01-01

    Objectives: Fenproporex is an amphetamine-based anorectic which is rapidly converted into amphetamine in vivo. Na+, K+-ATPase is a membrane-bound enzyme necessary to maintain neuronal excitability. Considering that the effects of fenproporex on brain metabolism are poorly known and that Na+, K+-ATPase is essential for normal brain function, this study sought to evaluate the effect of this drug on Na+, K+-ATPase activity in the hippocampus, hypothalamus, prefrontal cortex, and striatum of youn...

  18. Postirradiation changes in the systems of active ion transport. Na,K-ATPase of neurons in neuroglia

    International Nuclear Information System (INIS)

    Shainskaya, A.M.; Dvoretskij, A.I.; Valetova, Yu.O.

    1989-01-01

    A study was made of the effect of X-radiation (0.31 C/kg and 3.875 C/kg) on Na,K-ATPase in fractions enriched with neurons and neuroglia. The results show the impairment of the neuronal-glial relationship in Na,K-ATPase activity. The most important differences in the pattern of changes in Na,K-ATPase system of brain cells were followed up after irradiaion with lethal and sublethal doses

  19. Regulation of potassium dependent ATPase (kdp) operon of Deinococcus radiodurans.

    Science.gov (United States)

    Dani, Pratiksha; Ujaoney, Aman Kumar; Apte, Shree Kumar; Basu, Bhakti

    2017-01-01

    The genome of D. radiodurans harbors genes for structural and regulatory proteins of Kdp ATPase, in an operon pattern, on Mega plasmid 1. Organization of its two-component regulatory genes is unique. Here we demonstrate that both, the structural as well as regulatory components of the kdp operon of D. radiodurans are expressed quickly as the cells experience potassium limitation but are not expressed upon increase in osmolarity. The cognate DNA binding response regulator (RR) effects the expression of kdp operon during potassium deficiency through specific interaction with the kdp promoter. Deletion of the gene encoding RR protein renders the mutant D. radiodurans (ΔRR) unable to express kdp operon under potassium limitation. The ΔRR D. radiodurans displays no growth defect when grown on rich media or when exposed to oxidative or heat stress but shows reduced growth following gamma irradiation. The study elucidates the functional and regulatory aspects of the novel kdp operon of this extremophile, for the first time.

  20. Active transport of Na+ by reconstituted Na,K-ATPase

    International Nuclear Information System (INIS)

    Boldyrev, A.A.; Svinukhova, I.A.

    1987-01-01

    The ability of ATP, CTP, ITP, GTP, and UTP to support ouabain-sensitive accumulation of Na + by proteoliposomes with a reconstituted Na/K-pump was investigated. At a low [Na + ]/[K + ] ratio in the medium (20 mM/50 mM), a correlation is observed between the proton-accepting capacity of the nucleotide and its effectiveness as a substrate of active transport. To test the hypothesis of the importance of the presence of a negative charge in the 1-position of the purine (3-pyrimidine) base of the nucleotide for mutual transitions between the Na- and K-conformations of Na,K-ATPase they used two analogs of ATP: N 1 -hydroxy-ATP, possessing proton acceptor capacity, and N 1 -methoxy-ATP, in the molecule of which the negative charge is quenched by a methyl group. The first substrate supports active accumulation of Na + in proteoliposomes at the same rate as ATP, whereas the second substrate is relatively ineffective

  1. Æstetik er tilbage som nøgle til at forstå samtiden

    DEFF Research Database (Denmark)

    Sharma, Devika

    2013-01-01

    Casper Christensen er gakket, Hello Kitty er nuttet og konceptkunsten er interessant. Her er tre nye kategorier at forstå senkapitalismen med. I anledning af Sianne Ngais bog "Our aesthetic categories: zany, cute, interesting" .......Casper Christensen er gakket, Hello Kitty er nuttet og konceptkunsten er interessant. Her er tre nye kategorier at forstå senkapitalismen med. I anledning af Sianne Ngais bog "Our aesthetic categories: zany, cute, interesting" ....

  2. Novel Role for Na,K-ATPase in Phosphatidylinositol 3-Kinase Signaling and Suppression of Cell Motility

    OpenAIRE

    Barwe, Sonali P.; Anilkumar, Gopalakrishnapillai; Moon, Sun Y.; Zheng, Yi; Whitelegge, Julian P.; Rajasekaran, Sigrid A.; Rajasekaran, Ayyappan K.

    2005-01-01

    The Na,K-ATPase, consisting of α- and β-subunits, regulates intracellular ion homeostasis. Recent studies have demonstrated that Na,K-ATPase also regulates epithelial cell tight junction structure and functions. Consistent with an important role in the regulation of epithelial cell structure, both Na,K-ATPase enzyme activity and subunit levels are altered in carcinoma. Previously, we have shown that repletion of Na,K-ATPase β1-subunit (Na,K-β) in highly motile Moloney sarcoma virus-transforme...

  3. Reconstruction of the complete ouabain-binding pocket of Na,K-ATPase in gastric H,K-ATPase by substitution of only seven amino acids

    NARCIS (Netherlands)

    Qiu, L.Y.; Krieger, E.; Schaftenaar, G.; Swarts, H.G.P.; Willems, P.H.G.M.; Pont, J.J.H.H.M. de; Koenderink, J.B.

    2005-01-01

    Although cardiac glycosides have been used as drugs for more than 2 centuries and their primary target, the sodium pump (Na, K-ATPase), has already been known for 4 decades, their exact binding site is still elusive. In our efforts to define the molecular basis of digitalis glycosides binding we

  4. Reconstruction of the complete ouabain-binding pocket of Na,K-ATPase in gastric H,K-ATPase by substitution of only seven amino acids.

    NARCIS (Netherlands)

    Qiu, L.; Krieger, E.; Schaftenaar, G.; Swarts, H.G.P.; Willems, P.H.G.M.; Pont, J.J.H.H.M. de; Koenderink, J.B.

    2005-01-01

    Although cardiac glycosides have been used as drugs for more than 2 centuries and their primary target, the sodium pump (Na,K-ATPase), has already been known for 4 decades, their exact binding site is still elusive. In our efforts to define the molecular basis of digitalis glycosides binding we

  5. Properties of the ATPase activity associated with peroxisome-enriched fractions from rat liver: comparison with mitochondrial F1F0-ATPase

    NARCIS (Netherlands)

    Wolvetang, E. J.; Wanders, R. J.; Schutgens, R. B.; Berden, J. A.; Tager, J. M.

    1990-01-01

    Highly purified peroxisomal fractions from rat liver contain ATPase activity (18.8 +/- 0.1 nmol/min per mg, n = 6). This activity is about 2% of that found in purified mitochondrial fractions. Measurement of marker enzyme activities and immunoblotting of the peroxisomal fraction with an antiserum

  6. Na,K-ATPase structure/function relationships probed by the denaturant urea.

    Science.gov (United States)

    Esmann, Mikael; Fedosova, Natalya U; Olesen, Claus

    2015-05-01

    Urea interacts with the Na,K-ATPase, leading to reversible as well as irreversible inhibition of the hydrolytic activity. The enzyme purified from shark rectal glands is more sensitive to urea than Na,K-ATPase purified from pig kidney. An immediate and reversible inhibition under steady-state conditions of hydrolytic activity at 37°C is demonstrated for the three reactions studied: the overall Na,K-ATPase activity, the Na-ATPase activity observed in the absence of K+ as well as the K+-dependent phosphatase reaction (K-pNPPase) seen in the absence of Na+. Half-maximal inhibition is seen with about 1M urea for shark enzyme and about 2M urea for pig enzyme. In the presence of substrates there is also an irreversible inhibition in addition to the reversible process, and we show that ATP protects against the irreversible inhibition for both the Na,K-ATPase and Na-ATPase reaction, whereas the substrate paranitrophenylphosphate leads to a slight increase in the rate of irreversible inhibition of the K-pNPPase. The rate of the irreversible inactivation in the absence of substrates is much more rapid for shark enzyme than for pig enzyme. The larger number of potentially urea-sensitive hydrogen bonds in shark enzyme compared to pig enzyme suggests that interference with the extensive hydrogen bonding network might account for the higher urea sensitivity of shark enzyme. The reversible inactivation is interpreted in terms of domain interactions and domain accessibilities using as templates the available crystal structures of Na,K-ATPase. It is suggested that a few interdomain hydrogen bonds are those mainly affected by urea during reversible inactivation. Copyright © 2014 Elsevier B.V. All rights reserved.

  7. Active ingredients in Chinese medicines promoting blood circulation as Na+/K+ -ATPase inhibitors.

    Science.gov (United States)

    Chen, Ronald J Y; Jinn, Tzyy-rong; Chen, Yi-ching; Chung, Tse-yu; Yang, Wei-hung; Tzen, Jason T C

    2011-02-01

    The positive inotropic effect of cardiac glycosides lies in their reversible inhibition on the membrane-bound Na(+)/K(+)-ATPase in human myocardium. Steroid-like compounds containing a core structure similar to cardiac glycosides are found in many Chinese medicines conventionally used for promoting blood circulation. Some of them are demonstrated to be Na(+)/K(+)-ATPase inhibitors and thus putatively responsible for their therapeutic effects via the same molecular mechanism as cardiac glycosides. On the other hand, magnesium lithospermate B of danshen is also proposed to exert its cardiac therapeutic effect by effectively inhibiting Na(+)/K(+)-ATPase. Theoretical modeling suggests that the number of hydrogen bonds and the strength of hydrophobic interaction between the effective ingredients of various medicines and residues around the binding pocket of Na(+)/K(+)-ATPase are crucial for the inhibitory potency of these active ingredients. Ginsenosides, the active ingredients in ginseng and sanqi, substantially inhibit Na(+)/K(+)-ATPase when sugar moieties are attached only to the C-3 position of their steroid-like structure, equivalent to the sugar position in cardiac glycosides. Their inhibitory potency is abolished, however, when sugar moieties are linked to C-6 or C-20 position of the steroid nucleus; presumably, these sugar attachments lead to steric hindrance for the entrance of ginsenosides into the binding pocket of Na(+)/K(+)-ATPase. Neuroprotective effects of cardiac glycosides, several steroid-like compounds, and magnesium lithospermate B against ischemic stroke have been accordingly observed in a cortical brain slice-based assay model, and cumulative data support that effective inhibitors of Na(+)/K(+)-ATPase in the brain could be potential drugs for the treatment of ischemic stroke.

  8. The promiscuous phosphomonoestearase activity of Archaeoglobus fulgidus CopA, a thermophilic Cu+ transport ATPase.

    Science.gov (United States)

    Bredeston, Luis M; González Flecha, F Luis

    2016-07-01

    Membrane transport P-type ATPases display two characteristic enzymatic activities: a principal ATPase activity provides the driving force for ion transport across biological membranes, whereas a promiscuous secondary activity catalyzes the hydrolysis of phosphate monoesters. This last activity is usually denoted as the phosphatase activity of P-ATPases. In the present study, we characterize the phosphatase activity of the Cu(+)-transport ATPase from Archaeglobus fulgidus (Af-CopA) and compare it with the principal ATPase activity. Our results show that the phosphatase turnover number was 20 times higher than that corresponding to the ATPase activity, but it is compensated by a high value of Km, producing a less efficient catalysis for pNPP. This secondary activity is enhanced by Mg(2+) (essential activator) and phospholipids (non-essential activator), and inhibited by salts and Cu(+). Transition state analysis of the catalyzed and noncatalyzed hydrolysis of pNPP indicates that Af-CopA enhances the reaction rates by a factor of 10(5) (ΔΔG(‡)=38 kJ/mol) mainly by reducing the enthalpy of activation (ΔΔH(‡)=30 kJ/mol), whereas the entropy of activation is less negative on the enzyme than in solution. For the ATPase activity, the decrease in the enthalpic component of the barrier is higher (ΔΔH(‡)=39 kJ/mol) and the entropic component is small on both the enzyme and in solution. These results suggest that different mechanisms are involved in the transference of the phosphoryl group of p-nitrophenyl phosphate and ATP. Copyright © 2016 Elsevier B.V. All rights reserved.

  9. Renal aging in WKY rats: changes in Na+,K+ -ATPase function and oxidative stress.

    Science.gov (United States)

    Silva, E; Pinto, V; Simão, S; Serrão, M P; Afonso, J; Amaral, J; Pinho, M J; Gomes, P; Soares-da-Silva, P

    2010-12-01

    It has been suggested that alterations in Na(+),K(+)-ATPase mediate the development of several aging-related pathologies, such as hypertension and diabetes. Thus, we evaluated Na(+),K(+)-ATPase function and H(2)O(2) production in the renal cortex and medulla of Wistar Kyoto (WKY) rats at 13, 52 and 91 weeks of age. Creatinine clearance, proteinuria, urinary excretion of Na(+) and K(+) and fractional excretion of Na(+) were also determined. The results show that at 91 weeks old WKY rats had increased creatinine clearance and did not have proteinuria. Despite aging having had no effect on urinary Na(+) excretion, urinary K(+) excretion was increased and fractional Na(+) excretion was decreased with age. In renal proximal tubules and isolated renal cortical cells, 91 week old rats had decreased Na(+),K(+)-ATPase activity when compared to 13 and 52 week old rats. In renal medulla, 91 week old rats had increased Na(+),K(+)-ATPase activity, paralleled by an increase in protein expression of α(1)-subunit of Na(+),K(+)-ATPase. In addition, renal H(2)O(2) production increased with age and at 91 weeks of age renal medulla H(2)O(2) production was significantly higher than renal cortex production. The present work demonstrates that although at 91 weeks of age WKY rats were able to maintain Na(+) homeostasis, aging was accompanied by alterations in renal Na(+),K(+)-ATPase function. The observed increase in oxidative stress may account, in part, for the observed changes. Possibly, altered Na(+),K(+)-ATPase renal function may precede the development of age-related pathologies and loss of renal function. Copyright © 2010 Elsevier Inc. All rights reserved.

  10. Myofibril ATPase activity of cardiac and skeletal muscle of exhaustively exercised rats.

    Science.gov (United States)

    Belcastro, A N; Turcotte, R; Rossiter, M; Secord, D; Maybank, P E

    1984-01-01

    The activation characteristics of Mg-ATP and Ca2+ on cardiac and skeletal muscle myofibril ATPase activity were studied in rats following a run to exhaustion. In addition, the effect of varying ionic strength was determined on skeletal muscle from exhausted animals. The exhausted group (E) ran at a speed of 25 m min-1 with an 8% incline. Myofibril ATPase activities for control (C) and E were determined with 1, 3 and 5 mM Mg-ATP and 1 and 10 microM Ca2+ at pH 7.0 and 30 degrees C. For control skeletal muscle, at 1 and 10 microM Ca2+, there was an increase in ATPase activity from 1 to 5 mM Mg-ATP (P less than 0.05). For E animals the myofibril ATPase activities at 10 microM Ca2+ and all Mg-ATP concentrations were similar to C (P greater than 0.05). At 1.0 microM Ca2+ and all Mg-ATP concentrations were similar to C (P greater than 0.05). At 1.0 microM Ca2+ the activities at 3 and 5 mM Mg-ATP were greater for the E animals (P less than 0.05). Increasing KCl concentrations resulted in greater inhibition for E animals. With cardiac muscle, the myofibril ATPase activities at 1.0 microM free Ca2+ were lower for E at all Mg-ATP levels (P less than 0.05). In contrast, at 10 microM Ca2+, the E group exhibited an elevated myofibril ATPase activity. The results indicate that Mg-ATP and Ca2+ activation of cardiac and skeletal muscle myofibril ATPase is altered with exhaustive exercise.

  11. Suppression of concentration quenching of Er-related luminescence in Er-doped GaN

    International Nuclear Information System (INIS)

    Chen Shaoqiang; Tomita, Shigeo; Kudo, Hiroshi; Akimoto, Katsuhiro; Dierre, Benjamin; Lee, Woong; Sekiguchi, Takashi

    2010-01-01

    Erbium-doped GaN with different doping concentrations were grown by ammonia-source molecular beam epitaxy. The intra-4f-shell transitions related green luminescence were observed by both photoluminescence (PL) and cathodoluminescence (CL) measurements. It was found that concentration quenching of Er-related luminescence was observed in PL measurements while not in CL measurements. The different excitation and relaxation processes are suggested as the cause of the concentration quenching characteristics between PL and CL. The strong Er-related CL intensity in highly doped GaN demonstrates that high energy excitation is a promising approach to suppress the concentration quenching in Er-doped GaN.

  12. Interlayer exchange coupling in Er|Tb superlattices mediated by short range incommensurate Er order

    International Nuclear Information System (INIS)

    Pfuhl, E; Brueckel, T; Voigt, J; Mattauch, S; Korolkov, D

    2010-01-01

    We study the magnetic correlations in Er|Tb superlattices by means of off-specular scattering of polarized neutrons. We show here the co-existence of inhomogeneous magnetic states: i) ferromagnetic order of moments within the Tb layers below 230 K (FM), correlation length of about 10 bilayer, ii) an incommensurate modulated magnetic order, restricted to single Er layers and iii) antiferromagnetic coupling of ferromagnetic layers below 70K (AFC). Polarised off-specular neutron scattering under grazing incidence reveals that i) magnetic fluctuations appear when the sample is cooled below 70 K, ii) these fluctuations lead to AFC, when the sample is cooled to 10 K, which iii) persists, when the sample is subsequently heated up to 45 K, while the order is not present during the cooling cycle. Also the short range incommensurate order changes accordingly, implying that the magnetic order in the Er layers mediates the interlayer coupling between ferromagnetic Tb layers.

  13. Activity determination of Na+ K+ - ATPase and Mg++ - ATPase enzymes in the gill of Poecilia vivipara (Osteichthyes, Cyprinodontiformes in different salinities

    Directory of Open Access Journals (Sweden)

    Marcelo da Cunha Amaral

    2001-03-01

    Full Text Available This work aimed to know the tolerance mechanisms through the salinity variation by Na+ K+ - ATPase and Mg++ - ATPase and enzymes encountered in the gills of Poecilia vivipara. In field, the presence of this species was observed in salinities of 0 and 28‰. In laboratory, these fish were maintained in aquarium with mean salinity of 30‰ and positive responses were obtained. Some adult specimens, collected in a lagoon of the Coqueiros Beach, were utilized as matrixes. In the experiments the specimens used were those born in the test aquarium. For each salinity studied three replicates were made with three specimens for each one. The alevins were maintained in salinities of 5, 10, 15, 20, 25, 30 and 35‰ during a month for adaptation. Gills were extracted in appropriate buffer for isolation of plasma membrane and used for specific dosage of the total enzymatic activity of Na+ K+ - ATPase and Mg++ - ATPase. The relation of alevins to their adaptation towards the salinity variation was also studied. The activity of the two enzymes showed a different result. The major expression of Na+ K+ - ATPase was observed in 20‰ (35 µmoles Pi.mg protein.h-1, the best salinity to cultivate P. vivipara.Este trabalho teve como objetivo conhecer os mecanismos de tolerância às variações de salinidade, pelas enzimas Mg++ - ATPase e Na+ K+ - ATPase, encontrada nas brânquias de Poecilia vivipara. No campo, foi observada a presença desta espécie em salinidades entre 0 e 28‰. No laboratório, os indivíduos foram mantidos em salinidade de 30‰ e responderam positivamente. Os indivíduos adultos, coletados em uma lagoa na praia dos Coqueiros, foram utilizados como matrizes. Nos experimentos foram usados alevinos que nasceram nos aquários testes. Para cada salinidade estudada foram feitas três réplicas com três espécimens em cada uma. Os alevinos foram mantidos em salinidades de 5, 10, 15, 20, 25, 30 e 35‰, durante um mês para total adaptação. As br

  14. Effect of the unfolded protein response on ER protein export: a potential new mechanism to relieve ER stress.

    Science.gov (United States)

    Shaheen, Alaa

    2018-05-05

    The unfolded protein response (UPR) is an adaptive cellular response that aims to relieve endoplasmic reticulum (ER) stress via several mechanisms, including inhibition of protein synthesis and enhancement of protein folding and degradation. There is a controversy over the effect of the UPR on ER protein export. While some investigators suggested that ER export is inhibited during ER stress, others suggested the opposite. In this article, their conflicting studies are analyzed and compared in attempt to solve this controversy. The UPR appears indeed to enhance ER export, possibly via multiple mechanisms. However, another factor, which is the integrity of the folding machinery/environment inside ER, determines whether ER export will appear increased or decreased during experimentation. Also, different methods of stress induction appear to have different effects on ER export. Thus, improvement of ER export may represent a new mechanism by which the UPR alleviates ER stress. This may help researchers to understand how the UPR works inside cells and how to manipulate it to alter cell fate during stress, either to promote cell survival or death. This may open up new approaches for the treatment of ER stress-related diseases.

  15. Stabilization of the H,K-ATPase M5M6 membrane hairpin by K+ ions. Mechanistic significance for p2-type atpases.

    Science.gov (United States)

    Gatto, C; Lutsenko, S; Shin, J M; Sachs, G; Kaplan, J H

    1999-05-14

    The integral membrane protein, the gastric H,K-ATPase, is an alpha-beta heterodimer, with 10 putative transmembrane segments in the alpha-subunit and one such segment in the beta-subunit. All transmembrane segments remain within the membrane domain following trypsinization of the intact gastric H,K-ATPase in the presence of K+ ions, identified as M1M2, M3M4, M5M6, and M7, M8, M9, and M10. Removal of K+ ions from this digested preparation results in the selective loss of the M5M6 hairpin from the membrane. The release of the M5M6 fragment is directed to the extracellular phase as evidenced by the accumulation of the released M5M6 hairpin inside the sealed inside out vesicles. The stabilization of the M5M6 hairpin in the membrane phase by the transported cation as well as loss to the aqueous phase in the absence of the transported cation has been previously observed for another P2-type ATPase, the Na, K-ATPase (Lutsenko, S., Anderko, R., and Kaplan, J. H. (1995) Proc. Natl. Acad. Sci. U. S. A. 92, 7936-7940). Thus, the effects of the counter-transported cation on retention of the M5M6 segment in the membrane as compared with the other membrane pairs may be a general feature of P2-ATPase ion pumps, reflecting a flexibility of this region that relates to the mechanism of transport.

  16. Changes in acetylcholinesterase, Na+,K+-ATPase, and Mg2+-ATPase activities in the frontal cortex and the hippocampus of hyper- and hypothyroid adult rats.

    Science.gov (United States)

    Carageorgiou, Haris; Pantos, Constantinos; Zarros, Apostolos; Stolakis, Vasileios; Mourouzis, Iordanis; Cokkinos, Dennis; Tsakiris, Stylianos

    2007-08-01

    The thyroid hormones (THs) are crucial determinants of normal development and metabolism, especially in the central nervous system. The metabolic rate is known to increase in hyperthyroidism and decrease in hypothyroidism. The aim of this work was to investigate how changes in metabolism induced by THs could affect the activities of acetylcholinesterase (AChE), (Na+,K+)- and Mg2+-adenosinetriphosphatase (ATPase) in the frontal cortex and the hippocampus of adult rats. Hyperthyroidism was induced by subcutaneous administration of thyroxine (25 microg/100 g body weight) once daily for 14 days, and hypothyroidism was induced by oral administration of propylthiouracil (0.05%) for 21 days. All enzyme activities were evaluated spectrophotometrically in the homogenated brain regions of 10 three-animal pools. A region-specific behavior was observed concerning the examined enzyme activities in hyper- and hypothyroidism. In hyperthyroidism, AChE activity was significantly increased only in the hippocampus (+22%), whereas Na+,K+-ATPase activity was significantly decreased in the hyperthyroid rat hippocampus (-47%) and remained unchanged in the frontal cortex. In hypothyroidism, AChE activity was significantly decreased in the frontal cortex (-23%) and increased in the hippocampus (+21%). Na+,K+-ATPase activity was significantly decreased in both the frontal cortex (-35%) and the hippocampus (-43%) of hypothyroid rats. Mg2+-ATPase remained unchanged in the regions of both hyper- and hypothyroid rat brains. Our data revealed that THs affect the examined adult rat brain parameters in a region- and state-specific way. The TH-reduced Na+,K+-ATPase activity may increase the synaptic acetylcholine release and, thus, modulate AChE activity. Moreover, the above TH-induced changes may affect the monoamine neurotransmitter systems in the examined brain regions.

  17. On the Use of an ER-213 Detonator to Establish a Baseline for the ER-486

    Energy Technology Data Exchange (ETDEWEB)

    Thomas, Keith A. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Liechty, Gary H. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Jaramillo, Dennis C. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Munger, Alan C. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); McHugh, Douglas C. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Kennedy, James E. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

    2014-08-19

    This report documents a series of tests using a TSD-115 fireset coupled with an ER-213, a gold exploding bridgewire (EBW) detonator. These tests were designed to fire this EBW with a smaller fireset to obtain current and voltage data as well as timing information at voltage levels below, above, and throughout the threshold firing region. This study could then create a database for comparison to our current ER-486 EBW development, which is designed to be a lower voltage (<500V) device.

  18. GR and ER co-activation alters the expression of differentiation genes and associates with improved ER+ breast cancer outcome

    Science.gov (United States)

    West, Diana C.; Pan, Deng; Tonsing-Carter, Eva Y.; Hernandez, Kyle M.; Pierce, Charles F.; Styke, Sarah C.; Bowie, Kathleen R.; Garcia, Tzintzuni I.; Kocherginsky, Masha; Conzen, Suzanne D.

    2016-01-01

    In estrogen receptor (ER)-negative breast cancer (BC), high tumor glucocorticoid receptor (GR) expression has been associated with a relatively poor outcome. In contrast, using a meta-analysis of several genomic datasets, here we find that tumor GR mRNA expression is associated with improved ER+ relapse-free survival (RFS) (independently of progesterone receptor (PR) expression). To understand the mechanism by which GR expression is associated with a better ER+ BC outcome, the global effect of GR-mediated transcriptional activation in ER+ BC cells was studied. Analysis of GR chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) in ER+/GR+ MCF-7 cells revealed that upon co-activation of GR and ER, GR chromatin association became enriched at proximal promoter regions. Furthermore, following ER activation, increased GR chromatin association was observed at ER, FOXO, and AP1 response elements. In addition, ER associated with GR response elements, suggesting that ER and GR interact in a complex. Co-activation of GR and ER resulted in increased expression (relative to ER activation alone) of transcripts that encode proteins promoting cellular differentiation (e.g. KDM4B, VDR) and inhibiting the Wnt-signaling pathway (IGFBP4). Finally, expression of these individual pro-differentiation genes was associated with significantly improved RFS in ER+ BC patients. Together, these data suggest that the co-expression and subsequent activity of tumor cell GR and ER contribute to the less aggressive natural history of early-stage BC by coordinating the altered expression of genes favoring differentiation. Implications The interaction between estrogen and glucocorticoid receptor activity highlights the importance of context-dependent nuclear receptor function in cancer. PMID:27141101

  19. COPII-Dependent ER Export: A Critical Component of Insulin Biogenesis and β-Cell ER Homeostasis.

    Science.gov (United States)

    Fang, Jingye; Liu, Ming; Zhang, Xuebao; Sakamoto, Takeshi; Taatjes, Douglas J; Jena, Bhanu P; Sun, Fei; Woods, James; Bryson, Tim; Kowluru, Anjaneyulu; Zhang, Kezhong; Chen, Xuequn

    2015-08-01

    Pancreatic β-cells possess a highly active protein synthetic and export machinery in the endoplasmic reticulum (ER) to accommodate the massive production of proinsulin. ER homeostasis is vital for β-cell functions and is maintained by the delicate balance between protein synthesis, folding, export, and degradation. Disruption of ER homeostasis by diabetes-causing factors leads to β-cell death. Among the 4 components to maintain ER homeostasis in β-cells, the role of ER export in insulin biogenesis is the least understood. To address this knowledge gap, the present study investigated the molecular mechanism of proinsulin ER export in MIN6 cells and primary islets. Two inhibitory mutants of the secretion-associated RAS-related protein (Sar)1 small GTPase, known to specifically block coat protein complex II (COPII)-dependent ER export, were overexpressed in β-cells using recombinant adenoviruses. Results from this approach, as well as small interfering RNA-mediated Sar1 knockdown, demonstrated that defective Sar1 function blocked proinsulin ER export and abolished its conversion to mature insulin in MIN6 cells, isolated mouse, and human islets. It is further revealed, using an in vitro vesicle formation assay, that proinsulin was packaged into COPII vesicles in a GTP- and Sar1-dependent manner. Blockage of COPII-dependent ER exit by Sar1 mutants strongly induced ER morphology change, ER stress response, and β-cell apoptosis. These responses were mediated by the PKR (double-stranded RNA-dependent kinase)-like ER kinase (PERK)/eukaryotic translation initiation factor 2α (p-eIF2α) and inositol-requiring protein 1 (IRE1)/x-box binding protein 1 (Xbp1) pathways but not via activating transcription factor 6 (ATF6). Collectively, results from the study demonstrate that COPII-dependent ER export plays a vital role in insulin biogenesis, ER homeostasis, and β-cell survival.

  20. Inhibition of advanced glycation endproduct (AGE) rescues against streptozotocin-induced diabetic cardiomyopathy: Role of autophagy and ER stress.

    Science.gov (United States)

    Pei, Zhaohui; Deng, Qinqin; Babcock, Sara A; He, Emily Y; Ren, Jun; Zhang, Yingmei

    2018-03-01

    Diabetes mellitus leads to oxidative stress and contractile dysfunction in the heart. Although several rationales have been speculated, the precise mechanism behind diabetic cardiomyopathy remains elusive. This study was designed to assess the role of inhibition of advanced glycation endproducts (AGE) in streptozotocin (STZ)-induced diabetic cardiac dysfunction. Cardiac contractile function was assessed in normal C57BL/6 and STZ (200mg/kg, single injection and maintained for 2 wks)-induced diabetic mice treated with or without the AGE inhibitor aminoguanidine (50mg/kg/d in drinking water) for 2 weeks using echocardiography and IonOptix MyoCam techniques. Diabetes compromised cardiac contractile function shown as reduced fractional shortening and ejection fraction, enlarged left ventricular end systolic/diastolic diameters, decreased peak shortening, maximal velocity of shortening/relengthening, prolonged shortening and relengthening duration as well as impaired intracellular Ca 2+ homeostasis, the effects of which were alleviated or reversed by aminoguanidine treatment. Diabetes also inhibited autophagy, increased ER stress and phosphorylation of pro-hypertrophic signaling molecules Akt and mTOR, the effect of which was reversed by aminoguanidine. In vitro study revealed that methylglyoxal-derived AGE (MG-AGE) incubation in isolated cardiomyocytes promoted oxidation of sarco(endo)plasmic reticulum Ca 2+ -ATPase (SERCA2a) and production of superoxide, the effects of which were negated by the autophagy inducer rapamycin, the ER stress chaperone TUDCA or the antioxidant N-acetylcysteine. Taken together, these data revealed that inhibition of AGE formation rescues against experimental diabetes-induced cardiac remodeling and contractile dysfunction possible through regulation of autophagy and ER stress. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Hvor langt er PISA fra Danmark?

    DEFF Research Database (Denmark)

    Dolin, Jens; Krogh, Lars Brian

    2008-01-01

    PISA 2006 science testen i en dansk kontekst. I det første delprojekt sammenlignes PISA science frameworket med de danske mål for naturfagsuddannelserne. I det andet delprojekt undersøges hvorledes PISA testformatet er i overensstemmelse med evalueringspraksis i naturfagene i den danske folkeskole. I...... det tredje delprojekt sammenlignes elevers score i PISA science testen med deres resultater i en mere dialogbaseret evalueringssituation.   De tre delundersøgelser betjener sig af forskellige metoder, nemlig henholdsvis tekstanalyse, spørgeskemaundersøgelse/ interview samt en egentlig feltvalidering...

  2. BOREAS Level-0 ER-2 Navigation Data

    Science.gov (United States)

    Strub, Richard; Dominguez, Roseanne; Newcomer, Jeffrey A.; Hall, Forrest G. (Editor)

    2000-01-01

    The BOREAS Staff Science effort covered those activities that were BOREAS community-level activities or required uniform data collection procedures across sites and time. These activities included the acquisition, processing, and archiving of aircraft navigation/attitude data to complement the digital image data. The level-0 ER-2 navigation data files contain aircraft attitude and position information acquired during the digital image and photographic data collection missions. Temporally, the data were acquired from April to September 1994. Data were recorded at intervals of 5 seconds. The data are stored in tabular ASCII files.

  3. Photoluminescence of Er in SiOx

    International Nuclear Information System (INIS)

    Wan Jun; Sheng Chi; Lu Fang; Gong Dawei; Fan Yongliang; Lin Feng; Wang Xun

    1998-01-01

    Erbium-doped SiO x is prepared by molecular beam epitaxy. The influence of Er on the incorporation of O is studied by using Auger spectroscopy. Photoluminescence (PL) peaks around the wave-length of 1.53 μm have been observed within the temperature range of 18 to 300 K after annealing. The relationship between PL intensity and annealing temperature is discussed. The temperature dependence of the PL intensity shows an exponential decay with an activation energy of 12 meV at low temperatures ( 100 K)

  4. Direct interaction of the bacteriophage SPP1 packaging ATPase with the portal protein.

    Science.gov (United States)

    Oliveira, Leonor; Cuervo, Ana; Tavares, Paulo

    2010-03-05

    DNA packaging in tailed bacteriophages and other viruses requires assembly of a complex molecular machine at a specific vertex of the procapsid. This machine is composed of the portal protein that provides a tunnel for DNA entry, an ATPase that fuels DNA translocation (large terminase subunit), and most frequently, a small terminase subunit. Here we characterized the interaction between the terminase ATPase subunit of bacteriophage SPP1 (gp2) and the procapsid portal vertex. We found, by affinity pulldown assays with purified proteins, that gp2 interacts with the portal protein, gp6, independently of the terminase small subunit gp1, DNA, or ATP. The gp2-procapsid interaction via the portal protein depends on gp2 concentration and requires the presence of divalent cations. Competition experiments showed that isolated gp6 can only inhibit gp2-procapsid interactions and DNA packaging at gp6:procapsid molar ratios above 10-fold. Assays with gp6 carrying mutations in distinct regions of its structure that affect the portal-induced stimulation of ATPase and DNA packaging revealed that none of these mutations impedes gp2-gp6 binding. Our results demonstrate that the SPP1 packaging ATPase binds directly to the portal and that the interaction is stronger with the portal embedded in procapsids. Identification of mutations in gp6 that allow for assembly of the ATPase-portal complex but impair DNA packaging support an intricate cross-talk between the two proteins for activity of the DNA translocation motor.

  5. Tributyltin (TBT) inhibition of oligomycin-sensitive Mg-ATPase activity in mussel mitochondria.

    Science.gov (United States)

    Pagliarani, Alessandra; Bandiera, Patrizia; Ventrella, Vittoria; Trombetti, Fabiana; Pirini, Maurizio; Nesci, Salvatore; Borgatti, Anna Rosa

    2008-06-01

    Tributyltin (TBT), one of the most toxic lipophilic aquatic pollutants, can be efficiently incorporated from sea water and sediments by filter-feeding molluscs. As far as we are aware TBT effects on the mitochondrial oligomycin-sensitive Mg-ATPase, the enzymatic core of energy production and a known target of TBT toxicity in mammals, have not been yet investigated in molluscs; thus the hydrolytic capability of the mitochondrial complex in the presence of micromolar concentrations of TBT was assayed in the mussel Mytilus galloprovincialis. Gill and mantle ATPase activities were progressively depressed by increasing TBT doses up to a maximal inhibition (82% in the gills and 74% in the mantle) at 0.62 microM TBT. Non-cooperative inhibition kinetics (n(H) approximately -1) and a non-competitive mechanism with respect to ATP substrate were pointed out. The mitochondrial Mg-ATPase susceptivity to TBT in the marine mussel was consistent with the formation of a TBT-Mg-ATPase complex, apparently more stable in the gills than in the mantle. The complex shape of the dose-response curve and the partial release of Mg-ATPase inhibition within the 0.6-34.4 microM TBT range suggest multiple interactions of TBT with the enzyme complex putatively related to its molecular mechanism of toxicity.

  6. ATPase and morphologic changes induced by UVB on Langerhans cells in guinea pigs

    International Nuclear Information System (INIS)

    Hanau, D.; Fabre, M.; Lepoittevin, J.P.; Stampf, J.L.; Grosshans, E.; Benezra, C.

    1985-01-01

    The authors have devised, in guinea pigs, an improved ATPase technique which enables one to proceed from light to electron microscope study while preserving, on the ultrastructural level, the various membranous structures, in particular the Langerhans cell (LC) granules. Using this method, they have been able to confirm the action of acute, low-dose UVB on the surface enzymatic marker, ATPase. Moreover, this study has shown that the ATPase-negative LC contain abnormal LC granules or, more often, are deficient in LC granules. In a previous work, the authors have shown that, after epicutaneous application of a hapten, one successively observes an extensive adsorptive pinocytosis process, the disappearance of the membranous ATPase system, and the appearance of LC granules in the cytoplasm. Therefore, the authors may suppose that, after UVB irradiation, the disappearance of the ATPase system and/or the possible alteration of the adsorptive pinocytosis process interrupts or alters the formation of LC granules. These successive events might play a vital role in the formation of the hapten--carrier protein-Ia antigen complex. In their absence in a large number of LC, following UV irradiation, epicutaneous application of a hapten would lead to the development of a state of immune tolerance

  7. Cryo-EM studies of the structure and dynamics of vacuolar-type ATPases

    Science.gov (United States)

    Mazhab-Jafari, Mohammad T.; Rubinstein, John L.

    2016-01-01

    Electron cryomicroscopy (cryo-EM) has significantly advanced our understanding of molecular structure in biology. Recent innovations in both hardware and software have made cryo-EM a viable alternative for targets that are not amenable to x-ray crystallography or nuclear magnetic resonance (NMR) spectroscopy. Cryo-EM has even become the method of choice in some situations where x-ray crystallography and NMR spectroscopy are possible but where cryo-EM can determine structures at higher resolution or with less time or effort. Rotary adenosine triphosphatases (ATPases) are crucial to the maintenance of cellular homeostasis. These enzymes couple the synthesis or hydrolysis of adenosine triphosphate to the use or production of a transmembrane electrochemical ion gradient, respectively. However, the membrane-embedded nature and conformational heterogeneity of intact rotary ATPases have prevented their high-resolution structural analysis to date. Recent application of cryo-EM methods to the different types of rotary ATPase has led to sudden advances in understanding the structure and function of these enzymes, revealing significant conformational heterogeneity and characteristic transmembrane α helices that are highly tilted with respect to the membrane. In this Review, we will discuss what has been learned recently about rotary ATPase structure and function, with a particular focus on the vacuolar-type ATPases. PMID:27532044

  8. The ATPases of cohesin interface with regulators to modulate cohesin-mediated DNA tethering

    Science.gov (United States)

    Çamdere, Gamze; Guacci, Vincent; Stricklin, Jeremiah; Koshland, Douglas

    2015-01-01

    Cohesin tethers together regions of DNA, thereby mediating higher order chromatin organization that is critical for sister chromatid cohesion, DNA repair and transcriptional regulation. Cohesin contains a heterodimeric ATP-binding Cassette (ABC) ATPase comprised of Smc1 and Smc3 ATPase active sites. These ATPases are required for cohesin to bind DNA. Cohesin’s DNA binding activity is also promoted by the Eco1 acetyltransferase and inhibited by Wpl1. Recently we showed that after cohesin stably binds DNA, a second step is required for DNA tethering. This second step is also controlled by Eco1 acetylation. Here, we use genetic and biochemical analyses to show that this second DNA tethering step is regulated by cohesin ATPase. Furthermore, our results also suggest that Eco1 promotes cohesion by modulating the ATPase cycle of DNA-bound cohesin in a state that is permissive for DNA tethering and refractory to Wpl1 inhibition. DOI: http://dx.doi.org/10.7554/eLife.11315.001 PMID:26583750

  9. Characterization of ATPase activity of the AAA ARC from Bifidobacterium longum subsp. infantis.

    Science.gov (United States)

    Guzmán-Rodríguez, Mabel; de la Rosa, Ana Paulina Barba; Santos, Leticia

    2015-01-01

    Bifidobacteria are considered to be probiotics that exist in the large intestine and are helpful to maintain human health. Oral administration of bifidobacteria may be effective in improving the intestinal flora and environment, stimulating the immune response and possibly preventing cancer. However, for consistent and positive results, further well-controlled studies are urgently needed to describe the basic mechanisms of this microorganism. Analysis of the proteasome-lacking Bifidobacterium longum genome reveals that it possesses a gene, IPR003593 AAA ATPase core, which codes a 56 kDa protein containing one AAA ATPase domain. Phylogenetic classification made by CLANS, positioned this sequence into the ARC divergent branch of the AAA ATPase family of proteins. N-terminal analysis of the sequence indicates this protein is closely related to other ATPases such as the Rhodococcus erythropolis ARC, Archaeoglobus fulgidus PAN, Mycobacterium tuberculosis Mpa and the human proteasomal Rpt1 subunit. This gene was cloned, the full-length recombinant protein was overexpressed in Escherichia coli, purified as a high-molecular size complex and named Bl-ARC. Enzymatic characterization showed that Bl-ARC ATPase is active, Mg(+2)-dependent and sensitive to N-ethylmaleimide. Gene organization positions bl-arc in a region flanked by a cluster of genes that includes pup, dop and pafA genes. These findings point to a possible function as a chaperone in the degradation pathway via pupylation.

  10. Hormonal regulation of Na+-K+-ATPase in cultured epithelial cells

    International Nuclear Information System (INIS)

    Johnson, J.P.; Jones, D.; Wiesmann, W.P.

    1986-01-01

    Aldosterone and insulin stimulate Na + transport through mechanisms involving protein synthesis. Na + -K + -ATPase has been implicated in the action of both hormones. The authors examined the effect of aldosterone and insulin on Na + -K + -ATPase in epithelial cells in culture derived from toad urinary bladder (TB6C) and toad kidney (A6). Aldosterone, but not insulin, increases short-circuit current (I/sub sc/) in TB6C cells. Aldosterone increases Na + -K + -[ 32 P]ATPase activity after 18 h of incubation, but no effect can be seen at 3 and 6 h. Amiloride, which inhibits aldosterone-induced increases in I/sub sc/, has no effect on either basal or aldosterone stimulated enzyme activity. Both aldosterone and insulin increase I/sub sc/ in A6 cells and when added together are synergistic. Aldosterone stimulates enzyme activity in A6 cells, but insulin alone has no effect. However, aldosterone and insulin together stimulate enzyme activity more than aldosterone alone. It appears that stimulation of Na + -K + -ATPase activity is involved in aldosterone action in both cell lines but does not appear to be due to increased Na + entry, since enhanced enzyme activity is not inhibited by amiloride. In contrast, insulin alone has no direct effect on Na + -K + -ATPase, although the increased enzyme activity following both agents in combination may explain their synergism on I/sub sc/

  11. Isolation and characterization of a specific endogenous Na+, K+-ATPase inhibitor from bovine adrenal

    International Nuclear Information System (INIS)

    Tamura, M.; Lam, T.T.; Inagami, T.

    1988-01-01

    In order to identify a specific endogenous Na + ,K + -ATPase inhibitor which could possibly be related to salt-dependent hypertension, the authors looked for substances in the methanol extract of bovine whole adrenal which show all of the following properties: (i) inhibitory activity for Na + ,K + -ATPase; (ii) competitive displacing activity against [ 3 H]ouabain binding to the enzyme; (iii) inhibitory activity for 86 Rb uptake into intact human erythrocytes; and (iv) cross-reactivity with sheep anti-digoxin-specific antibody. After stepwise fractionation of the methanol extract of bovine adrenal glands by chromatography on a C 18 open column, a 0-15% acetonitrile fraction was fractionated by high-performance liquid chromatography on a Zorbax octadecylsilane column. One of the most active fractions in 0-15% acetonitrile was found to exhibit all of the four types of the activities. It was soluble in water and was distinct from various substances which have been known to inhibit Na + ,K + -ATPase. These results strongly suggest that this water-soluble nonpeptidic Na + ,K + -ATPase inhibitor may be a specific endogenous regulator for the ATPase

  12. Both ATPase sites of Escherichia coli UvrA have functional roles in nucleotide excision repair

    International Nuclear Information System (INIS)

    Thiagalingam, S.; Grossman, L.

    1991-01-01

    The roles of the two tandemly arranged putative ATP binding sites of Escherichia coli UvrA in UvrABC endonuclease-mediated excision repair were analyzed by site-directed mutagenesis and biochemical characterization of the representative mutant proteins. Evidence is presented that UvrA has two functional ATPase sites which coincide with the putative ATP binding motifs predicted from its amino acid sequence. The individual ATPase sites can independently hydrolyze ATP. The C-terminal ATPase site has a higher affinity for ATP than the N-terminal site. The invariable lysine residues at the ends of the glycine-rich loops of the consensus Walker type A motifs are indispensable for ATP hydrolysis. However, the mutations at these lysine residues do not significantly affect ATP binding. UvrA, with bound ATP, forms the most favored conformation for DNA binding. The initial binding of UvrA to DNA is chiefly at the undamaged sites. In contrast to the wild type UvrA, the ATPase site mutants bind equally to damaged and undamaged sites. Dissociation of tightly bound nucleoprotein complexes from the undamaged sites requires hydrolysis of ATP by the C-terminal ATPase site of UvrA. Thus, both ATP binding and hydrolysis are required for the damage recognition step enabling UvrA to discriminate between damaged and undamaged sites on DNA

  13. Inhibition of ecto-ATPase activities impairs HIV-1 infection of macrophages.

    Science.gov (United States)

    Schachter, Julieta; Delgado, Kelly Valcárcel; Barreto-de-Souza, Victor; Bou-Habib, Dumith Chequer; Persechini, Pedro Muanis; Meyer-Fernandes, José Roberto

    2015-05-01

    Nucleotides and nucleosides are secreted into extracellular media at different concentrations as a consequence of different physiologic and pathological conditions. Ecto-nucleotidases, enzymes present on the surface of most cells, hydrolyze these extracellular nucleotides and reduce the concentration of them, thus affecting the activation of different nucleotide and nucleoside receptors. Also, ecto-nucleotidases are present in a number of microorganisms and play important roles in host-pathogen interactions. Here, we characterized the ecto-ATPase activities present on the surface of HIV-1 particle and human macrophages as well. We found that the kinetic properties of HIV-1 and macrophage ecto-ATPases are similar, suggesting that the enzyme is the same. This ecto-ATPase activity was increased in macrophages infected in vitro with HIV-1. Using three different non-related ecto-ATPase inhibitors-POM-1, ARL67156 and BG0-we showed that the inhibition of these macrophage and viral ecto-ATPase activities impairs HIV-1 infection. In addition, we also found that elevated extracellular concentrations of ATP inhibit HIV-1 production by infected macrophages. Copyright © 2014 Elsevier GmbH. All rights reserved.

  14. Subcellular distribution of calcium-binding proteins and a calcium-ATPase in canine pancreas

    International Nuclear Information System (INIS)

    Nigam, S.K.; Towers, T.

    1990-01-01

    Using a 45Ca blot-overlay assay, we monitored the subcellular fractionation pattern of several Ca binding proteins of apparent molecular masses 94, 61, and 59 kD. These proteins also appeared to stain blue with Stains-All. Additionally, using a monoclonal antiserum raised against canine cardiac sarcoplasmic reticulum Ca-ATPase, we examined the subcellular distribution of a canine pancreatic 110-kD protein recognized by this antiserum. This protein had the same electrophoretic mobility as the cardiac protein against which the antiserum was raised. The three Ca binding proteins and the Ca-ATPase cofractionated into the rough microsomal fraction (RM), previously shown to consist of highly purified RER, in a pattern highly similar to that of the RER marker, ribophorin I. To provide further evidence for an RER localization, native RM were subjected to isopycnic flotation in sucrose gradients. The Ca binding proteins and the Ca-ATPase were found in dense fractions, along with ribophorin I. When RM were stripped of ribosomes with puromycin/high salt, the Ca binding proteins and the Ca-ATPase exhibited a shift to less dense fractions, as did ribophorin I. We conclude that, in pancreas, the Ca binding proteins and Ca-ATPase we detect are localized to the RER (conceivably a subcompartment of the RER) or, possibly, a structure intimately associated with the RER

  15. Phenylarsine Oxide Inhibits the Fusicoccin-Induced Activation of Plasma Membrane H+-ATPase1

    Science.gov (United States)

    Olivari, Claudio; Albumi, Cristina; Pugliarello, Maria Chiara; De Michelis, Maria Ida

    2000-01-01

    To investigate the mechanism by which fusicoccin (FC) induces the activation of the plasma membrane (PM) H+-ATPase, we used phenylarsine oxide (PAO), a known inhibitor of protein tyrosine-phosphatases. PAO was supplied in vivo in the absence or presence of FC to radish (Raphanus sativus L.) seedlings and cultured Arabidopsis cells prior to PM extraction. Treatment with PAO alone caused a slight decrease of PM H+-ATPase activity and, in radish, a decrease of PM-associated 14-3-3 proteins. When supplied prior to FC, PAO drastically inhibited FC-induced activation of PM H+-ATPase, FC binding to the PM, and the FC-induced increase of the amount of 14-3-3 associated with the PM. On the contrary, PAO was completely ineffective on all of the above-mentioned parameters when supplied after FC. The H+-ATPase isolated from PAO-treated Arabidopsis cells maintained the ability to respond to FC if supplied with exogenous, nonphosphorylated 14-3-3 proteins. Altogether, these results are consistent with a model in which the dephosphorylated state of tyrosine residues of a protein(s), such as 14-3-3 protein, is required to permit FC-induced association between the 14-3-3 protein and the PM H+-ATPase. PMID:10677439

  16. Effects of Na/K-ATPase and its ligands on bone marrow stromal cell differentiation

    Directory of Open Access Journals (Sweden)

    Moustafa Sayed

    2014-07-01

    Full Text Available Endogenous ligands of Na/K-ATPase have been demonstrated to increase in kidney dysfunction and heart failure. It is also reported that Na/K-ATPase signaling function effects stem cell differentiation. This study evaluated whether Na/K-ATPase activation through its ligands and associated signaling functions affect bone marrow stromal cells (BMSCs, also known as bone marrow-derived mesenchymal stem cells differentiation capacity. BMSCs were isolated from male Sprague–Dawley rats and cultured in minimal essential medium alpha (MEM-α supplemented with 15% Fetal Bovine serum (FBS. The results showed that marinobufagenin (MBG, a specific Na/K-ATPase ligand, potentiated rosiglitazone-induced adipogenesis in these BMSCs. Meanwhile, it attenuated BMSC osteogenesis. Mechanistically, MBG increased CCAAT/enhancer binding protein alpha (C/EBPα protein expression through activation of an extracellular regulated kinase (ERK signaling pathway, which leads to enhanced rosiglitazone-induced adipogenesis. Inhibition of ERK activation by U0126 blocks the effect of MBG on C/EBPα expression and on rosiglitazone-induced adipogenesis. Reciprocally, MBG reduced runt-related transcription factor 2 (RunX2 expression, which resulted in the inhibition of osteogenesis induced by β-glycerophosphate/ascorbic acid. MBG also potentiated rosiglitazone-induced adipogenesis in 3T3-L1 cells and in mouse BMSCs. These results suggest that Na/K-ATPase and its signaling functions are involved in the regulation of BMSCs differentiation.

  17. Electrostatic control by lipids upon the membrane-bound (Na+ + K+)-ATPase.

    Science.gov (United States)

    Ahrens, M L

    1981-04-06

    In this paper, the membrane-bound (Na+ + K+)-ATPase from bovine brain is shown to be controlled by electrostatic alterations of the charged lipids surrounding the enzyme. The properties under investigation are the enzymatic activity, activation energy and the response of the enzymatic system to temperature. Arrhenius plots of the ATPase activity are biphasic with a break at temperature Ti. The temperature Ti, the activation energies at temperatures above and below Ti, and the enzymatic activity at any constant temperature have been shown to depend upon the concentrations of alkali and alkaline-earth metal ions in the solution. These electrolyte dependencies are ascribed to changes of electrostatic conditions at the lipids surrounding the ATPase. If the higher electrostatic screening ability of divalent ions is taken into account, the results in the presence of mono- and divalent ions become virtually the same. As a result of this work, it is concluded that electrostatic alterations are transmitted to the ATPase from the lipids of the membrane in which the enzyme is embedded. Inhibition and activation of the enzyme by mono-and divalent metal ions may thus be explained without any auxiliary hypothesis, particularly without postulating specific binding sites for the different ionic species at the protein. In addition, the specific lipid requirement of the ATPase may be understood better in the light of this interpretation.

  18. An Arginine Finger Regulates the Sequential Action of Asymmetrical Hexameric ATPase in the Double-Stranded DNA Translocation Motor.

    Science.gov (United States)

    Zhao, Zhengyi; De-Donatis, Gian Marco; Schwartz, Chad; Fang, Huaming; Li, Jingyuan; Guo, Peixuan

    2016-10-01

    Biological motors are ubiquitous in living systems. Currently, how the motor components coordinate the unidirectional motion is elusive in most cases. Here, we report that the sequential action of the ATPase ring in the DNA packaging motor of bacteriophage ϕ29 is regulated by an arginine finger that extends from one ATPase subunit to the adjacent unit to promote noncovalent dimer formation. Mutation of the arginine finger resulted in the interruption of ATPase oligomerization, ATP binding/hydrolysis, and DNA translocation. Dimer formation reappeared when arginine mutants were mixed with other ATPase subunits that can offer the arginine to promote their interaction. Ultracentrifugation and virion assembly assays indicated that the ATPase was presenting as monomers and dimer mixtures. The isolated dimer alone was inactive in DNA translocation, but the addition of monomer could restore the activity, suggesting that the hexameric ATPase ring contained both dimer and monomers. Moreover, ATP binding or hydrolysis resulted in conformation and entropy changes of the ATPase with high or low DNA affinity. Taking these observations together, we concluded that the arginine finger regulates sequential action of the motor ATPase subunit by promoting the formation of the dimer inside the hexamer. The finding of asymmetrical hexameric organization is supported by structural evidence of many other ATPase systems showing the presence of one noncovalent dimer and four monomer subunits. All of these provide clues for why the asymmetrical hexameric ATPase gp16 of ϕ29 was previously reported as a pentameric configuration by cryo-electron microscopy (cryo-EM) since the contact by the arginine finger renders two adjacent ATPase subunits closer than other subunits. Thus, the asymmetrical hexamer would appear as a pentamer by cryo-EM, a technology that acquires the average of many images. Copyright © 2016 Zhao et al.

  19. Bcl-2 associated athanogene 5 (Bag5) is overexpressed in prostate cancer and inhibits ER-stress induced apoptosis

    International Nuclear Information System (INIS)

    Bruchmann, Anja; Roller, Corinna; Walther, Tamara Vanessa; Schäfer, Georg; Lehmusvaara, Sara; Visakorpi, Tapio; Klocker, Helmut; Cato, Andrew C B; Maddalo, Danilo

    2013-01-01

    The Bag (Bcl-2 associated athanogene) family of proteins consists of 6 members sharing a common, single-copied Bag domain through which they interact with the molecular chaperone Hsp70. Bag5 represents an exception in the Bag family since it consists of 5 Bag domains covering the whole protein. Bag proteins like Bag1 and Bag3 have been implicated in tumor growth and survival but it is not known whether Bag5 also exhibits this function. Bag5 mRNA and protein expression levels were investigated in prostate cancer patient samples using real-time PCR and immunoblot analyses. In addition immunohistological studies were carried out to determine the expression of Bag5 in tissue arrays. Analysis of Bag5 gene expression was carried out using one-way ANOVA and Bonferroni’s Multiple Comparison test. The mean values of the Bag5 stained cells in the tissue array was analyzed by Mann-Whitney test. Functional studies of the role of Bag5 in prostate cancer cell lines was performed using overexpression and RNA interference analyses. Our results show that Bag5 is overexpressed in malignant prostate tissue compared to benign samples. In addition we could show that Bag5 levels are increased following endoplasmic reticulum (ER)-stress induction, and Bag5 relocates from the cytoplasm to the ER during this process. We also demonstrate that Bag5 interacts with the ER-resident chaperone GRP78/BiP and enhances its ATPase activity. Bag5 overexpression in 22Rv.1 prostate cancer cells inhibited ER-stress induced apoptosis in the unfolded protein response by suppressing PERK-eIF2-ATF4 activity while enhancing the IRE1-Xbp1 axis of this pathway. Cells expressing high levels of Bag5 showed reduced sensitivity to apoptosis induced by different agents while Bag5 downregulation resulted in increased stress-induced cell death. We have therefore shown that Bag5 is overexpressed in prostate cancer and plays a role in ER-stress induced apoptosis. Furthermore we have identified GRP78/BiP as a novel

  20. Cadmium, ATPase-P, yeast. From transport to toxicity

    International Nuclear Information System (INIS)

    Gardarin, Aurelie

    2007-01-01

    Two projects has been developed during my PhD. One consisting in the functional study of CadA, the Cd 2+ -ATPase from Listeria monocytogenes, the other one was focused on the toxicity of cadmium and the associated response of the yeast Saccharomyces cerevisiae. This two studies used a a phenotype of sensitivity to cadmium induced by CadA expression in yeast. This phenotype was used as a screening tool to identify essential amino acids of Cd transport by CadA and to study cadmium toxicity and the corresponding yeast cellular response. CadA actively transports Cd using ATP hydrolysis as energy source. Directed mutagenesis of the membranous polar, sulphur and charged amino-acids revealed that Cd transport pathway implied four transmembrane segments (Tm) and more precisely the cysteine C 354 , C 356 and proline P 355 of the CPC motif located in Tm6, aspartate D 692 in Tm8, glutamate E 164 in Tm4 and methionine M 149 in Tm5. From our studies, 2 Cd ions would be translocated for each hydrolysis ATP. Expression of CadA in the yeast Saccharomyces cerevisiae induces an hypersensitivity to Cd. A wild type cell can grow up to 100 μm cadmium whereas CadA expressing yeast cannot grow with 1 μm cadmium in the culture medium. This cadmium sensitivity was due to the localisation of CadA in the endoplasmic reticulum membrane. Transport of cadmium in this compartment produces an accumulation of mis-folded proteins that induces the Unfolded Protein Response (UPR). As UPR also occurs in a wild type yeast exposed to low Cd concentration, one can point out endoplasmic reticulum as a extremely sensitive cellular compartment. UPR also appears as an early response to Cd as it happens far before any visible signs of toxicity. (author) [fr

  1. Er internethandel kommet for at blive? Eller er det bare krisen som kradser?

    DEFF Research Database (Denmark)

    Ivang, Reimer

    2012-01-01

    I de seneste dage har Jyllandsposten sat fokus på, at omsætningen i den danske detailhandel falder, der fyres, der spares, og det bliver endnu værre i de kommende måneder og år. I artiklen udråbes finanskrisen, som den store skurk, men er det nu den fulde sandhed?...

  2. Minimising Immunohistochemical False Negative ER Classification Using a Complementary 23 Gene Expression Signature of ER Status

    DEFF Research Database (Denmark)

    Li, Qiyuan; Eklund, Aron Charles; Birkbak, Nicolai Juul

    2010-01-01

    with clinical outcome. METHODOLOGY/PRINCIPAL FINDINGS: Firstly, ER status was discriminated by fitting the bimodal expression of ESR1 to a mixed Gaussian model. The discriminative power of ESR1 suggested bimodal expression as an efficient way to stratify breast cancer; therefore we identified a set of genes...

  3. Den der ser, er den, der ses

    DEFF Research Database (Denmark)

    Bjerrum, Troels

    2010-01-01

    Udgangspunktet for afhandlingen er at undersøge, hvordan unge udsatte mennesker kan opnå personlig forandring ved hjælp af åndedrætstræning og meditation. Afhandlingen indleder med at beskrive de unges liv i en globalisering. Mange unge mennesker uden en social ansvarlig baggrund har ikke lært...... at foretage sig nødvendige samfundsmæssige valg. På grund af nederlag i hjem, skole, uddannelse og erhverv søger unge fællesskaber i ligestillede miljøer svarende til egen negative forventningspraksis. Den unge praktiserer en vanedannende negativ livsførelse, som eskalerer til yderligere socialt forfald. Unge......, der har levet i kriminalitet, misbrug og medmisbrug i lang tid, vil ikke være i stand til at realisere sig positivt pga. en personlig indgroet diskvalificerende forestilling. Afhandlingen forklarer, hvorledes negative oplevelser forbliver spændinger i menneskers krop. Teorier af Maurice Merleau...

  4. Isolation, crystallization and crystal structure determination of bovine kidney Na(+),K(+)-ATPase.

    Science.gov (United States)

    Gregersen, Jonas Lindholt; Mattle, Daniel; Fedosova, Natalya U; Nissen, Poul; Reinhard, Linda

    2016-04-01

    Na(+),K(+)-ATPase is responsible for the transport of Na(+) and K(+) across the plasma membrane in animal cells, thereby sustaining vital electrochemical gradients that energize channels and secondary transporters. The crystal structure of Na(+),K(+)-ATPase has previously been elucidated using the enzyme from native sources such as porcine kidney and shark rectal gland. Here, the isolation, crystallization and first structure determination of bovine kidney Na(+),K(+)-ATPase in a high-affinity E2-BeF3(-)-ouabain complex with bound magnesium are described. Crystals belonging to the orthorhombic space group C2221 with one molecule in the asymmetric unit exhibited anisotropic diffraction to a resolution of 3.7 Å with full completeness to a resolution of 4.2 Å. The structure was determined by molecular replacement, revealing unbiased electron-density features for bound BeF3(-), ouabain and Mg(2+) ions.

  5. Partial purification and properties of adenosine triphosphatase (ATPase) from liver fluke Fasciola hepatica.

    Science.gov (United States)

    Hassan, Husain; Abeer, Ali

    2014-01-01

    The adenosine triphosphatase (ATP phosphohydrolase, EC 3.6.1.3.;ATPase) is a membrane -bound enzyme which transport protons across the plasma membrane using ATP as an energy source. The adenosine triphosphatase (ATPase ; EC: 3.6.1.3) was extracted from membrane preparations of adult Fasciola hepatica by chloroform treatment and purified by means of ammonium sulphate fractionation, gel filtration on sephadex G-200 and DEAE- Cellulose chromatography. The molecular weight was calculated to be 305.000 dalton by gel filtration. Kinetic experiments demonstrated a biphasic linear lineweaver - burk relationship (km=0.142 and 1.66 mM) thus revealing the existence of two substrate binding enzyme sites. In our study revealed that partial inhibition of Mg²⁺ dependent purified enzyme by oligomycin suggest the absence of mitochondrial ATPase in F. hepatica.

  6. Cellular localization of Na(+), K(+)-ATPase in the mammalian vestibular system

    Science.gov (United States)

    Kerr, T. P.

    1984-01-01

    Two different, but complementary, procedures for cellular localization of Na+, K+-ATPase in the guinea pig vestibular system were employed. One of these techniques, devised by Stirling, depends upon the well documented ability of the specific inhibitor ouabain to bind selectively to Na+,K+-ATPase, blocking catalytic activity. Microdisected vestibular tissues are incubated with tritium-labelled (3H-) ouabain, and regions with a high concentration of Na+,K+-ATPase are subsequently identified by light microscope autoradiography. A second method, originated by Ernst, detects inorganic phosphate released from an artificial substrate (nitrophenyl phosphate) by catalytic activity of the enzyme. In the presence of strontium ion, phosphate is precipitated near regions of high activity, then converted to a product which may finally be visualized in the electron microscope. This cytochemical enzymatic reaction is inhibited by ouabain.

  7. Nonspecific nature of the vanadate inhibition of rat ileal (Na, K)-ATPase

    International Nuclear Information System (INIS)

    Hajjar, J.J.; Rowe, W.A.; Tomicic, T.K.

    1988-01-01

    Vanadate has been suggested as an intracellular regulator of (Na+ + K+)-ATPase. To test this hypothesis the authors examined the stimulatory and inhibitory effects of vanadate on 86 Rb efflux and influx (measurements of the activity of the Na-pump) in rat ileum under conditions of normal, reduced and increased (Na+ + K+)-ATPase activity. The half maximal inhibition of the Rb efflux and the half maximal inhibition of the Rb influx were not different in the three conditions tested. This suggests that vanadate does not have a regulatory effect on the activity of the Na-K-transport enzyme. The vanadate effect seem rather, to be nonspecific in terms of being unrelated, on a mole per mole basis, to the activity of the (Na+ + K+)-ATPase enzyme

  8. Lens ion transport: from basic concepts to regulation of Na,K-ATPase activity

    Science.gov (United States)

    Delamere, Nicholas A.; Tamiya, Shigeo

    2009-01-01

    In the late 1960s, studies by George Duncan explained many of the basic principles that underlie lens ion homeostasis. The experiments pointed to a permeability barrier close to the surface of the lens and illustrated the requirement for continuous Na,K-ATPase-mediated active sodium extrusion. Without active sodium extrusion, lens sodium and calcium content increases resulting in lens swelling and deterioration of transparency. Later, Duncan's laboratory discovered functional muscarinic and purinergic receptors at the surface of the lens. Recent studies using intact lens suggest purinergic receptors might be involved in short-term regulation of Na,K-ATPase in the epithelium. Purinergic receptor agonists ATP and UTP selectively activate certain Src family tyrosine kinases and stimulate Na,K-ATPase activity. This might represent part of a control mechanism capable of adjusting, perhaps fine tuning, lens ion transport machinery. PMID:18614168

  9. Nonspecific nature of the vanadate inhibition of rat ileal (Na, K)-ATPase

    Energy Technology Data Exchange (ETDEWEB)

    Hajjar, J.J.; Rowe, W.A.; Tomicic, T.K.

    1988-01-01

    Vanadate has been suggested as an intracellular regulator of (Na+ + K+)-ATPase. To test this hypothesis the authors examined the stimulatory and inhibitory effects of vanadate on /sup 86/Rb efflux and influx (measurements of the activity of the Na-pump) in rat ileum under conditions of normal, reduced and increased (Na+ + K+)-ATPase activity. The half maximal inhibition of the Rb efflux and the half maximal inhibition of the Rb influx were not different in the three conditions tested. This suggests that vanadate does not have a regulatory effect on the activity of the Na-K-transport enzyme. The vanadate effect seem rather, to be nonspecific in terms of being unrelated, on a mole per mole basis, to the activity of the (Na+ + K+)-ATPase enzyme.

  10. Atrial Na,K-ATPase increase and potassium dysregulation accentuate the risk of postoperative atrial fibrillation

    DEFF Research Database (Denmark)

    Tran, Cao Thach; Schmidt, Thomas Andersen; Christensen, John Brochorst

    2009-01-01

    BACKGROUND: Postoperative atrial fibrillation is a common complication to cardiac surgery. Na,K-ATPase is of major importance for the resting membrane potential and action potential. The purpose of the present study was to evaluate the importance of Na,K-ATPase concentrations in human atrial...... biopsies and plasma potassium concentrations for the development of atrial fibrillation. METHODS: Atrial myocardial biopsies were obtained from 67 patients undergoing open chest cardiac surgery. Na,K-ATPase was quantified using vanadate-facilitated 3H-ouabain binding. Plasma potassium concentration....../g wet weight (n = 33), p = 0.03]. Also with multivariable analysis, 3H-ouabain-binding site concentration was significantly associated with the development of atrial fibrillation. High increase in plasma potassium concentration during the perioperative period and surgery was associated...

  11. Palytoxin isolated from marine coelenterates. The inhibitory action on (Na,K)-ATPase.

    Science.gov (United States)

    Ishida, Y; Takagi, K; Takahashi, M; Satake, N; Shibata, S

    1983-07-10

    Palytoxin (PTX), C129H223N3O54, a highly toxic substance isolated from zoanthids of Palythoa tuberculosa, inhibited (Na,K)-ATPase (ATP phosphohydrolase, EC 3.6.1.3) prepared from guinea pig heart and hog cerebral cortex in a dose-dependent manner at concentrations greater than 10(-8) M. In the presence of Na (100 mM) and K (20 mM), PTX showed potency nearly equal to that of ouabain. When the ATPase was activated by the various Na concentrations at a constant K concentration, both PTX and ouabain inhibited the ATPase activity noncompetitively. On the other hand, when K concentration was changed at a constant Na concentration, PTX caused a competitive inhibition in all ranges of K concentrations employed, whereas ouabain caused a competitive inhibition at low concentrations and a noncompetitive inhibition at high concentrations.

  12. Vejen til helvede er brolagt med frelste intentioner

    DEFF Research Database (Denmark)

    Vestenskov, David

    2013-01-01

    Det åbenlyse spørgsmål omkring, hvorfor vi er i Afghanistan (eller hvorfor vi bliver?) er bestemt et relevant spørgsmål, men at klæbe et prædikat på om krigens manglende moralske troværdighed kræver mere end blot sure opstød. Er vi netop ikke blevet i Afghanistan grundet moralske principper?...

  13. Employer branding er også intern kommunikation

    DEFF Research Database (Denmark)

    Hansen, Heidi

    2016-01-01

    I videnssamfundet og serviceøkonomien er medarbejderne virksomhedens vigtigste ressource, men milleniumgenerationen er ikke trofaste overfor deres arbejdsplads – de har typisk 15-20 jobs i løbet af deres livstid. Et stærkt employer brand bidrager til at fastholde medarbejdere – og dermed...... kompetencer – i virksomheden, og medarbejderne er dermed en central målgruppe for employer branding....

  14. The ER in 3-D: a multifunctional dynamic membrane network

    OpenAIRE

    Friedman, Jonathan R.; Voeltz, Gia K.

    2011-01-01

    The endoplasmic reticulum (ER) is a large, singular, membrane-bound organelle that has an elaborate 3-D structure with a diversity of structural domains. It contains regions that are flat and cisternal, ones that are highly curved and tubular, and others adapted to form contact with nearly every other organelle and with the plasma membrane. ER 3-D structure is determined by both integral ER membrane proteins and by interactions with the cytoskeleton. Here, we describe some of the factors that...

  15. Evaluating the Potential Bioactivity of a Novel Compound ER1626

    OpenAIRE

    Wang, Lijun; Zeng, Yanyan; Wang, Tianling; Liu, Hongyi; Xiao, Hong; Xiang, Hua

    2014-01-01

    Background ER1626, a novel compound, is a derivate of indeno-isoquinoline ketone. This study was designed to evaluate the biological activity and potential anti-tumor mechanism of ER1626. Method MTT assay, scratch assay and flow cytometry were used to determine cell proliferation, cell migration and cell cycle distribution as well as cell apoptosis on human breast cancer MCF-7 cells and endometrial cancer Ishikawa cells. We also explored the antiangiogenic effect of ER1626 on HUVEC cells and ...

  16. Hvorfor er danske fødevarer dyre?

    DEFF Research Database (Denmark)

    Hansen, Henning Otte

    2015-01-01

    De danske fødevarepriser er generelt høje set i forhold til resten af EU og også i forhold til lande uden for EU. Overordnet set er der en række forklaringer på internationale prisforskelle, og der er også flere mulige årsager til de høje danske fødevarepriser. Forskelle i moms, kvalitet...

  17. A conformation-specific interhelical salt bridge in the K+ binding site of gastric H,K-ATPase

    NARCIS (Netherlands)

    Koenderink, J.B.; Swarts, H.G.P.; Willems, P.H.G.M.; Krieger, E.; Pont, J.J.H.H.M. de

    2004-01-01

    Homology modeling of gastric H, K-ATPase based on the E-2 model of sarcoplasmic reticulum Ca2+-ATPase (Toyoshima, C., and Nomura, H. (2002) Nature 392, 835-839) revealed the presence of a single high-affinity binding site for K+ and an E-2 form-specific salt bridge between Glu(820) (M6) and Lys(791)

  18. A conformation-specific interhelical salt bridge in the K+ binding site of gastric H,K-ATPase.

    NARCIS (Netherlands)

    Koenderink, J.B.; Swarts, H.G.P.; Willems, P.H.G.M.; Krieger, E.; Pont, J.J.H.H.M. de

    2004-01-01

    Homology modeling of gastric H,K-ATPase based on the E2 model of sarcoplasmic reticulum Ca2+-ATPase (Toyoshima, C., and Nomura, H. (2002) Nature 392, 835-839) revealed the presence of a single high-affinity binding site for K+ and an E2 form-specific salt bridge between Glu820 (M6) and Lys791 (M5).

  19. Amino acid substitutions of Na,K-ATPase conferring decreased sensitivity to cardenolides in insects compared to mammals

    NARCIS (Netherlands)

    Dalla, S.; Swarts, H.G.P.; Koenderink, J.B.; Dobler, S.

    2013-01-01

    Mutagenesis analyses and a recent crystal structure of the mammalian Na,K-ATPase have identified amino acids which are responsible for high affinity binding of cardenolides (such as ouabain) which at higher doses block the enzyme in the phosphorylated state. Genetic analysis of the Na,K-ATPase of

  20. Structure-function relationships of Na+, K+, ATP, or Mg2+ binding and energy transduction in Na,K-ATPase

    DEFF Research Database (Denmark)

    Jorgensen, Peter L.; Pedersen, Per Amstrup

    2000-01-01

    Na,K-ATPase; Mutagenesis; Na+ binding; K+ binding; Tl+ binding; Mg2+ binding; ATP binding; Cation binding site; Energy transduction......Na,K-ATPase; Mutagenesis; Na+ binding; K+ binding; Tl+ binding; Mg2+ binding; ATP binding; Cation binding site; Energy transduction...

  1. Mechanism and significance of P4 ATPase-catalyzed lipid transport: lessons from a Na+/K+-pump

    NARCIS (Netherlands)

    Puts, C.F.; Holthuis, J.C.M.

    2009-01-01

    Members of the P4 subfamily of P-type ATPases are believed to catalyze phospholipid transport across membrane bilayers, a process influencing a host of cellular functions. Atomic structures and functional analysis of P-type ATPases that pump small cations and metal ions revealed a transport

  2. H+ V-ATPase-Energized Transporters in Brush Border Membrane Vesicles from Whole Larvae of Aedes Aegypti

    Science.gov (United States)

    Brush Border Membrane vesicles (BBMVs) from Whole larvae of Aedes aegypti (AeBBMVWs ) contain an H+ V-ATPase (V), a Na+/H+ antiporter, NHA1 (A) and a Na+-coupled, nutrient amino acid transporter, NAT8 (N), VAN for short. All V-ATPase subunits are present in the Ae. aegypti genome and in the vesicles...

  3. Relationship between serum leptin levels, ATPase activity of erythrocyte membrance and development of diabetic nephropathy in patients with DM2

    International Nuclear Information System (INIS)

    Wang Yuming

    2009-01-01

    Objective: To study the possible mechanism of development of nephrosis affected by changes of serum leptin levels and alteration of activities of Na + K + -ATPase and Ca 2+ Mg 2+ -ATPase of erythrocyte membrane in patients with type 2 diabetes(DM2). Methods: Serum leptin levels (with RIA) and erythrocyte membrane Na + K + -ATPase and Ca 2+ Mg 2+ -ATPase activitities (with Reinila method) were determined in 40 DM2 patients without nephropathy, 32 DM2 patients with nephropathy and 35 controls. Results Serum leptin levels were significantly higher in the diabetics as a whole than those in controls (P + K + -ATPase and Ca 2+ Mg 2+ -ATPase activities were significantly lower (P<0.01). Among the diabetic patients, the serum leptin levels in patients without nephrosis (P<0.05), but the RBC membrance ATPase activities were significantly lower(P<0.05). Conclusion: Development of type 2 diabetes nephrosis might be correlated with the high serum leptin level and decreased ATPase activities of erythrocite membrane. (authors)

  4. α3Na+/K+-ATPase deficiency causes brain ventricle dilation and abrupt embryonic motility in zebrafish

    DEFF Research Database (Denmark)

    Doğanli, Canan; Beck, Hans Christian; Ribera, Angeles B

    2013-01-01

    Na(+)/K(+)-ATPases are transmembrane ion pumps that maintain ion gradients across the basolateral plasma membrane in all animal cells to facilitate essential biological functions. Mutations in the Na(+)/K(+)-ATPase α3 subunit gene (ATP1A3) cause rapid-onset dystonia-parkinsonism, a rare movement ...

  5. Evaluation of Na+, K+-ATPase activity in the brain of young rats after acute administration of fenproporex.

    Science.gov (United States)

    Rezin, Gislaine T; Scaini, Giselli; Gonçalves, Cinara L; Ferreira, Gabriela K; Cardoso, Mariane R; Ferreira, Andréa G K; Cunha, Maira J; Schmitz, Felipe; Varela, Roger B; Quevedo, João; Wyse, Angela T S; Streck, Emilio L

    2014-01-01

    Fenproporex is an amphetamine-based anorectic which is rapidly converted into amphetamine in vivo. Na+, K+-ATPase is a membrane-bound enzyme necessary to maintain neuronal excitability. Considering that the effects of fenproporex on brain metabolism are poorly known and that Na+, K+-ATPase is essential for normal brain function, this study sought to evaluate the effect of this drug on Na+, K+-ATPase activity in the hippocampus, hypothalamus, prefrontal cortex, and striatum of young rats. Young male Wistar rats received a single injection of fenproporex (6.25, 12.5, or 25 mg/kg intraperitoneally) or polysorbate 80 (control group). Two hours after the last injection, the rats were killed by decapitation and the brain was removed for evaluation of Na+, K+-ATPase activity. Fenproporex decreased Na+, K+-ATPase activity in the striatum of young rats at doses of 6.25, 12.5, and 25 mg/kg and increased enzyme activity in the hypothalamus at the same doses. Na+, K+-ATPase activity was not affected in the hippocampus or prefrontal cortex. Fenproporex administration decreased Na+, K+-ATPase activity in the striatum even in low doses. However, in the hypothalamus, Na+, K+-ATPase activity was increased. Changes in this enzyme might be the result of the effects of fenproporex on neuronal excitability.

  6. Evaluation of Na+, K+-ATPase activity in the brain of young rats after acute administration of fenproporex

    Directory of Open Access Journals (Sweden)

    Gislaine T. Rezin

    2014-05-01

    Full Text Available Objectives: Fenproporex is an amphetamine-based anorectic which is rapidly converted into amphetamine in vivo. Na+, K+-ATPase is a membrane-bound enzyme necessary to maintain neuronal excitability. Considering that the effects of fenproporex on brain metabolism are poorly known and that Na+, K+-ATPase is essential for normal brain function, this study sought to evaluate the effect of this drug on Na+, K+-ATPase activity in the hippocampus, hypothalamus, prefrontal cortex, and striatum of young rats. Methods: Young male Wistar rats received a single injection of fenproporex (6.25, 12.5, or 25 mg/kg intraperitoneally or polysorbate 80 (control group. Two hours after the last injection, the rats were killed by decapitation and the brain was removed for evaluation of Na+, K+-ATPase activity. Results: Fenproporex decreased Na+, K+-ATPase activity in the striatum of young rats at doses of 6.25, 12.5, and 25 mg/kg and increased enzyme activity in the hypothalamus at the same doses. Na+, K+-ATPase activity was not affected in the hippocampus or prefrontal cortex. Conclusion: Fenproporex administration decreased Na+, K+-ATPase activity in the striatum even in low doses. However, in the hypothalamus, Na+, K+-ATPase activity was increased. Changes in this enzyme might be the result of the effects of fenproporex on neuronal excitability.

  7. General and specific lipid-protein interactions in Na,K-ATPase.

    Science.gov (United States)

    Cornelius, F; Habeck, M; Kanai, R; Toyoshima, C; Karlish, S J D

    2015-09-01

    The molecular activity of Na,K-ATPase and other P2 ATPases like Ca(2+)-ATPase is influenced by the lipid environment via both general (physical) and specific (chemical) interactions. Whereas the general effects of bilayer structure on membrane protein function are fairly well described and understood, the importance of the specific interactions has only been realized within the last decade due particularly to the growing field of membrane protein crystallization, which has shed new light on the molecular details of specific lipid-protein interactions. It is a remarkable observation that specific lipid-protein interactions seem to be evolutionarily conserved, and conformations of specifically bound lipids at the lipid-protein surface within the membrane are similar in crystal structures determined with different techniques and sources of the protein, despite the rather weak lipid-protein interaction energy. Studies of purified detergent-soluble recombinant αβ or αβFXYD Na,K-ATPase complexes reveal three separate functional effects of phospholipids and cholesterol with characteristic structural selectivity. The observations suggest that these three effects are exerted at separate binding sites for phophatidylserine/cholesterol (stabilizing), polyunsaturated phosphatidylethanolamine (stimulatory), and saturated PC or sphingomyelin/cholesterol (inhibitory), which may be located within three lipid-binding pockets identified in recent crystal structures of Na,K-ATPase. The findings point to a central role of direct and specific interactions of different phospholipids and cholesterol in determining both stability and molecular activity of Na,K-ATPase and possible implications for physiological regulation by membrane lipid composition. This article is part of a special issue titled "Lipid-Protein Interactions." Copyright © 2015 Elsevier B.V. All rights reserved.

  8. Regulation of renal Na+-K-ATPase in the rat: role of increased potassium transport

    International Nuclear Information System (INIS)

    Mujais, S.K.; Chekal, M.A.; Hayslett, J.P.; Katz, A.I.

    1986-01-01

    The purpose of this study was to characterize the alterations in collecting tubule Na + -K + -ATPase activity produced by sustained increments in dietary potassium in the rat and to evaluate the role of aldosterone in their generation. In adrenal-intact animals, feeding a high-potassium diet or administration of a high physiological dose of aldosterone, which simulates the delivery rate of this hormone during potassium loading, caused marked increments in Na + -K + -ATPase activity in the cortical collecting tubule (CCT) but had no effect on the enzyme in the inner stripe of the medullary collecting tubule (MCT). A significant increase in enzyme activity was also observed after smaller dietary potassium increments and after 4 days of dietary potassium load. In adrenalectomized rats provided with physiological replacement doses of corticosterone and aldosterone, Na + -K + -ATPase activity in both CCT and MCT was similar to that of adrenal-intact controls but remained unchanged after 7 days on the potassium-enriched (10-fold) diet. In contrast, adrenalectomized animals receiving the high physiological dose of aldosterone displayed an increase in Na + -K + -ATPase activity of CCT comparable with that of adrenal-intact animals, whereas the enzyme activity in the MCT was unaffected. In conclusion, 1) following chronic potassium loading Na + -K + -ATPase activity increases significantly in the CCT with no change in its activity in the inner stripe of the MCT; 2) this increase in enzyme activity occurs in a time-dependent fashion and in proportion to the potassium load; and 3) the stimulation of Na + -K + -ATPase activity in adrenal-replaced rats is facilitated by augmented levels of aldosterone, such as those actually observed in adrenal-intact rats subjected to chronic potassium loading

  9. Characterization of detergent-solubilized sarcoplasmic reticulum Ca2+-ATPase by high-performance liquid chromatography

    International Nuclear Information System (INIS)

    Andersen, J.P.; Vilsen, B.; Nielsen, H.; Moller, J.V.

    1986-01-01

    Sarcoplasmic reticulum Ca 2+ -ATPase solubilized by the nonionic detergent octaethylene glycol monododecyl ether was studied by molecular sieve high-performance liquid chromatography (HPLC) and analytical ultracentrifugation. Significant irreversible aggregation of soluble Ca 2+ -ATPase occurred within a few hours in the presence of ≤ 50 μM Ca 2+ . The aggregates were inactive and were primarily held together by hydrophobic forces. In the absence of reducing agent, secondary formation of disulfide bonds occurred. The stability of the inactive dimer upon dilution permitted unambiguous assignment of its elution position and sedimentation coefficient. At high 45 Ca 2+ concentration (500 μM), monomeric Ca 2+ -ATPase was stable for several house. Reversible self-association induced by variation in protein, detergent, and lipid concentrations was studied by large-zone HPLC. The association constant for dimerization of active Ca 2+ -ATPase was found to be 10 5 -10 6 M -1 depending on the detergent concentration. More detergent was bound to monomeric than to dimeric Ca 2+ -ATPase, even above the critical micellar concentration of the detergent. Binding of Ca 2+ and 48 V vanadate as well as ATP-dependent phosphorylation was studied in monomeric and in reversibly associated dimeric preparations. In both forms, two high-affinity Ca 2+ binding sites per phosphorylation site existed. The delipidated monomer purified by HPLC was able to form ADP-insensitive phosphoenzyme and to bind ATP and vanadate simultaneously. The results suggest that formation of Ca 2+ -ATPase oligomers in the membrane is governed by nonspecific forces (low affinity) and that each polypeptide chain constitutes a functional unit

  10. V-ATPase is a candidate therapeutic target for Ewing sarcoma.

    Science.gov (United States)

    Avnet, Sofia; Di Pompo, Gemma; Lemma, Silvia; Salerno, Manuela; Perut, Francesca; Bonuccelli, Gloria; Granchi, Donatella; Zini, Nicoletta; Baldini, Nicola

    2013-08-01

    Suppression of oxidative phosphorylation combined with enhanced aerobic glycolysis and the resulting increased generation of protons are common features of several types of cancer. An efficient mechanism to escape cell death resulting from intracellular acidification is proton pump activation. In Ewing sarcoma (ES), although the tumor-associated chimeric gene EWS-FLI1 is known to induce the accumulation of hypoxia-induced transcription factor HIF-1α, derangements in metabolic pathways have been neglected so far as candidate pathogenetic mechanisms. In this paper, we observed that ES cells simultaneously activate mitochondrial respiration and high levels of glycolysis. Moreover, although the most effective detoxification mechanism of proton intracellular storage is lysosomal compartmentalization, ES cells show a poorly represented lysosomal compartment, but a high sensitivity to the anti-lysosomal agent bafilomycin A1, targeting the V-ATPase proton pump. We therefore investigated the role of V-ATPase in the acidification activity of ES cells. ES cells with the highest GAPDH and V-ATPase expression also showed the highest acidification rate. Moreover, the localization of V-ATPase was both on the vacuolar and the plasma membrane of all ES cell lines. The acidic extracellular pH that we reproduced in vitro promoted high invasion ability and clonogenic efficiency. Finally, targeting V-ATPase with siRNA and omeprazole treatments, we obtained a significant selective reduction of tumor cell number. In summary, glycolytic activity and activation of V-ATPase are crucial mechanisms of survival of ES cells and can be considered as promising selective targets for the treatment of this tumor. Copyright © 2013 Elsevier B.V. All rights reserved.

  11. Binding of the Inhibitor Protein IF1 to Bovine F1-ATPase

    Science.gov (United States)

    Bason, John V.; Runswick, Michael J.; Fearnley, Ian M.; Walker, John E.

    2011-01-01

    In the structure of bovine F1-ATPase inhibited with residues 1–60 of the bovine inhibitor protein IF1, the α-helical inhibitor interacts with five of the nine subunits of F1-ATPase. In order to understand the contributions of individual amino acid residues to this complex binding mode, N-terminal deletions and point mutations have been introduced, and the binding properties of each mutant inhibitor protein have been examined. The N-terminal region of IF1 destabilizes the interaction of the inhibitor with F1-ATPase and may assist in removing the inhibitor from its binding site when F1Fo-ATPase is making ATP. Binding energy is provided by hydrophobic interactions between residues in the long α-helix of IF1 and the C-terminal domains of the βDP-subunit and βTP-subunit and a salt bridge between residue E30 in the inhibitor and residue R408 in the C-terminal domain of the βDP-subunit. Several conserved charged amino acids in the long α-helix of IF1 are also required for establishing inhibitory activity, but in the final inhibited state, they are not in contact with F1-ATPase and occupy aqueous cavities in F1-ATPase. They probably participate in the pathway from the initial interaction of the inhibitor and the enzyme to the final inhibited complex observed in the structure, in which two molecules of ATP are hydrolysed and the rotor of the enzyme turns through two 120° steps. These findings contribute to the fundamental understanding of how the inhibitor functions and to the design of new inhibitors for the systematic analysis of the catalytic cycle of the enzyme. PMID:21192948

  12. Regulation of renal Na -K-ATPase in the rat: role of increased potassium transport

    Energy Technology Data Exchange (ETDEWEB)

    Mujais, S.K.; Chekal, M.A.; Hayslett, J.P.; Katz, A.I.

    1986-08-01

    The purpose of this study was to characterize the alterations in collecting tubule Na -K -ATPase activity produced by sustained increments in dietary potassium in the rat and to evaluate the role of aldosterone in their generation. In adrenal-intact animals, feeding a high-potassium diet or administration of a high physiological dose of aldosterone, which simulates the delivery rate of this hormone during potassium loading, caused marked increments in Na -K -ATPase activity in the cortical collecting tubule (CCT) but had no effect on the enzyme in the inner stripe of the medullary collecting tubule (MCT). A significant increase in enzyme activity was also observed after smaller dietary potassium increments and after 4 days of dietary potassium load. In adrenalectomized rats provided with physiological replacement doses of corticosterone and aldosterone, Na -K -ATPase activity in both CCT and MCT was similar to that of adrenal-intact controls but remained unchanged after 7 days on the potassium-enriched (10-fold) diet. In contrast, adrenalectomized animals receiving the high physiological dose of aldosterone displayed an increase in Na -K -ATPase activity of CCT comparable with that of adrenal-intact animals, whereas the enzyme activity in the MCT was unaffected. In conclusion, 1) following chronic potassium loading Na -K -ATPase activity increases significantly in the CCT with no change in its activity in the inner stripe of the MCT; 2) this increase in enzyme activity occurs in a time-dependent fashion and in proportion to the potassium load; and 3) the stimulation of Na -K -ATPase activity in adrenal-replaced rats is facilitated by augmented levels of aldosterone, such as those actually observed in adrenal-intact rats subjected to chronic potassium loading.

  13. Forbruget af træer og buske i Danmark

    DEFF Research Database (Denmark)

    Jensen, Jan Svejgaard; Laursen, Allan Bach; Kjær, Erik Dahl

    Træer og buske er vigtige elementer i byen og landskabet, og i Danmark bruges der mellem 60 og 80 millioner vedplanter hvert år. Forbruget har været konstant stigende og der er sket store forskydninger i forbrugsmønstret i retning af større fokus på mangfoldighed og autencitet. Undersøgelsen...... klarlægger forbrugsmønstret for træer og buske og undersøger, hvad brugerne lægger vægt på i deres valg af planter....

  14. Preparation of Erbium-169 (169Er) Using Natural Erbium Target

    International Nuclear Information System (INIS)

    Azmairit Aziz; Nana Suherman

    2009-01-01

    The therapeutic radiopharmaceuticals which is labelled by β-particle emission are now increasingly used in nuclear medicine. Erbium-169 ('1 69 Er) is one of radioisotopes that can be used for radiation synovectomy (radio synovectomy) in the treatment of inflammatory joint diseases (arthritis) due to its β- particle emission (T 1/2 =9.4 days, E β maximum =0.34 MeV). The preliminary study on preparation of 169 Er by using natural erbium oxide (Er 2 O 3 ) target irradiated at TRIGA 2000 Bandung reactor has been carried out. The irradiated target was dissolved in hydrochloric acid solution and gentle warming. The optimum condition of 169 Er preparation was obtained by dissolution of 169 Er 2 O 3 by using 1N HCl solution. The radiochemical purity of 169 ErCl 3 was determined by paper chromatography, thin layer chromatography and paper electrophoresis techniques. The solution of 169 ErCl 3 formed was obtained with the pH of 1.5 – 2, clear, with the specific activity of 0.48 – 0.71 MBq/mg Er. The solution has the radiochemical purity of 98.32 ± 1.28% and the radionuclide purity of 99.98%. Study on the stability of 169 ErCl 3 solution showed that the solution was still stable for 4 days at room temperature with the radiochemical purity more than 95%. (author)

  15. Pæren er faldet langt fra stammen

    DEFF Research Database (Denmark)

    Haarder, Jon Helt

    2013-01-01

    Roman: Hassan Preislers vellykkede debutroman udleverer ikke bare multikulti-industrien. Den er også en rablende diagnosticering af det moderne menneskes livsvilkår HASSAN PREISLER BRUN MANDS BYRDE 224 sider, 249,95 Lindhardt og Ringhof Er udkommet 4......Roman: Hassan Preislers vellykkede debutroman udleverer ikke bare multikulti-industrien. Den er også en rablende diagnosticering af det moderne menneskes livsvilkår HASSAN PREISLER BRUN MANDS BYRDE 224 sider, 249,95 Lindhardt og Ringhof Er udkommet 4...

  16. Er Web 2.0 klar til mainstream?

    DEFF Research Database (Denmark)

    Ivang, Reimer

    2009-01-01

    BLOG: Spørgsmålene der relateres til Web 2.0 er mange. Men en af de mest signifikante er om netop din virksomhed skal anvende Web 2.0 teknologier? Hvad kan I få ud af det?......BLOG: Spørgsmålene der relateres til Web 2.0 er mange. Men en af de mest signifikante er om netop din virksomhed skal anvende Web 2.0 teknologier? Hvad kan I få ud af det?...

  17. Evaluating the potential bioactivity of a novel compound ER1626.

    Science.gov (United States)

    Wang, Lijun; Zeng, Yanyan; Wang, Tianling; Liu, Hongyi; Xiao, Hong; Xiang, Hua

    2014-01-01

    ER1626, a novel compound, is a derivate of indeno-isoquinoline ketone. This study was designed to evaluate the biological activity and potential anti-tumor mechanism of ER1626. MTT assay, scratch assay and flow cytometry were used to determine cell proliferation, cell migration and cell cycle distribution as well as cell apoptosis on human breast cancer MCF-7 cells and endometrial cancer Ishikawa cells. We also explored the antiangiogenic effect of ER1626 on HUVEC cells and chicken embryos. The expression of estrogen receptor protein was investigated with western-blot analysis. ER1626 down-regulated the expression of estrogen receptor α protein and up-regulated β protein in MCF-7 and Ishikawa cells. The value of IC50 of ER1626 on MCF-7 and Ishikawa cells were respectively 8.52 and 3.08 µmol/L. Meanwhile, ER1626 decreased VEGF secretion of MCF-7 and Ishikawa cells, disturbed the formation of VEGF-stimulated tubular structure in HUVEC cells, and inhibited the angiogenesis on the chicken chorioallantoic membrane. Scratch assay revealed that ER1626 suppressed the migration of MCF-7, Ishikawa and HUVEC cells. In addition to induction tumor cell apoptosis, ER1626 arrested cell cycle in G1/G0 phase in MCF-7 cells and G2/M phase in Ishikawa cells. In conclusion, our results demonstrated that ER1626 has favorable bioactivities to be a potential candidate against breast cancer and angiogenesis.

  18. Evaluating the potential bioactivity of a novel compound ER1626.

    Directory of Open Access Journals (Sweden)

    Lijun Wang

    Full Text Available ER1626, a novel compound, is a derivate of indeno-isoquinoline ketone. This study was designed to evaluate the biological activity and potential anti-tumor mechanism of ER1626.MTT assay, scratch assay and flow cytometry were used to determine cell proliferation, cell migration and cell cycle distribution as well as cell apoptosis on human breast cancer MCF-7 cells and endometrial cancer Ishikawa cells. We also explored the antiangiogenic effect of ER1626 on HUVEC cells and chicken embryos. The expression of estrogen receptor protein was investigated with western-blot analysis.ER1626 down-regulated the expression of estrogen receptor α protein and up-regulated β protein in MCF-7 and Ishikawa cells. The value of IC50 of ER1626 on MCF-7 and Ishikawa cells were respectively 8.52 and 3.08 µmol/L. Meanwhile, ER1626 decreased VEGF secretion of MCF-7 and Ishikawa cells, disturbed the formation of VEGF-stimulated tubular structure in HUVEC cells, and inhibited the angiogenesis on the chicken chorioallantoic membrane. Scratch assay revealed that ER1626 suppressed the migration of MCF-7, Ishikawa and HUVEC cells. In addition to induction tumor cell apoptosis, ER1626 arrested cell cycle in G1/G0 phase in MCF-7 cells and G2/M phase in Ishikawa cells.In conclusion, our results demonstrated that ER1626 has favorable bioactivities to be a potential candidate against breast cancer and angiogenesis.

  19. Regulatory assembly of the vacuolar proton pump VoV1-ATPase in yeast cells by FLIM-FRET

    Science.gov (United States)

    Ernst, Stefan; Batisse, Claire; Zarrabi, Nawid; Böttcher, Bettina; Börsch, Michael

    2010-02-01

    We investigate the reversible disassembly of VOV1-ATPase in life yeast cells by time resolved confocal FRET imaging. VOV1-ATPase in the vacuolar membrane pumps protons from the cytosol into the vacuole. VOV1-ATPase is a rotary biological nanomotor driven by ATP hydrolysis. The emerging proton gradient is used for secondary transport processes as well as for pH and Ca2+ homoeostasis in the cell. The activity of the VOV1-ATPase is regulated through assembly / disassembly processes. During starvation the two parts of VOV1-ATPase start to disassemble. This process is reversed after addition of glucose. The exact mechanisms are unknown. To follow the disassembly / reassembly in vivo we tagged two subunits C and E with different fluorescent proteins. Cellular distributions of C and E were monitored using a duty cycle-optimized alternating laser excitation scheme (DCO-ALEX) for time resolved confocal FRET-FLIM measurements.

  20. MsmK, an ATPase, Contributes to Utilization of Multiple Carbohydrates and Host Colonization of Streptococcus suis.

    Science.gov (United States)

    Tan, Mei-Fang; Gao, Ting; Liu, Wan-Quan; Zhang, Chun-Yan; Yang, Xi; Zhu, Jia-Wen; Teng, Mu-Ye; Li, Lu; Zhou, Rui

    2015-01-01

    Acquisition and metabolism of carbohydrates are essential for host colonization and pathogenesis of bacterial pathogens. Different bacteria can uptake different lines of carbohydrates via ABC transporters, in which ATPase subunits energize the transport though ATP hydrolysis. Some ABC transporters possess their own ATPases, while some share a common ATPase. Here we identified MsmK, an ATPase from Streptococcus suis, an emerging zoonotic bacterium causing dead infections in pigs and humans. Genetic and biochemistry studies revealed that the MsmK was responsible for the utilization of raffinose, melibiose, maltotetraose, glycogen and maltotriose. In infected mice, the msmK-deletion mutant showed significant defects of survival and colonization when compared with its parental and complementary strains. Taken together, MsmK is an ATPase that contributes to multiple carbohydrates utilization and host colonization of S. suis. This study gives new insight into our understanding of the carbohydrates utilization and its relationship to the pathogenesis of this zoonotic pathogen.

  1. Pro-contractile action of the Na,K-ATPase/Src-kinase signaling pathway in the vascular wall

    DEFF Research Database (Denmark)

    Bouzinova, Elena; Aalkjær, Christian; Matchkov, Vladimir

    Aim: Na,K-ATPase is essential for maintaining the transmembrane ion gradient and might initiate various intracellular signaling. These signals possibly act through a modification of the local ion concentrations or via Src-kinase activation. It is known that inhibition of the α-2 isoform of Na......,K-ATPase by ouabain elevates blood pressure. Consequently, ouabain was shown to potentiate arterial contraction in vitro. In contrast, we have demonstrated that siRNA-induced down-regulation of the α-2 isoform Na,K-ATPase expression reduced arterial sensitivity to agonist stimulation and prevented the effect......) phosphorylation assay. Down-regulation of the α-2 isoform Na,K-ATPase prevented the inhibitory effect of Src inhibitors on arterial contraction. Conclusions: The pro-contractile action of ouabain-sensitive Na,K-ATPase inhibition is associated with Src-kinase inhibition suggesting the role of this signaling...

  2. Data on Na,K-ATPase in primary cultures of renal proximal tubule cells treated with catecholamines

    Directory of Open Access Journals (Sweden)

    Mary Taub

    2016-03-01

    Full Text Available This data article is concerned with chronic regulation of Na,K-ATPase by catecholamines. After a chronic treatment, inhibition of Na,K-ATPase activity was observed in cultures with dopamine, while a stimulation was observed in cultures treated with norepinephrine. Following a chronic incubation with guanabenz, an α adrenergic agonist, an increase in Na,K-ATPase α and β subunit mRNAs was observed. This data supports the research article entitled, “Renal proximal tubule Na, K-ATPase is controlled by CREB regulated transcriptional coactivators as well as salt inducible kinase 1” (Taub et al. 2015 [1]. Keywords: Catecholamines, Kidney, Proximal tubule, Na,K-ATPase, Chronic

  3. Density gradient localization of vanadate- and NO-3-sensitive ATPase from sterile cultures of Spirodela polyrrhiza (L. Schleiden

    Directory of Open Access Journals (Sweden)

    Józef Buczek

    2014-01-01

    Full Text Available The present work deals with the separation and some characteristics of ATPase activities bound with plant membanes prepared from sterile cultures of Spirodela polyrrhiza. The membrane-bound ATPases were separated on sucrose gradients and distinguished by membrane density and sensitivity to several inhibitors. The results showed that N0-3-sensitive ATPase activity associated with the tonoplast was localized at a sucrose density between 1.095-1.117 g•cm-3. The vanadate-sensitive ATPase activity bound with the plasma membrane showed a density between 1.127-1.151 g•cm-3. Both ATPases were insensitive to azide and oligomycin and were separable from markers for mitochondria.

  4. The α2Na+/K+-ATPase is critical for skeletal and heart muscle function in zebrafish

    DEFF Research Database (Denmark)

    Doganli, Canan; Kjaer-Sørensen, Kasper; Knoeckel, Christopher

    2012-01-01

    The Na+/K+-ATPase generates ion gradients across the plasma membrane, essential for multiple cellular functions. In mammals, four different Na+/K+-ATPase α-subunit isoforms are associated with characteristic cell-type expression profiles and kinetics. We found the zebrafish α2Na+/K+-ATPase associ...

  5. Teaching Glycoproteins with a Classical Paper: Knowledge and Methods in the Course of an Exciting Discovery--The story of Discovering HK-ATPase [Beta]-Subunit

    Science.gov (United States)

    Zhu, Lixin

    2008-01-01

    To integrate research into the teaching of glycoproteins, the story of discovering hydrogen-potassium ATPase (HK-ATPase) [beta] subunit is presented in a way covering all the important teaching points. The interaction between the HK-ATPase [alpha] subunit and a glycoprotein of 60-80 kDa was demonstrated to support the existence of the [beta]…

  6. Phe783, Thr797, and Asp804 in transmembrane hairpin M5-M6 of Na+,K+-ATPase play a key role in ouabain binding.

    NARCIS (Netherlands)

    Qiu, L.Y.; Koenderink, J.B.; Swarts, H.G.P.; Willems, P.H.G.M.; Pont, J.J.H.H.M. de

    2003-01-01

    Ouabain is a glycoside that binds to and inhibits the action of Na+,K+-ATPase. Little is known, however, about the specific requirements of the protein surface for glycoside binding. Using chimeras of gastric H+,K+-ATPase and Na+,K+-ATPase, we demonstrated previously that the combined presence of

  7. Identification of Na+/K+-ATPase inhibitors in bovine plasma as fatty acids and hydrocarbons

    DEFF Research Database (Denmark)

    Tal, D M; Yanuck, M D; Van Hall, Gerrit

    1989-01-01

    A preparative purification of endogenous inhibitors of the Na+/K+-ATPase has been carried out from bovine blood. Dried plasma was deproteinized, hexane-extracted and desalted, followed by further purification through a series of reverse-phase HPLC fractionations. Fractions active in inhibiting Na...... ouabain, and in addition it enhanced ouabain binding at high dilutions. These properties are indicative of nonspecific interactions with the Na+/K+-ATPase. The active fraction was identified by TLC, HPLC, NMR, GLC and GC-MS, to be a mixture of three unesterified fatty acids, mainly oleic acid (72...

  8. Effect of TGFβ on Na{sup +}/K{sup +} ATPase activity in megakaryocytes

    Energy Technology Data Exchange (ETDEWEB)

    Hosseinzadeh, Zohreh; Schmid, Evi; Shumilina, Ekaterina [Department of Physiology, University of Tübingen (Germany); Laufer, Stefan [Pharmaceutical Chemistry, University of Tübingen (Germany); Borst, Oliver; Gawaz, Meinrad [Cardiology and Cardiovascular Medicine, University of Tübingen (Germany); Lang, Florian, E-mail: florian.lang@uni-tuebingen.de [Department of Physiology, University of Tübingen (Germany)

    2014-09-26

    Highlights: • TGFß1 markedly up-regulates Na{sup +}/K{sup +} ATPase in megakaryocytes. • The effect is abrogated by p38-MAP kinase inhibitor skepinone. • The effect is abrogated by SGK inhibitor EMD638683. • The effect is abrogated by NF-κB inhibitor wogonin. - Abstract: The Na{sup +}/K{sup +} ATPase generates the Na{sup +} and K{sup +} concentration gradients across the plasma membrane and is thus essential for cellular electrolyte homeostasis, cell membrane potential and cell volume maintenance. A powerful regulator of Na{sup +}/K{sup +} ATPase is the serum- and glucocorticoid-inducible kinase 1 (SGK1). The most powerful known regulator of SGK1 expression is TGFß1, which is pivotal in the regulation of megakaryocyte maturation and platelet formation. Signaling involved in the upregulation of SGK1 by TGFß1 includes p38 mitogen activated protein (MAP) kinase. SGK1 in turn phosphorylates the IκB kinase (IKKα/β), which phosphorylates the inhibitor protein IκBα thus triggering nuclear translocation of nuclear factor kappa B (NF-κB). The present study explored whether TGFβ influences Na{sup +}/K{sup +} ATPase activity in megakaryocytes, and if so, whether the effect of TGß1 requires p38 MAP kinase, SGK1 and/or NF-κB. To this end, murine megakaryocytes were treated with TGFß1 and Na{sup +}/K{sup +} ATPase activity determined from K{sup +} induced current utilizing whole cell patch clamp. The pump current (I{sub pump}) was determined in the absence and presence of Na{sup +}/K{sup +} ATPase inhibitor ouabain (100 μM). TGFß1 (60 ng/ml) was added in the absence or presence of p38 MAP kinase inhibitor skepinone-L (1 μM), SGK1 inhibitor EMD638683 (50 μM) or NF-κB inhibitor wogonin (50 nM). As a result, the I{sub pump} was significantly increased by pretreatment of the megakaryocytes with TGFß1, an effect reaching statistical significance within 16 and 24 h and virtually abrogated in the presence of skepinone-L, EMD638683 or wogonin. In conclusion

  9. Transient expression of P-type ATPases in tobacco epidermal cells

    DEFF Research Database (Denmark)

    Pedas, Lisbeth Rosager; Palmgren, Michael Broberg; Lopez Marques, Rosa Laura

    2016-01-01

    Transient expression in tobacco cells is a convenient method for several purposes such as analysis of protein-protein interactions and the subcellular localization of plant proteins. A suspension of Agrobacterium tumefaciens cells carrying the plasmid of interest is injected into the intracellula...... for example protein-protein interaction studies. In this chapter, we describe the procedure to transiently express P-type ATPases in tobacco epidermal cells, with focus on subcellular localization of the protein complexes formed by P4-ATPases and their β-subunits....

  10. Autoradiographic localization of Na+-K+-ATPase with 3H-ouabain

    International Nuclear Information System (INIS)

    Dormans, J.A.M.A.

    1976-01-01

    Using 3 H-ouabain as an inhibitor, the site of the Na + -K + -ATPase system in cells was determined autoradiographically. Experiments were performed woth guinea pig's kidney tissue. The application of light microscopical autoradiography to freeze-dried tissue showed that especially the distal tubule, and to a smaller extent the proximal tubule and the collecting tubule have Na + -K + -ATPase. Electron microscopical autoradiography showed that this activity is restricted to the baso-lateral plasmamembranes. The quantity of specific bound ouabain turns out to be correlated to the quantity of baso-lateral plasmamembrane's surface

  11. Effect of TGFβ on Na+/K+ ATPase activity in megakaryocytes

    International Nuclear Information System (INIS)

    Hosseinzadeh, Zohreh; Schmid, Evi; Shumilina, Ekaterina; Laufer, Stefan; Borst, Oliver; Gawaz, Meinrad; Lang, Florian

    2014-01-01

    Highlights: • TGFß1 markedly up-regulates Na + /K + ATPase in megakaryocytes. • The effect is abrogated by p38-MAP kinase inhibitor skepinone. • The effect is abrogated by SGK inhibitor EMD638683. • The effect is abrogated by NF-κB inhibitor wogonin. - Abstract: The Na + /K + ATPase generates the Na + and K + concentration gradients across the plasma membrane and is thus essential for cellular electrolyte homeostasis, cell membrane potential and cell volume maintenance. A powerful regulator of Na + /K + ATPase is the serum- and glucocorticoid-inducible kinase 1 (SGK1). The most powerful known regulator of SGK1 expression is TGFß1, which is pivotal in the regulation of megakaryocyte maturation and platelet formation. Signaling involved in the upregulation of SGK1 by TGFß1 includes p38 mitogen activated protein (MAP) kinase. SGK1 in turn phosphorylates the IκB kinase (IKKα/β), which phosphorylates the inhibitor protein IκBα thus triggering nuclear translocation of nuclear factor kappa B (NF-κB). The present study explored whether TGFβ influences Na + /K + ATPase activity in megakaryocytes, and if so, whether the effect of TGß1 requires p38 MAP kinase, SGK1 and/or NF-κB. To this end, murine megakaryocytes were treated with TGFß1 and Na + /K + ATPase activity determined from K + induced current utilizing whole cell patch clamp. The pump current (I pump ) was determined in the absence and presence of Na + /K + ATPase inhibitor ouabain (100 μM). TGFß1 (60 ng/ml) was added in the absence or presence of p38 MAP kinase inhibitor skepinone-L (1 μM), SGK1 inhibitor EMD638683 (50 μM) or NF-κB inhibitor wogonin (50 nM). As a result, the I pump was significantly increased by pretreatment of the megakaryocytes with TGFß1, an effect reaching statistical significance within 16 and 24 h and virtually abrogated in the presence of skepinone-L, EMD638683 or wogonin. In conclusion, TGFß1 is a powerful regulator of megakaryocytic Na + /K + ATPase activity

  12. Organophosphate inhibition of avian salt gland Na, K-ATPase activity

    Science.gov (United States)

    Eastin, W.C.; Fleming, W.J.; Murray, H.C.

    1982-01-01

    1. Adult black ducks (Anas rubripes) were given freshwater or saltwater (1.5% NaCl) for 11 days and half of each group was also given an organophosphate (17 p.p.m. fenthion) in the diet on days 6–11.2. After 11 days, ducks drinking saltrwater had lost more weight and had higher plasma Na and uric acid concentration and osmolalities than birds drinking freshwater.3. Saltwater treatment stimulated the salt gland to increased weight and Na, K-ATPase activity.4. Fenthion generally reduced plasma and brain cholinesterase activity and depressed cholinesterase and Na, K-ATPase activities in salt glands of birds drinking saltwater.

  13. ATP-ase positive cells in human oral mucosa transplanted to nude mice

    DEFF Research Database (Denmark)

    Dabelsteen, E; Kirkeby, S

    1981-01-01

    A model to study the differentiation of human oral epithelium in vivo utilizing transplantation of human tissue to nude mice has been described. Previous studies have described the epithelial cells in this model. In this study we demonstrate that 8 d after transplantation, Langerhans cells, ident......, identified as ATP-ase positive dendritic cells, have almost disappeared from the transplanted epithelium whereas at day 21 after transplantation such cells were abundant. It is suggested that the ATP-ase positive cells which reappear in the transplanted epithelium are of mouse origin....

  14. ATPase activity measurement of DNA replicative helicase from Bacillus stearothermophilus by malachite green method.

    Science.gov (United States)

    Yang, Mu; Wang, Ganggang

    2016-09-15

    The DnaB helicase from Bacillus stearothermophilus (DnaBBst) was a model protein for studying the bacterial DNA replication. In this work, a non-radioactive method for measuring ATPase activity of DnaBBst helicase was described. The working parameters and conditions were optimized. Furthermore, this method was applied to investigate effects of DnaG primase, ssDNA and helicase loader protein (DnaI) on ATPase activity of DnaBBst. Our results showed this method was sensitive and efficient. Moreover, it is suitable for the investigation of functional interaction between DnaB and related factors. Copyright © 2016 Elsevier Inc. All rights reserved.

  15. Effect of fullerene C(60 on ATPase activity and superprecipitation of skeletal muscle actomyosin

    Directory of Open Access Journals (Sweden)

    K. S. Andreichenko

    2013-04-01

    Full Text Available Creation of new biocompatible nanomaterials, which can exhibit the specific biological effects, is an important complex problem that requires the use of last accomplishments of biotechnology. The effect of pristine water-soluble fullerene C60 on ATPase activity and superprecipitation reaction of rabbit skeletal muscle natural actomyosin has been revealed, namely an increase of actomyosin superprecipitation and Мg2+, Са2+– and K+-ATPase activity by fullerene was investigated. We conclude that this finding offers a real possibility for the regulation of contraction-relaxation of skeletal muscle with fullerene C60.

  16. Aspects of gene structure and functional regulation of the isozymes of Na,K-ATPase

    DEFF Research Database (Denmark)

    Jorgensen, P.L.

    2001-01-01

    genomes, the genes of four alpha-subunit and at least three beta-subunit isoforms of Na,K-ATPase are identified and two gamma-subunits are expressed in kidney. The isoforms combine in a number of Na,K-ATPase isozymes that are expressed in a tissue and cell specific manner. Models of the molecular...... mechanism of regulation of these isozymes have become more reliable due to progress in understanding the three-dimensional protein structure and conformational transitions mediating transfer of energy from the P-domain to intramembrane Na+ and K+ binding sites....

  17. Structure and Function of Cu(I)- and Zn(II)-ATPases

    DEFF Research Database (Denmark)

    Sitsel, Oleg; Grønberg, Christina; Autzen, Henriette Elisabeth

    2015-01-01

    Copper and zinc are micronutrients essential for the function of many enzymes while also being toxic at elevated concentrations. Cu(I)- and Zn(II)-transporting P-type ATPases of subclass 1B are of key importance for the homeostasis of these transition metals, allowing ion transport across cellular...... membranes at the expense of ATP. Recent biochemical studies and crystal structures have significantly improved our understanding of the transport mechanisms of these proteins, but many details about their structure and function remain elusive. Here we compare the Cu(I)- and Zn(II)-ATPases, scrutinizing...

  18. Regulation of plant plasma membrane H+- and Ca2+-ATPases by terminal domains

    DEFF Research Database (Denmark)

    Bækgaard, Lone; Fuglsang, Anja Thoe; Palmgren, Michael Gjedde

    2005-01-01

    In the last few years, major progress has been made to elucidate the structure, function, and regulation of P-type plasma membrane H(+)-and Ca(2+)-ATPases. Even though a number of regulatory proteins have been identified, many pieces are still lacking in order to understand the complete regulatory...... mechanisms of these pumps. In plant plasma membrane H(+)- and Ca(2+)-ATPases, autoinhibitory domains are situated in the C- and N-terminal domains, respectively. A model for a common mechanism of autoinhibition is discussed....

  19. Na,K-ATPase biostimulation by low-energy laser irradiation: comparative effects in membrane, solubilized and proteoliposomes enzyme

    Energy Technology Data Exchange (ETDEWEB)

    Rigos, C.F.; Tedesco, A.C.; Ciancaglini, P. [Universidade de Sao Paulo (USP), Ribeirao Preto, SP (Brazil). Faculdade de Filosofia, Ciencias e Letras. Dept. de Quimica; Santos, H.L. [Universidade Federal de Sao Joao Del Rei (UFSJ), MG (Brazil)

    2008-07-01

    Full text: The mechanism of laser irradiation action on living cells is not yet understood. The role of membrane ATPases as possible targets has been analyzed. In our group we have been working with Na,K-ATPase. This enzyme is a member of the P-type family of active cation transport proteins. Thus, the aim of the present work was to investigate the effect of low-energy laser irradiation (685 nm, 35 mW) on the ATPase activity of different forms of the Na,K-ATPase. Membrane-bound and solubilized (ab)2 form of Na,K-ATPase was obtained from the rabbit kidney and DPPC:DPPE-proteoliposomes were prepared by the co-solubilization method. Irradiations were carried out at 685 nm. The ATPase activity of the membrane fraction was not altered with exposition to irradiation doses between 4 and 24 J/c m2. With irradiation doses ranging from 32 to 40 J/c m2, a 28% increase on the ATPase activity was observed while when using up to 50 J/c m2 no additional enhancement was observed. When bio stimulation was done using the purified or the reconstituted enzyme, an increase of about 36-40% on the ATPase activity was observed using only 4-8 J/c m2. With irradiation above these values (24 J/c m2) no additional increase in the activity appeared. These studies revealed that the bio stimulation of ATPase activity from different forms of the Na,K -ATPase is dose dependent in different ranges of irradiation exposure. The stimulation promoted by visible laser doses was modulated and the process was reverted after 2 h for the enzyme present in the membrane and after about 5 h for the solubilized or the reconstituted in DPPC:DPPE-liposomes.

  20. Na,K-ATPase biostimulation by low-energy laser irradiation: comparative effects in membrane, solubilized and proteoliposomes enzyme

    International Nuclear Information System (INIS)

    Rigos, C.F.; Tedesco, A.C.; Ciancaglini, P.

    2008-01-01

    Full text: The mechanism of laser irradiation action on living cells is not yet understood. The role of membrane ATPases as possible targets has been analyzed. In our group we have been working with Na,K-ATPase. This enzyme is a member of the P-type family of active cation transport proteins. Thus, the aim of the present work was to investigate the effect of low-energy laser irradiation (685 nm, 35 mW) on the ATPase activity of different forms of the Na,K-ATPase. Membrane-bound and solubilized (ab)2 form of Na,K-ATPase was obtained from the rabbit kidney and DPPC:DPPE-proteoliposomes were prepared by the co-solubilization method. Irradiations were carried out at 685 nm. The ATPase activity of the membrane fraction was not altered with exposition to irradiation doses between 4 and 24 J/c m2. With irradiation doses ranging from 32 to 40 J/c m2, a 28% increase on the ATPase activity was observed while when using up to 50 J/c m2 no additional enhancement was observed. When bio stimulation was done using the purified or the reconstituted enzyme, an increase of about 36-40% on the ATPase activity was observed using only 4-8 J/c m2. With irradiation above these values (24 J/c m2) no additional increase in the activity appeared. These studies revealed that the bio stimulation of ATPase activity from different forms of the Na,K -ATPase is dose dependent in different ranges of irradiation exposure. The stimulation promoted by visible laser doses was modulated and the process was reverted after 2 h for the enzyme present in the membrane and after about 5 h for the solubilized or the reconstituted in DPPC:DPPE-liposomes

  1. Structure of the /sup 168/Er nucleus and the /sup 166/Er(t,p)/sup 168/Er reaction in terms of the sdg interacting boson model

    Energy Technology Data Exchange (ETDEWEB)

    Akiyama, Y.; Heyde, K.; Arima, A.; Yoshinaga, N.

    1986-05-29

    Extending the interacting boson model by incorporating besides s and d, also the g-boson, we can describe the population of positive parity states of /sup 168/Er in the /sup 166/Er(t,P)/sup 168/Er reaction rather well. In particular, the excitation of I,Ksub(i)sup(..pi..) = 4,3/sub 1//sup +/; 2,2/sub 2//sup +/; 0,0/sub 3//sup +/ and 0,0/sub 4//sup +/ states is much improved over the sd-IBM appraoch.

  2. Large magnetoresistance in Er7Rh3

    International Nuclear Information System (INIS)

    Sengupta, Kaushik; Sampathkumaran, E.V.

    2005-01-01

    The compound Er 2 Rh 3 has been known to order antiferromagnetically below (T N =14K), and to exhibit a change in the sign of temperature coefficient of electrical resistivity (ρ) in the paramagnetic state around 120 K. Here we report the influence of external magnetic field (H) on the ρ(T) behavior of this compound (1.8-300 K). While the ρ behavior in the paramagnetic state, qualitatively speaking, is found to be robust to the application of H, the magnitude of the magnetoresistance (MR) is significant for moderate applications of H, even at temperatures for above T N untypical of metallic systems. In addition, large values are observed in the magnetically ordered state. (author)

  3. Final Technical Report for Award # ER64999

    Energy Technology Data Exchange (ETDEWEB)

    Metcalf, William W. [University of Illinois

    2014-10-08

    This report provides a summary of activities for Award # ER64999, a Genomes to Life Project funded by the Office of Science, Basic Energy Research. The project was entitled "Methanogenic archaea and the global carbon cycle: a systems biology approach to the study of Methanosarcina species". The long-term goal of this multi-investigator project was the creation of integrated, multiscale models that accurately and quantitatively predict the role of Methanosarcina species in the global carbon cycle under dynamic environmental conditions. To achieve these goals we pursed four specific aims: (1) genome sequencing of numerous members of the Order Methanosarcinales, (2) identification of genomic sources of phenotypic variation through in silico comparative genomics, (3) elucidation of the transcriptional networks of two Methanosarcina species, and (4) development of comprehensive metabolic network models for characterized strains to address the question of how metabolic models scale with genetic distance.

  4. ER@CEBAF: Modeling code developments

    Energy Technology Data Exchange (ETDEWEB)

    Meot, F. [Brookhaven National Lab. (BNL), Upton, NY (United States); Thomas Jefferson National Accelerator Facility (TJNAF), Newport News, VA (United States); Roblin, Y. [Brookhaven National Lab. (BNL), Upton, NY (United States); Thomas Jefferson National Accelerator Facility (TJNAF), Newport News, VA (United States)

    2016-04-13

    A proposal for a multiple-pass, high-energy, energy-recovery experiment using CEBAF is under preparation in the frame of a JLab-BNL collaboration. In view of beam dynamics investigations regarding this project, in addition to the existing model in use in Elegant a version of CEBAF is developed in the stepwise ray-tracing code Zgoubi, Beyond the ER experiment, it is also planned to use the latter for the study of polarization transport in the presence of synchrotron radiation, down to Hall D line where a 12 GeV polarized beam can be delivered. This Note briefly reports on the preliminary steps, and preliminary outcomes, based on an Elegant to Zgoubi translation.

  5. Nonrotational states of 165Er nuclei

    International Nuclear Information System (INIS)

    Bonch-Osmolovskaya, N.A.; Morozov, V.A.; Nesterenko, V.O.

    1985-01-01

    165 Er nucleus excited states have been analysed on the basis of available experimental data and calculations within the framework of a quasiparticle-phonon model. β-vibrational components of states are included into the calculations, the Pauli principle was taken account of consecutively. Taking account of the Pauli principle permitting to perform a more correct description of vibrational components resulted in a marked change of characteristics of a number of states especially in the case of 9/2 + [624]. The whole set of levels with 1/2 - [521], 297.4 keV; 1/2 - [530], 589.9 keV; 1/2 - [510], 920.7 keV; 3/2, 1/2 - [541], 1039 keV, is described satisfactorily. The calculation performed gives as a whole a better agreement with experiment as compared with earlier calculations of other authors

  6. ER and PR expression and survival after endometrial cancer.

    Science.gov (United States)

    Smith, Deborah; Stewart, Colin J R; Clarke, Edward M; Lose, Felicity; Davies, Claire; Armes, Jane; Obermair, Andreas; Brennan, Donal; Webb, Penelope M; Nagle, Christina M; Spurdle, Amanda B

    2018-02-01

    To measure association between endometrial carcinoma ER and PR status and endometrial cancer (EC) survival, accounting for inter-observer variation. The intensity and proportion of tumor cell expression of ER and PR in ECs were assessed independently and semi-quantitatively by two pathologists using digital images of duplicate tumor tissue microarrays (TMAs). Cases with inconsistent initial assessment were reviewed and final scoring agreed. The association between overall and EC-specific survival and hormone receptor expression (intensity, proportion and combined) was assessed using Cox regression analysis. The C-index was used to evaluate model discrimination with addition of ER and PR status. Tumor ER and PR analysis was possible in 659 TMAs from 255 patients, and in 459 TMAs from 243 patients, respectively. Initial ER and PR scoring was consistent in 82% and 80% of cases, respectively. In multivariate analyses decreased ER and PR expression was associated with increased tumor-related mortality. Associations reached statistical significance for ER proportion score (P=0.05), ER intensity score (P=0.003), and PR combined score (P=0.04). Decreased expression of combined ER/PR expression was associated with poorer EC-specific survival than decreased expression of either hormone receptor alone (P=0.005). However, hormone receptor status did not significantly improve mortality prediction in individual cases. ER and PR expression combined, using cut-points that capture variation in scoring and across cores, is significantly associated with EC-specific survival in analyses adjusting for known prognostic factors. However, at the individual level, ER and PR expression does not improve mortality prediction. Copyright © 2017 Elsevier Inc. All rights reserved.

  7. EC/β+ decay of 161Er

    International Nuclear Information System (INIS)

    Kalinnikov, V.G.; Ibraheem, Y.S.; Stegailov, V.I.

    2004-01-01

    Full text: The study of odd-proton nuclei in rare-earth region has been performed to get more information on the nuclear structure of these nuclei. In the framework of a program at the ISOL complex of YASNAPP-2 at JINR to study the decay of odd-nuclei, 161 Er has been investigated. The experiments have been focused on the 161 Ho level scheme populated in the EC/β + decay of 161 Er for which a few results have been previously reported [1,2]. Spectra of single γ-ray, γ-γ coincidence and internal conversion electrons were measured. There are appreciable disagreements between results of some γ-ray intensities obtained by us and in [2]. The greatest difference was for relative γ-ray intensity of the 11.28 keV, which has reported value I γ ∼ 10 (I γ 211 =1000) in [2], but this leads to an overpopulation of the 211 keV level by 14% of the decay [3]. The relative intensities of transitions with Eγ γ211 =1000). Inaccuracy of Iγ in [2] is probably due to a sudden change in the efficiency curve of their Ge(Li) detector in the low energy region while our HPGe detector feature a much smoother variation in the efficiency curve in this γ-ray energy region. Besides, the levels earlier introduced by us in [4], three new levels in 161 Ho were proposed from γ-γ coincidences at energies 693.2 keV, 859.6 keV, and 957.97 keV. Spins, parities and Nilsson quantum characteristics of some 161 Ho levels were established

  8. Ikon. iPod er kongen over mp3. Hvad er hemmeligheden?

    DEFF Research Database (Denmark)

    Dickson, Thomas

    2005-01-01

    iPod er blevet ikonet for det 21. århundrede. Men iPod var hverken den første mp3-afspiller eller den første veldesignede af slagsen. Før iPod så dagens lys var Bang & Olufsen på banen med sin næsten lige så lækre BeoSound2.......iPod er blevet ikonet for det 21. århundrede. Men iPod var hverken den første mp3-afspiller eller den første veldesignede af slagsen. Før iPod så dagens lys var Bang & Olufsen på banen med sin næsten lige så lækre BeoSound2....

  9. “Når man er tvunget ud i det, er det egentlig meget godt”

    DEFF Research Database (Denmark)

    Christiansen, René Boyer; Lilius, Karin Marianne; Petersen, Jørgen Haagen

    2018-01-01

    , er en nødvendighed for kompetenceudvikling understøttet af metoden lektionsstudier samt relevante stilladseringsværktøjer der kan rammesætte udviklingen af undervisningskompeten-cer – i dette tilfælde et såkaldt P-CKema. Desuden peger projektets empiri på at projekter der søger at udvikle...

  10. Operational durability of a giant ER valve for Braille display

    Science.gov (United States)

    Luning, Xu; Han, Li; Yufei, Li; Shen, Rong; Kunquan, Lu

    2017-05-01

    The compact configuration of giant ER (electrorheological) valves provides the possibility of realizing a full-page Braille display. The operational durability of ER valves is a key issue in fulfilling a Braille display. A giant ER valve was used to investigate the variations in pressure drops and critical pressure drops of the valves over a long period under some typical operational parameters. The results indicate that neither the pressure drops nor critical pressure drops of giant ER valves show apparent deterioration over a long period. Without ER fluid exchange, a blockage appears in the channel of the valve because the ER structures induced by an external electric field cannot be broken by the Brownian motion of hydraulic oil molecules when the external electric field is removed. Forcing ER fluid flow is an effective and necessary method to keep the channel of the valve unblocked. Thus the operational durability of the valve using giant ER fluids is able to meet the demands of Braille display.

  11. Er der forskel på grinder og grise?

    DEFF Research Database (Denmark)

    Heðinsdóttir, Katla; Gjerris, Mickey

    2015-01-01

    Når det gælder grindedrab, burde man måske spørge, om en tradition, som involverer smerte, angst og andre negative følelser for dyr, er værd at holde fast i, og om det ikke er muligt at fastholde en færøsk identitet uden grindefangst....

  12. Det man hører, er man selv

    DEFF Research Database (Denmark)

    Svømmekjær, Heidi Frank

    2012-01-01

    Katalog til udstillingen "Det man hører, er man selv" på Mediemuseet i Odense 7. september 2012 - 15. januar 2013.......Katalog til udstillingen "Det man hører, er man selv" på Mediemuseet i Odense 7. september 2012 - 15. januar 2013....

  13. Zebrafisken er et vigtigt forsøgsdyr

    DEFF Research Database (Denmark)

    Kjær-Sørensen, Kasper; Alstrup, Aage Kristian Olsen

    2017-01-01

    Det kan ved første øjekast undre, at en almindelig dansk akvariefisk er blevet forskernes favorit. Ikke desto mindre er det tilfældet for den lille zebrafisk (Danio rerio), som i disse år anvendes til at undersøge funktionen af gener, til at lave dyremodeller for menneskets sygdomme og til...

  14. Dietary toxins, endoplasmic reticulum (ER) stress and diabetes.

    Science.gov (United States)

    Hettiarachchi, Kalindi D; Zimmet, Paul Z; Myers, Mark A

    2008-05-01

    The incidence of Type 1 diabetes has been increasing at a rate too rapid to be due to changes in genetic risk. Instead changes in environmental factors are the likely culprit. The endoplasmic reticulum (ER) plays an important role in the production of newly synthesized proteins and interference with these processes leads to ER stress. The insulin-producing beta cells are particularly prone to ER stress as a result of their heavy engagement in insulin production. Increasing evidence suggests ER stress is central to initiation and progression of Type 1 diabetes. An early environmental exposure, such as toxins and viral infections, can impart a significant physiological load on beta cells to initiate abnormal processing of proinsulin, ER stress and insulin secretory defects. Release of altered proinsulin from the beta cells early in life may trigger autoimmunity in those with genetic susceptibility leading to cytokine-induced nitric oxide production and so exacerbating ER stress in beta cells, ultimately leading to apoptosis of beta cells and diabetes. Here we suggest that ER stress is an inherent cause of beta cell dysfunction and environmental factors, in particular dietary toxins derived from Streptomyces in infected root vegetables, can impart additional stress that aggravates beta cell death and progression to diabetes. Furthermore, we propose that the increasing incidence of Type 1 diabetes may be accounted for by increased dietary exposure to ER-stress-inducing Streptomyces toxins.

  15. Hvilke planter er hjemmehørende i Danmark?

    DEFF Research Database (Denmark)

    Erik, Buchwald,; Wind, Peter; Bruun, Hans Henrik

    2013-01-01

    Der har i de senere år været debat om, hvorvidt konkrete plantearter - fx Ær (Acer pseudoplatanus) og Gyvel (Cytisus scoparius) - er hjemmehørende arter i Danmark eller indførte fremmede (Buchwald 2008, 20110: Friis 2010: Møller 2011; Bruun2011). En medvirkende årsag til diskussionerne er, at derr...

  16. Giv Kejseren hvad Kejserens er - på langt sigt

    DEFF Research Database (Denmark)

    Sestoft, Carsten

    2007-01-01

    Fra OECD over EU til det danske videnskabsministerium er der en mængde magtfulde instanser, som i det seneste årti har stillet en række nye krav til universiteterne. Kravene kan i korthed sammenfattes som en fordring om samfundsmæssig og økonomisk relevans. Artiklen argumenterer for, at det er le...

  17. Ny vaccine mod ledbetændelse er ikke effektiv

    DEFF Research Database (Denmark)

    Nielsen, Elisabeth Okholm; Lauritsen, Klara Tølbøll

    2013-01-01

    En ny mulighed for at vaccinere mod mykoplasma-ledbetændelse er undersøgt hos en slagtesvineproducent. Vaccinen kunne desværre ikke forebygge halthed eff ektivt.......En ny mulighed for at vaccinere mod mykoplasma-ledbetændelse er undersøgt hos en slagtesvineproducent. Vaccinen kunne desværre ikke forebygge halthed eff ektivt....

  18. Hvad er fantastisk ved HBO-serien Game of Thrones?

    DEFF Research Database (Denmark)

    Schubart, Rikke

    2013-01-01

    Blogindlæg på videnskab.dk, kaldt "Hvad er fantastisk ved HBO-serien Game of Thrones?" om HBO tv-serien Game of Thrones......Blogindlæg på videnskab.dk, kaldt "Hvad er fantastisk ved HBO-serien Game of Thrones?" om HBO tv-serien Game of Thrones...

  19. Din tandbørste og pejsen er online

    DEFF Research Database (Denmark)

    Damsgaard, Jan

    2016-01-01

    Det er snart definitivt slut med de store mysterier om flys forsvinden og massive eftersøgninger efter overlevende. Danske Naviair er i disse år med til at sende 66 satellitter i kredsløb om Jorden. Så kan alle kommercielle fly følges, uanset hvor på jordkloden de befinder sig....

  20. Danskhed er pizza, kebab og flæskesteg

    DEFF Research Database (Denmark)

    Mehlsen, Camilla

    2010-01-01

    Dannebrog. Kongehuset. Flæskesteg. Folkeskolen. Fodbold og øl. En lang række symboler og fortællinger kendetegner danskhed, men det er en myte, at danskerne er en homogen gruppe, fortæller professor Richard Jenkins....