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Sample records for active inorganic phosphate

  1. High inorganic phosphate causes DNMT1 phosphorylation and subsequent fibrotic fibroblast activation.

    PubMed

    Tan, Xiaoying; Xu, Xingbo; Zeisberg, Elisabeth M; Zeisberg, Michael

    2016-04-08

    Phosphate is an essential constituent of critical cellular functions including energy metabolism, nucleic acid synthesis and phosphorylation-dependent cell signaling. Increased plasma phosphate levels are an independent risk factor for lowered life-expectancy as well as for heart and kidney failure. Nevertheless, direct cellular effects of elevated phosphate concentrations within the microenvironment are poorly understood and have been largely neglected in favor of phosphor-regulatory hormones. Because interstitial fibrosis is the common determinant of chronic progressive kidney disease, and because fibroblasts are major mediators of fibrogenesis, we here explored the effect of high extracellular phosphate levels on renal fibroblasts. We demonstrate that high inorganic phosphate directly induces fibrotic fibroblast activation associated with increased proliferative activity, increased expression of α-smooth muscle actin and increased synthesis of type I collagen. We further demonstrate that such fibroblast activation is dependent on phosphate influx, aberrant phosphorylation of DNA methyltransferase DNMT1 and aberrant CpG island promoter methylation. In summary, our studies demonstrate that elevated phosphate concentrations induce pro-fibrotic fibroblast activation independent of phospho-regulatory hormones.

  2. Depupylase Dop Requires Inorganic Phosphate in the Active Site for Catalysis.

    PubMed

    Bolten, Marcel; Vahlensieck, Christian; Lipp, Colette; Leibundgut, Marc; Ban, Nenad; Weber-Ban, Eilika

    2017-03-10

    Analogous to eukaryotic ubiquitination, proteins in actinobacteria can be post-translationally modified in a process referred to as pupylation, the covalent attachment of prokaryotic ubiquitin-like protein Pup to lysine side chains of the target protein via an isopeptide bond. As in eukaryotes, an opposing activity counteracts the modification by specific cleavage of the isopeptide bond formed with Pup. However, the enzymes involved in pupylation and depupylation have evolved independently of ubiquitination and are related to the family of ATP-binding and hydrolyzing carboxylate-amine ligases of the glutamine synthetase type. Furthermore, the Pup ligase PafA and the depupylase Dop share close structural and sequence homology and have a common evolutionary history despite catalyzing opposing reactions. Here, we investigate the role played by the nucleotide in the active site of the depupylase Dop using a combination of biochemical experiments and X-ray crystallographic studies. We show that, although Dop does not turn over ATP stoichiometrically with substrate, the active site nucleotide species in Dop is ADP and inorganic phosphate rather than ATP, and that non-hydrolyzable analogs of ATP cannot support the enzymatic reaction. This finding suggests that the catalytic mechanism is more similar to the mechanism of the ligase PafA than previously thought and likely involves the transient formation of a phosphorylated Pup-intermediate. Evidence is presented for a mechanism where the inorganic phosphate acts as the nucleophilic species in amide bond cleavage and implications for Dop function are discussed.

  3. Trypanosoma rangeli: differential expression of ecto-phosphatase activities in response to inorganic phosphate starvation.

    PubMed

    Dick, Claudia Fernanda; Dos-Santos, André Luiz Araújo; Fonseca-de-Souza, André L; Rocha-Ferreira, Juliana; Meyer-Fernandes, José Roberto

    2010-04-01

    In this work, we showed that living cells of Trypanosoma rangeli express different ecto-phosphatase activities in response to different inorganic phosphate (Pi) concentrations in the culture medium. The ecto-phosphatase activity from T. rangeli grown at low-Pi concentration was inhibited by the increase of the pH, while the ecto-phosphatase of the cells grown at high Pi concentration was not modulated by the change of the pH of the medium. Okadaic acid inhibited only the ecto-phosphatase activity from cells grown at low-Pi concentration but not the ecto-phosphatase activity from cells grown at high-Pi concentration. Accordingly, phosphatase activity from T. rangeli grown at low Pi concentration was able to hydrolyze P-serine and P-threonine at high rate but not P-tyrosine. The phosphatase activity from T. rangeli grown at high-Pi concentration was able to hydrolyze P-serine, P-threonine and P-tyrosine with the same rate. The addition of anterior midgut homogenate of Rhodnius prolixus on the epimastigotes suspension inhibited the enzyme activity of T. rangeli grown at low-Pi concentration. On the other hand, anterior midgut homogenate had no effect in the ecto-phosphatase of T. rangeli maintained at high-Pi concentration. Altogether, the results described here indicate that ecto-phosphatase activities hydrolyzing phosphorylated compounds present in the extracellular medium of T. rangeli are regulated by the external Pi concentration.

  4. Measurements of metabolically active inorganic phosphate in plants growing in natural and agronomic settings and under water stress. [Stromal Phosphate

    SciTech Connect

    Sharkey, T.D.

    1988-01-01

    At high rates of photosynthesis, the conflicting requirements of adenosine triphosphate (ATP) synthesis for phosphate and starch and sucrose synthesis for low phosphate, may limit the overall rate of photosynthesis. This is called feedback limitation of photosynthesis. A nonaqueous fractionation technique was used to measure stromal phosphate levels without contamination from vacuolar phosphate. Under normal conditions the stromal phosphate level was found to be 7mM. Under feedback limited photosynthesis, this value dropped to <1mM. In a related study, the effect of water stress on photosynthesis was examined. Water stress was shown to cause a decrease in total leaf photosynthesis, due not to a total loss of photosynthetic ability, but rather due to photosynthesis only occurring in patches of the leaf. Water stress was shown to cause a reduction in starch and sucrose synthesis. Since this decline can be reversed by increasing the CO{sub 2} level around the plant, this is proposed to be due to closing of stomata due to the water stress. (MHB)

  5. A Mg(2+)-dependent ecto-phosphatase activity on the external surface of Trypanosoma rangeli modulated by exogenous inorganic phosphate.

    PubMed

    Fonseca-de-Souza, André L; Dick, Claudia Fernanda; Dos Santos, André Luiz Araújo; Meyer-Fernandes, José Roberto

    2008-08-01

    In this work, we characterized a Mg(2+)-dependent ecto-phosphatase activity present in live Trypanosoma rangeli epimastigotes. This enzyme showed capacity to hydrolyze the artificial substrate for phosphatases, p-nitrophenylphosphate (p-NPP). At saturating concentration of p-NPP, half-maximal p-NPP hydrolysis was obtained with 0.23mM Mg(2+). Ca(2+) had no effect on the basal phosphatase activity, could not substitute Mg(2+) as an activator and in contrast inhibited the p-NPP hydrolysis stimulated by Mg(2+). The dependence on p-NPP concentration showed a normal Michaelis-Menten kinetics for this phosphatase activity with values of V(max) of 8.94+/-0.36 nmol p-NP x h(-1) x 10(-7) cells and apparent K(m) of 1.04+/-0.16 mM p-NPP. Mg(2+)-dependent ecto-phosphatase activity was stimulated by the alkaline pH range. Experiments using inhibitors, such as, sodium fluoride, sodium orthovanadate and ammonium molybdate, inhibited the Mg(2+)-dependent ecto-phosphatase activity. Inorganic phosphate (Pi), a product of phosphatases, inhibited reversibly in 50% this activity. Okadaic acid and microcystin-LR, specific phosphoserine/threonine phosphatase inhibitors, inhibited significantly the Mg(2+)-dependent ecto-phosphatase activity. In addition, this phosphatase activity was able to recognize as substrates only o-phosphoserine and o-phosphothreonine, while o-phosphotyrosine was not a good substrate for this phosphatase. Epimastigote forms of T. rangeli exhibit a typical growth curve, achieving the stationary phase around fifth or sixth day and the Mg(2+)-dependent ecto-phosphatase activity decreased around 10-fold with the cell growth progression. Cells maintained at Pi-deprived medium (2 mM Pi) present Mg(2+)-dependent ecto-phosphatase activity approximately threefold higher than that maintained at Pi-supplemented medium (50 mM Pi).

  6. The effects of chronic inorganic and organic phosphate exposure on bactericidal activity of the coelomic fluid of the sea urchin Lytechinus variegatus (Lamarck) (Echinodermata: Echinoidea).

    PubMed

    Böttger, S Anne; McClintock, James B

    2009-07-01

    The sea urchin Lytechinus variegatus can survive chronic exposure to sodium phosphate (inorganic phosphate) concentrations as high as 3.2 mg L-1, and triethyl phosphate (organic phosphate) concentrations of 1000 mg L-1. However, chronic exposure to low (0.8 mg L-1 inorganic and 10 mg L-1 organic phosphate), medium (1.6 mg L-1 inorganic and 100 mg L-1 organic phosphate) or high (3.2 mg L-1 inorganic and 1000 mg L-1 organic phosphate) sublethal concentrations of these phosphates inhibit bactericidal clearance of the marine bacterium Vibrio sp. Bacteria were exposed to coelomic fluid collected from individuals maintained in either artificial seawater, or three concentrations of either inorganic phosphate or organic phosphate. Sterile marine broth, natural seawater and cell free coelomic fluid (cfCF) were employed as controls. Bacterial survival indices were measured at 0, 24 and 48 h periods once a week for four weeks. Bacteria were readily eliminated from the whole coelomic fluid (wCF) of individuals maintained in artificial seawater. Individuals maintained in inorganic phosphates were able to clear bacteria following a two week exposure period, while individuals maintained at even low concentrations of organic phosphates failed to clear all bacteria from their coelomic fluid. Exposure to phosphates represses antimicrobial defenses and may ultimately compromise survival of L. variegatus in the nearshore environment.

  7. Inorganic phosphate inhibits sympathetic neurotransmission in canine saphenous veins

    SciTech Connect

    Edoute, Y.; Vanhoutte, P.M.; Shepherd, J.T.

    1987-01-01

    Inorganic phosphate has been proposed as the initiator of metabolic vasodilatation in active skeletal muscle. The present study was primarily designed to determine if this substance has an inhibitory effect on adrenergic neurotransmission. Rings of canine saphenous veins were suspended for isometric tension recording in organ chambers. A comparison was made of the ability of inorganic phosphate (3 to 14 mM) to relax rings contracted to the same degree by electrical stimulation, exogenous norepinephrine, and prostaglandin F/sub 2..cap alpha../. The relaxation during electrical stimulation was significantly greater at all concentrations of phosphate. In strips of saphenous veins previously incubated with (/sup 3/H)norepinephrine, the depression of the contractile response caused by phosphate during electrical stimulated was accompanied by a significant reduction in the overflow of labeled neurotransmitter. Thus inorganic phosphate inhibits sympathetic neurotransmission and hence may have a key role in the sympatholysis in the active skeletal muscles during exercise. By contrast, in this preparation, it has a modest direct relaxing action on the vascular smooth muscle.

  8. Inorganic Phosphate and Sulfate Transport in S. cerevisiae.

    PubMed

    Samyn, D R; Persson, B L

    2016-01-01

    Inorganic ions such as phosphate and sulfate are essential macronutrients required for a broad spectrum of cellular functions and their regulation. In a constantly fluctuating environment microorganisms have for their survival developed specific nutrient sensing and transport systems ensuring that the cellular nutrient needs are met. This chapter focuses on the S. cerevisiae plasma membrane localized transporters, of which some are strongly induced under conditions of nutrient scarcity and facilitate the active uptake of inorganic phosphate and sulfate. Recent advances in studying the properties of the high-affinity phosphate and sulfate transporters by means of site-directed mutagenesis have provided further insight into the molecular mechanisms contributing to substrate selectivity and transporter functionality of this important class of membrane transporters.

  9. THE DISTRIBUTION OF INORGANIC PHOSPHATE IN AMPHIBIAN MUSCLE

    PubMed Central

    Briner, G. P.; Simon, Shirley E.; Shaw, F. H.

    1958-01-01

    The Na+, K+, and inorganic phosphate levels of the plasma and sartorius muscle of the toad Bufo marinus were determined. Soaking in normal Ringer brought about the usual cation shifts, but did not alter the level of inorganic phosphate in the cell. Increases in the external phosphate level brought about an increase in the internal phosphate, but the apparent phosphate space of muscle is somewhat smaller than the apparent Cl- space. Phosphate spaces were compared with inulin spaces and were found to be significantly greater. Alteration of the H+ concentration of the high phosphate Ringer did not alter the partition of phosphate across the cell membrane. These results have been found to be consistent with the theory of a three compartment system for muscle, wherein the tissue is assumed to consist of an extracellular phase, and two intracellular phases. The inorganic phosphate of the cell is assumed to be adsorbed onto the "ordered phase," and increments in organic phosphate found on raising the external level are assumed to take place in the "free intracellular phase." PMID:13514009

  10. Inorganic Phosphate Prevents Erk1/2 and Stat3 Activation and Improves Sensitivity to Doxorubicin of MDA-MB-231 Breast Cancer Cells.

    PubMed

    Sapio, Luigi; Sorvillo, Luca; Illiano, Michela; Chiosi, Emilio; Spina, Annamaria; Naviglio, Silvio

    2015-09-01

    Due to its expression profile, triple-negative breast cancer (TNBC) is refractory to the most effective targeted therapies available for breast cancer treatment. Thus, cytotoxic chemotherapy represents the mainstay of treatment for early and metastatic TNBC. Therefore, it would be greatly beneficial to develop therapeutic approaches that cause TNBC cells to increase their sensitivity to cytotoxic drugs. Inorganic phosphate (Pi) is emerging as an important signaling molecule in many cell types. Interestingly, it has been shown that Pi greatly enhances the sensitivity of human osteosarcoma cell line (U2OS) to doxorubicin. We investigated the effects of Pi on the sensitivity of TNBC cells to doxorubicin and the underlying molecular mechanisms, carrying out flow cytometry-based assays of cell-cycle progression and cell death, MTT assays, direct cell number counting and immunoblotting experiments. We report that Pi inhibits the proliferation of triple-negative MDA-MB-231 breast cancer cells mainly by slowing down cell cycle progression. Interestingly, we found that Pi strongly increases doxorubicin-induced cytotoxicity in MDA-MB-231 cells by apoptosis induction, as revealed by a marked increase of sub-G1 population, Bcl-2 downregulation, caspase-3 activation and PARP cleavage. Remarkably, Pi/doxorubicin combination-induced cytotoxicity was dynamically accompanied by profound changes in Erk1/2 and Stat3 protein and phosphorylation levels. Altogether, our data enforce the evidence of Pi acting as a signaling molecule in MDA-MB-231 cells, capable of inhibiting Erk and Stat3 pathways and inducing sensitization to doxorubicin of TNBC cells, and suggest that targeting Pi levels at local sites might represent the rationale for developing effective and inexpensive strategies for improving triple-negative breast cancer therapy.

  11. Precaecal phosphorus digestibility of inorganic phosphate sources in male broilers

    PubMed Central

    Bikker, P.; Spek, J. W.; Van Emous, R. A.; Van Krimpen, M. M.

    2016-01-01

    Abstract The aim of this study, comprising two experiments, was (1) to determine in Experiment 1 the relationship of incremental dietary P (phosphorus) content on precaecal digestible P in male broilers and (2) to determine in Experiment 2 the precaecal P digestibility of various inorganic P sources at marginal levels of P supply.In Experiment 1, a total of 260 male Ross 308 broilers were divided into groups of 10 birds per pen resulting in 8 replicates for treatment 1 and 6 replicates for treatments 2–4. Experimental diets were formulated to contain 4 incremental concentrations of digestible P by means of increasing concentrations of monocalcium phosphate (MCP). In the second experiment, 480-d-old male Ross 308 broilers were divided in groups of 12 birds per pen resulting in 16 replicates for the basal diet and 6 replicates for each test diet. A total of 4 inorganic P sources, MCP, monodicalcium phosphate (MDCP), dicalcium phosphate (DCP) and defluorinated phosphate (DFP) were added to the basal diet to determine the precaecal P digestibility. Three of the 4 inorganic P sources (MCP, MDCP and DCP) represented a mix of batches from different producers. At the end of both experiments, the chyme of the posterior part of the small intestine was collected. Digestibility of P and Ca was determined using titanium dioxide as indigestible marker.In Experiment 1, a reduction in precaecal digestibility of P was observed above an estimated precaecal digestible dietary P concentration of 4.8 g/kg.The precaecal P digestibility of the tested inorganic P sources in Experiment 2 was 78.3% for MCP, 59.0% for DCP, 70.7% for MDCP and 31.5% for DFP. PMID:27635437

  12. Precaecal phosphorus digestibility of inorganic phosphate sources in male broilers.

    PubMed

    Bikker, P; Spek, J W; Van Emous, R A; Van Krimpen, M M

    2016-12-01

    The aim of this study, comprising two experiments, was (1) to determine in Experiment 1 the relationship of incremental dietary P (phosphorus) content on precaecal digestible P in male broilers and (2) to determine in Experiment 2 the precaecal P digestibility of various inorganic P sources at marginal levels of P supply. In Experiment 1, a total of 260 male Ross 308 broilers were divided into groups of 10 birds per pen resulting in 8 replicates for treatment 1 and 6 replicates for treatments 2-4. Experimental diets were formulated to contain 4 incremental concentrations of digestible P by means of increasing concentrations of monocalcium phosphate (MCP). In the second experiment, 480-d-old male Ross 308 broilers were divided in groups of 12 birds per pen resulting in 16 replicates for the basal diet and 6 replicates for each test diet. A total of 4 inorganic P sources, MCP, monodicalcium phosphate (MDCP), dicalcium phosphate (DCP) and defluorinated phosphate (DFP) were added to the basal diet to determine the precaecal P digestibility. Three of the 4 inorganic P sources (MCP, MDCP and DCP) represented a mix of batches from different producers. At the end of both experiments, the chyme of the posterior part of the small intestine was collected. Digestibility of P and Ca was determined using titanium dioxide as indigestible marker. In Experiment 1, a reduction in precaecal digestibility of P was observed above an estimated precaecal digestible dietary P concentration of 4.8 g/kg. The precaecal P digestibility of the tested inorganic P sources in Experiment 2 was 78.3% for MCP, 59.0% for DCP, 70.7% for MDCP and 31.5% for DFP.

  13. Dual mechanism of ion permeation through VDAC revealed with inorganic phosphate ions and phosphate metabolites.

    PubMed

    Krammer, Eva-Maria; Vu, Giang Thi; Homblé, Fabrice; Prévost, Martine

    2015-01-01

    In the exchange of metabolites and ions between the mitochondrion and the cytosol, the voltage-dependent anion channel (VDAC) is a key element, as it forms the major transport pathway for these compounds through the mitochondrial outer membrane. Numerous experimental studies have promoted the idea that VDAC acts as a regulator of essential mitochondrial functions. In this study, using a combination of molecular dynamics simulations, free-energy calculations, and electrophysiological measurements, we investigated the transport of ions through VDAC, with a focus on phosphate ions and metabolites. We showed that selectivity of VDAC towards small anions including monovalent phosphates arises from short-lived interactions with positively charged residues scattered throughout the pore. In dramatic contrast, permeation of divalent phosphate ions and phosphate metabolites (AMP and ATP) involves binding sites along a specific translocation pathway. This permeation mechanism offers an explanation for the decrease in VDAC conductance measured in the presence of ATP or AMP at physiological salt concentration. The binding sites occur at similar locations for the divalent phosphate ions, AMP and ATP, and contain identical basic residues. ATP features a marked affinity for a central region of the pore lined by two lysines and one arginine of the N-terminal helix. This cluster of residues together with a few other basic amino acids forms a "charged brush" which facilitates the passage of the anionic metabolites through the pore. All of this reveals that VDAC controls the transport of the inorganic phosphates and phosphate metabolites studied here through two different mechanisms.

  14. Research and application of method of oxygen isotope of inorganic phosphate in Beijing agricultural soils.

    PubMed

    Tian, Liyan; Guo, Qingjun; Zhu, Yongguan; He, Huijun; Lang, Yunchao; Hu, Jian; Zhang, Han; Wei, Rongfei; Han, Xiaokun; Peters, Marc; Yang, Junxing

    2016-12-01

    Phosphorus (P) in agricultural ecosystems is an essential and limited element for plants and microorganisms. However, environmental problems caused by P accumulation as well as by P loss have become more and more serious. Oxygen isotopes of phosphate can trace the sources, migration, and transformation of P in agricultural soils. In order to use the isotopes of phosphate oxygen, appropriate extraction and purification methods for inorganic phosphate from soils are necessary. Here, we combined two different methods to analyze the oxygen isotopic composition of inorganic phosphate (δ(18)OP) from chemical fertilizers and different fractions (Milli-Q water, 0.5 mol L(-1) NaHCO3 (pH = 8.5), 0.1 mol L(-1) NaOH and 1 mol L(-1) HCl) of agricultural soils from the Beijing area. The δ(18)OP results of the water extracts and NaHCO3 extracts in most samples were close to the calculated equilibrium value. These phenomena can be explained by rapid P cycling in soils and the influence of chemical fertilizers. The δ(18)OP value of the water extracts and NaHCO3 extracts in some soil samples below the equilibrium value may be caused by the hydrolysis of organic P fractions mediated by extracellular enzymes. The δ(18)OP values of the NaOH extracts were above the calculated equilibrium value reflecting the balance state between microbial uptake of phosphate and the release of intracellular phosphate back to the soil. The HCl extracts with the lowest δ(18)OP values and highest phosphate concentrations indicated that the HCl fraction was affected by microbial activity. Hence, these δ(18)Op values likely reflected the oxygen isotopic values of the parent materials. The results suggested that phosphate oxygen isotope analyses could be an effective tool in order to trace phosphate sources, transformation processes, and its utilization by microorganisms in agricultural soils.

  15. Biological effects of inorganic phosphate: potential signal of toxicity.

    PubMed

    Hong, Seong-Ho; Park, Sung-Jin; Lee, Somin; Kim, Sanghwa; Cho, Myung-Haing

    2015-02-01

    Inorganic phosphate (Pi) plays crucial roles in several biological processes and signaling pathways. Pi uptake is regulated by sodium-dependent phosphate (Na/Pi) transporters (NPTs). Moreover, Pi is used as a food additive in food items such as sausages, crackers, dairy products, and beverages. However, the high serum concentration of phosphate (> 5.5 mg/dL) can cause adverse renal effects, cardiovascular effects including vascular or valvular calcification, and stimulate bone resorption. In addition, Pi can also alter vital cellular signaling, related to cell growth and cap-dependent protein translation. Moreover, intake of dietary Pi, whether high (1.0%) or low (0.1%), affects organs in developing mice, and is related to tumorigenesis in mice. The recommended dietary allowance (RDA) of Pi is the daily dietary intake required to maintain levels above the lower limit of the range of normal serum Pi concentration (2.7 mg/dL) for most individuals (97-98%). Thus, adequate intake of Pi (RDA; 700 mg/day) and maintenance of normal Pi concentration (2.7-4.5 mg/dL) are important for health and prevention of diseases caused by inadequate Pi intake.

  16. Dual Mechanism of Ion Permeation through VDAC Revealed with Inorganic Phosphate Ions and Phosphate Metabolites

    PubMed Central

    Krammer, Eva-Maria; Vu, Giang Thi; Homblé, Fabrice; Prévost, Martine

    2015-01-01

    In the exchange of metabolites and ions between the mitochondrion and the cytosol, the voltage-dependent anion channel (VDAC) is a key element, as it forms the major transport pathway for these compounds through the mitochondrial outer membrane. Numerous experimental studies have promoted the idea that VDAC acts as a regulator of essential mitochondrial functions. In this study, using a combination of molecular dynamics simulations, free-energy calculations, and electrophysiological measurements, we investigated the transport of ions through VDAC, with a focus on phosphate ions and metabolites. We showed that selectivity of VDAC towards small anions including monovalent phosphates arises from short-lived interactions with positively charged residues scattered throughout the pore. In dramatic contrast, permeation of divalent phosphate ions and phosphate metabolites (AMP and ATP) involves binding sites along a specific translocation pathway. This permeation mechanism offers an explanation for the decrease in VDAC conductance measured in the presence of ATP or AMP at physiological salt concentration. The binding sites occur at similar locations for the divalent phosphate ions, AMP and ATP, and contain identical basic residues. ATP features a marked affinity for a central region of the pore lined by two lysines and one arginine of the N-terminal helix. This cluster of residues together with a few other basic amino acids forms a “charged brush” which facilitates the passage of the anionic metabolites through the pore. All of this reveals that VDAC controls the transport of the inorganic phosphates and phosphate metabolites studied here through two different mechanisms. PMID:25860993

  17. Inorganic phosphate homeostasis. Renal adaptation to the dietary intake in intact and thyroparathyroidectomized rats.

    PubMed Central

    Tröhler, U; Bonjour, J P; Fleisch, H

    1976-01-01

    The possibility of renal tubular adaptation to variations in dietary inorganic phosphate (Pi) was investigated in intact and thyroparathyroidectomized (TPTX) rats pair-fed diets containing low, normal, and high amounts of Pi for periods up to 10 days. Clearances were measured before and during active i.v. infusions with Pi in conscious animals. Thus tubular reabsorption of phosphate (TRPi) could be assessed over a wide range of plasma phosphate concentrations ([Pi]P1). It was found that the renal tubule could adapt its capacity to transport Pi according to the dietary Pi: TRPi was always higher, for a given [Pi]P1, in the animals fed low than in those fed higher Pi diets. This diet-induced modification also occurred in the absence of thyroparathyroid glands, in the presence of the same calcemia and urinary pH, and during marked extracellular volume expansion. A time-course study in rats TPTX both before and during the administration of the experimental diets showed that a difference in the tubular handling of Pi was detectable as early as 3 days after switching the animals from a normal to low- or high-Pi diets. These results indicate that factors other than parathyroid hormone are implicated in the tubular response to variations in the dietary intake of inorganic phosphate. PMID:3518

  18. Effect of altitude on brain intracellular pH and inorganic phosphate levels

    PubMed Central

    Shi, Xian-Feng; Carlson, Paul J.; Kim, Tae-Suk; Sung, Young-Hoon; Hellem, Tracy L.; Fiedler, Kristen K.; Kim, Seong-Eun; Glaeser, Breanna; Wang, Kristina; Zuo, Chun S.; Jeong, Eun-Kee; Renshaw, Perry F.; Kondo, Douglas G.

    2015-01-01

    Normal brain activity is associated with task-related pH changes. Although central nervous system syndromes associated with significant acidosis and alkalosis are well understood, the effects of less dramatic and chronic changes in brain pH are uncertain. One environmental factor known to alter brain pH is the extreme, acute change in altitude encountered by mountaineers. However, the effect of long-term exposure to moderate altitude has not been studied. The aim of this two-site study was to measure brain intracellular pH and phosphate-bearing metabolite levels at two altitudes in healthy volunteers, using phosphorus-31 magnetic resonance spectroscopy (31P-MRS). Increased brain pH and reduced inorganic phosphate (Pi) levels were found in healthy subjects who were long-term residents of Salt Lake City, UT (4720 ft/1438 m), compared with residents of Belmont, MA (20 ft/6 m). Brain intracellular pH at the altitude of 4720 ft was more alkaline than that observed near sea level. In addition, the ratio of inorganic phosphate to total phosphate signal also shifted toward lower values in the Salt Lake City region compared with the Belmont area. These results suggest that long-term residence at moderate altitude is associated with brain chemical changes. PMID:24768210

  19. Effect of altitude on brain intracellular pH and inorganic phosphate levels.

    PubMed

    Shi, Xian-Feng; Carlson, Paul J; Kim, Tae-Suk; Sung, Young-Hoon; Hellem, Tracy L; Fiedler, Kristen K; Kim, Seong-Eun; Glaeser, Breanna; Wang, Kristina; Zuo, Chun S; Jeong, Eun-Kee; Renshaw, Perry F; Kondo, Douglas G

    2014-06-30

    Normal brain activity is associated with task-related pH changes. Although central nervous system syndromes associated with significant acidosis and alkalosis are well understood, the effects of less dramatic and chronic changes in brain pH are uncertain. One environmental factor known to alter brain pH is the extreme, acute change in altitude encountered by mountaineers. However, the effect of long-term exposure to moderate altitude has not been studied. The aim of this two-site study was to measure brain intracellular pH and phosphate-bearing metabolite levels at two altitudes in healthy volunteers, using phosphorus-31 magnetic resonance spectroscopy ((31)P-MRS). Increased brain pH and reduced inorganic phosphate (Pi) levels were found in healthy subjects who were long-term residents of Salt Lake City, UT (4720ft/1438m), compared with residents of Belmont, MA (20ft/6m). Brain intracellular pH at the altitude of 4720ft was more alkaline than that observed near sea level. In addition, the ratio of inorganic phosphate to total phosphate signal also shifted toward lower values in the Salt Lake City region compared with the Belmont area. These results suggest that long-term residence at moderate altitude is associated with brain chemical changes.

  20. Calcium-Activated Phosphate Uptake in Contracting Corn Mitochondria 1

    PubMed Central

    Truelove, B.; Hanson, J. B.

    1966-01-01

    The phosphate inhibition of succinate-powered contraction in corn mitochondria can be reversed with calcium. Associated with this reversal is an accumulation of phosphate and calcium. Both ions are essential for accumulation, although strontium will partially substitute for calcium. Arsenate does not substitute for phosphate except in producing the inhibition of contraction. The antibiotics oligomycin and aurovertin do not block the phosphate inhibition of contraction or the calcium-activated phosphate uptake associated with the release of the inhibition. Dinitrophenol uncouples the phosphate uptake but permits full contraction. Calcium promotes inorganic phosphate accumulation in root tissue as well as in mitochondria. The results are discussed from the viewpoint of theories of calcium reaction with high energy intermediates of oxidative phosphorylation. It is concluded that calcium probably reacts with X∼P in corn mitochondria, rather than with X∼I as with animal mitochondria. PMID:16656343

  1. Inorganic chemically active adsorbents (ICAAs)

    SciTech Connect

    Ally, M.R.; Tavlarides, L.

    1997-10-01

    Oak Ridge National Laboratory (ORNL) researchers are developing a technology that combines metal chelation extraction technology and synthesis chemistry. They begin with a ceramic substrate such as alumina, titanium oxide or silica gel because they provide high surface area, high mechanical strength, and radiolytic stability. One preparation method involves silylation to hydrophobize the surface, followed by chemisorption of a suitable chelation agent using vapor deposition. Another route attaches newly designed chelating agents through covalent bonding by the use of coupling agents. These approaches provide stable and selective, inorganic chemically active adsorbents (ICAAs) tailored for removal of metals. The technology has the following advantages over ion exchange: (1) higher mechanical strength, (2) higher resistance to radiation fields, (3) higher selectivity for the desired metal ion, (4) no cation exchange, (5) reduced or no interference from accompanying anions, (6) faster kinetics, and (7) easy and selective regeneration. Target waste streams include metal-containing groundwater/process wastewater at ORNL`s Y-12 Plant (multiple metals), Savannah River Site (SRS), Rocky Flats (multiple metals), and Hanford; aqueous mixed wastes at Idaho National Engineering Laboratory (INEL); and scrubber water generated at SRS and INEL. Focus Areas that will benefit from this research include Mixed Waste, and Subsurface Contaminants.

  2. Availability of phosphorus from selected inorganic phosphate to juvenile abalone, Haliotis discus hannai ino.

    NASA Astrophysics Data System (ADS)

    Bei-Ping, Tan; Kang-Sen, Mai; Wei, Xu

    2002-06-01

    The availability of phosphorus to juvenile abalone, Haliotis discus hannai from primary, secondary and tertiary calcium phosphate, primary sodium or potassium phosphate separately or in combination was determined in a 120-day feeding trial. Seven semi-purified diets were formulated to contain relatively constant dietary phosphorus, ranging from 0.78%-0.82%. The concentrations of dietary phosphorus were presumed to be slightly below the requirement for abalone. A brown alga, Laminaria japonica, was used as a control diet. Abalone juveniles of silimar size (average weight 1.18 g; average shell length 18.74 mm) were distributed in a single-pass, flow-through system using a completely randomized design with eight treatments and three replicates each treatment. The survival ranged from 71.1% to 81.1%, and was not significantly (P>0.05) related to dietary treatment. However, the weight gain rate (41.72%-65.02%), daily increment in shell length (36.87-55.07 μm) and muscle RNA-DNA ratio (3.44-4.69) were significantly (P<0.05) affected by dietary treatment. Soft body alkaline phosphatase activity (10.9-19.8 U/g wet tissue) and carcass levels of lipid (7.71%-9.33%) and protein (46.68%-49.35%) were significantly (P<0.05) responsive to available phosphorus of the diets. Dietary treatment had significant effect (P<0.05) on concentrations of phosphorus in the whole body (WB) and soft body (SB). Apparent digestibility coefficients (45%-97%) of phosphorus were also significantly (P<0.05) different between dietary treatments. The results indicated that among these inorganic compounds, primary calcium, potassium and sodium phosphates separately or in combination could be utilized effectively by juvenile abalone as dietary phosphorus sources (availability ranging from 87%-97%). However, secondary and tertiary calcium phosphates were found to be low in availability, the values being 45% and 77%, repectively.

  3. Solubilization of insoluble inorganic phosphate by Burkholderia cepacia DA23 isolated from cultivated soil

    PubMed Central

    Song, Ok-Ryul; Lee, Seung-Jin; Lee, Yong-Seok; Lee, Sang-Cheol; Kim, Keun-Ki; Choi, Yong-Lark

    2008-01-01

    A mineral phosphate solubilizing bacterium, Burkholderia cepacia DA23 has been isolated from cultivated soils. Phosphate-solubilizing activities of the strain against three types of insoluble phosphate were quantitatively determined. When 3% of glucose concentration was used for carbon source, the strain had a marked mineral phosphate-solubilizing activity. Mineral phosphate solubilization was directly related to the pH drop by the strain. Analysis of the culture medium by high pressure liquid chromatography identified gluconic acid as the main organic acid released by Burkholderia cepacia DA23. Gluconic acid production was apparently the result of the glucose dehydrogenase activity and glucose dehydrogenase was affected by phosphate regulation. PMID:24031195

  4. Novel organic polymer-inorganic hybrid material zinc poly(styrene-phenylvinylphosphonate)-phosphate prepared with a simple method

    SciTech Connect

    Huang Jing; Fu Xiangkai; Wang Gang; Miao Qiang

    2011-09-15

    A novel type of organic polymer-inorganic hybrid material layered crystalline zinc poly(styrene-phenylvinylphosphonate)-phosphate (ZnPS-PVPP) was synthesized under mild conditions in the absence of any template. And the ZnPS-PVPP were characterized by FT-IR, diffusion reflection UV-vis, AAS, N{sub 2} volumetric adsorption, SEM, TEM and TG. Notably, this method was entirely different from the traditional means used for preparing other zinc phosphonate. Moreover, it could be deduced that ZnPS-PVPP possessed the potential applications for catalyst supports. In the initial catalytic tests, the catalysts immobilized onto ZnPS-PVPP showed comparable or higher activity and enantioselectivity with that of catalysts reported by our group in the asymmetric epoxidation of unfunctional olefins. - Graphical Abstract: Zinc poly(styrene-phenylvinylphosphonate)-phosphate was a novel type of layered crystalline organic polymer-inorganic hybrid material prepared under mild conditions without addition of any template and could be used as heterogeneous catalyst supports. Highlights: > New types of layered crystalline inorganic-organic polymer hybrid materials zinc poly(styrene-phenylvinylphosphonate-phosphate(ZnPS-PVPP)). > ZnPS-PVPP prepared under mild condition without adding of any template. > Immobilized chiral salen Mn (III) catalysts on ZnPS-PVPP supports show comparative activity and enantioselectivity with that of on ZSPP or ZPS-PVPA.

  5. Identification of Phosphorylation Sites Altering Pollen Soluble Inorganic Pyrophosphatase Activity.

    PubMed

    Eaves, Deborah J; Haque, Tamanna; Tudor, Richard L; Barron, Yoshimi; Zampronio, Cleidiane G; Cotton, Nicholas P J; de Graaf, Barend H J; White, Scott A; Cooper, Helen J; Franklin, F Christopher H; Harper, Jeffery F; Franklin-Tong, Vernonica E

    2017-03-01

    Protein phosphorylation regulates numerous cellular processes. Identifying the substrates and protein kinases involved is vital to understand how these important posttranslational modifications modulate biological function in eukaryotic cells. Pyrophosphatases catalyze the hydrolysis of inorganic phosphate (PPi) to inorganic phosphate Pi, driving biosynthetic reactions; they are essential for low cytosolic inorganic phosphate. It was suggested recently that posttranslational regulation of Family I soluble inorganic pyrophosphatases (sPPases) may affect their activity. We previously demonstrated that two pollen-expressed sPPases, Pr-p26.1a and Pr-p26.1b, from the flowering plant Papaver rhoeas were inhibited by phosphorylation. Despite the potential significance, there is a paucity of data on sPPase phosphorylation and regulation. Here, we used liquid chromatographic tandem mass spectrometry to map phosphorylation sites to the otherwise divergent amino-terminal extensions on these pollen sPPases. Despite the absence of reports in the literature on mapping phosphorylation sites on sPPases, a database survey of various proteomes identified a number of examples, suggesting that phosphorylation may be a more widely used mechanism to regulate these enzymes. Phosphomimetic mutants of Pr-p26.1a/b significantly and differentially reduced PPase activities by up to 2.5-fold at pH 6.8 and 52% in the presence of Ca(2+) and hydrogen peroxide over unmodified proteins. This indicates that phosphoregulation of key sites can inhibit the catalytic responsiveness of these proteins in concert with key intracellular events. As sPPases are essential for many metabolic pathways in eukaryotic cells, our findings identify the phosphorylation of sPPases as a potential master regulatory mechanism that could be used to attenuate metabolism.

  6. On the effect of the injection of potassium phosphate in vivo inducing the precipitation of serum calcium with inorganic phosphate

    PubMed Central

    Soares, Alcimar B; Ticianeli, José G; Soares, Letícia B M; Amaro, George

    2013-01-01

    High concentrations of inorganic phosphate (Pi) resulted from the hydrolysis of ATP is strongly associated to the weakness of the contractile mechanism of muscles due to its attractiveness to calcium. The majority of the experiments to study such effect are conducted in vitro. This work investigates the effects of different concentrations of Pi, induced by the injection of potassium phosphate in live animals, in the precipitation with serum calcium and the generation of calcium phosphate composites. The experiments were also designed to find out the ideal amount of potassium phosphate to induce an effective reaction. Potassium phosphate was injected in Wistar rats, randomly separated and distributed into seven groups. Group I was injected with 0.5 ml of saline solution (control) and groups II through VII were injected with 0.5, 1.5, 2.5, 5.0, 7.5 and 10.0 mg/kg of potassium phosphate, respectively. Blood collected from the inferior vena cava was submitted to biochemical analyses to measure the concentrations of calcium, Pi, urea and creatinine. The results showed that Pi, induced by the injection of potassium phosphate in live animals, causes precipitation with serum calcium, with statistically significant differences between the control and the treatment groups for doses up to 5.0 mg/kg. No statistically significant differences were found between the different doses and the concentration of urea and creatinine in the plasma. We conclude that potassium phosphate can be used to induce serum calcium precipitation in-vivo, with minor effects on other physiological variables, and the ideal dose to do so is 5.0 mg/kg. PMID:24379908

  7. The effects of inorganic phosphate and arsenate on both passive muscle visco-elasticity and maximum Ca2+ activated tension in chemically skinned rat fast and slow twitch muscle fibres.

    PubMed

    Mutungi, Gabriel

    2003-01-01

    The effects of adding either 25 mM inorganic phosphate (Pi) or its structural analogue arsenate (ASi) on both the maximum Ca2+ activated tension (Po) and passive muscle visco-elasticity (P2 tension) were investigated at 10 degrees C, using segments of single, chemically skinned rat muscle fibres. Whilst the results confirmed some previous findings on the effects of Pi on Po, they also showed that the addition of 25 mM ASi led to a large (approximately 50%) but completely reversible depression of Po in both the fast and slow twitch rat muscle fibres. Moreover, the depression of Po by ASi was greater at low than at high pH values. Examined in the presence of Dextran T-500, the passive tension and sarcomere length responses to a ramp stretch were found to be qualitatively and quantitatively similar to those previously reported in intact rat muscle fibres. Thus, the tension response to a ramp stretch, in the presence and absence of either 25 mM Pi or ASi, consisted of a viscous (P1), a visco-elastic (P2) and an elastic (P3) tension. However, the addition of either 25 mM Pi or ASi led to approximately 15-18% increase in the amplitude of the visco-elastic (P2) tension but had little or no effect on the amplitudes of the other two tension components (viscous, P1 and elastic, P3 tensions). Furthermore, neither compound significantly altered the relaxation rate of the passive muscle visco-elasticity (P2 tension). These results show that Po (arising from cycling cross-bridges) and passive muscle visco-elasticity (P2 tension) are affected differently by both Pi and ASi and suggest that they may not share a common structural basis. The possibility that passive muscle visco-elasticity (P2 tension) arises from the gap-(titin) filament (as suggested previously by Mutungi and Ranatunga, 1996b J Physiol 496: 827-837) and that Pi and ASi increase its amplitude by interacting with the PEVK region of the filament are discussed.

  8. Availability of phosphorus from selected inorganic phosphate to juvenile abalone, Haliotis discus hannai ino.

    NASA Astrophysics Data System (ADS)

    Tan, Bei-Ping; Mai, Kang-Sen; Xu, Wei

    2002-06-01

    The availability of phosphorus to juvenile abalone, Haliotis discus hannai from primary, secondary and tertiary calcium phosphate, primary sodium or potassium phosphate separately or in combination was determined in a 120-day feeding trial. Seven semi-purified diets were formulated to contain relatively constant dietary phosphorus, ranging from 0.78% 0.82%. The concentrations of dietary phosphorus were presumed to be slightly below the requirement for abalone. A brown alga, Laminaria japonica, was used as a control diet. Abalone juveniles of silimar size (average weight 1.18 g; average shell length 18.74 mm) were distributed in a single-pass, flow-through system using a completely randomized design with eight treatments and three replicates each treatment. The survival ranged from 71.1% to 81.1%, and was not significantly (P>0.05) related to dietary treatment. However, the weight gain rate (41.72% 65.02%), daily increment in shell length (36.87 55.07 μm) and muscle RNA-DNA ratio (3.44 4.69) were significantly (P<0.05) affected by dietary treatment. Soft body alkaline phosphatase activity (10.9 19.8 U/g wet tissue) and carcass levels of lipid (7.71% 9.33%) and protein (46.68% 49.35%) were significantly (P<0.05) responsive to available phosphorus of the diets. Dietary treatment had significant effect (P<0.05) on concentrations of phosphorus in the whole body (WB) and soft body (SB). Apparent digestibility coefficients (45% 97%) of phosphorus were also significantly (P<0.05) different between dietary treatments. The results indicated that among these inorganic compounds, primary calcium, potassium and sodium phosphates separately or in combination could be utilized effectively by juvenile abalone as dietary phosphorus sources (availability ranging from 87% 97%). However, secondary and tertiary calcium phosphates were found to be low in availability, the values being 45% and 77%, repectively.

  9. A precise method for the analysis of d18O of dissolved inorganic phosphate in seawater

    USGS Publications Warehouse

    McLaughlin, K.; Silva, S.; Kendall, C.; Stuart-Williams, Hilary; Paytan, A.

    2004-01-01

    A method for preparation and analysis of the oxygen isotope composition (d18O) of dissolved inorganic phosphate (DIP) has been developed and preliminary results for water samples from various locations are reported. Phosphate is extracted from seawater samples by coprecipitation with magnesium hydroxide. Phosphate is further purified through a series of precipitations and resin separation and is ultimately converted to silver phosphate. Silver phosphate samples are pyrolitically decomposed to carbon monoxide and analyzed for d18O. Silver phosphate samples weighing 0.7 mg (3.5 mol oxygen) can be analyzed routinely with an average standard deviation of about 0.3. There is no isotope fractionation during extraction and blanks are negligible within analytical error. Reproducibility was determined for both laboratory standards and natural samples by multiple analyses. A comparison between filtered and unfiltered natural seawater samples was also conducted and no appreciable difference was observed for the samples tested. The d18O values of DIP in seawater determined using this method range from 18.6 to 22.3, suggesting small but detectable natural variability in seawater. For the San Francisco Bay estuary DIP d18O is more variable, ranging from 11.4 near the San Joaquin River to 20.1 near the Golden Gate Bridge, and was well correlated with salinity, phosphate concentration, and d18O of water.

  10. Controlled formation of calcium-phosphate-based hybrid mesocrystals by organic-inorganic co-assembly.

    PubMed

    Zhai, Halei; Chu, Xiaobin; Li, Li; Xu, Xurong; Tang, Ruikang

    2010-11-01

    An understanding of controlled formation of biomimetic mesocrystals is of great importance in materials chemistry and engineering. Here we report that organic-inorganic hybrid plates and even mesocrystals can be conveniently synthesized using a one-pot reaction in a mixed system of protein (bovine serum albumin (BSA)), surfactant (sodium bis(2-ethylhexyl) sulfosuccinate (AOT)) and supersaturated calcium phosphate solution. The morphologies of calcium-phosphate-based products are analogous to the general inorganic crystals but they have abnormal and interesting substructures. The hybrids are constructed by the alternate stacking of organic layer (thickness of 1.31 nm) and well-crystallized inorganic mineral layer (thickness of 2.13 nm) at the nanoscale. Their morphologies (spindle, rhomboid and round) and sizes (200 nm-2 μm) can be tuned gradually by changing BSA, AOT and calcium phosphate concentrations. This modulation effect can be explained by a competition between the anisotropic and isotropic assembly of the ultrathin plate-like units. The anisotropic assembly confers mesocrystal characteristics on the hybrids while the round ones are the results of isotropic assembly. However, the basic lamellar organic-inorganic substructure remains unchanged during the hybrid formation, which is a key factor to ensure the self-assembly from molecule to micrometre scale. A morphological ternary diagram of BSA-AOT-calcium phosphate is used to describe this controlled formation process, providing a feasible strategy to prepare the required materials. This study highlights the cooperative effect of macromolecule (frame structure), small biomolecule (binding sites) and mineral phase (main component) on the generation and regulation of biomimetic hybrid mesocrystals.

  11. Controlled formation of calcium-phosphate-based hybrid mesocrystals by organic-inorganic co-assembly

    NASA Astrophysics Data System (ADS)

    Zhai, Halei; Chu, Xiaobin; Li, Li; Xu, Xurong; Tang, Ruikang

    2010-11-01

    An understanding of controlled formation of biomimetic mesocrystals is of great importance in materials chemistry and engineering. Here we report that organic-inorganic hybrid plates and even mesocrystals can be conveniently synthesized using a one-pot reaction in a mixed system of protein (bovine serum albumin (BSA)), surfactant (sodium bis(2-ethylhexyl) sulfosuccinate (AOT)) and supersaturated calcium phosphate solution. The morphologies of calcium-phosphate-based products are analogous to the general inorganic crystals but they have abnormal and interesting substructures. The hybrids are constructed by the alternate stacking of organic layer (thickness of 1.31 nm) and well-crystallized inorganic mineral layer (thickness of 2.13 nm) at the nanoscale. Their morphologies (spindle, rhomboid and round) and sizes (200 nm-2 μm) can be tuned gradually by changing BSA, AOT and calcium phosphate concentrations. This modulation effect can be explained by a competition between the anisotropic and isotropic assembly of the ultrathin plate-like units. The anisotropic assembly confers mesocrystal characteristics on the hybrids while the round ones are the results of isotropic assembly. However, the basic lamellar organic-inorganic substructure remains unchanged during the hybrid formation, which is a key factor to ensure the self-assembly from molecule to micrometre scale. A morphological ternary diagram of BSA-AOT-calcium phosphate is used to describe this controlled formation process, providing a feasible strategy to prepare the required materials. This study highlights the cooperative effect of macromolecule (frame structure), small biomolecule (binding sites) and mineral phase (main component) on the generation and regulation of biomimetic hybrid mesocrystals.

  12. Supplementation of inorganic phosphate enhancing the removal efficiency of tannery sludge-borne Cr through bioleaching.

    PubMed

    Zheng, Guanyu; Zhou, Lixiang

    2011-10-15

    Four inorganic mineral nutrients including NH4+, K+, Mg2+ and soluble inorganic phosphate (Pi) were investigated to reveal the potential limiting nutrients for tannery sludge bioleaching process driven by Acidithiobacillus species, and the feasibility of supplementing the limiting nutrients to accelerate tannery sludge bioleaching was studied in the present study. It was found that the concentration of Pi was lower than 3.5 mg/L throughout the whole bioleaching process, which is the most probable restricting nutrient for tannery sludge bioleaching. Further experiments revealed that the deficiency of Pi could seriously influence the growth of Acidithiobacillus thiooxidans and lower its oxidization capacity for S0, and the limiting concentration of Pi for the growth of A. thiooxidans was 6 mg/L. The low concentration of soluble Pi in sludge matrix was resulted from the extremely strong sorbing/binding capacity of tannery sludge for phosphate. The supplementation of more than 1.6 g/L KH2PO4 into tannery sludge bioleaching system could effectively stimulate the growth of Acidithiobacillus species, enhance Cr removal rate and further shorten tannery sludge bioleaching period from 10 days to 7 days. Therefore, inorganic phosphate supplementation is an effective and feasible method to accelerate tannery sludge bioleaching process, and the optimum dosage of KH2PO4 was 1.6 g/L for tannery sludge with 5.1% of total solids.

  13. Heat Stress Affects Pi-related Genes Expression and Inorganic Phosphate Deposition/Accumulation in Barley

    PubMed Central

    Pacak, Andrzej; Barciszewska-Pacak, Maria; Swida-Barteczka, Aleksandra; Kruszka, Katarzyna; Sega, Pawel; Milanowska, Kaja; Jakobsen, Iver; Jarmolowski, Artur; Szweykowska-Kulinska, Zofia

    2016-01-01

    Phosphorus (P) in plants is taken from soil as an inorganic phosphate (Pi) and is one of the most important macroelements in growth and development. Plants actively react to Pi starvation by the induced expression of Pi transporters, MIR399, MIR827, and miR399 molecular sponge – IPS1 genes and by the decreased expression of the ubiquitin-conjugating enzyme E2 (PHOSPHATE2 – PHO2) and Pi sensing and transport SPX-MFS genes. The PHO2 protein is involved in the degradation of Pi transporters PHT1;1 (from soil to roots) and PHO1 (from roots to shoots). The decreased expression of PHO2 leads to Pi accumulation in shoots. In contrast, the pho1 mutant shows a decreased level of Pi concentration in shoots. Finally, Pi starvation leads to decreased Pi concentration in all plant tissues. Little is known about plant Pi homeostasis in other abiotic stress conditions. We found that, during the first hour of heat stress, Pi accumulated in barley shoots but not in the roots, and transcriptomic data analysis as well as RT-qPCR led us to propose an explanation for this phenomenon. Pi transport inhibition from soil to roots is balanced by lower Pi efflux from roots to shoots directed by the PHO1 transporter. In shoots, the PHO2 mRNA level is decreased, leading to an increased Pi level. We concluded that Pi homeostasis in barley during heat stress is maintained by dynamic changes in Pi-related genes expression. PMID:27446155

  14. Heat Stress Affects Pi-related Genes Expression and Inorganic Phosphate Deposition/Accumulation in Barley.

    PubMed

    Pacak, Andrzej; Barciszewska-Pacak, Maria; Swida-Barteczka, Aleksandra; Kruszka, Katarzyna; Sega, Pawel; Milanowska, Kaja; Jakobsen, Iver; Jarmolowski, Artur; Szweykowska-Kulinska, Zofia

    2016-01-01

    Phosphorus (P) in plants is taken from soil as an inorganic phosphate (Pi) and is one of the most important macroelements in growth and development. Plants actively react to Pi starvation by the induced expression of Pi transporters, MIR399, MIR827, and miR399 molecular sponge - IPS1 genes and by the decreased expression of the ubiquitin-conjugating enzyme E2 (PHOSPHATE2 - PHO2) and Pi sensing and transport SPX-MFS genes. The PHO2 protein is involved in the degradation of Pi transporters PHT1;1 (from soil to roots) and PHO1 (from roots to shoots). The decreased expression of PHO2 leads to Pi accumulation in shoots. In contrast, the pho1 mutant shows a decreased level of Pi concentration in shoots. Finally, Pi starvation leads to decreased Pi concentration in all plant tissues. Little is known about plant Pi homeostasis in other abiotic stress conditions. We found that, during the first hour of heat stress, Pi accumulated in barley shoots but not in the roots, and transcriptomic data analysis as well as RT-qPCR led us to propose an explanation for this phenomenon. Pi transport inhibition from soil to roots is balanced by lower Pi efflux from roots to shoots directed by the PHO1 transporter. In shoots, the PHO2 mRNA level is decreased, leading to an increased Pi level. We concluded that Pi homeostasis in barley during heat stress is maintained by dynamic changes in Pi-related genes expression.

  15. H(+)-dependent inorganic phosphate uptake in Trypanosoma brucei is influenced by myo-inositol transporter.

    PubMed

    Russo-Abrahão, Thais; Koeller, Carolina Macedo; Steinmann, Michael E; Silva-Rito, Stephanie; Marins-Lucena, Thaissa; Alves-Bezerra, Michele; Lima-Giarola, Naira Ligia; de-Paula, Iron Francisco; Gonzalez-Salgado, Amaia; Sigel, Erwin; Bütikofer, Peter; Gondim, Katia Calp; Heise, Norton; Meyer-Fernandes, José Roberto

    2017-04-01

    Trypanosoma brucei is an extracellular protozoan parasite that causes human African trypanosomiasis or "sleeping sickness". During the different phases of its life cycle, T. brucei depends on exogenous inorganic phosphate (Pi), but little is known about the transport of Pi in this organism. In the present study, we showed that the transport of (32)Pi across the plasma membrane follows Michaelis-Menten kinetics and is modulated by pH variation, with higher activity at acidic pH. Bloodstream forms presented lower Pi transport in comparison to procyclic forms, that displayed an apparent K0.5 = 0.093 ± 0.008 mM. Additionally, FCCP (H(+)-ionophore), valinomycin (K(+)-ionophore) and SCH28080 (H(+), K(+)-ATPase inhibitor) inhibited the Pi transport. Gene Tb11.02.3020, previously described to encode the parasite H(+):myo-inositol transporter (TbHMIT), was hypothesized to be potentially involved in the H(+):Pi cotransport because of its similarity with the Pho84 transporter described in S. cerevisiae and other trypanosomatids. Indeed, the RNAi mediated knockdown remarkably reduced TbHMIT gene expression, compromised cell growth and decreased Pi transport by half. In addition, Pi transport was inhibited when parasites were incubated in the presence of concentrations of myo-inositol that are above 300 μM. However, when expressed in Xenopus laevis oocytes, two-electrode voltage clamp experiments provided direct electrophysiological evidence that the protein encoded by TbHMIT is definitely a myo-inositol transporter that may be only marginally affected by the presence of Pi. These results confirmed the presence of a Pi carrier in T. brucei, similar to the H(+)-dependent inorganic phosphate system described in S. cerevisiae and other trypanosomatids. This transport system contributes to the acquisition of Pi and may be involved in the growth and survival of procyclic forms. In summary, this work presents the first description of a Pi transport system in T. brucei.

  16. Inorganic phosphate (Pi) modulates the expression of key regulatory proteins of the inorganic pyrophosphate (PPi) metabolism in TGF-β1-stimulated chondrocytes.

    PubMed

    Hamade, Tala; Bianchi, Arnaud; Sebillaud, Sylvie; Netter, Patrick; Jouzeau, Jean-Yves; Cailotto, Frédéric

    2010-01-01

    The balance between extracellular inorganic phosphate (ePi) and extracellular inorganic pyrophosphate (ePPi) is controlled by four membrane proteins: the transporters ANK (exporting PPi outside the cells) and PiT-1 (importing ePi into the cells), and the enzymes PC-1 (generating ePPi from nucleotides) and Tissue Non-specific Alkaline Phosphatase (TNAP, hydrolyzing ePPi into ePi). TGF-β1 was shown to stimulate ANK and PC-1 expression in articular chondrocytes, and subsequent ePPi level, as well as to increase ePi uptake by inducing PiT-1 expression in a chondrogenic cell line. Thus, we investigated the ability of ePi to modulate the effect of TGF-β1 on the regulatory proteins of the ePi/ePPi balance in chondrocytes. In the pathophysiological range of 0.01-1 mM, ePi was inactive by itself but potentiated the stimulatory effects of TGF-β1 on ANK, PC-1 or PiT-1 mRNA (RT-qPCR) and protein (Western blot) levels. PC-1 activity was also increased by TGF-β1 and further potentiated by ePi supplementation. TNAP mRNA and activity became undetectable in response to TGF-β1. These data suggest that ePi could increase ePPi level by changing the control of ANK and PC-1 expression by TGF-β1, further highlighting an adaptative regulation of the Pi/PPi balance to prevent basic calcium phosphate deposition into the joints.

  17. Colorimetric estimation of inorganic phosphate in colored and/or turbid biological samples: assay of phosphohydrolases.

    PubMed

    Upreti, G C

    1984-03-01

    A simple method of inorganic phosphate determination for colored and/or turbid biological samples is described. The procedure is mild, and so is suitable for routine phosphohydrolase assays. Following deproteinization by ice-cold trichloroacetic (or silicotungstic) acid, the sample was treated with acid-washed charcoal to remove interference due to color. The phosphate in the colorless supernatant was assayed either by measuring the phosphomolybdate spectrophotometrically at 310 nm, following its extraction in organic solvents or by a modified Fiske and Subbarow method. The turbidity interference in the latter case was eliminated either by centrifugation, by sodium dodecyl sulfate treatment, or by extraction of reduced phosphomolybdate blue color by cyclohexanone. Though deproteinization by silicotungstic acid eliminated the turbidity problem, its use in conjunction with charcoal treatment was not convenient.

  18. Evaluation of a low temperature hardening Inorganic Phosphate Cement for high-temperature applications

    SciTech Connect

    Alshaaer, M.; Cuypers, H.; Mosselmans, G.; Rahier, H.; Wastiels, J.

    2011-01-15

    Phase and mechanical changes of Inorganic Phosphate Cement (IPC) are identified along with changes in macro properties as functions of temperature and time. In addition to amorphous phases, the presence of significant amounts of brushite and wollastonite in the reference IPC is confirmed using X-ray diffraction. The thermal behavior of IPC up to 1000 {sup o}C shows that contraction of the solid phase in IPC due to chemical transformations causes reduction in the volume of the material. Also the ongoing meta-stable calcium phosphate transformations and reactions over a long time contribute significantly to the phase instability of the material at ambient conditions. It is found that the strength of IPC increases with ageing at ambient conditions but the formation microcracks below 105 {sup o}C causes a sharp reduction in the mechanical performance of IPC. According to the results obtained by Mercury intrusion porosimetry, the pore system of the reference IPC is dominated by mesopores.

  19. A kinetic study of the effects of phosphate and organic phosphates on the activity of phosphoenolpyruvate carboxylase from Crassula argentea.

    PubMed

    Meyer, C R; Rustin, P; Wedding, R T

    1989-05-15

    The effects of phosphate and several phosphate-containing compounds on the activity of purified phosphoenolpyruvate carboxylase (PEPC) from the crassulacean acid metabolism plant, Crassula argentea, were investigated. When assayed at subsaturating phosphoenolpyruvate (PEP) concentrations, low concentrations of most of the compounds tested were found to stimulate PEPC activity. This activation, variable in extent, was found in all cases to be competitive with glucose 6-phosphate (Glc-6-P) stimulation, suggesting that these effectors bind to the Glc-6-P site. At higher concentrations, depending upon the effector molecule studied, deactivation, inhibition, or no response was observed. More detailed studies were performed with Glc-6-P, AMP, phosphoglycolate, and phosphate. AMP had previously been shown to be a specific ligand for the Glc-6-P site. The main effect of Glc-6-P and AMP on the kinetic parameters was to decrease the apparent Km and increase Vmax/Km. AMP also caused a decrease in the Vmax of the reaction. In contrast, phosphoglycolate acted essentially as a competitive inhibitor increasing the apparent Km for PEP and decreasing Vmax/Km. Inorganic phosphate had a biphasic effect on the kinetic parameters, resulting in a transient decrease in Km followed by an increase of the apparent Km for PEP with increasing concentration of phosphate. The Vmax also was decreased with increasing phosphate concentrations. Further, the enzyme appeared to respond to the complex of phosphate with magnesium. In the presence of a saturating concentration of AMP, no activation but rather inhibition was observed with increasing phosphate concentration. This is consistent with the binding of phosphate to two separate sites--the Glc-6-P activation site and an inhibitory site, a phenomenon that may be occurring with other phosphate containing compounds. High concentrations of phosphate with magnesium were found to protect enzyme activity when PEPC, previously shown to contain an

  20. Enzyme activity in dialkyl phosphate ionic liquids

    SciTech Connect

    Thomas, M.F.; Dunn, J.; Li, L.-L.; Handley-Pendleton, J. M.; van der lelie, D.; Wishart, J. F.

    2011-12-01

    The activity of four metagenomic enzymes and an enzyme cloned from the straw mushroom, Volvariellavolvacea were studied in the following ionic liquids, 1,3-dimethylimidazolium dimethyl phosphate, [mmim][dmp], 1-ethyl-3-methylimidazolium dimethyl phosphate, [emim][dmp], 1-ethyl-3-methylimidazolium diethyl phosphate, [emim][dep] and 1-ethyl-3-methylimidazolium acetate, [emim][OAc]. Activity was determined by analyzing the hydrolysis of para-nitrobenzene carbohydrate derivatives. In general, the enzymes were most active in the dimethyl phosphate ionic liquids, followed by acetate. Generally speaking, activity decreased sharply for concentrations of [emim][dep] above 10% v/v, while the other ionic liquids showed less impact on activity up to 20% v/v.

  1. Interstitial Inorganic Phosphate as a Tumor Microenvironment Marker for Tumor Progression

    PubMed Central

    Bobko, Andrey A.; Eubank, Timothy D.; Driesschaert, Benoit; Dhimitruka, Ilirian; Evans, Jason; Mohammad, Rahman; Tchekneva, Elena E.; Dikov, Mikhail M.; Khramtsov, Valery V.

    2017-01-01

    Noninvasive in vivo assessment of chemical tumor microenvironment (TME) parameters such as oxygen (pO2), extracellular acidosis (pHe), and concentration of interstitial inorganic phosphate (Pi) may provide unique insights into biological processes in solid tumors. In this work, we employ a recently developed multifunctional trityl paramagnetic probe and electron paramagnetic resonance (EPR) technique for in vivo concurrent assessment of these TME parameters in various mouse models of cancer. While the data support the existence of hypoxic and acidic regions in TME, the most dramatic differences, about 2-fold higher concentrations in tumors vs. normal tissues, were observed for interstitial Pi - the only parameter that also allowed for discrimination between non-metastatic and highly metastatic tumors. Correlation analysis between [Pi], pO2, pHe and tumor volumes reveal an association of high [Pi] with changes in tumor metabolism and supports different mechanisms of protons and Pi accumulation in TME. Our data identifies interstitial inorganic phosphate as a new TME marker for tumor progression. Pi association with tumor metabolism, buffer-mediated proton transport, and a requirement of high phosphorus content for the rapid growth in the “growth rate hypothesis” may underline its potential role in tumorigenesis and tumor progression. PMID:28117423

  2. Utilization of titanium oxide-like compound as an inorganic phosphate adsorbent for the control of serum phosphate level in chronic renal failure.

    PubMed

    Tamagawa, Kazuhiko; Nakayama-Imaohji, Haruyuki; Wakimoto, Shin; Ichimura, Minoru; Kuwahara, Tomomi

    2010-08-01

    Hyperphosphatemia adversely affects the prognosis of patients with chronic renal failure (CRF). We synthesized a titanium oxide-like compound (TAP) as a phosphate adsorbent for treatment of hyperphosphatemia in CFR patients. We evaluated the ability of TAP to adsorb inorganic phosphate in vitro and in vivo. TAP was shown to contain sulfate and hydroxyl groups by thermal analysis, which probably involved in phosphate adsorption through an ionic exchange mechanism. TAP constantly adsorbed phosphate (66.20-72.84 mg/g TAP) over a wide pH range (1.22-7.27) in vitro. To evaluate the phosphate binding potential of TAP in vivo, adenine-induced CRF rats were fed AIN-76 diet containing 3% TAP, 10% TAP, 3% sevelamer hydrochloride (clinical phosphate adsorbent), or 3% calcium carbonate, and serum levels of phosphate and calcium and urinary phosphate were compared with those in untreated CRF rats. Orally administered TAP showed the inhibitory effect on serum phosphate level in adenine-induced CRF rats, which was equivalent to that of sevelamer hydrochloride. These results indicate that TAP is a useful alternative phosphate-binder with fewer side effects than sevelamer hydrochloride and calcium carbonate.

  3. Estimation of standardized phosphorus retention for inorganic phosphate sources in broilers.

    PubMed

    Liu, S B; Xie, J J; Lu, L; Li, S F; Zhang, L Y; Jiang, Y; Luo, X G

    2013-08-01

    Two experiments were conducted to estimate standardized P retention (SPR) values of dicalcium phosphate (DCP), monocalcium phosphate (MCP), and monopotassium phosphate (MKP) in broilers. In total, ninety-six 22-d-old male broilers with similar BW (780 g average) were used in each experiment. The chicks were randomly allotted to 1 of 4 treatments (P-free, DCP, MCP, or MKP diets) with 6 replicate cages of 4 chicks each in a completely randomized design. After 3-d acclimation, chicks were fasted for 24 h and then fed P-free, DCP, MCP, or MKP diets for 4 h in Exp. 1 or 72 h in Exp. 2. Excreta samples were collected for a total of 28 or 52 h (24 or 48 h after feed withdrawal) in Exp. 1 and 96 or 120 h (24 or 48 h after feed withdrawal) in Exp. 2, respectively. The excreta collection time of 52 h in Exp. 1 or 96 h in Exp. 2 was adequate for the estimation of SPR. The estimated basal endogenous P losses (EPL) in chicks fed the P-free diet were 109 ± 4 mg/52 h per bird and 49.2 ± 4.0 mg/96 h per bird in Exp. 1 and 2, respectively. The SPR values of inorganic phosphate sources corrected by the above basal EPL differed (P < 0.001) in Exp. 2 but not in Exp. 1. However, these SPR values were very similar between the 2 experiments with 68.7, 69.8, or 76.6% in Exp. 1 and 71.8, 70.6, or 78.3% in Exp. 2 for DCP, MCP, or MKP, respectively. The results from the current study indicated that, compared with the 72-h feeding and 96-h excreta collection procedure, the 4-h feeding and 52-h excreta collection procedure was a relatively quicker time- and labor-saving method for estimating the SPR values of inorganic P sources in broilers. The estimated SPR values of commonly used inorganic P sources (MCP and DCP) were about 70%.

  4. Characterization of the interaction between chitosan and inorganic sodium phosphates by means of rheological and optical microscopy studies.

    PubMed

    Casettari, Luca; Cespi, Marco; Palmieri, Giovanni Filippo; Bonacucina, Giulia

    2013-01-16

    The physicochemical and rheological properties of chitosan and two different inorganic sodium phosphate dispersions (NaH(2)PO(4) and Na(3)PO(4)) were investigated in order to elucidate the role of different factors, such as ratios between polymer and sodium inorganic phosphates, different pHs and storage stability, on the gelling properties of chitosan. This was deemed opportune since physico-chemical characterizations of chitosan in the literature often appear incomplete and questionable. We also compared the elastic modulus values of the different chitosan/inorganic phosphate systems and examined their behaviour through optical microscopy analyses. The most efficient formulations that showed a thermogelling capacity with a significant gel transition behaviour after 24h were the NaH(2)PO(4)/chitosan and Na(3)PO(4)/chitosan systems at ratio 2 and pH 7.0. These results confirmed the importance of the pH value and ratio selection for the final systems.

  5. Effects of inorganic phosphate on the light dependent thylakoid energization of intact spinach chloroplasts

    SciTech Connect

    Heineke, D.; Heldt, H.W. ); Stitt, M. )

    1989-09-01

    The light dependent energization of the thylakoid membrane was analyzed in isolated intact spinach (Spinacia oleracea L.) chloroplasts incubated with different concentrations of inorganic phosphate (Pi). Two independent methods were used: (a) the accumulation of ({sup 14}C)5,5-dimethyl-2,4-oxazolidinedione and ({sup 14}C)methylamine; (b) the energy dependent chlorophyll fluorescence quenching. The inhibition of CO{sub 2} fixation by superoptimal medium Pi or by adding glyceraldehyde - an inhibitor of the Calvin cycle - leads to an increased energization of the thylakoid membrane; however, the membrane energization decreases when chloroplasts are inhibited by suboptimal Pi. This specific low phosphate effect could be partially reversed by adding oxaloacetate, which regenerates the electron acceptor NADP{sup +} and stimulates linear electron transport. The energization seen in low Pi is, however, always lower than in superoptimal Pi, even in the presence of oxaloacetate. Energization recovers in the presence of low amounts of N,N{prime}-dicyclohexylcarbodiimide, which reacts with proton channels including the coupling factor 1 ATP synthase. N,N{prime}-Dicyclohexylcarbodiimide has no effect on energization of chloroplasts in superoptimal Pi. These results suggest there is a specific low phosphate proton leak in the thylakoids, and its origin is discussed.

  6. Influence of inorganic phosphate and pH on sarcoplasmic reticular ATPase in skinned muscle fibres of Xenopus laevis.

    PubMed

    Stienen, G J; Papp, Z; Zaremba, R

    1999-08-01

    1. The influence of 30 mM inorganic phosphate (Pi) and pH (6.2-7.4) on the rate of ATP utilization was determined in mechanically skinned bundles of myofibrils from the iliofibularis muscle of Xenopus laevis at approximately 5 C. 2. BDM (2,3-butanedione monoxime; 10 mM) depressed isometric force production and actomyosin (AM) ATPase activity equally. Therefore sarcoplasmic reticular (SR) ATPase activity could be determined by extrapolation of the total ATPase activity to zero force. 3. The SR ATPase activity without added Pi at pH 7.1 was 42 +/- 2 % of the total ATPase activity. Addition of 30 mM Pi reduced SR ATPase activity slightly, by 9 +/- 5 %, and depressed force by 62 +/- 2 % and AM ATPase activity by 21 +/- 6 %. 4. At pH 6.2, force, SR ATPase activity and AM ATPase activity were reduced by 21 +/- 5, 61 +/- 5 and 10 +/- 4 % of their respective values at pH 7.1. 5. The SR ATPase activity at 30 mM Pi and pH 6.2 was reduced markedly to 20 +/- 6 % of the value under control conditions, suggesting that the maximum rate of Ca2+ uptake during muscle fatigue was strongly depressed. This reduction was larger than expected on the basis of the effects of Pi and pH alone.

  7. A simple method for tuning the glass transition process in inorganic phosphate glasses

    NASA Astrophysics Data System (ADS)

    Fulchiron, René; Belyamani, Imane; Otaigbe, Joshua U.; Bounor-Legaré, Véronique

    2015-02-01

    The physical modification of glass transition temperature (Tg) and properties of materials via blending is a common practice in industry and academia and has a large economic advantage. In this context, simple production of hitherto unattainable new inorganic glass blends from already existing glass compositions via blending raises much hope with the potential to provide new glasses with new and improved properties, that cannot be achieved with classical glass synthesis, for a plethora of applications such as computers screens, glass-to-metal seals, and storage materials for nuclear wastes. Here, we demonstrate that blends of the specific glass compositions studied are miscible in all proportions, an unreported phenomenon in hard condensed matter like glass. Interestingly, excellent agreement was found between the obtained data and calculated Tgs from theoretical equations (Supplementary information) for predicting the composition dependence of Tg for miscible blends with weak but significant specific interactions between the blend components. That this blending method is at present not applied to inorganic glasses reflects the fact that water and chemically resistant phosphate glasses with relatively low Tgs have become available only recently.

  8. A simple method for tuning the glass transition process in inorganic phosphate glasses

    PubMed Central

    Fulchiron, René; Belyamani, Imane; Otaigbe, Joshua U.; Bounor-Legaré, Véronique

    2015-01-01

    The physical modification of glass transition temperature (Tg) and properties of materials via blending is a common practice in industry and academia and has a large economic advantage. In this context, simple production of hitherto unattainable new inorganic glass blends from already existing glass compositions via blending raises much hope with the potential to provide new glasses with new and improved properties, that cannot be achieved with classical glass synthesis, for a plethora of applications such as computers screens, glass-to-metal seals, and storage materials for nuclear wastes. Here, we demonstrate that blends of the specific glass compositions studied are miscible in all proportions, an unreported phenomenon in hard condensed matter like glass. Interestingly, excellent agreement was found between the obtained data and calculated Tgs from theoretical equations (Supplementary information) for predicting the composition dependence of Tg for miscible blends with weak but significant specific interactions between the blend components. That this blending method is at present not applied to inorganic glasses reflects the fact that water and chemically resistant phosphate glasses with relatively low Tgs have become available only recently. PMID:25666949

  9. The effect of toxins on inorganic phosphate release during actin polymerization.

    PubMed

    Vig, Andrea; Ohmacht, Róbert; Jámbor, Eva; Bugyi, Beáta; Nyitrai, Miklós; Hild, Gábor

    2011-05-01

    During the polymerization of actin, hydrolysis of bound ATP occurs in two consecutive steps: chemical cleavage of the high-energy nucleotide and slow release of the γ-phosphate. In this study the effect of phalloidin and jasplakinolide on the kinetics of P(i) release was monitored during the formation of actin filaments. An enzyme-linked assay based spectrophotometric technique was used to follow the liberation of inorganic phosphate. It was verified that jasplakinolide reduced the P(i) release in the same way as phalloidin. It was not possible to demonstrate long-range allosteric effects of the toxins by release of P(i) from F-actin. The products of ATP hydrolysis were released by denaturation of the actin filaments. HPLC analysis of the samples revealed that the ATP in the toxin-bound region was completely hydrolysed into ADP and P(i). The effect of both toxins can be sufficiently explained by local and mechanical blockade of P(i) dissociation.

  10. Three-step preparation and purification of phosphorus-33-labeled creatine phosphate of high specific activity

    SciTech Connect

    Savabi, F.; Geiger, P.J.; Bessman, S.P.

    1984-03-01

    Rabbit heart mitochondria were used as a source of enzymes for the synthesis of phosphorus-labeled creatine phosphate. This method is based on the coupled reaction between mitochondrial oxidative phosphorylation and mitochondrial-bound creatine kinase. It is possible to convert more than 90% of the inorganic phosphate (P/sub i/) to creatine phosphate. The method used only small amounts of adenine nucleotides which led to a product with only slight nucleotide contamination. This could be removed by activated charcoal extraction. For further purification, a method for the removal of residual P/sub i/ is described. 20 references.

  11. Effect of decreasing dietary phosphorus supply on net recycling of inorganic phosphate in lactating dairy cows.

    PubMed

    Puggaard, L; Kristensen, N B; Sehested, J

    2011-03-01

    Five ruminally cannulated lactating Holstein cows, fitted with permanent indwelling catheters in the mesenteric vein, hepatic vein, portal vein, and an artery were used to study intestinal absorption and net recycling of inorganic phosphate (P(i)) to the gastrointestinal tract. Treatments were low P (LP; 2.4 g of P/kg of DM) and high P (HP; 3.4 g of P/kg of DM). The dietary total P (tP) concentrations were obtained by replacing 0.50% calcium carbonate in the LP diet with 0.50% monocalcium phosphate in the HP diet. Diets were fed for 14 d and cows were sampled on d 14 in each period. Cows were fed restrictively, resulting in equal dry matter intakes as well as milk, fat, and protein yields between treatments. Net P(i) recycling (primarily salivary) was estimated as the difference between net portal plasma flux (net absorption of P(i)) and apparently digested tP (feed - fecal tP difference). Phosphorus intake, apparently digested tP, and fecal tP excretion decreased with LP. An effect of decreased tP intake on net portal plasma flux of P(i) could not be detected. However, despite numerically minute net fluxes across the liver, the net splanchnic flux of P(i) was less in LP compared with that in HP. Though arterial plasma P(i) concentration decreased, net P(i) recycling was not decreased when tP intake was decreased, and recycling of P(i) was maintained at the expense of deposition of P(i) in bones. Data are not consistent with salivary P(i) secretion being the primary regulator of P(i) homeostasis at low tP intakes. On the contrary, maintaining salivary P(i) recycling at low tP intakes indicates that rumen function was prioritized at the expense of bone P reserves.

  12. High Inorganic Phosphate Intake Promotes Tumorigenesis at Early Stages in a Mouse Model of Lung Cancer

    PubMed Central

    Lee, Somin; Kim, Ji-Eun; Hong, Seong-Ho; Lee, Ah-Young; Park, Eun-Jung; Seo, Hwi Won; Chae, Chanhee; Doble, Philip; Bishop, David; Cho, Myung-Haing

    2015-01-01

    Inorganic phosphate (Pi) is required by all living organisms for the development of organs such as bone, muscle, brain, and lungs, regulating the expression of several critical genes as well as signal transduction. However, little is known about the effects of prolonged dietary Pi consumption on lung cancer progression. This study investigated the effects of a high-phosphate diet (HPD) in a mouse model of adenocarcinoma. K-rasLA1 mice were fed a normal diet (0.3% Pi) or an HPD (1% Pi) for 1, 2, or 4 months. Mice were then sacrificed and subjected to inductively coupled plasma mass/optical emission spectrometry and laser ablation inductively coupled plasma mass-spectrometry analyses, western blot analysis, histopathological, immunohistochemical, and immunocytochemical analyses to evaluate tumor formation and progression (including cell proliferation, angiogenesis, and apoptosis), changes in ion levels and metabolism, autophagy, epithelial-to-mesenchymal transition, and protein translation in the lungs. An HPD accelerated tumorigenesis, as evidenced by increased adenoma and adenocarcinoma rates as well as tumor size. However, after 4 months of the HPD, cell proliferation was arrested, and marked increases in liver and lung ion levels and in energy production via the tricarboxylic acid cycle in the liver were observed, which were accompanied by increased autophagy and decreased angiogenesis and apoptosis. These results indicate that an HPD initially promotes but later inhibits lung cancer progression because of metabolic adaptation leading to tumor cell quiescence. Moreover, the results suggest that carefully regulated Pi consumption are effective in lung cancer prevention. PMID:26285136

  13. Determination of pre-cecal phosphorus digestibility of inorganic phosphates and bone meal products in broilers.

    PubMed

    van Harn, J; Spek, J W; van Vuure, C A; van Krimpen, M M

    2017-02-22

    A broiler study was performed to determine the pre-cecal phosphorus (P) digestibility of 5 P sources, 3 from animal (Delfos, Calfos, and porcine bone meal) and 2 of inorganic (monocalcium phosphate [MCP] and dicalcium phosphate [DCP]) origin. Delfos is processed from bones resulting in a dicalcium phosphate product, and Calfos is processed from bones in which part of the gelatin is removed but in which the hydroxy-apatite matrix is preserved. During the first 14 d, birds were housed in floor pens bedded with wood shavings and received a commercial starter diet. At d 14, broilers were randomly assigned to pens (0.9 m2, 10 birds/pen) with a slatted floor. From d 14 onwards, one of the 6 experimental diets (a basal diet, and 5 diets containing the P sources) was provided. Test diets were replicated 6 times, and the basal diet 8 times. Electron microscopy images of test products were made in order to verify whether the spatial structure of the test products could be related to the pre-cecal P digestibility of the same products. Diets met or exceeded CVB (2011) requirements for all nutrients except for P and were formulated to contain a calcium to total P ratio of between 1.4 and 1.6 and a minimal amount of phytate P. Diets contained 5 g/kg titanium oxide as a marker to determine digestibility of P. At d 24 all birds were euthanized, after which the content of the terminal part of the ileum was sampled. The P digestibility was calculated by linear regression according to World's Poultry Science Association (WPSA) protocol for determination of pre-cecal P digestibility. Pre-cecal P digestibility of MCP, DCP, Delfos, Calfos, and porcine bone meal was 88.5, 82.4, 94.5, 86.9, and 78.2%, respectively. Based on visual inspection of electron microscopy images of test products, the spatial structure of the test products might be related to P digestibility. It is concluded that processing of bone meal increases the pre-cecal P digestibility in broilers.

  14. Thoroughbred blood serum inorganic phosphate concentrations in relation to feeding regime and racing performance.

    PubMed

    Denny, J E

    1987-06-01

    Horses receiving a pelleted or cubed dietary supplementation with roughage, have serum inorganic phosphate (SIP) concentrations consistently below an accepted mean of 1,032 mmol l-1 or 3, 1 mg dl-1. Further, it has been reported that the best eight, two-year-old Irish Thoroughbred track performers of 51 horses tested over a 10 month period, had significantly lower SIP concentrations than the worst eight track performers. In an endeavour to assess any nutritive effect on SIP concentrations and also to assess any effect of SIP concentrations on track performance, metabolic blood profiles from 303 horses in training at the Summerveld Training centre in Natal, were evaluated for various blood parameters over a two year period. Of these 303 profiles, 264 were analysed for SIP concentrations. These horses were on three known feeding regimes viz. Feed 1--cube feeding plus hay; Feed 2--oats, wheaten bran and greens plus hay; Feed 3--Mixed feeding regime of feeds 1 and 2; Feed 4--unknown regime. Dry matter intake varied between 2 and 2.5% of estimated bodymass and in the Feed 1 regime, the proportion of cubed supplement in the diet was increased from 30 to 70% as the training programme progressed. Statistical analysis of SIP concentrations showed that horses on the Feed 1 regime had significantly lower SIP concentrations than horses on the other feed regimes. Of the 303 profiles, 224 could be identified with actual races.(ABSTRACT TRUNCATED AT 250 WORDS)

  15. Plasma Calcium, Inorganic Phosphate and Magnesium During Hypocalcaemia Induced by a Standardized EDTA Infusion in Cows

    PubMed Central

    Mellau, LSB; Jørgensen, RJ; Enemark, JMD

    2001-01-01

    The intravenous Na2EDTA infusion technique allows effective specific chelation of circulating Ca2+ leading to a progressive hypocalcaemia. Methods previously used were not described in detail and results obtained by monitoring total and free ionic calcium were not comparable due to differences in sampling and analysis. This paper describes a standardized EDTA infusion technique that allowed comparison of the response of calcium, phosphorus and magnesium between 2 groups of experimental cows. The concentration of the Na2EDTA solution was 0.134 mol/l and the flow rate was standardized at 1.2 ml/kg per hour. Involuntary recumbency occurred when ionised calcium dropped to 0.39 – 0.52 mmol/l due to chelation. An initial fast drop of ionized calcium was observed during the first 20 min of infusion followed by a fluctuation leading to a further drop until recumbency. Pre-infusion [Ca2+] between tests does not correlate with the amount of EDTA required to induce involuntary recumbence. Total calcium concentration measured by atomic absorption remained almost constant during the first 100 min of infusion but declined gradually when the infusion was prolonged. The concentration of inorganic phosphate declined gradually in a fluctuating manner until recumbency. Magnesium concentration remained constant during infusion. Such electrolyte responses during infusion were comparable to those in spontaneous milk fever. The standardized infusion technique might be useful in future experimental studies. PMID:11503370

  16. Two different mechanisms mediate chemotaxis to inorganic phosphate in Pseudomonas aeruginosa

    PubMed Central

    Rico-Jiménez, Miriam; Reyes-Darias, Jose Antonio; Ortega, Álvaro; Díez Peña, Ana Isabel; Morel, Bertrand; Krell, Tino

    2016-01-01

    Inorganic phosphate (Pi) is a central signaling molecule that modulates virulence in various pathogens. In Pseudomonas aeruginosa, low Pi concentrations induce transcriptional alterations that increase virulence. Also, under low Pi levels, P. aeruginosa exhibits Pi chemotaxis—a process mediated by the two non-paralogous receptors CtpH and CtpL. Here we show that the two receptors operate via different mechanisms. We demonstrate that the ligand binding domain (LBD) of CtpH but not CtpL binds Pi directly. We identify the periplasmic ligand binding protein PstS as the protein that binds in its Pi loaded state to CtpL, resulting in receptor stimulation. PstS forms part of the Pi transporter and has thus a double function in Pi transport and chemotaxis. The affinity of Pi for CtpH was modest whereas that for PstS very high, which may explain why CtpH and CtpL mediate chemotaxis to high and low Pi concentrations, respectively. The pstS/ctpH double mutant was almost devoid of Pi taxis, indicating that PstS is the only CtpL Pi-shuttle. Chemotaxis mechanisms based on indirect ligand recognition were unambiguously identified in enterobacteria. The discovery of a similar mechanism in a different bacterial order, involving a different chemoreceptor type and chemoeffector suggests that such systems are widespread. PMID:27353565

  17. Extracellular inorganic phosphate regulates gibbon ape leukemia virus receptor-2/phosphate transporter mRNA expression in rat bone marrow stromal cells.

    PubMed

    Wada, Keinoshin; Mizuno, Morimichi; Komori, Takahide; Tamura, Masato

    2004-01-01

    In mammalian cells, several observations indicate not only that phosphate transport probably regulates local inorganic phosphate (Pi) concentration, but also that Pi affects normal cellular metabolism, which in turn regulates apoptosis and the process of mineralization. To elucidate how extracellular Pi regulates cellular functions of pre-osteoblastic cells, we investigated the expression of type III sodium (Na)-dependent Pi transporters in rat bone marrow stromal cells and ROB-C26 pre-osteoblastic cells. The mRNA expression level of gibbon ape leukemia virus receptor (Glvr)-2 was increased by the addition of Pi in rat bone marrow stromal cells, but not in ROB-C26 or normal rat kidney (NRK) cells. In contrast, the level of Glvr-1 mRNA was not altered by the addition of extracellular Pi in these cells. The induction of Glvr-2 mRNA by Pi was inhibited in the presence of cycloheximide (CHX). Moreover, mitogen-activated protein kinase (MEK) /extracellular-signal-regulated kinase (ERK) pathway inhibitors; U0126 (1.4-diamino-2, 3-dicyano-1, 4-bis [2-amino-phenylthio] butadiene) and PD98059 (2'-Amino-3'-methoxyflavone) inhibited inducible Glvr-2 mRNA expression, but p38 MEK inhibitor SB203580 [4-(4'-fluorophenyl)-2-(4'-methyl-sulfinylphenyl)-5-(4'pyridyl) imidazole] did not inhibit the induction of Glvr-2 mRNA expression, suggesting that extracellular Pi regulates de novo protein synthesis and MEK/ERK activity in rat bone marrow stromal cells, and through these, induction of Glvr-2 mRNA. Although Pi also induced osteopontin mRNA expression in rat bone marrow stromal cells but not in ROB-C26 and NRK cells, changes in cell viability with the addition of Pi were similar in both cell types. These data indicate that extracellular Pi regulates Glvr-2 mRNA expression, provide insights into possible mechanisms whereby Pi may regulate protein phosphorylation, and suggest a potential role for the Pi transporter in rat bone marrow stromal cells.

  18. Can inorganic phosphate explain sag during unfused tetanic contractions of skeletal muscle?

    PubMed

    Smith, Ian C; Bellissimo, Catherine; Herzog, Walter; Tupling, A Russell

    2016-11-01

    We test the hypothesis that cytosolic inorganic phosphate (Pi) can account for the contraction-induced reductions in twitch duration which impair summation and cause force to decline (sag) during unfused tetanic contractions of fast-twitch muscle. A five-state model of crossbridge cycling was used to simulate twitch and unfused tetanic contractions. As Pi concentration ([Pi]) was increased from 0 to 30 mmol·L(-1), twitch duration decreased, with progressive reductions in sensitivity to Pi as [Pi] was increased. When unfused tetani were simulated with rising [Pi], sag was most pronounced when initial [Pi] was low, and when the magnitude of [Pi] increase was large. Fast-twitch extensor digitorum longus (EDL) muscles (sag-prone, typically low basal [Pi]) and slow-twitch soleus muscles (sag-resistant, typically high basal [Pi]) were isolated from 14 female C57BL/6 mice. Muscles were sequentially incubated in solutions containing either glucose or pyruvate to create typical and low Pi environments, respectively. Twitch duration was greater (P < 0.05) in pyruvate than glucose in both muscles. Stimuli applied at intervals approximately three times the time to peak twitch tension resulted in sag of 35.0 ± 3.7% in glucose and 50.5 ± 1.4% in pyruvate in the EDL (pyruvate > glucose; P < 0.05), and 3.9 ± 0.3% in glucose and 37.8 ± 2.7% in pyruvate in the soleus (pyruvate > glucose; P < 0.05). The influence of Pi on crossbridge cycling provides a tenable mechanism for sag. Moreover, the low basal [Pi] in fast-twitch relative to slow-twitch muscle has promise as an explanation for the fiber-type dependency of sag.

  19. Seminal Plasma pH, Inorganic Phosphate, Total and Ionized Calcium Concentrations In The Assessment of Human Spermatozoa Function

    PubMed Central

    Banjoko, S. Olatunbosun; Adeseolu, Fasiu O.

    2013-01-01

    Introduction: Fertilization in humans is dependent on viability of the male spermatozoa among other factors and there have been conflicting reports on the role of pH, calcium and phosphate concentrations in sperm function. This study therefore aimed to investigate seminal plasma pH, inorganic phosphate, total and ionized calcium concentrations relative to spermatozoa function. Material and Methods: Seminal plasma concentrations of pH, total calcium, ionized calcium (Ca++); inorganic phosphate, motility and spermatozoa count were determined in 80 males by standard methods. Results: Forty-nine of the subjects had normal spermatozoa motility (> 60%) and 31 had hypomotility (< 60%). The hypomotility group exhibited lower calcium ion (Ca2+) concentrations; 0.19+0.01mmol/L compared with normal motility group; 0.24+0.01mmol/L (p<0.001) the latter also had significantly higher inorganic phosphate; 7.83+1.27 while the former had 5.64+1.62mmol/L (p= 0.004). The mean spermatozoa counts for hypomotility and normal motility group were 42.0 ± 13 x 106 , 72.35 + 20 x 106 respectively (p< 0.001). No significant differences were observed in pH, volume of ejaculate and total calcium concentration between the hypomotility and normal motility groups The mean concentrations of pH were 7.51 ± 0.02 and 7.54 ± 0.03 respectively (p= 0.21) and total calcium; 3.10 ± 0.12 and 3.36 ± 0.14mmol/L respectively (p= 0.16 ). There was a significant difference in percentage of abnormal forms in both groups with hypomotile group having 36% compared to mormal motility group with 5% (p< 0.05). Conclusion: Correlations were observed between seminal concentrations of calcium ions, inorganic phosphate, spermatozoa count and motility but not with total calcium concentrations and pH and therefore should be considered in understanding male infertility and preparation of media for sperm preservation for in vitro fertilization. PMID:24392378

  20. Effect of various vitamin D metabolites on serum calcium and inorganic phosphate in the freshwater snake Natrix piscator.

    PubMed

    Srivastav, A K; Srivastav, S K; Singh, S; Norman, A W

    1995-10-01

    Vitamin D3 (650 pmol and 6.50 nmol/100 g body wt), 25-hydroxyvitamin D (650 pmol and 6.50 nmol/100 g body wt), and 1,25-dihydroxyvitamin D (65 pmol and 650 pmol/100 g body wt) were administered daily to the freshwater snake Natrix piscator for 15 days. Both serum calcium and inorganic phosphate levels were increased significantly in all of the treated groups. This is the first report of hypercalcemia and hyperphosphatemia in reptiles induced by 25-hydroxyvitamin D and 1,25-dihydroxyvitamin D.

  1. Activation by Gene Amplification of pitB, Encoding a Third Phosphate Transporter of Escherichia coli K-12

    PubMed Central

    Hoffer, Sally M.; Schoondermark, Paul; van Veen, Hendrik W.; Tommassen, Jan

    2001-01-01

    Two systems for the uptake of inorganic phosphate (Pi) in Escherichia coli, PitA and Pst, have been described. A revertant of a pitA pstS double mutant that could grow on Pi was isolated. We demonstrate that the expression of a new Pi transporter, PitB, is activated in this strain by a gene amplification event. PMID:11443103

  2. High Inorganic Triphosphatase Activities in Bacteria and Mammalian Cells: Identification of the Enzymes Involved

    PubMed Central

    Lakaye, Bernard; Servais, Anne-Catherine; Scholer, Georges; Fillet, Marianne; Elias, Benjamin; Derochette, Jean-Michel; Crommen, Jacques; Wins, Pierre; Bettendorff, Lucien

    2012-01-01

    Background We recently characterized a specific inorganic triphosphatase (PPPase) from Nitrosomonas europaea. This enzyme belongs to the CYTH superfamily of proteins. Many bacterial members of this family are annotated as predicted adenylate cyclases, because one of the founding members is CyaB adenylate cyclase from A. hydrophila. The aim of the present study is to determine whether other members of the CYTH protein family also have a PPPase activity, if there are PPPase activities in animal tissues and what enzymes are responsible for these activities. Methodology/Principal Findings Recombinant enzymes were expressed and purified as GST- or His-tagged fusion proteins and the enzyme activities were determined by measuring the release of inorganic phosphate. We show that the hitherto uncharacterized E. coli CYTH protein ygiF is a specific PPPase, but it contributes only marginally to the total PPPase activity in this organism, where the main enzyme responsible for hydrolysis of inorganic triphosphate (PPPi) is inorganic pyrophosphatase. We further show that CyaB hydrolyzes PPPi but this activity is low compared to its adenylate cyclase activity. Finally we demonstrate a high PPPase activity in mammalian and quail tissue, particularly in the brain. We show that this activity is mainly due to Prune, an exopolyphosphatase overexpressed in metastatic tumors where it promotes cell motility. Conclusions and General Significance We show for the first time that PPPase activities are widespread in bacteria and animals. We identified the enzymes responsible for these activities but we were unable to detect significant amounts of PPPi in E. coli or brain extracts using ion chromatography and capillary electrophoresis. The role of these enzymes may be to hydrolyze PPPi, which could be cytotoxic because of its high affinity for Ca2+, thereby interfering with Ca2+ signaling. PMID:22984449

  3. Substrate activity of synthetic formyl phosphate in the reaction catalyzed by formyltetrahydrofolate synthetase

    SciTech Connect

    Smithers, G.W.; Jahansouz, H.; Kofron, J.L.; Himes, R.H.; Reed, G.H.

    1987-06-30

    Formyl phosphate, a putative enzyme-bound intermediate in the reaction catalyzed by formyltetrahydrofolate synthetase (EC 6.3.4.3), was synthesized from formyl fluoride and inorganic phosphate, and the product was characterized by /sup 31/P, /sup 1/H, and /sup 13/C nuclear magnetic resonance (NMR). Measurement of hydrolysis rates by /sup 31/P NMR indicates that formyl phosphate is particularly labile, with a half-life of 48 min in a buffered neutral solution at 20 /sup 0/C. At pH 7, hydrolysis occurs with P-O bond cleavage, as demonstrated by /sup 18/O incorporation from H/sub 2//sup 18/O into P/sub i/, while at pH 1 and pH 13 hydrolysis occurs with C-O bond cleavage. The substrate activity of formyl phosphate was tested in the reaction catalyzed by formyltetrahydrofolate synthetase isolated from Clostridium cylindrosporum. Formyl phosphate supports the reaction in both the forward and reverse directions. Thus, N/sup 10/-formyltetrahydrofolate is produced from tetrahydrofolate and formyl phosphate in a reaction mixture that contains enzyme, Mg(II), and ADP, and ATP is produced from formyl phosphate and ADP with enzyme, Mg(II), and tetrahydrofolate present. The requirements for ADP and for tetrahydrofolate as cofactors in these reactions are consistent with previous steady-state kinetic and isotope exchange studies, which demonstrated that all substrate subsites must be occupied prior to catalysis. The k/sub cat/ values for both the forward and reverse directions, with formyl phosphate as the substrate, are much lower than those for the normal forward and reverse reactions. Kinetic analysis of the formyl phosphate supported reactions indicates that the low steady-state rates observed for the synthetic intermediate are most likely due to the sequential nature of the normal reaction.

  4. Influence of pH and inorganic phosphate on toxicity of zinc to Arthrobacter sp. isolated from heavy-metal-contaminated sediments.

    PubMed

    Moberly, James G; Staven, Ari; Sani, Rajesh K; Peyton, Brent M

    2010-10-01

    Because of its high solubility over a wide range of pH conditions, zinc is found in many natural and human-impacted systems. Zinc speciation is critical in assessing zinc toxicity to microorganisms because it varies considerably with pH and is dependent on other aqueous constituents. Combined results of thermodynamic modeling, statistical analysis, and batch culture studies using Arthrobacter sp. JM018 suggest that the toxic species may not be solely limited to the free ion, but also includes ZnHPO(4)(0)(aq). Cellular uptake of ZnHPO(4)(0)(aq) through the inorganic phosphate transporter (Pit family), which requires a neutral metal phosphate complex for phosphate transport, may explain the observed toxicity. Based on visual MINTEQ (v3.0) modeling, at 50 μM total zinc, ZnHPO(4)(0)(aq) constitutes 33, 70, and 76% of the neutral metal phosphate pool at pH 6, 7, and 8, respectively. At 50 μM total zinc, cultures supplied with organic phosphate (glycerol-3-phosphate) show no significant response to pH (p = 0.13) while inhibition of inorganic phosphate-supplemented cultures, whose neutral metal phosphates are increasingly dominated by ZnHPO(4)(0)(aq), show significant pH dependence (p = 9.45 × 10(-7)). Using sodium to decrease the distribution of ZnHPO(4)(0)(aq) in the neutral metal phosphate pool also decreased the pH dependent toxicity, further supporting this mechanism. These findings show the important role of minor zinc species in organism toxicity and have wider implications because the Pit inorganic phosphate transport system is widely distributed in Bacteria, Archaea, and Eukarya.

  5. 2,3-diphosphoglycerate phosphatase activity of phosphoglycerate mutase: stimulation by vanadate and phosphate

    SciTech Connect

    Stankiewicz, P.J.; Gresser, M.J.; Tracey, A.S.; Hass, L.F.

    1987-03-10

    The binding of inorganic vanadate (V/sub i/) to rabbit muscle phosphoglycerate mutase (PGM), studied by using /sup 51/V nuclear magnetic resonance spectroscopy, shows a sigmoidal dependence on vanadate concentration with a stoichiometry of four vanadium atoms per PGM molecule at saturating (V/sub i/). The data are consistent with binding of one divanadate ion to each of the two subunits of PGM in a noncooperative manner with an intrinsic dissociation constant of 4 x 10/sup -6/ M. The relevance of this result to other studies which have shown that the V/sub i/-stimulated 2,3-diphosphoglycerate (2,3-DPG) phosphatase activity of PGM has a sigmoidal dependence on (V/sub i/) with a Hill coefficient of 2.0 is discussed. At pH 7.0, inorganic phosphate has little effect on the 2,3-DPG phosphatase activity of PGM, even at concentrations as high as 50 mM. Similarly, 25 ..mu..M V/sub i/ has little effect on the phosphatase activity. However, in the presence of 25 ..mu..M V/sub i/, a phosphate concentration of 20 mM increases the phosphatase activity by more than 3-fold. This behavior is rationalized in terms of activation of the phosphatase activity by a phosphate/vanadate mixed anhydride. This interpretation is supported by the observation of strong activation of the phosphatase activity by inorganic pyrophosphate. A molecular mechanism for the observed effects of vanadate is proposed, and the relevance of this study to the possible use of vanadate as a therapeutic agent for the treatment of sickle cell anemia is discussed.

  6. Phosphate addition enhanced soil inorganic nutrients to a large extent in three tropical forests.

    PubMed

    Zhu, Feifei; Lu, Xiankai; Liu, Lei; Mo, Jiangming

    2015-01-21

    Elevated nitrogen (N) deposition may constrain soil phosphorus (P) and base cation availability in tropical forests, for which limited evidence have yet been available. In this study, we reported responses of soil inorganic nutrients to full factorial N and P treatments in three tropical forests different in initial soil N status (N-saturated old-growth forest and two less-N-rich younger forests). Responses of microbial biomass, annual litterfall production and nutrient input were also monitored. Results showed that N treatments decreased soil inorganic nutrients (except N) in all three forests, but the underlying mechanisms varied depending on forests: through inhibition on litter decomposition in the old-growth forest and through Al(3+) replacement of Ca(2+) in the two younger forests. In contrast, besides great elevation in soil available P, P treatments induced 60%, 50%, 26% increases in sum of exchangeable (K(+)+Ca(2+)+Mg(2+)) in the old-growth and the two younger forests, respectively. These positive effects of P were closely related to P-stimulated microbial biomass and litter nutrient input, implying possible stimulation of nutrient return. Our results suggest that N deposition may result in decreases in soil inorganic nutrients (except N) and that P addition can enhance soil inorganic nutrients to support ecosystem processes in these tropical forests.

  7. Structure of human farnesyl pyrophosphate synthase in complex with an aminopyridine bisphosphonate and two molecules of inorganic phosphate

    SciTech Connect

    Park, Jaeok; Lin, Yih-Shyan; Tsantrizos, Youla S.; Berghuis, Albert M.

    2014-02-19

    A co-crystal structure of human farnesyl pyrophosphate synthase in complex with an aminopyridine bisphosphonate, YS0470, and two molecules of inorganic phosphate has been determined. The identity of the phosphate ligands was confirmed by anomalous diffraction data. Human farnesyl pyrophosphate synthase (hFPPS) produces farnesyl pyrophos@@phate, an isoprenoid essential for a variety of cellular processes. The enzyme has been well established as the molecular target of the nitrogen-containing bisphosphonates (N-BPs), which are best known for their antiresorptive effects in bone but are also known for their anticancer properties. Crystal structures of hFPPS in ternary complexes with a novel bisphosphonate, YS0470, and the secondary ligands inorganic phosphate (P{sub i}), inorganic pyrophosphate (PP{sub i}) and isopentenyl pyrophosphate (IPP) have recently been reported. Only the co-binding of the bisphosphonate with either PP{sub i} or IPP resulted in the full closure of the C-@@terminal tail of the enzyme, a conformational change that is required for catalysis and that is also responsible for the potent in vivo efficacy of N-BPs. In the present communication, a co-crystal structure of hFPPS in complex with YS0470 and two molecules of P{sub i} is reported. The unusually close proximity between these ligands, which was confirmed by anomalous diffraction data, suggests that they interact with one another, with their anionic charges neutralized in their bound state. The structure also showed the tail of the enzyme to be fully disordered, indicating that simultaneous binding of two P{sub i} molecules with a bisphosphonate cannot induce the tail-closing conformational change in hFPPS. Examination of homologous FPPSs suggested that this ligand-dependent tail closure is only conserved in the mammalian proteins. The prevalence of P{sub i}-bound hFPPS structures in the PDB raises a question regarding the in vivo relevance of P{sub i} binding to the function of the enzyme.

  8. Optimization and validation of a rapid method to determine citrate and inorganic phosphate in milk by capillary electrophoresis.

    PubMed

    Izco, J M; Tormo, M; Harris, A; Tong, P S; Jimenez-Flores, R

    2003-01-01

    Quantification of phosphate and citrate compounds is very important because their distribution between soluble and colloidal phases of milk and their interactions with milk proteins influence the stability and some functional properties of dairy products. The aim of this work was to optimize and validate a capillary electrophoresis method for the rapid determination of these compounds in milk. Various parameters affecting analysis have been optimized, including type, composition, and pH of the electrolyte, and sample extraction. Ethanol, acetonitrile, sulfuric acid, water at 50 degrees C or at room temperature were tested as sample buffers (SB). Water at room temperature yielded the best overall results and was chosen for further validation. The extraction time was checked and could be shortened to less than 1 min. Also, sample preparation was simplified to pipet 12 microl of milk into 1 ml of water containing 20 ppm of tartaric acid as an internal standard. The linearity of the method was excellent (R2 > 0.999) with CV values of response factors <3%. The detection limits for phosphate and citrate were 5.1 and 2.4 nM, respectively. The accuracy of the method was calculated for each compound (103.2 and 100.3%). In addition, citrate and phosphate content of several commercial milk samples were analyzed by this method, and the results deviated less than 5% from values obtained when analyzing the samples by official methods. To study the versatility of the technique, other dairy productssuch as cream cheese, yogurt, or Cheddar cheese were analyzed and accuracy was similar to milk in all products tested. The procedure is rapid and offers a very fast and simple sample preparation. Once the sample has arrived at the laboratory, less than 5 min (including handling, preparation, running, integration, and quantification) are necessary to determine the concentration of citric acid and inorganic phosphate. Because of the speed and accuracy of this method, it is promising as an

  9. Crystallization and preliminary X-ray diffraction analysis of a high-affinity phosphate-binding protein endowed with phosphatase activity from Pseudomonas aeruginosa PAO1.

    PubMed

    Djeghader, Ahmed; Gotthard, Guillaume; Suh, Andrew; Gonzalez, Daniel; Scott, Ken; Chabriere, Eric; Elias, Mikael

    2013-10-01

    In prokaryotes, phosphate starvation induces the expression of numerous phosphate-responsive genes, such as the pst operon including the high-affinity phosphate-binding protein (PBP or pstS) and alkaline phosphatases such as PhoA. This response increases the cellular inorganic phosphate import efficiency. Notably, some Pseudomonas species secrete, via a type-2 secretion system, a phosphate-binding protein dubbed LapA endowed with phosphatase activity. Here, the expression, purification, crystallization and X-ray data collection at 0.87 Å resolution of LapA are described. Combined with biochemical and enzymatic characterization, the structure of this intriguing phosphate-binding protein will help to elucidate the molecular origin of its phosphatase activity and to decipher its putative role in phosphate uptake.

  10. The effect of nitrite on aerobic phosphate uptake and denitrifying activity of phosphate-accumulating organisms.

    PubMed

    Yoshida, Y; Takahashi, K; Saito, T; Tanaka, K

    2006-01-01

    An anaerobic/aerobic/anoxic/aerobic sequencing batch reactor (SBR) was operated with municipal wastewater to investigate the effect of nitrite on biological phosphorus removal (BPR). When nitrite accumulated, aerobic phosphate uptake activity significantly decreased and, in case of hard exposure to nitrite, BPR severely deteriorated. The interesting observation was that the relative anoxic activity of phosphate accumulating organisms (PAOs) increased after nitrite exposure. Moreover batch tests of aerobic phosphate uptake in the presence/absence of nitrite indicated that PAOs with the higher relative anoxic activity are less sensitive to nitrite exposure. From these results, we concluded that BPR is sensitive to nitrite exposure, but BPR containing PAOs with the higher relative anoxic activity is possibly more stable against nitrite than BPR containing PAOs with the lower relative anoxic activity.

  11. Long-term nitrogen fertilization decreased the abundance of inorganic phosphate solubilizing bacteria in an alkaline soil

    NASA Astrophysics Data System (ADS)

    Zheng, Bang-Xiao; Hao, Xiu-Li; Ding, Kai; Zhou, Guo-Wei; Chen, Qing-Lin; Zhang, Jia-Bao; Zhu, Yong-Guan

    2017-02-01

    Inorganic phosphate solubilizing bacteria (iPSB) are essential to facilitate phosphorus (P) mobilization in alkaline soil, however, the phylogenetic structure of iPSB communities remains poorly characterized. Thus, we use a reference iPSB database to analyze the distribution of iPSB communities based on 16S rRNA gene illumina sequencing. Additionally, a noval pqqC primer was developed to quantify iPSB abundance. In our study, an alkaline soil with 27-year fertilization treatment was selected. The percentage of iPSB was 1.10~2.87% per sample, and the dominant iPSB genera were closely related to Arthrobacter, Bacillus, Brevibacterium and Streptomyces. Long-term P fertilization had no significant effect on the abundance of iPSB communities. Rather than P and potassium (K) additions, long-term nitrogen (N) fertilization decreased the iPSB abundance, which was validated by reduced relative abundance of pqqC gene (pqqC/16S). The decreased iPSB abundance was strongly related to pH decline and total N increase, revealing that the long-term N additions may cause pH decline and subsequent P releases relatively decreasing the demands of the iPSB community. The methodology and understanding obtained here provides insights into the ecology of inorganic P solubilizers and how to manipulate for better P use efficiency.

  12. Long-term nitrogen fertilization decreased the abundance of inorganic phosphate solubilizing bacteria in an alkaline soil

    PubMed Central

    Zheng, Bang-Xiao; Hao, Xiu-Li; Ding, Kai; Zhou, Guo-Wei; Chen, Qing-Lin; Zhang, Jia-Bao; Zhu, Yong-Guan

    2017-01-01

    Inorganic phosphate solubilizing bacteria (iPSB) are essential to facilitate phosphorus (P) mobilization in alkaline soil, however, the phylogenetic structure of iPSB communities remains poorly characterized. Thus, we use a reference iPSB database to analyze the distribution of iPSB communities based on 16S rRNA gene illumina sequencing. Additionally, a noval pqqC primer was developed to quantify iPSB abundance. In our study, an alkaline soil with 27-year fertilization treatment was selected. The percentage of iPSB was 1.10~2.87% per sample, and the dominant iPSB genera were closely related to Arthrobacter, Bacillus, Brevibacterium and Streptomyces. Long-term P fertilization had no significant effect on the abundance of iPSB communities. Rather than P and potassium (K) additions, long-term nitrogen (N) fertilization decreased the iPSB abundance, which was validated by reduced relative abundance of pqqC gene (pqqC/16S). The decreased iPSB abundance was strongly related to pH decline and total N increase, revealing that the long-term N additions may cause pH decline and subsequent P releases relatively decreasing the demands of the iPSB community. The methodology and understanding obtained here provides insights into the ecology of inorganic P solubilizers and how to manipulate for better P use efficiency. PMID:28181569

  13. Calcium phosphate-based organic-inorganic hybrid nanocarriers with pH-responsive on/off switch for photodynamic therapy.

    PubMed

    Nomoto, Takahiro; Fukushima, Shigeto; Kumagai, Michiaki; Miyazaki, Kozo; Inoue, Aki; Mi, Peng; Maeda, Yoshinori; Toh, Kazuko; Matsumoto, Yu; Morimoto, Yuji; Kishimura, Akihiro; Nishiyama, Nobuhiro; Kataoka, Kazunori

    2016-05-26

    Photodynamic therapy (PDT) is a promising treatment modality for malignant tumors in a light-selective manner. To improve the PDT efficacy, numerous kinds of nanocarriers have been developed to deliver photosensitizers (PSs) selectively into the tumor through leaky tumor-associated vasculature. However, the corresponding prolonged retention of the nanocarrier in the bloodstream may lead to unfavorable photochemical damage to normal tissues such as skin. Here, we report an organic-inorganic hybrid nanocarrier with a pH-responsive on/off switch of PDT efficacy. This hybrid nanocarrier is constructed by hydrothermal synthesis after simple mixing of calcium/phosphate ions, chlorin e6 (amphiphilic low molecular weight PS), and poly(ethylene glycol)-b-poly(aspartic acid) (PEG-PAsp) copolymers in an aqueous solution. The hybrid nanocarrier possesses a calcium phosphate (CaP) core encapsulating the PSs, which is surrounded by a PEG shielding layer. Under physiological conditions (pH 7.4), the nanocarrier suppressed the photochemical activity of PS by lowering the access of oxygen molecules to the incorporated PS, while PDT efficacy was restored in a pH-responsive manner because of the dissolution of CaP and eventual recovery of access between the oxygen and the PS. Owing to this switch, the nanocarrier reduced the photochemical damage in the bloodstream, while it induced effective PDT efficacy inside the tumor cell in response to the acidic conditions of the endo-/lysosomes.

  14. An experimental-computer modeling study of inorganic phosphates surface adsorption on hydroxyapatite particles.

    PubMed

    Rivas, Manuel; Casanovas, Jordi; del Valle, Luis J; Bertran, Oscar; Revilla-López, Guillermo; Turon, Pau; Puiggalí, Jordi; Alemán, Carlos

    2015-06-07

    The adsorption of orthophosphate, pyrophosphate, triphosphate and a trisphosphonate onto hydroxyapatite has been examined using experiments and quantum mechanical calculations. Adsorption studies with FTIR and X-ray photoelectron spectroscopies have been performed considering both crystalline hydroxyapatite (HAp) and amorphous calcium phosphate particles, which were specifically prepared and characterized for this purpose. Density functional theory (DFT) calculations have been carried out considering the (100) and (001) surfaces of HAp, which were represented using 1 × 2 × 2 and 3 × 3 × 1 slab models, respectively. The adsorption of phosphate onto the two crystallographic surfaces is very much favored from an energetic point of view, which is fully consistent with current interpretations of the HAp growing process. The structures calculated for the adsorption of pyrophosphate and triphosphate evidence that this process is easier for the latter than for the former. Thus, the adsorption of pyrophosphate is severely limited by the surface geometry while the flexibility of triphosphate allows transforming repulsive electrostatic interactions into molecular strain. On the other hand, calculations predict that the trisphosphonate only adsorbs onto the (001) surface of HAp. Theoretical predictions are fully consistent with experimental data. Thus, comparison of DFT results and spectroscopic data suggests that the experimental conditions used to prepare HAp particles promote the predominance of the (100) surface. Accordingly, experimental identification of the adsorption of trisphosphonate onto such crystalline particles is unclear while the adsorption of pyrophosphate and triphosphate is clearly observed.

  15. Hydrophobic and Metallophobic Surfaces: Highly Stable Non-wetting Inorganic Surfaces Based on Lanthanum Phosphate Nanorods

    PubMed Central

    Sankar, Sasidharan; Nair, Balagopal N.; Suzuki, Takehiro; Anilkumar, Gopinathan M.; Padmanabhan, Moothetty; Hareesh, Unnikrishnan Nair S.; Warrier, Krishna G.

    2016-01-01

    Metal oxides, in general, are known to exhibit significant wettability towards water molecules because of the high feasibility of synergetic hydrogen-bonding interactions possible at the solid-water interface. Here we show that the nano sized phosphates of rare earth materials (Rare Earth Phosphates, REPs), LaPO4 in particular, exhibit without any chemical modification, unique combination of intrinsic properties including remarkable hydrophobicity that could be retained even after exposure to extreme temperatures and harsh hydrothermal conditions. Transparent nanocoatings of LaPO4 as well as mixture of other REPs on glass surfaces are shown to display notable hydrophobicity with water contact angle (WCA) value of 120° while sintered and polished monoliths manifested WCA greater than 105°. Significantly, these materials in the form of coatings and monoliths also exhibit complete non-wettability and inertness towards molten metals like Ag, Zn, and Al well above their melting points. These properties, coupled with their excellent chemical and thermal stability, ease of processing, machinability and their versatile photo-physical and emission properties, render LaPO4 and other REP ceramics utility in diverse applications. PMID:26955962

  16. Hydrophobic and Metallophobic Surfaces: Highly Stable Non-wetting Inorganic Surfaces Based on Lanthanum Phosphate Nanorods

    NASA Astrophysics Data System (ADS)

    Sankar, Sasidharan; Nair, Balagopal N.; Suzuki, Takehiro; Anilkumar, Gopinathan M.; Padmanabhan, Moothetty; Hareesh, Unnikrishnan Nair S.; Warrier, Krishna G.

    2016-03-01

    Metal oxides, in general, are known to exhibit significant wettability towards water molecules because of the high feasibility of synergetic hydrogen-bonding interactions possible at the solid-water interface. Here we show that the nano sized phosphates of rare earth materials (Rare Earth Phosphates, REPs), LaPO4 in particular, exhibit without any chemical modification, unique combination of intrinsic properties including remarkable hydrophobicity that could be retained even after exposure to extreme temperatures and harsh hydrothermal conditions. Transparent nanocoatings of LaPO4 as well as mixture of other REPs on glass surfaces are shown to display notable hydrophobicity with water contact angle (WCA) value of 120° while sintered and polished monoliths manifested WCA greater than 105°. Significantly, these materials in the form of coatings and monoliths also exhibit complete non-wettability and inertness towards molten metals like Ag, Zn, and Al well above their melting points. These properties, coupled with their excellent chemical and thermal stability, ease of processing, machinability and their versatile photo-physical and emission properties, render LaPO4 and other REP ceramics utility in diverse applications.

  17. Effect of inorganic salts, soaps and detergents on dissolution and larvicidal activity of alginate formulation of Bacillus sphaericus.

    PubMed

    Vijayan, V; Balaraman, K

    1995-03-01

    Various inorganic salts and commonly used soaps and detergents were tested in the laboratory for their effect on the dissolution and larvicidal residual activity of a slow-release alginate encapsulated granular formation of Bacillus sphaericus. Fluoride, chloride and sulphate salts and a detergent powder affected the residual activity of this formulation drastically by rupturing it but did not effect its larvicidal activity. Nitrates and phosphates of sodium and potassium also had the same effect but to a moderate level. The safest concentration of these water impurities for effective functioning of the alginate encapsulated B. sphaericus formulation have been determined.

  18. Phosphate oxygen isotope ratio proxy for specific microbial activity in marine sediments (Peru Margin)

    NASA Astrophysics Data System (ADS)

    Liang, Y.; Blake, R. E.

    2005-12-01

    Oxygen (O) isotope ratios of biogenic apatites have been widely used as paleotemperature and environmental geochemical proxies. With improved knowledge of the phosphate O isotope effects of different P cycling pathways, the δ18O value of inorganic phosphate (δ18OP) has been proposed as a useful proxy and tracer of biological reactions and P cycling in natural environments[1,2,3,4]. Being the only way of removing P from oceanic water, sedimentary P burial is one of the most important processes during biogeochemical cycling of P. The high concentrations of organic matter and pronounced microbial activity at ODP Site 1230 along the Peru Margin result in unusually high interstitial water phosphate concentrations, which provides a unique opportunity to use δ18OP to investigate inorganic phosphate (Pi) regeneration and P cycling pathways in marine sediments. The isotopic measurements of both dissolved inorganic phosphate (DIP) and bulk sediment Pi show that DIP δ18OP values are affected by three different processes, which are all induced by specific microbial activities present in the sediments. In sediments at ~ 65 to 120 mbsf, porewater DIP is derived from dissolved organophosphorus compounds (DOP) through enzymatic degradation pathways, evidenced by both DIP δ18OP values and interstitial water chemistry. Measured porewater DIP δ18OP values also suggest that 4 to 8% of interstitial water DIP reflects regeneration of Pi from Porg by microbially-synthesized enzymes. Throughout the sediment column and especially at ~ 120 to 150 mbsf, DIP is released from the sediments by microbially-induced reductive dissolution of Fe-oxides, which contributes to the overall high DIP concentrations at Site 1230. The third and dominant process controlling measured DIP δ18OP values is microbial turnover of regenerated Pi. The presence of high microbial activities in organic-rich Site 1230 sediments promotes the remobilization of P and affects marine P cycling by potentially enhancing

  19. Stable organic-inorganic hybrid of polyaniline/α-zirconium phosphate for efficient removal of organic pollutants in water environment.

    PubMed

    Wang, Lei; Wu, Xi-Lin; Xu, Wei-Hong; Huang, Xing-Jiu; Liu, Jin-Huai; Xu, An-Wu

    2012-05-01

    In this article, organic-inorganic hybrid materials of polyaniline/α-zirconium phosphate (PANI/α-ZrP) was synthesized by in situ oxidative polymerization reaction and characterized by Fourier transformed infrared (FTIR), field-emission scanning electron microscopic (FE-SEM) and X-ray diffraction (XRD). The results showed that polyaniline (PANI) was successfully grown on the surface of α-zirconium phosphate (α-ZrP) nanoplates. The PANI/α-ZrP nanocomposites were further applied to remove methyl orange (MO), which was used as a model of organic pollutants in aqueous solution. A synergistic effect of PANI and α-ZrP on promoting the adsorption removal of MO was observed. The PANI/α-ZrP nanocomposites exhibited excellent maximum adsorption capacity toward MO (377.46 mg g(-1)), which is superior to that of PANI nanotubes (254.15 mg g(-1)) and much higher than that of many other adsorbents. The adsorption isotherms of MO can be well-fitted with the Langmuir model and the adsorption kinetics follows the pseudo-second-order model. MO adsorption decreased with increasing solution pH at pH > 4.0 implying that MO adsorption on PANI/α-ZrP may via electrostatic interactions between amine and imine groups on the surface of PANI/α-ZrP and MO molecules. This study implies that the hybrid materials of PANI/α-ZrP can be suggested as potential adsorbents to remove organic dyes from large volumes of aqueous solutions.

  20. Dipolar Coupling Information in Multispin Systems: Application of a Compensated REDOR NMR Approach to Inorganic Phosphates

    NASA Astrophysics Data System (ADS)

    Chan, Jerry C. C.; Eckert, Hellmut

    2000-12-01

    Anexperimental strategy has been developed for measuring multiple dipole-dipole interactions in inorganic compounds using the technique of rotational echo double resonance (REDOR) NMR. Geometry-independent information about the dipole couplings between the observe nuclear species S (arbitrary quantum number) and the heteronuclear species I (spin-{1}/{2}) can be conveniently obtained from the experimental curve of ΔS/S0 versus dipolar evolution time by limiting the analysis to the initial data range 0 < ΔS/S0 < 0.30. Numerical simulations have been carried out on a three-spin system of type SI2 in order to assess the effect of the I-I homonuclear dipole-dipole coupling and the influence of experimental imperfections such as finite pulse length and misadjustments of the 180° pulses applied to the I-spin species. The simulations show further that within the initial data range the effects of such misadjustments can be internally compensated by a modified sequence having an additional 180° pulse on the I channel in the middle of the dipolar evolution periods. Experimental 27Al{31P} REDOR results on the multispin systems Al(PO3)3, AlPO4, [AlPO4]12(C3H7)4NF, and Na3PO4 confirm the general utility of this approach. Thus, for applications to unknown systems the compensation strategy obviates calibration procedures with model compounds.

  1. Inorganic Corrosion-Inhibitive Pigments for High-Temperature Alkali-activated Well Casing Foam Cement

    SciTech Connect

    Sugama, T.; Pyatina, T.

    2014-11-14

    This study evaluates inorganic pigments for improving carbon steel (CS) brine-corrosion protection by the sodium metasilicate-activated calcium aluminate cement/Fly Ash blend at 300°C. Calcium borosilicate (CBS) and zinc phosphate, significantly improved CS corrosion-protection by decreasing cement’s permeability for corrosive ions and inhibiting anodic corrosion. An amorphous Na2O-Al2O3-SiO2-H2O phase tightly attached to CS surface formed at 300oC in CBS-modified cement pore solution. The corrosion rate of the CS covered with this phase was nearly 4-fold lower than in the case of nonmodified cement pore solution where the major phase formed on the surface of CS was crystalline analcime.

  2. Inorganic Corrosion-Inhibitive Pigments for High-Temperature Alkali-activated Well Casing Foam Cement

    SciTech Connect

    Sugama, T.; Pyatina, T.

    2014-11-01

    This study evaluates inorganic pigments for improving carbon steel (CS) brine-corrosion protection by the sodium metasilicate-activated calcium aluminate cement/Fly Ash blend at 300°C. Calcium borosilicate (CBS) and zinc phosphate, significantly improved CS corrosion-protection by decreasing cement’s permeability for corrosive ions and inhibiting anodic corrosion. An amorphous Na2O-Al2O3-SiO2-H2O phase tightly attached to CS surface formed at 300oC in CBS-modified cement pore solution. The corrosion rate of the CS covered with this phase was nearly 4-fold lower than in the case of nonmodified cement pore solution where the major phase formed on the surface of CS was crystalline analcime.

  3. Hybrid organic-inorganic crystals based on ammonium dihydrogen phosphate and ammonium salicylate

    NASA Astrophysics Data System (ADS)

    Voronov, A. P.; Salo, V. I.; Puzikov, V. M.; Babenko, G. N.; Roshal, A. D.; Tkachenko, V. F.

    2011-11-01

    ADP-NH 4Sal hybrid crystals are grown from aqueous solutions. The influence of the acidity of the mixed solution on the conditions of co-crystallization of the components is studied. The spectral and scintillation characteristics are determined. Co-crystallization of ammonium salicylate (NH 4Sal) and ammonium dihydrogen phosphate (ADP, NH 4H 2PO 4) is shown to be feasible, the structure of the doping addition being defined by the solution рН. In basic and weak acidic media the hybrid crystals ADP:NH 4Sal are formed in which salicylate anions are located in the interplanar space between the {110}-type planes in the lattice of ADP. The luminescence spectra contain an emission band maximum with λ max=360 nm. In acidic solutions there are ADP:HSal crystals in which salicylic acid molecules captured by the growth macrosteps are located in the interplanar space of the prismatic {100} and pyramidal {101} growth sectors. The luminescence band undergoes bathochromic shift to λmax=400 nm. The sensitivity of ADP:NH 4Sal scintillation crystals to fast neutrons depends on the concentration of ammonium salicylate in ADP matrix. The highest neutron sensitivity is characteristic of the co-doped ADP:NH 4Sal/Tl scintillation crystals.

  4. Influence of an organic and an inorganic additive on the crystallization of dicalcium phosphate dihydrate

    NASA Astrophysics Data System (ADS)

    Anee, T. K.; Meenakshi Sundaram, N.; Arivuoli, D.; Ramasamy, P.; Narayana Kalkura, S.

    2005-12-01

    Dicalcium phosphate dihydrate (DCPD) was crystallized by single diffusion method under physiological pH, in the presence of cobalt and malic acid. The morphology, composition and microstructure of the grown crystals were analyzed using EDTA titration, UV-Visible, X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), atomic absorption spectroscopy (AAS) and scanning electron microscopy (SEM). These analyses showed that the grown crystals were Co 2+-doped DCPD. The temperature has influenced the dopant entry into DCPD crystals, but the dopant content and temperature were having not much influence on the crystal morphology. In pathological crystallization, the size and morphology are very important since they decide the mode of treatment to adopt. The morphological changes arise when the growth assay is doped with cobalt and malic acid. The effect of malic acid on the crystallization was highly specific, adsorbing on certain crystal faces during growth and producing different morphologies. At higher concentration, the morphology showed a feature frequently encountered in biomineralization, the orientational ordering in [0 0 1], leading to elongated crystals.

  5. Proteomic Analysis Provides New Insights in Phosphorus Homeostasis Subjected to Pi (Inorganic Phosphate) Starvation in Tomato Plants (Solanum lycopersicum L.)

    PubMed Central

    Muneer, Sowbiya; Jeong, Byoung Ryong

    2015-01-01

    Phosphorus is a major nutrient acquired by plants via high-affinity inorganic phosphate (Pi) transporters. To determine the adaptation and homeostasis strategy to Pi starvation, we compared the proteome analysis of tomato leaves that were treated with and without Pi (as KH2PO4) for 10 days. Among 600 reproducible proteins on 2-DE gels 46 of them were differentially expressed. These proteins were involved in major metabolic pathways, including photosynthesis, transcriptional/translational regulations, carbohydrate/energy metabolism, protein synthesis, defense response, and other secondary metabolism. The results also showed that the reduction in photosynthetic pigments lowered P content under –Pi treatments. Furthermore, high-affinity Pi transporters (lePT1 and lePT2) expressed in higher amounts under –Pi treatments. Also, the accumulation of Pi transporters was observed highly in the epidermis and palisade parenchyma under +Pi treatments compared to –Pi treatments. Our data suggested that tomato plants developed reactive oxygen species (ROS) scavenging mechanisms to cope with low Pi content, including the up-regulation of proteins mostly involved in important metabolic pathways. Moreover, Pi-starved tomato plants increased their internal Pi utilization efficiency by increasing the Pi transporter genes and their rational localization. These results thus provide imperative information about how tomato plants respond to Pi starvation and its homeostasis. PMID:26222137

  6. Inactivation of highly activated spinach leaf sucrose-phosphate synthase by dephosphorylation. [Spinacia oleracea

    SciTech Connect

    Huber, J.L. ); Huber, S.C. North Carolina State Univ., Raleigh ); Hite, D.R.C.; Outlaw, W.H. Jr. )

    1991-01-01

    Spinach (Spinacia oleracea L.) leaf sucrose-phosphate synthase (SPS) can be phosphorylated and inactivated in vitro with ({gamma}-{sup 32}P)ATP. Thus, it was surprising to find that SPS, extracted from leaves fed mannose in the light to highly activate the enzyme, could be inactivated in an ATP-independent manner when desalted crude extracts were preincubated at 25{degrees}C before assay. The spontaneous inactivation involved a loss in activity measured with limiting substrate concentrations in the presence of the inhibitor, Pi, without affecting maximum catalytic activity. The spontaneous inactivation was unaffected by exogenous carrier proteins and protease inhibitors, but was inhibited by inorganic phosphate, fluoride, and molybdate, suggesting that a phosphatase may be involved. Okadaic acid, a potent inhibitor of mammalian type 1 and 2A protein phosphatases, had no effect up to 5 micromolar. Inactivation was stimulated about twofold by exogenous Mg{sup 2+} and was relatively insensitive to Ca{sup 2+} and to pH over the range pH 6.5 to 8.5. Radioactive phosphate incorporated into SPS during labeling of excised leaves with ({sup 32}P)Pi (initially in the dark and then in the light with mannose) was lost with time when desalted crude extracts were incubated at 25 C, and the loss in radiolabel was substantially reduced by fluoride. These results provide direct evidence for action of an endogenous phosphatase(s) using SPS as substrate.

  7. Structural basis for glucose-6-phosphate activation of glycogen synthase

    SciTech Connect

    Baskaran, Sulochanadevi; Roach, Peter J.; DePaoli-Roach, Anna A.; Hurley, Thomas D.

    2010-11-22

    Regulation of the storage of glycogen, one of the major energy reserves, is of utmost metabolic importance. In eukaryotes, this regulation is accomplished through glucose-6-phosphate levels and protein phosphorylation. Glycogen synthase homologs in bacteria and archaea lack regulation, while the eukaryotic enzymes are inhibited by protein kinase mediated phosphorylation and activated by protein phosphatases and glucose-6-phosphate binding. We determined the crystal structures corresponding to the basal activity state and glucose-6-phosphate activated state of yeast glycogen synthase-2. The enzyme is assembled into an unusual tetramer by an insertion unique to the eukaryotic enzymes, and this subunit interface is rearranged by the binding of glucose-6-phosphate, which frees the active site cleft and facilitates catalysis. Using both mutagenesis and intein-mediated phospho-peptide ligation experiments, we demonstrate that the enzyme's response to glucose-6-phosphate is controlled by Arg583 and Arg587, while four additional arginine residues present within the same regulatory helix regulate the response to phosphorylation.

  8. Inactivation of Highly Activated Spinach Leaf Sucrose-Phosphate Synthase by Dephosphorylation 1

    PubMed Central

    Huber, Joan L.; Hite, Daniel R. C.; Outlaw, William H.; Huber, Steven C.

    1991-01-01

    Spinach (Spinacia oleracea L.) leaf sucrose-phosphate synthase (SPS) can be phosphorylated and inactivated in vitro with [γ-32P]ATP (JLA Huber, SC Huber, TH Nielsen [1989] Arch Biochem Biophys 270: 681-690). Thus, it was surprising to find that SPS, extracted from leaves fed mannose in the light to highly activate the enzyme, could be inactivated in an ATP-independent manner when desalted crude extracts were preincubated at 25°C before assay. The “spontaneous” inactivation involved a loss in activity measured with limiting substrate concentrations in the presence of the inhibitor, Pi, without affecting maximum catalytic activity. The spontaneous inactivation was unaffected by exogenous carrier proteins and protease inhibitors, but was inhibited by inorganic phosphate, fluoride, and molybdate, suggesting that a phosphatase may be involved. Okadaic acid, a potent inhibitor of mammalian type 1 and 2A protein phosphatases, had no effect up to 5 micromolar. Inactivation was stimulated about twofold by exogenous Mg2+ and was relatively insensitive to Ca2+ and to pH over the range pH 6.5 to 8.5. Radioactive phosphate incorporated into SPS during labeling of excised leaves with [32P]Pi (initially in the dark and then in the light with mannose) was lost with time when desalted crude extracts were incubated at 25°C, and the loss in radiolabel was substantially reduced by fluoride. These results provide direct evidence for action of an endogenous phosphatase(s) using SPS as substrate. We postulate that highly activated SPS contains phosphorylated residue(s) that increase activation state, and that spontaneous inactivation occurs by removal of these phosphate group(s). Inactivation of SPS in vivo caused by feeding uncouplers to darkened leaf tissue that had previously been fed mannose in the dark, may occur by this mechanism. However, there is no evidence that this mechanism is involved in light-dark regulation of SPS in vivo. PMID:16667968

  9. Effects of inorganic phosphate analogues on stiffness and unloaded shortening of skinned muscle fibres from rabbit.

    PubMed Central

    Chase, P B; Martyn, D A; Kushmerick, M J; Gordon, A M

    1993-01-01

    1. We examined the effects of aluminofluoride (AlFx) and orthovanadate (Vi), tightly binding analogues of orthophosphate (Pi), on the mechanical properties of glycerinated fibres from rabbit psoas muscle. Maximum Ca(2+)-activated force, stiffness, and unloaded shortening velocity (Vus) were measured under conditions of steady-state inhibition (up to 1 mM of inhibitor) and during the recovery from inhibition. 2. Stiffness was measured using either step or sinusoidal (1 kHz) changes in fibre length. Sarcomere length was monitored continuously by helium-neon laser diffraction during maximum Ca2+ activation. Stiffness was determined from the changes in sarcomere length and the corresponding changes in force. Vus was measured using the slack test method. 3. AlF chi and Vi each reversibly inhibited force, stiffness and Vus. Actively cycling cross-bridges were required for reversal of these inhibitory effects. Recovery from inhibition by AlF chi was 3- to 4-fold slower than that following removal of V1. 4. At various degrees of inhibition, AlF chi and Vi both inhibited steady-state isometric force more than either Vus or stiffness. For both AlF chi and Vi, the relatively greater inhibition of force over stiffness persisted during recovery from steady-state inhibition. We interpret these results to indicate that the cross-bridges with AlF chi or Vi bound are analogous to those which occur early in the cross-bridge cycle. PMID:8487194

  10. Phosphate-Modified Nucleotides for Monitoring Enzyme Activity.

    PubMed

    Ermert, Susanne; Marx, Andreas; Hacker, Stephan M

    2017-04-01

    Nucleotides modified at the terminal phosphate position have been proven to be interesting entities to study the activity of a variety of different protein classes. In this chapter, we present various types of modifications that were attached as reporter molecules to the phosphate chain of nucleotides and briefly describe the chemical reactions that are frequently used to synthesize them. Furthermore, we discuss a variety of applications of these molecules. Kinase activity, for instance, was studied by transfer of a phosphate modified with a reporter group to the target proteins. This allows not only studying the activity of kinases, but also identifying their target proteins. Moreover, kinases can also be directly labeled with a reporter at a conserved lysine using acyl-phosphate probes. Another important application for phosphate-modified nucleotides is the study of RNA and DNA polymerases. In this context, single-molecule sequencing is made possible using detection in zero-mode waveguides, nanopores or by a Förster resonance energy transfer (FRET)-based mechanism between the polymerase and a fluorophore-labeled nucleotide. Additionally, fluorogenic nucleotides that utilize an intramolecular interaction between a fluorophore and the nucleobase or an intramolecular FRET effect have been successfully developed to study a variety of different enzymes. Finally, also some novel techniques applying electron paramagnetic resonance (EPR)-based detection of nucleotide cleavage or the detection of the cleavage of fluorophosphates are discussed. Taken together, nucleotides modified at the terminal phosphate position have been applied to study the activity of a large diversity of proteins and are valuable tools to enhance the knowledge of biological systems.

  11. Inorganic phosphate induces cancer cell mediated angiogenesis dependent on forkhead box protein C2 (FOXC2) regulated osteopontin expression.

    PubMed

    Lin, Yiming; McKinnon, Kelly E; Ha, Shin Woo; Beck, George R

    2015-09-01

    Recent studies in both rodents and humans suggest that elevated serum phosphorus, in the context of normal renal function, potentiates, or exacerbates pathologies associates with cardiovascular disease, bone metabolism, and cancer. Our recent microarray studies identified the potent stimulation of pro-angiogenic genes such as forkhead box protein C2 (FOXC2), osteopontin, and Vegfα, among others in response to elevated inorganic phosphate (Pi). Increased angiogenesis and neovascularization are important events in tumor growth and the progression to malignancy and FOXC2 has recently been identified as a potential transcriptional regulator of these processes. In this study we addressed the possibility that a high Pi environment would increase the angiogenic potential of cancer cells through a mechanism requiring FOXC2. Our studies utilized lung and breast cancer cell lines in combination with the human umbilical vascular endothelial cell (HUVEC) vessel formation model to better understand the mechanism(s) by which a high Pi environment might alter cancer progression. Exposure of cancer cells to elevated Pi stimulated expression of FOXC2 and conditioned medium from the Pi-stimulated cancer cells stimulated migration and tube formation in the HUVEC model. Mechanistically, we define the requirement of FOXC2 for Pi-induced osteopontin (OPN) expression and secretion from cancer cells as necessary for the angiogenic response. These studies reveal for the first time that cancer cells grown in a high Pi environment promote migration of endothelial cells and tube formation and in so doing identify a novel potential therapeutic target to reduce tumor progression.

  12. Association of serum inorganic phosphate with sex steroid hormones and vitamin D in a nationally representative sample of men.

    PubMed

    Wulaningsih, W; Van Hemelrijck, M; Michaelsson, K; Kanarek, N; Nelson, W G; Ix, J H; Platz, E A; Rohrmann, S

    2014-11-01

    Defects in bone regulatory pathways have been linked to chronic diseases including cardiovascular disease and cancer. In men, a link between bone metabolism and gonadal hormones has been suggested. However, to date, there is lack of evidence on the association between serum inorganic phosphate (Pi) and sex steroid hormones. The objective of this study was to investigate the association between Pi, sex steroid hormones and a known Pi metabolic regulator, vitamin D, in men in the National Health and Nutrition Examination Survey III (NHANES III). From NHANES III, we selected 1412 men aged 20+ who participated in the morning session of Phase I (1988-1991) with serum measurements of Pi, sex hormones, and vitamin D. Multivariable linear regression was used to calculate crude and geometric mean Pi by total and estimated free testosterone and estradiol, sex hormone-binding globulin, androstanediol glucuronide (AAG), and vitamin D. Similar analyses were performed while stratifying by race/ethnicity and vitamin D levels. We found a lack of statistically significant difference in geometric means of Pi across quintiles of concentrations of sex hormones, indicating a tight regulation of Pi. However, Pi levels were inversely associated with calculated free testosterone in non-Hispanic black men, with geometric mean levels of Pi of 1.16 and 1.02 ng/mL for those in the lowest and highest quintiles of free testosterone, respectively (p-trend < 0.05). A similar but weaker pattern was seen between total testosterone and Pi. An inverse association was also seen between AAG and Pi in men with vitamin D concentration below the median (<24.2 ng/mL). No associations were observed among men with vitamin D levels at or above the median. Our findings suggest a weak link among sex hormones, vitamin D, and Pi in men. The observed effects of race/ethnicity and vitamin D indicate a complex association involving various regulators of Pi homeostasis.

  13. Phosphate oxygen isotopic evidence for a temperate and biologically active Archaean ocean.

    PubMed

    Blake, Ruth E; Chang, Sae Jung; Lepland, Aivo

    2010-04-15

    Oxygen and silicon isotope compositions of cherts and studies of protein evolution have been interpreted to reflect ocean temperatures of 55-85 degrees C during the early Palaeoarchaean era ( approximately 3.5 billion years ago). A recent study combining oxygen and hydrogen isotope compositions of cherts, however, makes a case for Archaean ocean temperatures being no greater than 40 degrees C (ref. 5). Ocean temperature can also be assessed using the oxygen isotope composition of phosphate. Recent studies show that (18)O:(16)O ratios of dissolved inorganic phosphate (delta(18)O(P)) reflect ambient seawater temperature as well as biological processing that dominates marine phosphorus cycling at low temperature. All forms of life require and concentrate phosphorus, and as a result of biological processing, modern marine phosphates have delta(18)O(P) values typically between 19-26 per thousand (VSMOW), highly evolved from presumed source values of approximately 6-8 per thousand that are characteristic of apatite in igneous rocks and meteorites. Here we report oxygen isotope compositions of phosphates in sediments from the 3.2-3.5-billion-year-old Barberton Greenstone Belt in South Africa. We find that delta(18)O(P) values range from 9.3 per thousand to 19.9 per thousand and include the highest values reported for Archaean rocks. The temperatures calculated from our highest delta(18)O(P) values and assuming equilibrium with sea water with delta(18)O = 0 per thousand (ref. 12) range from 26 degrees C to 35 degrees C. The higher delta(18)O(P) values are similar to those of modern marine phosphate and suggest a well-developed phosphorus cycle and evolved biologic activity on the Archaean Earth.

  14. Development of an engineered soil bacterium enabling to convert both insoluble inorganic and organic phosphate into plant available phosphate and its use as a biofertilizer.

    PubMed

    Liu, Lili; Du, Wenya; Luo, Wenyu; Su, Yi; Hui, Jiejie; Ma, Shengwu

    2015-05-01

    Phosphorus (P) is one of the most important nutrient elements for plant growth and metabolism. We previously isolated a P-solubilizing bacterium 9320-SD with the ability to utilize inorganic P and convert it into plant-available P. The present study aims to enhance the P-solubilizing capacity of 9320-SD, as our long-term goal is to develop a more effective P-solubilizing bacterial strain for use as a biofertilizer. In this end, we introduced a bacterial phytase encoding gene into 9320-SD. One randomly selected transformant, SDLiuTP02, was examined for recombinant protein expression and phytase activity, and assessed for its ability to promote plant growth. Our results indicate that SDLiuTP02 is capable of expressing high levels of phytase activity. Importantly, corn seedlings treated with the SDLiuTP02 cell culture exhibited increased rates of photosynthesis, transpiration, and stomatal conductance as well as increased growth rate under laboratory conditions and increased growth rate in pot assays compared to seedlings treated with cell cultures of the parental strain 9320-SD. Field experiments further indicated that application of SDLiuTP02 promoted a greater growth rate in young cucumber plant and a higher foliar chlorophyll level in chop suey greens when compared to 9320-SD treated controls. These results indicate that SDLiuTP02 has the potential to be a more effective P biofertilizer to increase agricultural productivity.

  15. Vesicular-arbuscular mycorrhizae and the enzymatic utilization of inorganic phosphate by plant roots: Progress report 1985

    SciTech Connect

    Marx, D. H.

    1985-01-01

    It is well known that phosphorus absorption, especially from soil with low phosphorus levels, by plant roots can be enhanced by mycorrhizal infection. Root cortical cells colonized by vesicular-arbuscular mycorrhizal fungi (VAM) have higher concentrations of phosphorus than noninfected cells. Polyphosphate is the major phosphorus reserve in many fungi and is reported to be present abundantly in young and proliferating arbuscules. We propose that mycorrhizal polyphosphate can be utilized by the VAM-plant symbiont system as a phosphorus donor and an energy source in the glycolytic pathway, possibly after being hydrolyzed to pytrophosphate (PPi). The VAM systems of infected and noninfected roots of sweetgum (Liquidambar styraciflua L.) and onion (Allium cepa L. var. Texas Grand) were used to compare the activity of PPI-dependent phosphofructokinase (PFK), an enzyme utilizing PPi to convert frutose-6-phosphate into fructosel,6-bisphosphate. The ATP-PKF activity was measured also. 1 fig., 3 tabs.

  16. Carbon and Nitrogen Sources Influence Tricalcium Phosphate Solubilization and Extracellular Phosphatase Activity by Talaromyces flavus.

    PubMed

    Stefanoni Rubio, P J; Godoy, M S; Della Mónica, I F; Pettinari, M J; Godeas, A M; Scervino, J M

    2016-01-01

    The aim of this work was to study phosphate (P) solubilization (and the processes involved in this event) by Talaromyces flavus (BAFC 3125) as a function of carbon and/or nitrogen sources. P solubilization was evaluated in NBRIP media supplemented with different carbon (glucose, sorbitol, sucrose, and fructose) and nitrogen (L-asparagine, urea, ammonium sulfate (AS), and ammonium nitrate (AN) combinations. The highest P solubilization was related to the highest organic acid production (especially gluconic acid) and pH drop for those treatments where glucose was present. Also P solubilization was higher when an inorganic nitrogen source was supplemented to the media when compared to an organic one. Although not being present an organic P source, phosphatase activity was observed. This shows that P mineralization and P solubilization can occur simultaneously, and that P mineralization is not induced by the enzyme substrate. The combination that showed highest P solubilization was for AN-glucose. The highest acid phosphatase activity was for AS-fructose, while for alkaline phosphatase were for AS-fructose and AN-fructose. Acid phosphatase activity was higher than alkaline. P solubilization and phosphatase activity (acid and alkaline) were influenced by the different carbon-nitrogen combinations. A better understanding of phosphate-solubilizing fungi could bring a better use of soil P.

  17. Catalysis of hydrolysis and nucleophilic substitution at the P-N bond of phosphoimidazolide-activated nucleotides in phosphate buffers

    NASA Technical Reports Server (NTRS)

    Kanavarioti, A.; Rosenbach, M. T.

    1991-01-01

    Phosphoimidazolide-activated derivatives of guanosine and cytidine 5'-monophosphates, henceforth called ImpN's, exhibit enhanced rates of degradation in the presence of aqueous inorganic phosphate in the range 4.0 < or = pH < or = 8.6. This degradation is been attributed to (i) nucleophilic substitution of the imidazolide and (ii) catalysis of the P-N bond hydrolysis by phosphate. The first reaction results in the formation of nucleoside 5'-diphosphate and the second in nucleoside 5'-monophosphate. Analysis of the observed rates as well as the product ratios as a function of pH and phosphate concentration allow distinction between various mechanistic possibilities. The results show that both H2PO4- and HPO4(2-) participate in both hydrolysis and nucleophilic substitution. Statistically corrected biomolecular rate constants indicate that the dianion is 4 times more effective as a general base than the monoanion, and 8 times more effective as nucleophile. The low Bronsted value beta = 0.15 calculated for these phosphate species, presumed to act as general bases in facilitating water attack, is consistent with the fact that catalysis of the hydrolysis of the P-N bond in ImpN's has not been detected before. The beta nuc = 0.35 calculated for water, H2PO4-, HPO4(2-), and hydroxide acting as nucleophiles indicates a more associative transition state for nucleotidyl (O2POR- with R = nucleoside) transfers than that observed for phosphoryl (PO3(2-)) transfers (beta nuc = 0.25). With respect to the stability/reactivity of ImpN's under prebiotic conditions, our study shows that these materials would not suffer additional degradation due to inorganic phosphate, assuming the concentrations of phosphate, Pi, on prebiotic Earth were similar to those in the present oceans ([Pi] approximately 2.25 micromoles).

  18. Glycerolipid biosynthesis in Saccharomyces cerevisiae: sn-glycerol-3-phosphate and dihydroxyacetone phosphate acyltransferase activities.

    PubMed Central

    Schlossman, D M; Bell, R M

    1978-01-01

    Yeast acyl-coenzyme A:dihydroxyacetone-phosphate O-acyltransferase (DHAP acyltransferase; EC 2.3.1.42) was investigated to (i) determine whether its activity and that of acyl-coenzyme A:sn-glycerol-3-phosphate O-acyltransferase (glycerol-P acyltransferase; EC 2.3.1.15) represent dual catalytic functions of a single membranous enzyme, (ii) estimate the relative contributions of the glycerol-P and DHAP pathways for yeast glycerolipid synthesis, and (iii) evaluate the suitability of yeast for future genetic investigations of the eucaryotic glycerol-P and DHAP acyltransferase activities. The membranous DHAP acyltransferase activity showed an apparent Km of 0.79 mM for DHAP, with a Vmax of 5.3 nmol/min per mg, whereas the glycerol-P acyltransferase activity showed an apparent Km of 0.05 mM for glycerol-P, with a Vmax of 3.4 nmol/min per mg. Glycerol-P was a competitive inhibitor (Ki, 0.07 mM) of the DHAP acyltransferase activity, and DHAP was a competitive inhibitor (Ki, 0.91 mM) of the glycerol-P acyltransferase activity. The two acyltransferase activities exhibited marked similarities in their pH dependence, acyl-coenzyme A chain length preference and substrate concentration dependencies, thermolability, and patterns of inactivation by N-ethylmaleimide, trypsin, and detergents. Thus, the data strongly suggest that yeast glycerol-P and DHAP acyltransferase activities represent dual catalytic functions of a single membrane-bound enzyme. Furthermore, since no acyl-DHAP oxidoreductase activity could be detected in yeast membranes, the DHAP pathway for glycerolipid synthesis may not operate in yeast. PMID:25265

  19. Synthesis, characterization and antibacterial activity of cellulose acetate-tin (IV) phosphate nanocomposite.

    PubMed

    Rathore, Bhim Singh; Sharma, Gaurav; Pathania, Deepak; Gupta, Vinod Kumar

    2014-03-15

    Cellulose acetate-tin (IV) phosphate nanocomposite (CA/TPNC) was prepared using simple method at 0-1 pH. The nanocomposite ion exchanger was characterized using some techniques such as Fourier transform infrared spectroscopy (FTIR), transmission electron microscopy (TEM), scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDX), X-ray diffraction (XRD), and thermogravimetric analysis (TGA/DTA/DSC). The nanocomposite material was explored for different properties such as ion exchange capacity, pH titration, elution behavior, thermal stability, and distribution coefficient. The ion exchange capacity of CA/TPNC was found higher compared to their inorganic counterpart. The distribution coefficient studies of nanocomposite ion exchanger were investigated for different metal ions. On the basis of distribution coefficient studies CA/TPNC material was found more selective for Cd(2+) and Mg(2+). CA/TPNC ion exchange was explored for antibacterial activities against E. coli bacteria.

  20. Novel phosphate-activated macrophages prevent ectopic calcification by increasing extracellular ATP and pyrophosphate

    PubMed Central

    Villa-Bellosta, Ricardo; Hamczyk, Magda R.; Andrés, Vicente

    2017-01-01

    Purpose Phosphorus is an essential nutrient involved in many pathobiological processes. Less than 1% of phosphorus is found in extracellular fluids as inorganic phosphate ion (Pi) in solution. High serum Pi level promotes ectopic calcification in many tissues, including blood vessels. Here, we studied the effect of elevated Pi concentration on macrophage polarization and calcification. Macrophages, present in virtually all tissues, play key roles in health and disease and display remarkable plasticity, being able to change their physiology in response to environmental cues. Methods and results High-throughput transcriptomic analysis and functional studies demonstrated that Pi induces unpolarized macrophages to adopt a phenotype closely resembling that of alternatively-activated M2 macrophages, as revealed by arginine hydrolysis and energetic and antioxidant profiles. Pi-induced macrophages showed an anti-calcifying action mediated by increased availability of extracellular ATP and pyrophosphate. Conclusion We conclude that the ability of Pi-activated macrophages to prevent calcium-phosphate deposition is a compensatory mechanism protecting tissues from hyperphosphatemia-induced pathologic calcification. PMID:28362852

  1. Inorganic lead and calcium interact positively in activation of calmodulin.

    PubMed

    Kern, M; Wisniewski, M; Cabell, L; Audesirk, G

    2000-06-01

    Calmodulin is a ubiquitous calcium-binding protein that mediates many of the intracellular actions of Ca2+ ions. The calcium-binding sites of calmodulin consist of four EF-hand motifs; full activation of calmodulin normally occurs when all four sites are occupied by Ca2+. Inorganic lead (PY2+) has been shown to activate calmodulin at total lead concentrations similar to the concentrations of Ca2+ required for activation (Goldstein and Ar, 1983; Habermann et al., 1983), but the free Pb2+ concentrations required for calmodulin activation have not been determined. In addition, it is possible that activation may occur with different sites occupied by different divalent cations, for example Ca2+ and Pb2+. We investigated the ability of free Pb2+, alone or in combination with Ca2+, to activate calmodulin. In aqueous media, N-phenyl-1-naphthylamine (NPN) and 8-anilino-1-naphthalenesulfonate (ANS) show increased fluorescence when bound to hydrophobic regions of proteins. This increased fluorescence has been used to monitor the conformational change that occurs during calmodulin activation (LaPorte et al., 1980). In the presence of calmodulin, both Ca2+ and Pb2+ stimulated increased fluorescence of NPN and ANS. Threshold and EC50 free metal concentrations were approximately 100 nM and 450-500 nM, respectively, for Ca2+ and 100 pM and 400-550 pM, respectively, for Pb2+. Fluorescence was enhanced by combinations of low concentrations of free Ca2+ and Pb2+; for example, as little as 20 pM free Pb2+ enhanced fluorescence in combination with 200 nM free Ca2+. The activity of the PDE1 isoform of cyclic nucleotide phosphodiesterase is stimulated by Ca2+/calmodulin (Wang et al., 1990). In the presence of calmodulin, we found that Ca2+ and Pb2+ activated calmodulin-stimulated PDE activity, with threshold and EC50 free metal concentrations of approximately 200 nM and 1200 nM, respectively, for Ca2+ and 300 pM and 430 pM, respectively, for Pb2+. PDE activity was stimulated by

  2. Evaluation of commonly used methods for the analysis of acid-soluble phosphate in internationally traded inorganic fertilizers.

    PubMed

    Hall, William L; Siegel, Sanford

    2014-01-01

    Several methodologies are used throughout the world to determine phosphate concentration (measured as PO4 and expressed as % P2O5) in fertilizers. Concentrated phosphate materials, including diammonium phosphate (DAP) and monoammonium phosphate (MAP), are traded in large volumes (millions of metric tons) internationally. The International Fertilizer Association (IFA) identified a need to assess the methods currently being used to measure the phosphate content for suitability (scope), accuracy, and repeatability. Even small discrepancies in the expressed P2O5 content can have a major financial impact on buyers and sellers as contracts are settled and import regulations are imposed. The IFA's Technical Committee selected a working group to address issues dealing with harmonization of fertilizer sampling and analytical methodologies. The working group identified phosphate content in DAP and MAP fertilizers as a major concern for commerce. The working group initiated a method screening and comparison project to assess method performance and to determine which methods, if any, could be considered best practice methods and, therefore, be deemed acceptable for use by the industry. In order to systematically review the acceptability of methods for consideration, the task force developed an assessment protocol outlined in a white paper involving three steps: (1) compile all known relevant methods practiced in global fertilizer trade, (2) review and evaluate methods based upon specific evaluation criteria, and (3) compare the methods that most closely fit the evaluation criteria by multilaboratory analysis of unknown materials for accuracy and repeatability. Six methods were evaluated for analysis of total phosphate in concentrated phosphate products. From these methods, four were determined to be acceptable as best practice methods. The study members proposed three of the methods, while a fourth method was commonly used among the participating laboratories. This publication

  3. Faster and stronger manifestation of mitochondrial diseases in skeletal muscle than in heart related to cytosolic inorganic phosphate (Pi) accumulation.

    PubMed

    Korzeniewski, Bernard

    2016-08-01

    A model of the cell bioenergetic system was used to compare the effect of oxidative phosphorylation (OXPHOS) deficiencies in a broad range of moderate ATP demand in skeletal muscle and heart. Computer simulations revealed that kinetic properties of the system are similar in both cases despite the much higher mitochondria content and "basic" OXPHOS activity in heart than in skeletal muscle, because of a much higher each-step activation (ESA) of OXPHOS in skeletal muscle than in heart. Large OXPHOS deficiencies lead in both tissues to a significant decrease in oxygen consumption (V̇o2) and phosphocreatine (PCr) and increase in cytosolic ADP, Pi, and H(+) The main difference between skeletal muscle and heart is a much higher cytosolic Pi concentration in healthy tissue and much higher cytosolic Pi accumulation (level) at low OXPHOS activities in the former, caused by a higher PCr level in healthy tissue (and higher total phosphate pool) and smaller Pi redistribution between cytosol and mitochondria at OXPHOS deficiency. This difference does not depend on ATP demand in a broad range. A much greater Pi increase and PCr decrease during rest-to-moderate work transition in skeletal muscle at OXPHOS deficiencies than at normal OXPHOS activity significantly slows down the V̇o2 on-kinetics. Because high cytosolic Pi concentrations cause fatigue in skeletal muscle and can compromise force generation in skeletal muscle and heart, this system property can contribute to the faster and stronger manifestation of mitochondrial diseases in skeletal muscle than in heart. Shortly, skeletal muscle with large OXPHOS deficiencies becomes fatigued already during low/moderate exercise.

  4. Fluorometric method for inorganic pyrophosphatase activity detection and inhibitor screening based on click chemistry.

    PubMed

    Xu, Kefeng; Chen, Zhonghui; Zhou, Ling; Zheng, Ou; Wu, Xiaoping; Guo, Longhua; Qiu, Bin; Lin, Zhenyu; Chen, Guonan

    2015-01-06

    A fluorometric method for pyrophosphatase (PPase) activity detection was developed based on click chemistry. Cu(II) can coordinate with pyrophosphate (PPi), the addition of pyrophosphatase (PPase) into the above system can destroy the coordinate compound because PPase catalyzes the hydrolysis of PPi into inorganic phosphate and produces free Cu(II), and free Cu(II) can be reduced by sodium ascorbate (SA) to form Cu(I), which in turn initiates the ligating reaction between nonfluorescent 3-azidocoumarins and terminal alkynes to produce a highly fluorescent triazole complex, based on which, a simple and sensitive turn on fluorometric method for PPase can be developed. The fluorescence intensity of the system has a linear relationship with the logarithm of the PPase concentration in the range of 0.5 and 10 mU with a detection limit down to 0.2 mU (S/N = 3). This method is cost-effective and convenient without any labels or complicated operations. The proposed system was applied to screen the potential PPase inhibitor with high efficiency. The proposed method can be applied to diagnosis of PPase-related diseases.

  5. Effect of pH and coexisting anions on removal of phosphate from aqueous solutions by inorganic-based mesostructures.

    PubMed

    Choi, Jae-Woo; Choi, Yong-Soo; Hong, Seok-Won; Kim, Dong-Ju; Lee, Sang-Hyup

    2012-07-01

    This study investigated the effect of pH and the presence of coexisting (competitive) anions on the removal of phosphate by titanium mesostructures synthesized using do- or hexadecyltrimethylammonium bromide. To address these research objectives, experiments were conducted (1) under controlled initial pH values (2 to 10); and (2) through injection of nitrate, fluoride, chloride, or sulfate anions into a phosphate solution. Based on the experimental results, an initial of pH of 2 was found to be optimal for use of titanium mesostructures. The presence of fluoride anions in solution significantly decreased the removal efficiency of phosphate removal (3.56% at 3.95 mg/g). However, the addition of nitrate, chloride, and sulfate anions did not affect phosphate removal.

  6. Inorganic phosphate-triggered release of anti-cancer arsenic trioxide from a self-delivery system: an in vitro and in vivo study

    NASA Astrophysics Data System (ADS)

    Chen, Fei-Yan; Yi, Jing-Wei; Gu, Zhe-Jia; Tang, Bin-Bing; Li, Jian-Qi; Li, Li; Kulkarni, Padmakar; Liu, Li; Mason, Ralph P.; Tang, Qun

    2016-03-01

    On-demand drug delivery is becoming feasible via the design of either exogenous or endogenous stimulus-responsive drug delivery systems. Herein we report the development of gadolinium arsenite nanoparticles as a self-delivery platform to store, deliver and release arsenic trioxide (ATO, Trisenox), a clinical anti-cancer drug. Specifically, unloading of the small molecule drug is triggered by an endogenous stimulus: inorganic phosphate (Pi) in the blood, fluid, and soft or hard tissue. Kinetics in vitro demonstrated that ATO is released with high ON/OFF specificity and no leakage was observed in the silent state. The nanoparticles induced tumor cell apoptosis, and reduced cancer cell migration and invasion. Plasma pharmacokinetics verified extended retention time, but no obvious disturbance of phosphate balance. Therapeutic efficacy on a liver cancer xenograft mouse model was dramatically potentiated with reduced toxicity compared to the free drug. These results suggest a new drug delivery strategy which might be applied for ATO therapy on solid tumors.On-demand drug delivery is becoming feasible via the design of either exogenous or endogenous stimulus-responsive drug delivery systems. Herein we report the development of gadolinium arsenite nanoparticles as a self-delivery platform to store, deliver and release arsenic trioxide (ATO, Trisenox), a clinical anti-cancer drug. Specifically, unloading of the small molecule drug is triggered by an endogenous stimulus: inorganic phosphate (Pi) in the blood, fluid, and soft or hard tissue. Kinetics in vitro demonstrated that ATO is released with high ON/OFF specificity and no leakage was observed in the silent state. The nanoparticles induced tumor cell apoptosis, and reduced cancer cell migration and invasion. Plasma pharmacokinetics verified extended retention time, but no obvious disturbance of phosphate balance. Therapeutic efficacy on a liver cancer xenograft mouse model was dramatically potentiated with reduced

  7. ATP binding and hydrolysis steps of the uni-site catalysis by the mitochondrial F(1)-ATPase are affected by inorganic phosphate.

    PubMed

    Milgrom, Yakov M

    2010-10-01

    The effect of inorganic phosphate (P(i)) on uni-site ATP binding and hydrolysis by the nucleotide-depleted F(1)-ATPase from beef heart mitochondria (ndMF(1)) has been investigated. It is shown for the first time that P(i) decreases the apparent rate constant of uni-site ATP binding by ndMF(1) 3-fold with the K(d) of 0.38+/-0.14mM. During uni-site ATP hydrolysis, P(i) also shifts equilibrium between bound ATP and ADP+P(i) in the direction of ATP synthesis with the K(d) of 0.17+/-0.03mM. However, 10mM P(i) does not significantly affect ATP binding during multi-site catalysis.

  8. Inhibitory activity of phosphates on molds isolated from foods and food processing plants.

    PubMed

    Suárez, V B; Frisón, L; de Basílico, M Z; Rivera, M; Reinheimer, J A

    2005-11-01

    Six commercial phosphates were evaluated for inhibition of the growth of 17 molds isolated from food sources. The assays were performed at neutral and natural (without pH adjustment) pH values, and the molds were streaked on plate count agar with added phosphates. Phosphate concentrations of 0.1, 0.3, 0.5, 1.0, and 1.5% (wt/vol) were used, and the MIC was determined. The resistance of molds to phosphates depended on the species. At a neutral pH, Aspergillus ochraceus and Fusarium proliferatum were resistant to all phosphates at all concentrations assayed, and Byssochlamys nivea, Aureobasidium pullulans, and Penicillium glabrum were most sensitive. The most inhibitory phosphates were those with chain lengths greater than 15 phosphate units and the highest sequestering power. At natural pH values (resulting from dissolving the phosphate in the medium), inhibitory activity changed dramatically for phosphates that produced alkaline or acidic pH in the medium. Phosphates with alkaline pH values (sodium tripolyphosphate of high solubility, sodium tripolyphosphate, and sodium neutral pyrophosphate) were much more inhibitory than phosphates at a neutral pH, but sodium acid pyrophosphate (acidic pH) had decreased inhibitory activity. The results indicate that some phosphates could be used in the food industry to inhibit molds linked to food spoilage.

  9. Poly(alkylene phosphates): from synthetic models of biomacromolecules and biomembranes toward polymer-inorganic hybrids (mimicking biomineralization).

    PubMed

    Penczek, Stanislaw; Pretula, Julia; Kaluzynski, Krzysztof

    2005-01-01

    Syntheses of poly(alkylene phosphates), with repeating units having two or three methylene groups and phosphoryl groups and mimicking backbones of biomacromolecules, are reviewed. Two major methods elaborated in this laboratory, namely, ring-opening polymerization and transesterification, are described. The resulting polymers were used as carriers of cations (Ca2+ and Mg2+) in membrane processes and in controlling the crystallization of CaCO3, in a process related to biomineralization.

  10. Effect of aluminum phosphate on alkaline phosphatase activity of polyurethane foam immobilized cyanobacteria.

    PubMed

    Ramalingam, N; Prasanna, B Gowtham

    2006-09-01

    The impact of insoluble phosphorus such as aluminum and rock phosphate on alkaline phosphatase activity of polyurethane foam immobilized cyanobacteria was assessed. Polyurethane foam immobilized Nodularia recorded the highest alkaline phosphatase activity of 9.04 (m. mol p-nitrophenol released h(-1) mg(-1) protein) in vitro. A higher concentration of aluminum phosphate was recorded a 25% reduction in alkaline phosphatase activity, ammonia content, and available phosphorus in culture filtrate of polyurethane foam immobilized cyanobacteria. In general, immobilized cyanobacteria exhibited a higher alkaline phosphatase activity in rock phosphate than aluminum phosphate.

  11. Activation and fluoride-assisted phosphating of aluminum-silicon-coated steel.

    PubMed

    Schneider, Paul; Sigel, Reinhard; Lange, Miriam M; Beier, Frank; Renner, Frank U; Erbe, Andreas

    2013-05-22

    Phosphating is a crucial process in the corrosion protection of metals. Here, activation and fluoride-assisted tricationic phosphating is investigated on aluminum-silicon (AS) coated steel surfaces. Dynamic light scattering results from the activation bath show a bimodal size distribution, with hydrodynamic radii of ~400 nm and ~10 μm. For the smaller particle fraction, static light scattering results are consistent with the interpretation of disklike particles as scattering objects. Particles of the larger fraction sediment with time. In the presence of electrolyte, the scattering intensity from the larger particle fraction increases. Coagulation with time is suggested to be related to the decrease in activity of the activation bath. Scanning Auger microscopy (SAM) shows a higher phosphorus concentration after titanium phosphate activation in the Al-rich areas compared to the Si-rich areas of the AS coatings. There is no correlation between the size of the species in the activation bath, and the size of the phosphate-containing regions on the activated surface. Phosphating was performed in the presence of hexafluorosilicic acid, H2SiF6, ammonium hydrogen difluoride, NH4HF2, and both, at an initial pH of 2.5. The absence of crystals after phosphating with H2SiF6 is an indication that SiF6(2-) is the final product of the oxide dissolution in the presence of fluoride. In the presence of NH4HF2, the Si-rich regions of the surface are phosphated before the Si-poor (Al-rich) regions. Hence, the phosphate distribution after activation and after phosphating are opposite. These results show that a high surface concentration of phosphate after activation is not sufficient for a high coverage with phosphate crystals after phosphating.

  12. Biologic activity of cyclic and caged phosphates: a review.

    PubMed

    Lorke, Dietrich E; Stegmeier-Petroianu, Anka; Petroianu, Georg A

    2017-01-01

    The recognition in the early 1960s by Morifusa Eto that tri-o-cresyl phosphate (TOCP) is hydroxylated by the cytochrome P450 system to an intermediate that spontaneously cyclizes to a neurotoxic phosphate (saligenin phosphate ester) ignited the interest in this group of compounds. Only the ortho isomer can cyclize and clinically cause Organo Phosphate Induced Delayed Neurotoxicity (OPIDN); the meta and para isomers of tri-cresyl phosphate are not neuropathic because they are unable to form stable cyclic saligenin phosphate esters. This review identifies the diverse biological effects associated with various cyclic and caged phosphates and phosphonates and their possible use. Cyclic compounds that inhibit acetylcholine esterase (AChE), such as salithion, can be employed as pesticides. Others are neurotoxic, most probably because of inhibition of neuropathy target esterase (NTE). Cyclic phosphates that inhibit lipases, the cyclipostins, possibly represent promising therapeutic avenues for the treatment of type 2 diabetes mellitus and/or microbial infections; those compounds inhibiting β-lactamase may prevent bacterial resistance against β-lactam antibiotics. Naturally occurring cyclic phosphates, such as cyclic AMP, cyclic phosphatidic acid and the ryanodine receptor modulator cyclic adenosine diphosphate ribose, play an important physiological role in signal transduction. Moreover, some cyclic phosphates are GABA-antagonists, while others are an essential component of Molybdenum-containing enzymes. Some cyclic phosphates (cyclophosphamide, ifosfamide) are clinically used in tumor therapy, while the coupling of therapeutic agents with other cyclic phosphates (HepDirect® Technology) allows drugs to be targeted to specific organs. Possible clinical applications of these compounds are considered. Copyright © 2016 John Wiley & Sons, Ltd.

  13. Oxygen isotope ratios of PO4: an inorganic indicator of enzymatic activity and P metabolism and a new biomarker in the search for life.

    PubMed

    Blake, R E; Alt, J C; Martini, A M

    2001-02-27

    The distinctive relations between biological activity and isotopic effect recorded in biomarkers (e.g., carbon and sulfur isotope ratios) have allowed scientists to suggest that life originated on this planet nearly 3.8 billion years ago. The existence of life on other planets may be similarly identified by geochemical biomarkers, including the oxygen isotope ratio of phosphate (delta(18)O(p)) presented here. At low near-surface temperatures, the exchange of oxygen isotopes between phosphate and water requires enzymatic catalysis. Because enzymes are indicative of cellular activity, the demonstration of enzyme-catalyzed PO(4)-H(2)O exchange is indicative of the presence of life. Results of laboratory experiments are presented that clearly show that delta(18)O(P) values of inorganic phosphate can be used to detect enzymatic activity and microbial metabolism of phosphate. Applications of delta(18)O(p) as a biomarker are presented for two Earth environments relevant to the search for extraterrestrial life: a shallow groundwater reservoir and a marine hydrothermal vent system. With the development of in situ analytical techniques and future planned sample return strategies, delta(18)O(p) may provide an important biosignature of the presence of life in extraterrestrial systems such as that on Mars.

  14. Special Feature: Oxygen isotope ratios of PO4: An inorganic indicator of enzymatic activity and P metabolism and a new biomarker in the search for life

    NASA Astrophysics Data System (ADS)

    Blake, Ruth E.; Alt, Jeffrey C.; Martini, Anna M.

    2001-02-01

    The distinctive relations between biological activity and isotopic effect recorded in biomarkers (e.g., carbon and sulfur isotope ratios) have allowed scientists to suggest that life originated on this planet nearly 3.8 billion years ago. The existence of life on other planets may be similarly identified by geochemical biomarkers, including the oxygen isotope ratio of phosphate (18Op) presented here. At low near-surface temperatures, the exchange of oxygen isotopes between phosphate and water requires enzymatic catalysis. Because enzymes are indicative of cellular activity,the demonstration of enzyme-catalyzed PO4-H2O exchange is indicative of the presence of life. Results of laboratory experiments are presented that clearly show that δ18OP values of inorganic phosphate can be used to detect enzymatic activity and microbial metabolism of phosphate. Applications of δ18Op as a biomarker are presented for two Earth environments relevant to the search for extraterrestrial life: a shallow groundwater reservoir and a marine hydrothermal vent system. With the development of in situ analytical techniques and future planned sample return strategies, δ18Op may provide an important biosignature of the presence of life in extraterrestrial systems such as that on Mars.

  15. Oxygen isotope ratios of PO4: An inorganic indicator of enzymatic activity and P metabolism and a new biomarker in the search for life

    PubMed Central

    Blake, Ruth E.; Alt, Jeffrey C.; Martini, Anna M.

    2001-01-01

    The distinctive relations between biological activity and isotopic effect recorded in biomarkers (e.g., carbon and sulfur isotope ratios) have allowed scientists to suggest that life originated on this planet nearly 3.8 billion years ago. The existence of life on other planets may be similarly identified by geochemical biomarkers, including the oxygen isotope ratio of phosphate (δ18Op) presented here. At low near-surface temperatures, the exchange of oxygen isotopes between phosphate and water requires enzymatic catalysis. Because enzymes are indicative of cellular activity, the demonstration of enzyme-catalyzed PO4–H2O exchange is indicative of the presence of life. Results of laboratory experiments are presented that clearly show that δ18OP values of inorganic phosphate can be used to detect enzymatic activity and microbial metabolism of phosphate. Applications of δ18Op as a biomarker are presented for two Earth environments relevant to the search for extraterrestrial life: a shallow groundwater reservoir and a marine hydrothermal vent system. With the development of in situ analytical techniques and future planned sample return strategies, δ18Op may provide an important biosignature of the presence of life in extraterrestrial systems such as that on Mars. PMID:11226207

  16. Transformation of inorganic P fractions of soil and plant growth promotion by phosphate-solubilizing ability of Penicillium oxalicum I1.

    PubMed

    Gong, Mingbo; Du, Peng; Liu, Xue; Zhu, Changxiong

    2014-12-01

    The solubilization of tricalcium phosphate is often considered as the standard for screening of most phosphate-solubilizing microorganisms (PSMs). However, usually the effect of large-scale application of PSM on the promotion of crop growth varies. This study presents an efficient method for screening and testing phosphate-solubilizing fungus that enhance plant growth. A fungus Penicillium oxalicum I1 (P-I1) was isolated and identified that had high ability of phosphate-solubilization and could utilize maize root exudates as sources, and propagate well in vitro and in soil. P-I1 excreted oxalic acid and reached 593.9 μg/ml, and the pH value was decreased from 6.90 to 1.65 in 26 h. The amount of P-I1 increased by 48-fold in 28 d and was maintained for 49 d in soil. PSM showed selectivity on the transformation of the different forms of phosphorus, a wide range of insoluble phosphates, such as Ca₈H₂(PO₄)₆·5H₂O, AlPO₄, FePO₄, and Ca10(PO₄)₆(OH)₂, were converted to soluble CaHPO₄in soil, and CaHPO₄was also inhibited from being converted into insoluble phosphate by P-I1. The Ca₂-P content reached 27.11 μg/g soil on day 28 at 20°C, which increased by 110.32%, and plant growth promotion was tested and verified, the results showed that maize yield increased remarkably than control after inoculated P-I1, maize yield increased maximum by 14.47%. The data presented that P-I1 appear attractive for exploring their plant growth-promoting activity and potential field application.

  17. Succinyl-CoA synthetase is a phosphate target for the activation of mitochondrial metabolism.

    PubMed

    Phillips, Darci; Aponte, Angel M; French, Stephanie A; Chess, David J; Balaban, Robert S

    2009-08-04

    Succinyl-CoA synthetase (SCS) is the only mitochondrial enzyme capable of ATP production via substrate level phosphorylation in the absence of oxygen, but it also plays a key role in the citric acid cycle, ketone metabolism, and heme synthesis. Inorganic phosphate (P(i)) is a signaling molecule capable of activating oxidative phosphorylation at several sites, including NADH generation and as a substrate for ATP formation. In this study, it was shown that P(i) binds the porcine heart SCS alpha-subunit (SCSalpha) in a noncovalent manner and enhances its enzymatic activity, thereby providing a new target for P(i) activation in mitochondria. Coupling 32P labeling of intact mitochondria with SDS gel electrophoresis revealed that 32P labeling of SCSalpha was enhanced in substrate-depleted mitochondria. Using mitochondrial extracts and purified bacterial SCS (BSCS), we showed that this enhanced 32P labeling resulted from a simple binding of 32P, not covalent protein phosphorylation. The ability of SCSalpha to retain its 32P throughout the SDS denaturing gel process was unique over the entire mitochondrial proteome. In vitro studies also revealed a P(i)-induced activation of SCS activity by more than 2-fold when mitochondrial extracts and purified BSCS were incubated with millimolar concentrations of P(i). Since the level of 32P binding to SCSalpha was increased in substrate-depleted mitochondria, where the matrix P(i) concentration is increased, we conclude that SCS activation by P(i) binding represents another mitochondrial target for the P(i)-induced activation of oxidative phosphorylation and anaerobic ATP production in energy-limited mitochondria.

  18. Inorganic Polyphosphates Regulate Hexokinase Activity and Reactive Oxygen Species Generation in Mitochondria of Rhipicephalus (Boophilus) microplus Embryo

    PubMed Central

    Fraga, Amanda; Moraes, Jorge; da Silva, José Roberto; Costa, Evenilton P.; Menezes, Jackson; da Silva Vaz Jr, Itabajara; Logullo, Carlos; da Fonseca, Rodrigo Nunes; Campos, Eldo

    2013-01-01

    The physiological roles of polyphosphates (poly P) recently found in arthropod mitochondria remain obscure. Here, the possible involvement of poly P with reactive oxygen species generation in mitochondria of Rhipicephalus microplus embryos was investigated. Mitochondrial hexokinase and scavenger antioxidant enzymes, such as superoxide dismutase, catalase, and glutathione reductase were assayed during embryogenesis of R. microplus. The influence of poly P3 and poly P15 were analyzed during the period of higher enzymatic activity during embryogenesis. Both poly Ps inhibited hexokinase activity by up to 90% and, interestingly, the mitochondrial membrane exopolyphosphatase activity was stimulated by the hexokinase reaction product, glucose-6-phosphate. Poly P increased hydrogen peroxide generation in mitochondria in a situation where mitochondrial hexokinase is also active. The superoxide dismutase, catalase and glutathione reductase activities were higher during embryo cellularization, at the end of embryogenesis and during embryo segmentation, respectively. All of the enzymes were stimulated by poly P3. However, superoxide dismutase was not affected by poly P15, catalase activity was stimulated only at high concentrations and glutathione reductase was the only enzyme that was stimulated in the same way by both poly Ps. Altogether, our results indicate that inorganic polyphosphate and mitochondrial membrane exopolyphosphatase regulation can be correlated with the generation of reactive oxygen species in the mitochondria of R. microplus embryos. PMID:23983617

  19. Modeling the Thermodynamics of Mixed Organic-Inorganic Aerosols to Predict Water Activities and Phase Equilibria

    NASA Astrophysics Data System (ADS)

    Zuend, A.; Marcolli, C.; Luo, B.; Peter, T.

    2008-12-01

    Tropospheric aerosol particles contain mixtures of inorganic salts, acids, water, and a large variety of organic compounds. Interactions between these substances in liquid mixtures lead to discrepancies from ideal thermodynamic behavior. While the thermodynamics of aqueous inorganic systems at atmospheric temperatures are well established, little is known about the physicochemistry of mixed organic-inorganic particles. Salting-out and salting-in effects result from organic-inorganic interactions and are used to improve industrial separation processes. In the atmosphere, they may influence the aerosol phases. Liquid-liquid phase separations into a mainly polar (aqueous) and a less polar organic phase may considerably influence the gas/particle partitioning of semi-volatile substances compared to a single phase estimation. Moreover, the phases present in the aerosol define the reaction medium for heterogeneous and multiphase chemistry occurring in aerosol particles. A correct description of these phases is needed when gas- or cloud-phase reaction schemes are adapted to aerosols. Non-ideal thermodynamic behavior in mixtures is usually described by an expression for the excess Gibbs energy. We present the group-contribution model AIOMFAC (Aerosol Inorganic-Organic Mixtures Functional groups Activity Coefficients), which explicitly accounts for molecular interactions between solution constituents, both organic and inorganic, to calculate activities, chemical potentials and the total Gibbs energy of mixed systems. This model allows to compute vapor-liquid (VLE), liquid-liquid (LLE) and solid-liquid (SLE) equilibria within one framework. Focusing on atmospheric applications we considered eight different cations, five anions and a wide range of alcohols/polyols as organic compounds. With AIOMFAC, the activities of the components within an aqueous electrolyte solution are very well represented up to high ionic strength. We show that the semiempirical middle

  20. High levels of the type III inorganic phosphate transporter PiT1 (SLC20A1) can confer faster cell adhesion

    SciTech Connect

    Kongsfelt, Iben Boutrup; Byskov, Kristina; Pedersen, Lasse Ebdrup; Pedersen, Lene

    2014-08-01

    The inorganic phosphate transporter PiT1 (SLC20A1) is ubiquitously expressed in mammalian cells. We recently showed that overexpression of human PiT1 was sufficient to increase proliferation of two strict density-inhibited cell lines, murine fibroblastic NIH3T3 and pre-osteoblastic MC3T3-E1 cells, and allowed the cultures to grow to higher cell densities. In addition, upon transformation NIH3T3 cells showed increased ability to form colonies in soft agar. The cellular regulation of PiT1 expression supports that cells utilize the PiT1 levels to control proliferation, with non-proliferating cells showing the lowest PiT1 mRNA levels. The mechanism behind the role of PiT1 in increased cell proliferation is not known. We, however, found that compared to control cells, cultures of NIH3T3 cells overexpressing PiT1 upon seeding showed increased cell number after 24 h and had shifted more cells from G0/G1 to S+G2/M within 12 h, suggesting that an early event may play a role. We here show that expression of human PiT1 in NIH3T3 cells led to faster cell adhesion; this effect was not cell type specific in that it was also observed when expressing human PiT1 in MC3T3-E1 cells. We also show for NIH3T3 that PiT1 overexpression led to faster cell spreading. The final total numbers of attached cells did, however, not differ between cultures of PiT1 overexpressing cells and control cells of neither cell type. We suggest that the PiT1-mediated fast adhesion potentials allow the cells to go faster out of G0/G1 and thereby contribute to their proliferative advantage within the first 24 h after seeding. - Highlights: • Effects of elevated levels of the inorganic phosphate transporter PiT1 were studied. • The density-inhibited murine cell lines NIH3T3 and MC3T3-E1 showed faster adhesion. • NIH3T3 cells showed faster spreading. • We suggest that the faster adhesion/spreading contributes to faster proliferation.

  1. Novel, inorganic composites using porous, alkali-activated, aluminosilicate binders

    NASA Astrophysics Data System (ADS)

    Musil, Sean

    Geopolymers are an inorganic polymeric material composed of alumina, silica, and alkali metal oxides. Geopolymers are chemical and fire resistant, can be used as refractory adhesives, and are processed at or near ambient temperature. These properties make geopolymer an attractive choice as a matrix material for elevated temperature composites. This body of research investigated numerous different reinforcement possibilities and variants of geopolymer matrix material and characterized their mechanical performance in tension, flexure and flexural creep. Reinforcements can then be chosen based on the resulting properties to tailor the geopolymer matrix composites to a specific application condition. Geopolymer matrix composites combine the ease of processing of polymer matrix composites with the high temperature capability of ceramic matrix composites. This study incorporated particulate, unidirectional fiber and woven fiber reinforcements. Sodium, potassium, and cesium based geopolymer matrices were evaluated with cesium based geopolymer showing great promise as a high temperature matrix material. It showed the best strength retention at elevated temperature, as well as a very low coefficient of thermal expansion when crystallized into pollucite. These qualities made cesium geopolymer the best choice for creep resistant applications. Cesium geopolymer binders were combined with unidirectional continuous polycrystalline mullite fibers (Nextel(TM) 720) and single crystal mullite fibers, then the matrix was crystallized to form cubic pollucite. Single crystal mullite fibers were obtained by the internal crystallization method and show excellent creep resistance up to 1400°C. High temperature flexural strength and flexural creep resistance of pollucite and polycrystalline/single-crystal fibers was evaluated at 1000-1400°C.

  2. Functional expression of PHO1 to the Golgi and trans-Golgi network and its role in export of inorganic phosphate.

    PubMed

    Arpat, A Bulak; Magliano, Pasqualina; Wege, Stefanie; Rouached, Hatem; Stefanovic, Aleksandra; Poirier, Yves

    2012-08-01

    Arabidopsis thaliana PHO1 is primarily expressed in the root vascular cylinder and is involved in the transfer of inorganic phosphate (Pi) from roots to shoots. To analyze the role of PHO1 in transport of Pi, we have generated transgenic plants expressing PHO1 in ectopic A. thaliana tissues using an estradiol-inducible promoter. Leaves treated with estradiol showed strong PHO1 expression, leading to detectable accumulation of PHO1 protein. Estradiol-mediated induction of PHO1 in leaves from soil-grown plants, in leaves and roots of plants grown in liquid culture, or in leaf mesophyll protoplasts, was all accompanied by the specific release of Pi to the extracellular medium as early as 2-3 h after addition of estradiol. Net Pi export triggered by PHO1 induction was enhanced by high extracellular Pi and weakly inhibited by the proton-ionophore carbonyl cyanide m-chlorophenylhydrazone. Expression of a PHO1-GFP construct complementing the pho1 mutant revealed GFP expression in punctate structures in the pericycle cells but no fluorescence at the plasma membrane. When expressed in onion epidermal cells or in tobacco mesophyll cells, PHO1-GFP was associated with similar punctate structures that co-localized with the Golgi/trans-Golgi network and uncharacterized vesicles. However, PHO1-GFP could be partially relocated to the plasma membrane in leaves infiltrated with a high-phosphate solution. Together, these results show that PHO1 can trigger Pi export in ectopic plant cells, strongly indicating that PHO1 is itself a Pi exporter. Interestingly, PHO1-mediated Pi export was associated with its localization to the Golgi and trans-Golgi networks, revealing a role for these organelles in Pi transport.

  3. Boosting Proton Conductivity in Highly Robust 3D Inorganic Cationic Extended Frameworks through Ion Exchange with Dihydrogen Phosphate Anions.

    PubMed

    Xiao, Chengliang; Wang, Yaxing; Chen, Lanhua; Yin, Xuemiao; Shu, Jie; Sheng, Daopeng; Chai, Zhifang; Albrecht-Schmitt, Thomas E; Wang, Shuao

    2015-12-01

    The limited long-term hydrolytic stability of rapidly emerging 3D-extended framework materials (MOFs, COFs, MOPs, etc.) is still one of major barriers for their practical applications as new solid-state electrolytes in fuel cells. To obtain hydrolytically stable materials, two H2 PO4 (-) -exchanged 3D inorganic cationic extended frameworks (CEFs) were successfully prepared by a facile anion-exchange method. Both anion-exchanged CEFs (YbO(OH)P and NDTBP) show significantly enhanced proton conductivity when compared with the original materials (YbO(OH)Cl and NDTB) with an increase of up to four orders-of-magnitude, reaching 2.36×10(-3) and 1.96×10(-2)  S cm(-1) at 98 % RH and 85 °C for YbO(OH)P and NDTBP, respectively. These values are comparable to the most efficient proton-conducting MOFs. In addition, these two anion-exchanged materials are stable in boiling water, which originates from the strong electrostatic interaction between the H2 PO4 (-) anion and the cationic host framework, showing a clear advance over all the acid-impregnated materials (H2 SO4 @MIL-101, H3 PO4 @MIL-101, and H3 PO4 @Tp-Azo) as practical solid-state fuel-cell electrolytes. This work offers a new general and efficient approach to functionalize 3D-extended frameworks through an anion-exchange process and achieves water-stability with ultra-high proton conductivity above 10(-2)  S cm(-1) .

  4. Compound- and enzyme-specific phosphodiester hydrolysis mechanisms revealed by δ18O of dissolved inorganic phosphate: Implications for marine P cycling

    NASA Astrophysics Data System (ADS)

    Liang, Yuhong; Blake, Ruth E.

    2009-07-01

    We have studied the oxygen isotope signature of inorganic phosphate (P i) generated by hydrolysis of nucleic acid phosphodiester (P-diester) compounds by cell-free enzymes (Deoxyribonuclease 1, Phosphodiesterase 1, Alkaline phosphatase) and microbial cultures at natural isotopic abundances. We demonstrate that the diesterase-catalyzed hydrolytic step leads to incorporation of at least one water O into released P i for a total of two O atoms from water incorporated into P i released from P-diesters. In the presence of Phosphodiesterase 1, 16O is preferentially incorporated into nucleotides released from DNA; whereas 18O is preferentially incorporated into nucleotides released from RNA. A strong consistency between predicted O-isotope regeneration signatures based on results of cell-free enzyme experiments and measured isotopic signatures from independent experiments with E. coli cultures was observed and confirms proposed models for phosphoester hydrolysis. Results from these studies made at natural 18O abundance levels provide a new tool, enzyme-specific O-isotope fractionation, for investigations of organophosphate metabolism and phosphorus cycling pathways in natural aquatic systems.

  5. Determination of the kinetic parameters for phospholipase C (Bacillus cereus) on different phospholipid substrates using a chromogenic assay based on the quantitation of inorganic phosphate.

    PubMed

    Hergenrother, P J; Martin, S F

    1997-08-15

    The kinetic parameters of the phosphatidylcholine-preferring phospholipase C from Bacillus cereus (PLCBc) have been evaluated for phosphatidylcholine, phosphatidylethanolamine, and phosphatidylserine substrates with a new assay based on the quantitation of inorganic phosphate (Pi). Treatment of the phosphomonoester product of the PLCBc-catalyzed hydrolysis of these phospholipids with alkaline phosphatase releases Pi. This Pi forms a complex with ammonium molybdate that is then reduced by ascorbic acid to provide a blue molybdenum chromogen with an absorbance maximum at 700 nm. This highly sensitive assay may be used to determine accurately less than 5 nmol of Pi in solution. Performing the assay in 96-well plates provides a rapid and convenient method to evaluate a variety of phospholipids as substrates for PLCBc. The assay has been utilized to ascertain the kinetic constants for the PLCBc-catalyzed hydrolysis of 1,2-dihexanoyl-sn-glycero-3-phosphocholine, 1,2-dihexanoyl-sn-glycero-3-phosphoethanolamine, and 1,2-dihexanoyl-sn-glycero-3-phospho-L-serine. It is found that these compounds are substrates for the enzyme with their VmaxS being in the order of phosphatidylcholine > phosphatidylethanolamine > phosphatidylserine.

  6. Selective release of inorganic constituents in broiler manure biochars under different post-activation treatments

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Previous studies determined that poultry litter is a desirable feedstock for activated biochars with enhanced adsorption towards cations. Animal manures such as poultry litter contain a significant fraction of inorganic material that can significantly affect the final physical, chemical and adsorpt...

  7. DIFFERENTIAL ACTIVATION OF AP-1 IN HUMAN BLADDER EPITHELIAL CELLS BY INORGANIC AND METHYLATED ARSENICALS

    EPA Science Inventory

    Differential Activation of AP-1 in Human Bladder Epithelial Cells by Inorganic and Methylated Arsenicals

    Zuzana Drobna, Ilona Jaspers, David J. Thomas, and Miroslav Styblo

    ABSTRACT

    Epidemiological studies have linked chronic ingestion of drinking water contai...

  8. Protein-inorganic hybrid nanoflowers

    NASA Astrophysics Data System (ADS)

    Ge, Jun; Lei, Jiandu; Zare, Richard N.

    2012-07-01

    Flower-shaped inorganic nanocrystals have been used for applications in catalysis and analytical science, but so far there have been no reports of `nanoflowers' made of organic components. Here, we report a method for creating hybrid organic-inorganic nanoflowers using copper (II) ions as the inorganic component and various proteins as the organic component. The protein molecules form complexes with the copper ions, and these complexes become nucleation sites for primary crystals of copper phosphate. Interaction between the protein and copper ions then leads to the growth of micrometre-sized particles that have nanoscale features and that are shaped like flower petals. When an enzyme is used as the protein component of the hybrid nanoflower, it exhibits enhanced enzymatic activity and stability compared with the free enzyme. This is attributed to the high surface area and confinement of the enzymes in the nanoflowers.

  9. A thermodynamic model of mixed organic-inorganic aerosols to predict activity coefficients

    NASA Astrophysics Data System (ADS)

    Zuend, A.; Marcolli, C.; Luo, B. P.; Peter, T.

    2008-08-01

    Tropospheric aerosols contain mixtures of inorganic salts, acids, water, and a large variety of organic compounds. Interactions between these substances in liquid mixtures lead to discrepancies from ideal thermodynamic behaviour. By means of activity coefficients, non-ideal behaviour can be taken into account. We present here a thermodynamic model named AIOMFAC (Aerosol Inorganic-Organic Mixtures Functional groups Activity Coefficients) that is able to calculate activity coefficients covering inorganic, organic, and organic-inorganic interactions in aqueous solutions over a wide concentration range. This model is based on the activity coefficient model LIFAC by Yan et al. (1999) that we modified and reparametrised to better describe atmospherically relevant conditions and mixture compositions. Focusing on atmospheric applications we considered H+, Li+, Na+, K+, NH+4, Mg2+, Ca2+, Cl-, Br-, NO-3, HSO-4, and SO2-4 as cations and anions and a wide range of alcohols/polyols composed of the functional groups CHn and OH as organic compounds. With AIOMFAC, the activities of the components within an aqueous electrolyte solution are well represented up to high ionic strength. Most notably, a semi-empirical middle-range parametrisation of direct organic-inorganic interactions in alcohol+water+salt solutions strongly improves the agreement between experimental and modelled activity coefficients. At room temperature, this novel thermodynamic model offers the possibility to compute equilibrium relative humidities, gas/particle partitioning and liquid-liquid phase separations with high accuracy. In further studies, other organic functional groups will be introduced. The model framework is not restricted to specific ions or organic compounds and is therefore also applicable for other research topics.

  10. A thermodynamic model of mixed organic-inorganic aerosols to predict activity coefficients

    NASA Astrophysics Data System (ADS)

    Zuend, A.; Marcolli, C.; Luo, B. P.; Peter, Th.

    2008-03-01

    Tropospheric aerosols contain mixtures of inorganic salts, acids, water, and a large variety of organic compounds. Interactions between these substances in liquid mixtures lead to discrepancies from ideal thermodynamic behaviour. By means of activity coefficients, non-ideal behaviour can be taken into account. We present here a thermodynamic model named AIOMFAC (Aerosol Inorganic-Organic Mixtures Functional groups Activity Coefficients) that is able to calculate activity coefficients covering inorganic, organic, and organic-inorganic interactions in aqueous solutions over a wide concentration range. This model is based on the activity coefficient model LIFAC by Yan et al. (1999) that we modified and reparametrised to better describe atmospherically relevant conditions and mixture compositions. Focusing on atmospheric applications we considered H+, Li+, Na+, K+, NH4+, Mg2+, Ca2+, Cl-, Br-, NO3-, HSO4-, and SO42- as cations and anions and a wide range of alcohols/polyols composed of the functional groups CHn and OH as organic compounds. With AIOMFAC, the activities of the components within an aqueous electrolyte solution are well represented up to high ionic strength. Most notably, a semi-empirical middle-range parametrisation of direct organic-inorganic interactions in alcohol + water + salt solutions strongly improves the agreement between experimental and modelled activity coefficients. At room temperature, this novel thermodynamic model offers the possibility to compute equilibrium relative humidities, gas/particle partitioning and liquid-liquid phase separations with high accuracy. In further studies, other organic functional groups will be introduced. The model framework is not restricted to specific ions or organic compounds and is therefore also applicable for other research topics.

  11. Fluoride-containing bioactive glasses inhibit pentose phosphate oxidative pathway and glucose 6-phosphate dehydrogenase activity in human osteoblasts.

    PubMed

    Bergandi, Loredana; Aina, Valentina; Garetto, Stefano; Malavasi, Gianluca; Aldieri, Elisabetta; Laurenti, Enzo; Matera, Lina; Morterra, Claudio; Ghigo, Dario

    2010-02-12

    Bioactive glasses such as Hench's 45S5 (Bioglass) have applications to tissue engineering as well as bone repair, and the insertion of fluoride in their composition has been proposed to enhance their bioactivity. In view of a potential clinical application, we investigated whether fluoride-containing glasses exert toxic effects on human MG-63 osteoblasts, and whether and how fluoride, which is released in the cell culture medium, might play a role in such cytotoxicity. A 24h incubation with 50 microg/ml (12.5 microg/cm(2)) of fluoride-containing bioactive glasses termed HCaCaF(2) (F content: 5, 10 and 15 mol.%) caused the release of lactate dehydrogenase in the extracellular medium (index of cytotoxicity), the accumulation of intracellular malonyldialdehyde (index of lipoperoxidation), and the increase of glutathione consumption. Furthermore, fluoride-containing glasses inhibited the pentose phosphate oxidative pathway and the glucose 6-phosphate dehydrogenase activity. These effects are ascribable to the fluoride content/release of glass powders, since they were mimicked by NaF solutions and were prevented by dimethyl sulfoxide and tempol (two radical scavengers), by superoxide dismutase (a superoxide scavenger), and by glutathione (the most important intracellular antioxidant molecule), but not by apocynin (an inhibitor of NADPH oxidase). The presence of fluoride-containing glasses and NaF caused also the generation of reactive oxygen species, which was prevented by superoxide dismutase and catalase. The data suggest that fluoride released from glasses is the cause of MG-63 cell oxidative damage and is independent of NADPH oxidase activation. Our data provide a new mechanism to explain F(-) ions toxicity: fluoride could trigger, at least in part, an oxidative stress via inhibition of the pentose phosphate oxidative pathway and, in particular, through the oxidative inhibition of glucose 6-phosphate dehydrogenase.

  12. Lithium Iron Phosphate Cell Performance Evaluations for Lunar Extravehicular Activities

    NASA Technical Reports Server (NTRS)

    Reid, Concha

    2007-01-01

    Lithium-ion battery cells are being evaluated for their ability to provide primary power and energy storage for NASA s future Exploration missions. These missions include the Orion Crew Exploration Vehicle, the Ares Crew Launch Vehicle Upper Stage, Extravehicular Activities (EVA, the advanced space suit), the Lunar Surface Ascent Module (LSAM), and the Lunar Precursor and Robotic Program (LPRP), among others. Each of these missions will have different battery requirements. Some missions may require high specific energy and high energy density, while others may require high specific power, wide operating temperature ranges, or a combination of several of these attributes. EVA is one type of mission that presents particular challenges for today s existing power sources. The Portable Life Support System (PLSS) for the advanced Lunar surface suit will be carried on an astronaut s back during eight hour long sorties, requiring a lightweight power source. Lunar sorties are also expected to occur during varying environmental conditions, requiring a power source that can operate over a wide range of temperatures. Concepts for Lunar EVAs include a primary power source for the PLSS that can recharge rapidly. A power source that can charge quickly could enable a lighter weight system that can be recharged while an astronaut is taking a short break. Preliminary results of Al23 Ml 26650 lithium iron phosphate cell performance evaluations for an advanced Lunar surface space suit application are discussed in this paper. These cells exhibit excellent recharge rate capability, however, their specific energy and energy density is lower than typical lithium-ion cell chemistries. The cells were evaluated for their ability to provide primary power in a lightweight battery system while operating at multiple temperatures.

  13. Soil Inorganic Carbon in Deserts: Active Carbon Sink or Inert Reservoir?

    NASA Astrophysics Data System (ADS)

    Monger, H. C.; Cole, D. R.

    2011-12-01

    Soil inorganic carbon is the third largest C pool in the active global carbon cycle, containing at least 800 petagrams of carbon. Although carbonate dissolution-precipitation reactions have been understood for over a century, the role of soil inorganic carbon in carbon sequestration, and in particular pedogenic carbonate, is a deceptively complex process because it involves interdependent connections among climate, plants, microorganisms, silicate minerals, soil moisture, pH, and Ca supply via rain, dust, or in situ weathering. An understanding of soil inorganic carbon as a sink or reservoir also requires examination of the system at local to continental scales and at seasonal to millennial time scales. In desert soils studied in North America, carbon isotope ratios and radiocarbon dates were measured in combination with electron microscopy, lab and field experiments with biological calcite formation, and field measurements of carbon dioxide emissions. These investigations reveal that soil inorganic carbon is both an active sink and a inert reservoir depending on the spatial and temporal scale and source of calcium.

  14. Nostoc commune UTEX 584 gene expressing indole phosphate hydrolase activity in Escherichia coli.

    PubMed Central

    Xie, W Q; Whitton, B A; Simon, J W; Jäger, K; Reed, D; Potts, M

    1989-01-01

    A gene encoding an enzyme capable of hydrolyzing indole phosphate was isolated from a recombinant gene library of Nostoc commune UTEX 584 DNA in lambda gt10. The gene (designated iph) is located on a 2.9-kilobase EcoRI restriction fragment and is present in a single copy in the genome of N. commune UTEX 584. The iph gene was expressed when the purified 2.9-kilobase DNA fragment, free of any vector sequences, was added to a cell-free coupled transcription-translation system. A polypeptide with an Mr of 74,000 was synthesized when the iph gene or different iph-vector DNA templates were expressed in vitro. When carried by different multicopy plasmids and phagemids (pMP005, pBH6, pB8) the cyanobacterial iph gene conferred an Iph+ phenotype upon various strains of Escherichia coli, including a phoA mutant. Hydrolysis of 5-bromo-4-chloro-3-indolyl phosphate was detected in recombinant E. coli strains grown in phosphate-rich medium, and the activity persisted in assay buffers that contained phosphate. In contrast, indole phosphate hydrolase activity only developed in cells of N. commune UTEX 584, when they were partially depleted of phosphorus, and the activity associated with these cells was suppressed partially by the addition of phosphate to assay buffers. Indole phosphate hydrolase activity was detected in periplasmic extracts from E. coli (Iph+) transformants. Images PMID:2536677

  15. alpha-Tocopheryl phosphate – an active lipid mediator?

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The vitamin E (alpha-tocopherol, alphaT) derivative, alpha-tocopheryl phosphate (alphaTP), is detectable in small amounts in plasma, tissues, and cultured cells. Studies done in vitro and in vivo suggest that alphaT can become phosphorylated and alphaTP dephosphorylated, suggesting the existence of ...

  16. Joint effect of organic acids and inorganic salts on cloud droplet activation

    NASA Astrophysics Data System (ADS)

    Frosch, M.; Prisle, N. L.; Bilde, M.; Varga, Z.; Kiss, G.

    2010-07-01

    We have investigated CCN properties of internally mixed particles composed of one organic acid (oxalic acid, succinic acid, adipic acid, citric acid, cis-pinonic acid, or nordic reference fulvic acid) and one inorganic salt (sodium chloride or ammonium sulphate). Surface tension and water activity of aqueous model solutions with concentrations relevant for CCN activation were measured using a tensiometer and osmometry, respectively. The measurements were used to calculate Köhler curves, which were compared to measured critical supersaturations of particles with the same chemical compositions, determined with a cloud condensation nucleus counter. Surfactant surface partitioning was not accounted for. For the mixtures containing cis-pinonic acid or fulvic acid, a depression of surface tension was observed, but for the remaining mixtures the effect on surface tension was negligle at concentrations relevant for cloud droplet activation, and water activity was the more significant term in the Köhler equation. The surface tension depression of aqueous solutions containing both organic acid and inorganic salt was approximately the same as or smaller than that of aqueous solutions containing the same mass of the corresponding pure organic acids. Water activity was found to be highly dependent on the type and amount of inorganic salt. Sodium chloride was able to decrease water activity more than ammonium sulphate and both inorganic compounds had a higher effect on water activity than the studied organic acids, and increasing the mass ratio of the inorganic compound led to a decrease in water activity. Water activity measurements were compared to results from the E-AIM model and values estimated from both constant and variable van't Hoff factors to evaluate the performance of these approaches. The correspondence between measuments and estimates was overall good, except for highly concentrated solutions. Critical supersaturations calculated with Köhler theory based on

  17. Requirement of a soluble intracellular factor for activation of transient receptor potential A1 by pungent chemicals: role of inorganic polyphosphates.

    PubMed

    Kim, Donghee; Cavanaugh, Eric J

    2007-06-13

    Pungent chemicals such as allyl isothiocyanate (AITC), cinnamaldehyde, and allicin, produce nociceptive sensation by directly activating transient receptor potential A1 (TRPA1) expressed in sensory afferent neurons. In this study, we found that pungent chemicals added to the pipette or bath solution easily activated TRPA1 in cell-attached patches but failed to do so in inside-out or outside-out patches. Thus, a soluble cytosolic factor was required to activate TRPA1. N-Ethylmaleimide, (2-aminoethyl)-methane thiosulfonate, 2-aminoethoxydiphneyl borate, and trinitrophenol, compounds that are known to activate TRPA1, also failed to activate it in inside-out patches. To identify a factor that supports activation of TRPA1 by pungent chemicals, we screened approximately 30 intracellular molecules known to modulate ion channels. Among them, pyrophosphate (PPi) and polytriphosphate (PPPi) were found to support activation of TRPA1 by pungent chemicals. Structure-function studies showed that inorganic polyphosphates (polyP(n), where n = number of phosphates) with at least four phosphate groups were highly effective (polyP4 approximately = polyP65 approximately = polyP45 approximately = polyP25 > PPPi > PPi), with K(1/2) values ranging from 0.2 to 2.8 mM. Inositol-trisphosphate and inositol-hexaphosphate also partially supported activation of TRPA1 by AITC. ATP, GTP, and phosphatidylinositol-4,5-bisphosphate that have three phosphate groups did not support TRPA1 activation. TRPA1 recorded from cell bodies of trigeminal ganglion neurons showed similar behavior with respect to sensitivity to pungent chemicals; no activation was observed in inside-out patches unless a polyphosphate was present. These results show that TRPA1 requires an intracellular factor to adopt a functional conformation that is sensitive to pungent chemicals and suggest that polyphosphates may partly act as such a factor.

  18. Temperature Dependence of NMR Relaxation Times of Nucleoside Triphosphates and Inorganic Phosphate in the Isolated Perfused Rat Liver. Effect on Pi Compartmentation

    NASA Astrophysics Data System (ADS)

    Dufour, Sylvie; Thiaudière, Eric; Vidal, Giovanni; Gallis, Jean-Louis; Rousse, Nicole; Canioni, Paul

    1996-11-01

    The effect of temperature on31P NMR spectra from isolated perfused rat livers was studied at 9.4 T. Relaxation times (T1andT2) of nucleoside triphosphates (NTP) and inorganic phosphate (Pi) were determined at 37, 25, 15, and 4°C. Under hypothermic conditions, an unexpected apparent line sharpening in the Pi spectral region and a clear emergence of an additional Pi resonance were observed. This additional signal was assigned to mitochondrial Pi.T1values obtained for cytosolic and mitochondrial Pi at 4°C were 1.14 ± 0.24 s (n= 5) and 0.71 ± 0.18 s (n= 5), respectively. No significant mitochondrial contribution to the Pi resonance was observed at 37°C. Quantification of Pi and NTP liver contents at 37 and 4°C was performed by comparing the perfused liver spectrum and the corresponding perchloric acid extract spectrum. Under experimental conditions of low external Pi (0.12 mM), it was concluded that intracellular Pi was completely NMR-visible at 4 and 37°C. The observation of the mitochondrial Pi signal at 4°C was well explained by an increase in the Pi level within the matrix, in response to the mitochondrial swelling induced by hypothermia, as observed by electron microscopy.T2values for the cytosolic Pi at 37 and 4°C were 17 ± 4 ms (n= 8) and 22 ± 4 ms (n= 10), respectively. Comparison with measured linewidths indicated that line broadening for the main phosphorylated metabolites-including matrix Pi-was the result ofB0field inhomogeneity. The additional broadening of the cytosolic Pi resonance at 4 and 37°C was attributed to pH heterogeneity within the liver.

  19. Polycarboxylic acids as network modifiers for water durability improvement of inorganic-organic hybrid tin-silico-phosphate low-melting glasses

    SciTech Connect

    Menaa, Bouzid . E-mail: bouzidmenaa@noncry.kuicr.kyoto-u.ac.jp; Mizuno, Megumi; Takahashi, Masahide . E-mail: masahide@noncry.kuicr.kyoto-u.ac.jp; Tokuda, Yomei; Yoko, Toshinobu

    2006-02-15

    We investigated the water durability of the inorganic-organic hybrid tin-silico-phosphate glasses Me{sub 2}SiO-SnO-P{sub 2}O{sub 5} (Me designs the organic methyl group) doped with organic acids (salicylic acid (SA), tartaric acid (TA), citric acid (Canada) and butane tetracarboxylic acid (BTCA)) containing one or more of carboxylic groups per molecule. The structure, thermal properties and durability of the final glasses obtained via a non-aqueous acid-base reaction were discussed owing to the nature and the concentration of the acid added. {sup 29}Si magic angle spinning (MAS) NMR and {sup 31}P MAS NMR spectra, respectively, showed clearly a modification of the network in the host glass matrix of the Me{sub 2}SiO-SnO-P{sub 2}O{sub 5} system. The polycondensation enhancement to form -P-O-Si-O-P- linkages (PSP) and the increase of the Q {sup 2} unit (two bridging oxygens per phosphorus atom) over the Q {sup 3} unit (three bridging oxygens per phosphorus atom) as a function of the acid in the order SA

  20. Joint effect of organic acids and inorganic salts on cloud droplet activation

    NASA Astrophysics Data System (ADS)

    Frosch, M.; Prisle, N. L.; Bilde, M.; Varga, Z.; Kiss, G.

    2011-04-01

    We have investigated CCN properties of internally mixed particles composed of one organic acid (oxalic acid dihydrate, succinic acid, adipic acid, citric acid, cis-pinonic acid, or Nordic reference fulvic acid) and one inorganic salt (sodium chloride or ammonium sulphate). Surface tension and water activity of aqueous model solutions with concentrations relevant for CCN activation were measured using a tensiometer and osmometry, respectively. The measurements were used to calculate Köhler curves and critical supersaturations, which were compared to measured critical supersaturations of particles with the same chemical compositions, determined with a cloud condensation nucleus counter. Surfactant surface partitioning was not accounted for. For the aqueous solutions containing cis-pinonic acid and fulvic acid, a depression of surface tension was observed, but for the remaining solutions the effect on surface tension was negligible at concentrations relevant for cloud droplet activation. The surface tension depression of aqueous solutions containing both organic acid and inorganic salt was approximately the same as or smaller than that of aqueous solutions containing the same mass of the corresponding pure organic acids. Water activity was found to be highly dependent on the type and amount of inorganic salt. Sodium chloride was able to decrease water activity more than ammonium sulphate and both inorganic salts are predicted to have a smaller Raoult term than the studied organic acids. Increasing the mass ratio of the inorganic salt led to a decrease in water activity. Water activity measurements were compared to results from the E-AIM model and values estimated from both constant and variable van't Hoff factors. The correspondence between measurements and estimates was overall good, except for highly concentrated solutions. Critical supersaturations calculated with Köhler theory based on measured water activity and surface tension, but not accounting for surface

  1. 2-O-α-D-Glucosylglycerol Phosphorylase from Bacillus selenitireducens MLS10 Possessing Hydrolytic Activity on β-D-Glucose 1-Phosphate

    PubMed Central

    Nihira, Takanori; Saito, Yuka; Ohtsubo, Ken’ichi; Nakai, Hiroyuki; Kitaoka, Motomitsu

    2014-01-01

    The glycoside hydrolase family (GH) 65 is a family of inverting phosphorylases that act on α-glucosides. A GH65 protein (Bsel_2816) from Bacillus selenitireducens MLS10 exhibited inorganic phosphate (Pi)-dependent hydrolysis of kojibiose at the rate of 0.43 s−1. No carbohydrate acted as acceptor for the reverse phosphorolysis using β-d-glucose 1-phosphate (βGlc1P) as donor. During the search for a suitable acceptor, we found that Bsel_2816 possessed hydrolytic activity on βGlc1P with a kcat of 2.8 s−1; moreover, such significant hydrolytic activity on sugar 1-phosphate had not been reported for any inverting phosphorylase. The H218O incorporation experiment and the anomeric analysis during the hydrolysis of βGlc1P revealed that the hydrolysis was due to the glucosyl-transferring reaction to a water molecule and not a phosphatase-type reaction. Glycerol was found to be the best acceptor to generate 2-O-α-d-glucosylglycerol (GG) at the rate of 180 s−1. Bsel_2816 phosphorolyzed GG through sequential Bi-Bi mechanism with a kcat of 95 s−1. We propose 2-O-α-d-glucopyranosylglycerol: phosphate β-d-glucosyltransferase as the systematic name and 2-O-α-d-glucosylglycerol phosphorylase as the short name for Bsel_2816. This is the first report describing a phosphorylase that utilizes polyols, and not carbohydrates, as suitable acceptor substrates. PMID:24466148

  2. Composite block copolymer stabilized nanoparticles: simultaneous encapsulation of organic actives and inorganic nanostructures.

    PubMed

    Gindy, Marian E; Panagiotopoulos, Athanassios Z; Prud'homme, Robert K

    2008-01-01

    We describe the preparation and characterization of hybrid block copolymer nanoparticles (NPs) for use as multimodal carriers for drugs and imaging agents. Stable, water-soluble, biocompatible poly(ethylene glycol)-block-poly(epsilon-caprolactone) NPs simultaneously co-encapsulating hydrophobic organic actives (beta-carotene) and inorganic imaging nanostructures (Au) are prepared using the flash nanoprecipitation process in a multi-inlet vortex mixer. These composite nanoparticles (CNPs) are produced with tunable sizes between 75 nm and 275 nm, narrow particle size distributions, high encapsulation efficiencies, specified component compositions, and long-term stability. The process is tunable and flexible because it relies on the control of mixing and aggregation timescales. It is anticipated that the technique can be applied to a variety of hydrophobic active compounds, fluorescent dyes, and inorganic nanostructures, yielding CNPs for combined therapy and multimodal imaging applications.

  3. Synthesis of organic/inorganic hybrid gel with acid activated clay after γ-ray radiation.

    PubMed

    Kim, Donghyun; Lee, Hoik; Sohn, Daewon

    2014-08-01

    A hybrid gel was prepared from acid activated clay (AA clay) and acrylic acid by gamma ray irradiation. Irradiated inorganic particles which have peroxide groups act as initiator because it generates oxide radicals by increasing temperature. Inorganic nanoparticles which are rigid part in hybrid gel also contribute to increase the mechanical property as a crosslinker. We prepared two hybrid gels to compare the effect of acid activated treatment of clay; one is synthesized with raw clay particles and another is synthesized with AA clay particles. The composition and structure of AA clay particles and raw clay particles were confirmed by X-ray diffraction (XRD), X-ray fluorescence instrument and surface area analyzer. And chemical and physical property of hybrid gel with different ratios of acrylic acid and clay particle was tested by Raman spectroscope and universal testing machine (UTM). The synthesized hydrogel with 76% gel contents can elongated approximately 1000% of its original size.

  4. RNA 3'-terminal phosphate cyclase activity and RNA ligation in HeLa cell extract.

    PubMed Central

    Filipowicz, W; Konarska, M; Gross, H J; Shatkin, A J

    1983-01-01

    HeLa cell extract contains RNA ligase activity that converts linear polyribonucleotides to covalently closed circles. RNA substrates containing 2',3'-cyclic phosphate and 5'-hydroxyl termini are circularized by formation of a normal 3',5' phosphodiester bond. This activity differs from a previously described wheat germ RNA ligase which circularizes molecules with 2',3'-cyclic and 5' phosphate ends by a 2'-phosphomonester, 3',5'-phosphodiester linkage (Konarska et al., Nature 293, 112-116, 1981; Proc. Natl. Acad. Sci. USA 79, 1474-1478, 1982). The HeLa cell ligase can also utilize molecules with 3'-phosphate ends. However, in this case ligation is preceded by an ATP-dependent conversion of the 3'-terminal phosphate to the 2',3' cyclic form by a novel activity, RNA 3'-terminal phosphate cyclase. Both RNA ligase and RNA 3'-terminal phosphate cyclase activities are also present in extract of Xenopus oocyte nuclei, consistent with a role in RNA processing. Images PMID:6828385

  5. Inorganic nitrogen assimilation in yeasts: alteration in enzyme activities associated with changes in cultural conditions and growth phase.

    PubMed

    Thomulka, K W; Moat, A G

    1972-01-01

    Ammonia assimilation has been investigated in four strains of Saccharomyces cerevisiae by measuring, at intervals throughout the growth cycle, the activities of several enzymes concerned with inorganic ammonia assimilation. Enzyme activities in extracts of cells were compared after growth in complete and defined media. The effect of shift from growth in a complete to growth in a defined medium (and the reverse) was also determined. The absence of aspartase (EC 4.3.1.1, l-aspartate-ammonia lyase) activity, the low specific activities of alanine dehydrogenase, glutamine synthetase [EC 6.3.1.2, l-glutamate-ammonia ligase (ADP)], and the marked increase in activity of the nicotinamide adenine dinucleotide phosphate-linked glutamate dehydrogenase (NADP-GDH) [EC 1.4.1.4, l-glutamate:NADP-oxidoreductase (deaminating)] during the early stages of growth support the conclusion that yeasts assimilate ammonia primarily via glutamate. The NADP-GDH showed a rapid increase in activity just before the initiation of exponential growth, reached a maximum at the mid-exponential stage, and then gradually declined in activity in the stationary phase. The NADP-GDH reached a higher level of activity when the yeasts were grown on the defined medium as compared with complete medium. The nicotinamide adenine dinucleotide-linked glutamate dehydrogenase (NAD-GDH) [EC 1.4.1.2, l-glutamate:NAD-oxidoreductase (deaminating)] showed only slight increases in activity during the exponential phase of growth. There was an inverse relationship in that the NADP-GDH increased in activity as the NAD-GDH decreased. The NAD-GDH activity was higher after growth on the complete medium. The glutamate-oxaloacetate transaminase (EC 2.6.1.1. l-aspartate:2-oxoglutarate aminotransferase) activity rose and fell in parallel with the NADP-GDH, although its specific activity was somewhat lower. Although other ammonia-assimilatory enzymes were demonstrable, it seems unlikely that their combined activities could account

  6. Hydrous iron oxide modified diatomite as an active filtration medium for phosphate capture.

    PubMed

    Wang, Zhe; Lin, Yan; Wu, Deyi; Kong, Hainan

    2016-02-01

    A simple method to functionalize diatomite with hydrous iron oxide was attempted and its performance as a new active filtration material to remove and recover phosphate from water was investigated under varying solution conditions. The Langmuir phosphate adsorption capacity increased from 0.6 mgP/g for raw diatomite to 4.89, 14.71, 25.02 mgP/g for hydrous iron oxide modified diatomite (HIOMD), depending on the amount of iron loaded. Loading of hydrous iron oxide caused the increase in true and bulk density and a decline in filtration rate, but to a lesser extent. It was shown that the HIOMD product with suitable iron content could retain a good filtration performance with a greatly increased adsorption capacity for phosphate. The phosphate adsorption increased by decreasing pH and by increasing ionic strength at high pH levels. The adsorption process was interpreted by ligand exchange. Coexisting oxyanions of sulfate, nitrate, citrate, carbonate, silicate and humic acid showed different effects on phosphate fixation but it was presumed that their influence at their concentrations and pH levels commonly encountered in effluent or natural waters was limited, i.e., HIOMD had a reasonably good selectivity. Results in repeated adsorption, desorption and regeneration experiment showed that the adsorbed phosphate could be recovered and the material could be reused after regeneration. The column test showed that HIOMD could be potentially utilized as an adsorption filtration medium for phosphate removal and recovery from water.

  7. Looking for phosphate-accumulating bacteria in activated sludge processes: a multidisciplinary approach.

    PubMed

    Tarayre, Cédric; Charlier, Raphaëlle; Delepierre, Anissa; Brognaux, Alison; Bauwens, Julien; Francis, Frédéric; Dermience, Michaël; Lognay, Georges; Taminiau, Bernard; Daube, Georges; Compère, Philippe; Meers, Erik; Michels, Evi; Delvigne, Frank

    2017-01-29

    Over the past decades, an increasing need in renewable resources has progressively appeared. This trend concerns not only fossil fuels but also mineral resources. Wastewater and sewage sludge contain significant concentrations in phosphate and can be considered as a fertilizer source of the utmost importance. In wastewater treatment plants, the biological uptake of phosphate is performed by a specific microbiota: the phosphate-accumulating organisms. These microorganisms are recovered in sewage sludge. Here, we aimed to investigate the occurrence of phosphate accumulators in four wastewater treatment plants. A 16S metagenetic analysis identified the main bacterial phyla extracted from the aerobic treatment: α-Proteobacteria, β-Proteobacteria, and Sphingobacteria. An enrichment stage was performed to stimulate the specific growth of phosphate-accumulating bacteria in an acetate medium. An analysis of metabolic activities of sulfur and phosphorus highlighted strong modifications related to phosphorus and much less distinguishable effects with sulfur. A solid acetate medium containing 5-Br-4-Cl-3-indolyl phosphate was used to select potential phosphate-accumulating bacteria from the enriched consortia. The positive strains have been found to belong in the genera Acinetobacter, Corynebacterium, and Pseudomonas. Finally, electron microscopy was applied to the strains and allowed to confirm the presence of polyphosphate granules. Some of these bacteria contained granules the size of which exceeded 100 nm.

  8. Intrinsic activation: the relationship between biomass inorganic content and porosity formation during pyrolysis.

    PubMed

    Stratford, James P; Hutchings, Tony R; de Leij, Frans A A M

    2014-05-01

    The utility of pyrolytic carbons is closely related to their porosity and surface area, there is a clear benefit to the development of biomass pyrolysis processes which produce highly porous carbons. The results presented in this work demonstrate that by using biomass precursors with high inorganic content along with specified process conditions, carbons can be consistently produced with specific surface areas between 900 and 1600 m(2)/g. Results from 12 different source materials show that the formation of increased porosity in pyrolytic carbons is strongly associated with the presence of inorganic elements in the precursors including: magnesium, potassium and sulfur. It was found that pyrolysis of macro-algae can produce especially high specific surface area carbons (mean: 1500 m(2)/g), without externally applied activating agents. Using cheap readily available agricultural residues such as oilseed rape straw, pyrolytic carbons can be produced with specific surface areas of around 950 m(2)/g.

  9. The oxygen isotope composition of phosphate released from phytic acid by the activity of wheat and Aspergillus niger phytase

    NASA Astrophysics Data System (ADS)

    Sperber, C. v.; Tamburini, F.; Brunner, B.; Bernasconi, S. M.; Frossard, E.

    2015-03-01

    Phosphorus (P) is an essential nutrient for living organisms. Under P-limiting conditions plants and microorganisms can exude extracellular phosphatases that release inorganic phosphate (Pi) from organic phosphorus compounds (Porg). Phytic acid (IP6) is an important form of Porg in many soils. The enzymatic hydrolysis of IP6 by phytase yields plant available inorganic phosphate (Pi) and less phosphorylated inositol derivates as products. The hydrolysis of organic P-compounds by phosphatases leaves an isotopic imprint on the oxygen isotope composition (δ18O) of released Pi, which might be used to trace P in the environment. This study aims at determining the effect of phytase on the oxygen isotope composition of released Pi. For this purpose, enzymatic assays with histidine acid phytases from wheat and Aspergillus niger were prepared using IP6, adenosine 5'monophosphate (AMP) and glycerophosphate (GPO4) as substrates. For a comparison to the δ18O of Pi released by other extracellular enzymes, enzymatic assays with acid phosphatases from potato and wheat germ with IP6 as substrate were prepared. During the hydrolysis of IP6 by phytase, four Pi are released, and one oxygen atom from water is incorporated into each Pi. This incorporation of oxygen from water into Pi is subject to an apparent inverse isotopic fractionation (ϵ ∼ 6 to 10‰), which is similar to that imparted by acid phosphatase from potato during the hydrolysis of IP6 (ϵ ∼ 7‰) where less than three Pi are released. The incorporation of oxygen from water into Pi during the hydrolysis of AMP and GPO4 by phytase yielded a normal isotopic fractionation (ϵ ∼ -12‰), again similar to values reported for acid phosphatases from potato and wheat germ. We attribute this similarity in ɛ to the same amino acid sequence motif (RHGXRXP) at the active site of these enzymes, which leads to similar reaction mechanisms. We suggest that the striking substrate-dependency of

  10. A Relationship Between Microbial Activity in Soils and Phosphate Levels in Tributaries to Lake Champlain

    NASA Astrophysics Data System (ADS)

    Larose, R.; Lee, S.; Lane, T.

    2015-12-01

    Lake Champlain is a large natural freshwater lake. It forms the western boundary of Vermont and drains over half of the state. It is bordered by the state of New York on its western side and drains to the north into Quebec, Canada. Lake Champlain is the source of fresh drinking water for over quarter of a million people and provides for the livelihoods and recreational opportunities of many well beyond its borders. The health of this lake is important. During the summer month's algae blooms plague the lake. These unsightly growths, which affect other aquatic organisms, are the result of excess phosphate flowing into the lake from many sources. Examining whether there is a relationship between microbial activity in the soils bordering tributaries to Lake Champlain and phosphate levels in those tributaries sheds insight into the origins and paths by which phosphate moves into Lake Champlain. Understanding the how phosphate moves into the water system may assist in mitigation efforts.Total Phosphate levels and Total Suspended Solids were measured in second and third order streams in the Lake Champlain Basin over a three-year period. In addition microbial activity was measured within the toe, bank and upland riparian zone areas of these streams during the summer months. In general in areas showing greater microbial activity in the soil(s) there were increased levels of phosphate in the streams.

  11. Regeneration of spent powdered activated carbon saturated with inorganic ions by cavitation united with ion exchange method.

    PubMed

    Li, Gang; Gao, Hong; Li, Yansheng; Yang, Huixin

    2011-06-01

    Using ion exchange resin as transfer media, regenerate powdered activated carbon (PAC) adsorbed inorganic ions by cavitation to enhance the transfer; we studied how the regeneration time and the mass ratio of resin and PAC influence the regeneration rate respectively through re-adsorption. The result showed that the effective regeneration of PAC saturated with inorganic ions was above 90% using ion exchange resin as media and transfer carrier, the quantity of PAC did not reduced but activated in the process.

  12. Formation of directly mutagenic alpha-hydroxy-N-nitrosopiperidine phosphate ester by near-ultraviolet irradiation of N-nitrosopiperidine in phosphate buffer

    SciTech Connect

    Arimoto, S.; Shimada, H.; Ukawa, S.; Mochizuki, M.; Hayatsu, H. )

    1989-08-15

    Previously we found that direct-acting mutagens can be formed from N-nitrosodialkylamines on exposure to near-ultraviolet light in the presence of phosphates. We have now isolated the mutagenic photoproduct formed from N-nitrosopiperidine and inorganic phosphate and identified its structure as the phosphate ester of alpha-hydroxy-N-nitrosopiperidine. This reaction represents a new, non-enzymatic activation of promutagenic N-nitrosodialkylamines.

  13. Sphingomyelinase D activity in model membranes: structural effects of in situ generation of ceramide-1-phosphate.

    PubMed

    Stock, Roberto P; Brewer, Jonathan; Wagner, Kerstin; Ramos-Cerrillo, Blanca; Duelund, Lars; Jernshøj, Kit Drescher; Olsen, Lars Folke; Bagatolli, Luis A

    2012-01-01

    The toxicity of Loxosceles spider venom has been attributed to a rare enzyme, sphingomyelinase D, which transforms sphingomyelin to ceramide-1-phosphate. The bases of its inflammatory and dermonecrotic activity, however, remain unclear. In this work the effects of ceramide-1-phosphate on model membranes were studied both by in situ generation of this lipid using a recombinant sphingomyelinase D from the spider Loxosceles laeta and by pre-mixing it with sphingomyelin and cholesterol. The systems of choice were large unilamellar vesicles for bulk studies (enzyme kinetics, fluorescence spectroscopy and dynamic light scattering) and giant unilamellar vesicles for fluorescence microscopy examination using a variety of fluorescent probes. The influence of membrane lateral structure on the kinetics of enzyme activity and the consequences of enzyme activity on the structure of target membranes containing sphingomyelin were examined. The findings indicate that: 1) ceramide-1-phosphate (particularly lauroyl ceramide-1-phosphate) can be incorporated into sphingomyelin bilayers in a concentration-dependent manner and generates coexistence of liquid disordered/solid ordered domains, 2) the activity of sphingomyelinase D is clearly influenced by the supramolecular organization of its substrate in membranes and, 3) in situ ceramide-1-phosphate generation by enzymatic activity profoundly alters the lateral structure and morphology of the target membranes.

  14. Polymeric enzyme mimics: catalytic activity of ribose-containing polymers for a phosphate substrate.

    PubMed

    Han, Man Jung; Yoo, Kyung Soo; Kim, Young Heui; Chang, Ji Young

    2003-07-07

    The polymers containing ribose rings: poly(5'-acrylamido-5'-deoxy-1',2'-O-isopropylidene-alpha-D-ribose) (11), poly(5'-acrylamido-5'-deoxy-alpha-D-ribose) (12) and poly(5'-acrylamido-5'-deoxy-1'-O-methyl-D-ribose) (13) were prepared as enzyme mimics. Polymers 12 and 13 with free vic-cis-diol groups catalyzed the hydrolysis of phosphodiester (ethyl p-nitrophenyl phosphate and N-methylpyridinium 4-tert-butylcatechol cyclic phosphate) and phosphomonoester substrates with a rate acceleration of 10 approximately equal to 10(3) compared with the uncatalyzed reaction. They also catalyzed the reverse reactions, i.e., the esterification of phosphomonoester to phosphodiester and the phosphorylation of alcohols with phosphate ions. The catalytic activity was attributable to the vic-cis-diols of riboses on polymer chains, which formed hydrogen bonds with two phosphoryl oxygen atoms of phosphates so as to activate the phosphorus atoms to be attacked by nucleophiles. The catalytic activity was negligible for polymer 11 where vic-cis-diol groups were blocked with isopropylidene groups. The catalytic activity was attributable to the vic-cis-diols of riboses on polymer chains, which formed hydrogen bonds with two phosphoryl oxygen atoms of phosphates so as to activate the phosphorus atoms to be attacked by nucleophiles.

  15. Role of phosphate on stability and catalase mimetic activity of cerium oxide nanoparticles.

    PubMed

    Singh, Ragini; Singh, Sanjay

    2015-08-01

    Cerium oxide nanoparticles (CeNPs) have been recently shown to scavenge reactive oxygen and nitrogen species (ROS and RNS) in different experimental model systems. CeNPs (3+) and CeNPs (4+) have been shown to exhibit superoxide dismutase (SOD) and catalase mimetic activity, respectively. Due to their nanoscale dimension, CeNPs are expected to interact with the components of biologically relevant buffers and medium, which could alter their catalytic properties. We have demonstrated earlier that CeNPs (3+) interact with phosphate and lose the SOD activity. However, very little is known about the interaction of CeNPs (4+) with the phosphate and other anions, predominantly present in biological buffers and their effects on the catalase mimetic-activity of these nanoparticles. In this study, we report that catalase mimetic-activity of CeNPs (4+) is resistant to the phosphate anions, pH changes and composition of cell culture media. Given the abundance of phosphate anions in the biological system, it is likely that internalized CeNPs would be influenced by cytoplasmic and nucleoplasmic concentration of phosphate.

  16. Physicochemical characteristics and sorption capacities of heavy metal ions of activated carbons derived by activation with different alkyl phosphate triesters

    NASA Astrophysics Data System (ADS)

    Wang, Jing; Liu, Hai; Yang, Shaokun; Zhang, Jian; Zhang, Chenglu; Wu, Haiming

    2014-10-01

    Five alkyl phosphate triesters (APTEs), including trimethyl phosphate (TMP), triethyl phosphate (TEP), triisopropyl phosphate (TPP), tributyl phosphate (TBP) and trioctyl phosphate (TOP), were used as activating agents for preparing activated carbons (AC-APTEs) with high surface acidity and metal ion sorption capacity. N2 adsorption/desorption isotherms, surface morphologies, elemental compositions, results of Boehm's titration and sorption capacities of heavy metal ions of the carbons were investigated. AC-APTEs contained much more acidic groups and exhibited much less surface area (<500 m2/g) in comparison with activated carbon (AC-PPA, 1145 m2/g) obtained from phosphoric acid activation. For the AC-APTEs, AC-TOP had the highest surface area (488 m2/g), AC-TMP showed the highest yield (41.1%), and AC-TBP possessed the highest acidic groups (2.695 mmol/g), oxygen content (47.0%) and metal ion sorption capacities (40.1 mg/g for Ni(II) and 53.5 mg/g for Cd(II)). For the carbons, AC-APTEs showed much larger Ni(II) and Cd(II) sorption capacities than AC-PPA, except AC-TPP. The differences of the carbons in the physicochemical and sorption properties suggested surface chemistry of the carbons was the main factor influencing their sorption capacities whereas the pore structure played a secondary role.

  17. Biocalcite, a multifunctional inorganic polymer: Building block for calcareous sponge spicules and bioseed for the synthesis of calcium phosphate-based bone

    PubMed Central

    Schröder, Heinz C; Müller, Werner E G

    2014-01-01

    Summary Calcium carbonate is the material that builds up the spicules of the calcareous sponges. Recent results revealed that the calcium carbonate/biocalcite-based spicular skeleton of these animals is formed through an enzymatic mechanism, such as the skeleton of the siliceous sponges, evolutionarily the oldest animals that consist of biosilica. The enzyme that mediates the calcium carbonate deposition has been identified as a carbonic anhydrase (CA) and has been cloned from the calcareous sponge species Sycon raphanus. Calcium carbonate deposits are also found in vertebrate bones besides the main constituent, calcium phosphate/hydroxyapatite (HA). Evidence has been presented that during the initial phase of HA synthesis poorly crystalline carbonated apatite is deposited. Recent data summarized here indicate that during early bone formation calcium carbonate deposits enzymatically formed by CA, act as potential bioseeds for the precipitation of calcium phosphate mineral onto bone-forming osteoblasts. Two different calcium carbonate phases have been found during CA-driven enzymatic calcium carbonate deposition in in vitro assays: calcite crystals and round-shaped vaterite deposits. The CA provides a new target of potential anabolic agents for treatment of bone diseases; a first CA activator stimulating the CA-driven calcium carbonate deposition has been identified. In addition, the CA-driven calcium carbonate crystal formation can be frozen at the vaterite state in the presence of silintaphin-2, an aspartic acid/glutamic acid-rich sponge-specific protein. The discovery that calcium carbonate crystals act as bioseeds in human bone formation may allow the development of novel biomimetic scaffolds for bone tissue engineering. Na-alginate hydrogels, enriched with biosilica, have recently been demonstrated as a suitable matrix to embed bone forming cells for rapid prototyping bioprinting/3D cell printing applications. PMID:24991497

  18. Practical tethering of vitamin B1 on a silica surface via its phosphate group and evaluation of its activity.

    PubMed

    Vartzouma, Ch; Louloudi, M; Butler, I S; Hadjiliadis, N

    2002-03-07

    A convenient immobilization of thiamine pyrophosphate molecules on a silica surface through the phosphate group is developed, leading to a very active heterogenized biocatalyst for pyruvate decarboxylation.

  19. Mechanisms for solubilization of various insoluble phosphates and activation of immobilized phosphates in different soils by an efficient and salinity-tolerant Aspergillus niger strain An2.

    PubMed

    Li, Xiaolong; Luo, Lijin; Yang, Jinshui; Li, Baozhen; Yuan, Hongli

    2015-03-01

    Mechanisms for solubilization of different types of phosphates and activation of immobilized phosphates in different types of soils by an efficient fungal strain An2 were explored and evaluated in this study. An2 was isolated from a Chinese cabbage rhizosphere soil and identified as Aspergillus niger. It could fast release up to 1722, 2066, and 2356 mg L(-1) of soluble phosphorus (P) from 1 % Ca3(PO4)2, Mg3(PO4)2, and AlPO4 (Ca-P, Mg-P, and Al-P) and 215 and 179 mg L(-1) from 0.5 % FePO4 and rock phosphate (Fe-P and RP), respectively. HPLC assay demonstrated that An2 mainly secreted oxalic acid to solubilize Ca-P, Mg-P, Al-P, and Fe-P whereas secreted tartaric acid to solubilize RP. Furthermore, An2 could tolerate salinity up to 4 % NaCl without impairing its phosphate-solubilizing ability. The simulation experiments validated that An2 was able to effectively activate immobilized phosphates in general calcareous, acidic, as well as saline-alkali soils with high total P content. This study shows new insights into the mechanisms for microbial solubilization of different types of phosphates and supports the future application of strain An2 in different types of soils to effectively activate P for plants.

  20. Partially phosphorylated Pho4 activates transcription of a subset of phosphate-responsive genes.

    PubMed

    Springer, Michael; Wykoff, Dennis D; Miller, Nicole; O'Shea, Erin K

    2003-11-01

    A cell's ability to generate different responses to different levels of stimulus is an important component of an adaptive environmental response. Transcriptional responses are frequently controlled by transcription factors regulated by phosphorylation. We demonstrate that differential phosphorylation of the budding yeast transcription factor Pho4 contributes to differential gene expression. When yeast cells are grown in high-phosphate growth medium, Pho4 is phosphorylated on four critical residues by the cyclin-CDK complex Pho80-Pho85 and is inactivated. When yeast cells are starved for phosphate, Pho4 is dephosphorylated and fully active. In intermediate-phosphate conditions, a form of Pho4 preferentially phosphorylated on one of the four sites accumulates and activates transcription of a subset of phosphate-responsive genes. This Pho4 phosphoform binds differentially to phosphate-responsive promoters and helps to trigger differential gene expression. Our results demonstrate that three transcriptional outputs can be generated by a pathway whose regulation is controlled by one kinase, Pho80-Pho85, and one transcription factor, Pho4. Differential phosphorylation of Pho4 by Pho80-Pho85 produces phosphorylated forms of Pho4 that differ in their ability to activate transcription, contributing to multiple outputs.

  1. Partially Phosphorylated Pho4 Activates Transcription of a Subset of Phosphate-Responsive Genes

    PubMed Central

    Miller, Nicole

    2003-01-01

    A cell's ability to generate different responses to different levels of stimulus is an important component of an adaptive environmental response. Transcriptional responses are frequently controlled by transcription factors regulated by phosphorylation. We demonstrate that differential phosphorylation of the budding yeast transcription factor Pho4 contributes to differential gene expression. When yeast cells are grown in high-phosphate growth medium, Pho4 is phosphorylated on four critical residues by the cyclin–CDK complex Pho80–Pho85 and is inactivated. When yeast cells are starved for phosphate, Pho4 is dephosphorylated and fully active. In intermediate-phosphate conditions, a form of Pho4 preferentially phosphorylated on one of the four sites accumulates and activates transcription of a subset of phosphate-responsive genes. This Pho4 phosphoform binds differentially to phosphate-responsive promoters and helps to trigger differential gene expression. Our results demonstrate that three transcriptional outputs can be generated by a pathway whose regulation is controlled by one kinase, Pho80–Pho85, and one transcription factor, Pho4. Differential phosphorylation of Pho4 by Pho80–Pho85 produces phosphorylated forms of Pho4 that differ in their ability to activate transcription, contributing to multiple outputs. PMID:14624238

  2. A Facile Stable-Isotope Dilution Method for Determination of Sphingosine Phosphate Lyase Activity

    PubMed Central

    Suh, Jung H.; Eltanawy, Abeer; Rangan, Apoorva; Saba, Julie D.

    2015-01-01

    A new technique for quantifying sphingosine phosphate lyase activity in biological samples is described. In this procedure, 2-hydrazinoquinoline is used to convert (2E)-hexadecenal into the corresponding hydrazone derivative to improve ionization efficiency and selectivity of detection. Combined utilization of liquid chromatographic separation and multiple reaction monitoring-mass spectrometry allows for simultaneous quantification of the substrate S1P and product (2E)-hexadecenal. Incorporation of (2E)-d5-hexadecenal as an internal standard improves detection accuracy and precision. A simple one-step derivatization procedure eliminates the need for further extractions. Limits of quantification for (2E)-hexadecenal and sphingosine-1-phosphate are 100 and 50 fmol, respectively. The assay displays a wide dynamic detection range useful for detection of low basal sphingosine phosphate lyase activity in wild type cells, SPL-overexpressing cell lines, and wild type mouse tissues. Compared to current methods, the capacity for simultaneous detection of sphingosine-1-phosphate and (2E)-hexadecenal greatly improves the accuracy of results and shows excellent sensitivity and specificity for sphingosine phosphate lyase activity detection. PMID:26408264

  3. CADMIUM PHOSPHATE GLASS

    DOEpatents

    Carpenter, H.W.; Johnson, P.D.

    1963-04-01

    A method of preparing a cadmium phosphate glass that comprises providing a mixture of solid inorganic compounds of cadmuim and phosphate having vaporizable components and heating the resulting composition to a temperature of at least 850 un. Concent 85% C is presented. (AEC)

  4. Simultaneous removal of ammonium and phosphate by alkaline-activated and lanthanum-impregnated zeolite.

    PubMed

    He, Yinhai; Lin, Hai; Dong, Yingbo; Liu, Quanli; Wang, Liang

    2016-12-01

    Simultaneous ammonium and phosphate removal characteristics and mechanism, as well as the major influencing factors, such as pH, temperature and co-existing ions, onto NaOH-activated and lanthanum-impregnated zeolite (NLZ) were investigated. The phosphate adsorption increases from 0.2 mg g(-1) for natural zeolite up to 8.96 mg g(-1) for NLZ, while only a slight decrease on the ammonium adsorption capacity from 23.9 mg g(-1) for NaOH-activated zeolite to 21.2 mg g(-1) for NLZ was observed. The ammonium and phosphate adsorption showed little pH dependence in the range from pH 3 to 7, while it decreased sharply with the pH increased above pH 7. Adsorption of ammonium and phosphate could be well described by the pseudo-second-order model and the process was mainly governed by intra-particle diffusion. The Langmuir and Freundlich model can be acceptably applied to fit the experimental data, which suggested that adsorption was caused by both the monolayer and homogeneous coverage at specific and equal affinity sites available NLZ. The underlying mechanism for the specific adsorption of phosphate by NLZ was revealed with the aid of SEM-EDS, XPS, and FTIR analysis, and the formation of (LaO)(OH)PO2 was verified to be the dominant pathway for selective phosphate adsorption by lanthanum-impregnated zeolite. While the removal mechanism of ammonium could be well interpreted by SEM-EDS, FTIR and ICP analysis, and ion-exchange was expected to be the main removal process for ammonium. The results indicate that NLZ could efficiently and simultaneously remove low concentration of ammonium and phosphate from contaminated waters.

  5. Synthesis, characterization and osteoblastic activity of polycaprolactone nanofibers coated with biomimetic calcium phosphate.

    PubMed

    Mavis, Bora; Demirtaş, Tolga T; Gümüşderelioğlu, Menemşe; Gündüz, Güngör; Colak, Uner

    2009-10-01

    Immersion of electrospun polycaprolactone (PCL) nanofiber mats in calcium phosphate solutions similar to simulated body fluid resulted in deposition of biomimetic calcium phosphate layer on the nanofibers and thus a highly bioactive novel scaffold has been developed for bone tissue engineering. Coatings with adequate integrity, favorable chemistry and morphology were achieved in less than 6h of immersion. In the coating solutions, use of lower concentrations of phosphate sources with respect to the literature values (i.e., 3.62 vs. 10 mM) was substantiated by a thermodynamic modeling approach. Recipe concentration combinations that were away from the calculated dicalcium phosphate phase stability region resulted in micron-sized calcium phosphates with native nanostructures. While the nano/microstructure formed by the deposited calcium phosphate layer is controlled by increasing the solution pH to above 6.5 and increasing the duration of immersion experimentally, the nanostructure imposed by the dimensions of the fibers was controlled by the polymer concentration (12% w/v), applied voltage (25 kV) and capillary tip to collector distance (35 cm). The deposited coating increased quantitatively by extending the soak up to 6h. On the other hand, the porosity values attained in the scaffolds were around 87% and the biomimetic coatings did not alter the nanofiber mat porosities negatively since the deposition continued along the fibers after the first 2h. Upon confirming the non-toxic nature of the electrospun PCL nanofiber mats, the effects of different nano/microstructures formed were evaluated by the osteoblastic activity. The levels of both alkaline phosphatase activity and osteocalcin were found to be higher in the coated PCL nanofibers than in the uncoated PCL nanofibers, indicating that biomimetic calcium phosphate on PCL nanofibers supports osteoblastic differentiation.

  6. Enhanced ozone loss by active inorganic bromine chemistry in the tropical troposphere

    NASA Astrophysics Data System (ADS)

    Le Breton, Michael; Bannan, Thomas J.; Shallcross, Dudley E.; Khan, M. Anwar; Evans, Mathew J.; Lee, James; Lidster, Richard; Andrews, Stephen; Carpenter, Lucy J.; Schmidt, Johan; Jacob, Daniel; Harris, Neil R. P.; Bauguitte, Stephane; Gallagher, Martin; Bacak, Asan; Leather, Kimberley E.; Percival, Carl J.

    2017-04-01

    Bromine chemistry, particularly in the tropics, has been suggested to play an important role in tropospheric ozone loss although a lack of measurements of active bromine species impedes a quantitative understanding of its impacts. Recent modelling and measurements of bromine monoxide (BrO) by Wang et al. (2015) have shown current models under predict BrO concentrations over the Pacific Ocean and allude to a missing source of BrO. Here, we present the first simultaneous aircraft measurements of atmospheric bromine monoxide, BrO (a radical that along with atomic Br catalytically destroys ozone) and the inorganic Br precursor compounds HOBr, BrCl and Br2 over the Western Pacific Ocean from 0.5 to 7 km. The presence of 0.17-1.64 pptv BrO and 3.6-8 pptv total inorganic Br from these four species throughout the troposphere causes 10-20% of total ozone loss, and confirms the importance of bromine chemistry in the tropical troposphere; contributing to a 6 ppb decrease in ozone levels due to halogen chemistry. Observations are compared with a global chemical transport model and find that the observed high levels of BrO, BrCl and HOBr can be reconciled by active multiphase oxidation of halide (Br- and Cl-) by HOBr and ozone in cloud droplets and aerosols. Measurements indicate that 99% of the instantaneous free Br in the troposphere up to 8 km originates from inorganic halogen photolysis rather than from photolysis of organobromine species.

  7. Characteristics of Zinc Phosphate Coating Activated by Different Concentrations of Nickel Acetate Solution

    NASA Astrophysics Data System (ADS)

    Abdalla, Khalid; Zuhailawati, H.; Rahmat, Azmi; Azizan, A.

    2017-02-01

    Activation pretreatment with nickel acetate solution at various concentrations was performed prior to the phosphating step to enhance the corrosion resistance of carbon steel substrates. The activation solution was studied over various concentrations: 10, 50, and 100 g/L. The effects of these concentrations on surface characteristics and microstructural evolution of the coated samples were characterized by scanning electron microscopy and energy-dispersive spectroscopy. The electrochemical behavior was evaluated using potentiodynamic polarization curves, electrochemical impedance spectroscopy, and immersion test in a 3.5 pct NaCl solution. Significant increases in the nucleation sites and surface coverage of zinc phosphate coating were observed as the concentration of activation solution reached 50 g/L. The electrochemical analysis revealed that the activation treatment with 50 g/L nickel acetate solution significantly improved the protection ability of the zinc phosphate coating. The corrosion current density of activated phosphate coating with 50 g/L was reduced by 64.64 and 13.22 pct, compared to the coatings obtained with activation solutions of 10 and 100 g/L, respectively.

  8. Synthesis and characterization of inorganic polymers from the alkali activation of an aluminosilicate

    NASA Astrophysics Data System (ADS)

    González, C. P.; Montaño, A. M.; González, A. K.; Ríos, C. A.

    2014-06-01

    This paper presents the results of the synthesis and characterization of inorganic polymers (IP) from aluminosilicates: bentonite (BT) and pumice (PP). The synthesis of IP, was carried out by two methods involving alkaline activation, at room temperature and 80 ± 5 °C, using as activating agent sodium silicate both commercial and analytical (Na2SiO3). Sodium hydroxide (NaOH) at 3 M, 7 M and 12 M was added. A lower degree of polymerization was obtained by using analytical precursors subjected to room temperature and 80 ± 5°C. Replacement of heating by the use of the commercial activating agent with greater alkalinity allows the formation of a 3D network. The materials were structurally characterized by FTIR spectroscopy with Attenuated Reflectance (ATR), Scanning Electron Microscope (SEM) and X -ray diffraction (DRX).

  9. Immobilization of inorganic pyrophosphatase on nanodiamond particles retaining its high enzymatic activity.

    PubMed

    Rodina, Elena V; Valueva, Anastasiya V; Yakovlev, Ruslan Yu; Vorobyeva, Nataliya N; Kulakova, Inna I; Lisichkin, Georgy V; Leonidov, Nikolay B

    2015-12-21

    Nanodiamond (ND) particles are popular platforms for the immobilization of molecular species. In the present research, enzyme Escherichia coli inorganic pyrophosphatase (PPase) was immobilized on detonation ND through covalent or noncovalent bonding and its enzymatic activity was characterized. Factors affecting adsorption of PPase such as ND size and surface chemistry were studied. The obtained material is a submicron size association of ND particles and protein molecules in approximately equal amounts. Both covalently and noncovalently immobilized PPase retains a significant enzymatic activity (up to 95% of its soluble form) as well as thermostability. The obtained hybrid material has a very high enzyme loading capacity (∼1 mg mg(-1)) and may be considered as a promising delivery system of biologically active proteinaceous substances, particularly in the treatment of diseases such as calcium pyrophosphate crystal deposition disease and related pathologies. They can also be used as recoverable heterogeneous catalysts in the traditional uses of PPase.

  10. Microbial Group Specific Uptake Kinetics of Inorganic Phosphate and Adenosine-5′-Triphosphate (ATP) in the North Pacific Subtropical Gyre

    PubMed Central

    Björkman, Karin; Duhamel, Solange; Karl, David M.

    2012-01-01

    We investigated the concentration dependent uptake of inorganic phosphate (Pi) and adenosine-5′-triphosphate (ATP) in microbial populations in the North Pacific Subtropical Gyre (NPSG). We used radiotracers to measure substrate uptake into whole water communities, differentiated microbial size classes, and two flow sorted groups; Prochlorococcus (PRO) and non-pigmented bacteria (NPB). The Pi concentrations, uptake rates, and Pi pool turnover times (Tt) were (mean, ±SD); 54.9 ± 35.0 nmol L−1 (n = 22), 4.8 ± 1.9 nmol L−1 day−1 (n = 19), and 14.7 ± 10.2 days (n = 19), respectively. Pi uptake into >2 μm cells was on average 12 ± 7% (n = 15) of the total uptake. The kinetic response to Pi (10–500 nmol L−1) was small, indicating that the microorganisms were close to their maximum uptake velocity (Vmax). Vmax averaged 8.0 ± 3.6 nmol L−1 day−1 (n = 19) in the >0.2 μm group, with half saturation constants (Km) of 40 ± 28 nmol L−1 (n = 19). PRO had three times the cell specific Pi uptake rate of NPB, at ambient concentrations, but when adjusted to cells L−1 the rates were similar, and these two groups were equally competitive for Pi. The Tt of γ-P-ATP in the >0.2 μm group were shorter than for the Pi pool (4.4 ± 1.0 days; n = 6), but this difference diminished in the larger size classes. The kinetic response to ATP was large in the >0.2 μm class with Vmax exceeding the rates at ambient concentrations (mean 62 ± 27 times; n = 6) with a mean Vmax for γ-P-ATP of 2.8 ± 1.0 nmol L−1 day−1, and Km at 11.5 ± 5.4 nmol L−1 (n = 6). The NPB contribution to γ-P-ATP uptake was high (95 ± 3%, n = 4) at ambient concentrations but decreased to ∼50% at the highest ATP amendment. PRO had Km values 5–10 times greater than NPB. The above indicates that PRO and NPB were in close competition in terms of Pi acquisition

  11. Spatial and Temporal Variability in the Concentration and Turnover of the Inorganic Phosphate and Adenosine-5'-triphosphate pools in the North Pacific Subtropical Gyre

    NASA Astrophysics Data System (ADS)

    Björkman, Karin; Church, Matthew; Karl, David

    2015-04-01

    The microbial community's utilization of inorganic phosphate (Pi) and adenosine-5'-triphosphate (ATP) as a function of the Pi pool concentration was studied over a multi-year period at Station ALOHA (22.75˚N, 158˚W) in the North Pacific Subtropical Gyre (NPSG). Additionally, the spatial variability in these same properties was investigated along an east-west transect from California to Hawaii in the Fall of 2014. We used radiotracer techniques to determine the turnover times of the Pi or ATP pools respectively, and assessed the net production of dissolved organic phosphorus, and Pi hydrolysis rate from ATP. Pi concentrations in the upper water column at Station ALOHA are temporally highly dynamic, with periods of <10 nM-P to near 200 nM-P recorded within the top 50 m over the past decades of observations. During the California to Hawaii transect Pi concentrations showed a similarly large range (<10 to >200 nM-P), emphasizing the spatially and temporally mosaic nature of the upper ocean of this large biome. The Pi-pool turnover time ranged from a few hours to several weeks, and was strongly correlated with measured Pi pool concentrations (r2=0.8; n=30 Station ALOHA; n=15 transect). The calculated Pi uptake rates at Station ALOHA averaged 3.7±1.3 nM-P d-1 (n=30), reflecting the typically low maximum Pi uptake rates of the Prochlorococcus dominated community and the predominantly non-limiting Pi conditions. The Pi uptake rates along the transect were more variable than Station ALOHA (averaging 9.2±4.7 nM=P d-1, n=15), possibly due to a more diverse planktonic community structure, including stations with elevated concentrations of chlorophyll and primary productivity. The turnover time of the dissolved ATP pool was typically substantially shorter than for the Pi-pool (2-5 days at Station ALOHA; 0.3-2.5 days along the transect), likely reflecting its low nanomolar to picomolar ambient pool concentrations. However, at stations with the lowest SRP concentrations the

  12. 77 FR 5061 - Agency Information Collection Activities; Submission for OMB Review; Comment Request; Inorganic...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-02-01

    ...; Inorganic Arsenic Standard ACTION: Notice. SUMMARY: The Department of Labor (DOL) is submitting the..., ``Inorganic Arsenic Standard,'' to the Office of Management and Budget (OMB) for review and approval for... . SUPPLEMENTARY INFORMATION: The purpose of the Inorganic Arsenic Standard is to protect workers from the...

  13. TWEAK favors phosphate-induced calcification of vascular smooth muscle cells through canonical and non-canonical activation of NFκB

    PubMed Central

    Hénaut, L; Sanz, A B; Martin-Sanchez, D; Carrasco, S; Villa-Bellosta, R; Aldamiz-Echevarria, G; Massy, Z A; Sanchez-Nino, M D; Ortiz, A

    2016-01-01

    Vascular calcification (VC) is associated with increased cardiovascular mortality in aging, chronic kidney disease (CKD), type 2 diabetes mellitus (T2DM) and atherosclerosis. TNF-like weak inducer of apoptosis (TWEAK) recently emerged as a new biomarker for the diagnosis and prognosis of cardiovascular diseases. TWEAK binding to its functional receptor Fn14 was reported to promote several steps of atherosclerotic plaque progression. However, no information is currently available on the role of TWEAK/Fn14 on the development of medial calcification, which is highly prevalent in aging, CKD and T2DM. This study explored the involvement of TWEAK in human vascular smooth muscle cells (h-VSMCs) calcification in vitro. We report that TWEAK binding to Fn14 promotes inorganic phosphate-induced h-VSMCs calcification, favors h-VSMCs osteogenic transition, decreasing acta2 and myh11 and increasing bmp2 mRNA and tissue non-specific alkaline phosphatase (TNAP), and increases MMP9 activity. Blockade of the canonical NFκB pathway reduced by 80% TWEAK pro-calcific properties and decreased osteogenic transition, TNAP and MMP9 activity. Blockade of non-canonical NFκB signaling by a siRNA targeting RelB reduced by 20% TWEAK pro-calcific effects and decreased TWEAK-induced loss of h-VSMCs contractile phenotype and MMP9 activity, without modulating bmp2 mRNA or TNAP activity. Inhibition of ERK1/2 activation by a MAPK kinase inhibitor did not influence TWEAK pro-calcific properties. Our results suggest that TWEAK/Fn14 directly favors inorganic phosphate-induced h-VSMCs calcification by activation of both canonical and non-canonical NFκB pathways. Given the availability of neutralizing anti-TWEAK strategies, our study sheds light on the TWEAK/Fn14 axis as a novel therapeutic target in the prevention of VC. PMID:27441657

  14. A grit separation module for inorganic matter removal from activated sludge: investigation on characteristics of split sludge from the module.

    PubMed

    Chen, You-Peng; Guo, Jin-Song; Wang, Jing; Yan, Peng; Ji, Fang-Ying; Fang, Fang; Dong, Yang

    2016-12-01

    A grit separation module was developed to prevent the accumulation of inorganic solids in activated sludge systems, and it achieved effective separation of organic matter and inorganic solids. To provide technical and theoretical support for further comprehensive utilization of split sludge (underflow and overflow sludge from the separation module), the characteristics of split sludge were investigated. The settling and dewatering properties of the underflow sludge were excellent, and it had high inorganic matter content, whereas the overflow sludge had higher organic matter content. The most abundant inorganic constituent was SiO2 (59.34%), and SiO2, Al2O3, and Fe2O3 together accounted for 79.53% of the inorganic matter in the underflow sludge. The mass ratio of Fe2O3, CaO, and MgO to SiO2 and Al2O3 was 0.245 in the inorganic component of the underflow sludge. The underflow sludge had the beneficial characteristics of simple treatment and disposal, and it was suitable for use as a base raw material for ceramsite production. The overflow sludge with higher organic matter content was constantly returned from the separation module to the wastewater treatment system, gradually improving the volatile suspended solid/total suspended solid ratio of the activated sludge in the wastewater treatment system.

  15. Phosphate-Activated Cyclin-Dependent Kinase Stabilizes G1 Cyclin To Trigger Cell Cycle Entry

    PubMed Central

    Menoyo, S.; Ricco, N.; Bru, S.; Hernández-Ortega, S.; Escoté, X.; Aldea, M.

    2013-01-01

    G1 cyclins, in association with a cyclin-dependent kinase (CDK), are universal activators of the transcriptional G1-S machinery during entry into the cell cycle. Regulation of cyclin degradation is crucial for coordinating progression through the cell cycle, but the mechanisms that modulate cyclin stability to control cell cycle entry are still unknown. Here, we show that a lack of phosphate downregulates Cln3 cyclin and leads to G1 arrest in Saccharomyces cerevisiae. The stability of Cln3 protein is diminished in strains with low activity of Pho85, a phosphate-sensing CDK. Cln3 is an in vitro substrate of Pho85, and both proteins interact in vivo. More interestingly, cells that carry a CLN3 allele encoding aspartic acid substitutions at the sites of Pho85 phosphorylation maintain high levels of Cln3 independently of Pho85 activity. Moreover, these cells do not properly arrest in G1 in the absence of phosphate and they die prematurely. Finally, the activity of Pho85 is essential for accumulating Cln3 and for reentering the cell cycle after phosphate refeeding. Taken together, our data indicate that Cln3 is a molecular target of the Pho85 kinase that is required to modulate cell cycle entry in response to environmental changes in nutrient availability. PMID:23339867

  16. Combining phosphate and bacteria removal on chemically active filter membranes allows prolonged storage of drinking water.

    PubMed

    Rotzetter, A C C; Kellenberger, C R; Schumacher, C M; Mora, C; Grass, R N; Loepfe, M; Luechinger, N A; Stark, W J

    2013-11-13

    A chemically active filtration membrane with incorporated lanthanum oxide nanoparticles enables the removal of bacteria and phosphate at the same time and thus provides a simple device for preparation of drinking water and subsequent safe storage without using any kind of disinfectants.

  17. Determination of the active center in calcium-nickel phosphate dehydrogenation catalyst

    SciTech Connect

    Attali, S.; Vigouroux, B.; Lenzi, M.; Pescia, J.

    1980-06-01

    Determination of the active center in calcium-nickel phosphate dehydrogenation catalyst, used industrially in the dehydrogenation of butenes to butadiene, showed that a stable trivalent nickel ion is involved. Apparently, electrons generated in the first (oxidation) step of the reaction are eliminated by reducing the trivalent to divalent nickel which is reoxidized by protons. The results were obtained by propanol dehydration-dehydrogenation on calcium-nickel phosphate (Ca/sub 8/Ni(PO/sub 4//sub )/6) calcined at 400/sup 0/-900/sup 0/C and by ESR spectroscopy.

  18. Benzylic Phosphates in Friedel-Crafts Reactions with Activated and Unactivated Arenes: Access to Polyarylated Alkanes.

    PubMed

    Pallikonda, Gangaram; Chakravartya, Manab

    2016-03-04

    Easily reachable electron-poor/rich primary and secondary benzylic phosphates are suitably used as substrates for Friedel-Crafts benzylation reactions with only 1.2 equiv activated/deactivated arenes (no additional solvent) to access structurally and electronically diverse polyarylated alkanes with excellent yields and selectivities at room temperature. Specifically, diversely substituted di/triarylmethanes are generated within 2-30 min using this approach. A wide number of electron-poor polyarylated alkanes are easily accomplished through this route by just tuning the phosphates.

  19. Controlling the release of active compounds from the inorganic carrier halloysite

    NASA Astrophysics Data System (ADS)

    Tescione, F.; Buonocore, G. G.; Stanzione, M.; Oliviero, M.; Lavorgna, M.

    2014-05-01

    Halloysite (HNTs), a natural material characterized by a nanotube structure, has been used as an inorganic carrier of active compounds in several applications from medicine to anticorrosion coatings. In this present work, vanillin (VAN) used as a antimicrobial model, has been encapsulated within HNTs for exploiting its applicability in the active food packaging sector. The molecule release rate has been controlled by crosslinking at the tube ends the loaded vanillin with copper ions, thus producing a stopper network. The vanillin-loaded HNTs were characterized using transmission electron microscopy (TEM), Fourier transform infrared (FTIR) spectroscopy and thermo gravimetric analysis. The antimicrobial release kinetics from the loaded nanoparticles (VAN/HNTs) in water was investigated using UV-vis spectroscopy. The results show that the vanillin crosslinked with cupper ions is a feasible method to tailor the release rate of antimicrobial model from HTNs nanoparticles.

  20. Controlling the release of active compounds from the inorganic carrier halloysite

    SciTech Connect

    Tescione, F.; Buonocore, G. G.; Stanzione, M.; Oliviero, M.; Lavorgna, M.

    2014-05-15

    Halloysite (HNTs), a natural material characterized by a nanotube structure, has been used as an inorganic carrier of active compounds in several applications from medicine to anticorrosion coatings. In this present work, vanillin (VAN) used as a antimicrobial model, has been encapsulated within HNTs for exploiting its applicability in the active food packaging sector. The molecule release rate has been controlled by crosslinking at the tube ends the loaded vanillin with copper ions, thus producing a stopper network. The vanillin-loaded HNTs were characterized using transmission electron microscopy (TEM), Fourier transform infrared (FTIR) spectroscopy and thermo gravimetric analysis. The antimicrobial release kinetics from the loaded nanoparticles (VAN/HNTs) in water was investigated using UV-vis spectroscopy. The results show that the vanillin crosslinked with cupper ions is a feasible method to tailor the release rate of antimicrobial model from HTNs nanoparticles.

  1. Investigation into the Mode of Phosphate Activation in the 4-Hydroxy-4-Methyl-2-Oxoglutarate/4-Carboxy-4-Hydroxy-2-Oxoadipate Aldolase from Pseudomonas putida F1

    PubMed Central

    Mazurkewich, Scott; Seah, Stephen Y. K.

    2016-01-01

    The 4-hydroxy-4-methyl-2-oxoglutarate (HMG)/4-carboxy-4-hydroxy-2-oxoadipate (CHA) aldolase is the last enzyme of both the gallate and protocatechuate 4,5-cleavage pathways which links aromatic catabolism to central cellular metabolism. The enzyme is a class II, divalent metal dependent, aldolase which is activated in the presence of inorganic phosphate (Pi), increasing its turnover rate >10-fold. This phosphate activation is unique for a class II aldolase. The aldolase pyruvate methyl proton exchange rate, a probe of the general acid half reaction, was increased 300-fold in the presence of 1 mM Pi and the rate enhancement followed saturation kinetics giving rise to a KM of 397 ± 30 μM. Docking studies revealed a potential Pi binding site close to, or overlapping with, the proposed general acid water site. Putative Pi binding residues were substituted by site-directed mutagenesis which resulted in reductions of Pi activation. Significantly, the active site residue Arg-123, known to be critical for the catalytic mechanism of the enzyme, was also implicated in supporting Pi mediated activation. PMID:27741265

  2. Paraoxonase 1 activity in subchronic low-level inorganic arsenic exposure through drinking water.

    PubMed

    Afolabi, Olusegun K; Wusu, Adedoja D; Ogunrinola, Olufunmilayo O; Abam, Esther O; Babayemi, David O; Dosumu, Oluwatosin A; Onunkwor, Okechukwu B; Balogun, Elizabeth A; Odukoya, Olusegun O; Ademuyiwa, Oladipo

    2016-02-01

    Epidemiological evidences indicate close association between inorganic arsenic exposure via drinking water and cardiovascular diseases. While the exact mechanism of this arsenic-mediated increase in cardiovascular risk factors remains enigmatic, epidemiological studies indicate a role for paraoxonase 1 (PON1) in cardiovascular diseases. To investigate the association between inorganic arsenic exposure and cardiovascular diseases, rats were exposed to sodium arsenite (trivalent; 50, 100, and 150 ppm As) and sodium arsenate (pentavalent; 100, 150, and 200 ppm As) in their drinking water for 12 weeks. PON1 activity towards paraoxon (PONase) and phenylacetate (AREase) in plasma, lipoproteins, hepatic, and brain microsomal fractions were determined. Inhibition of PONase and AREase in plasma and HDL characterized the effects of the two arsenicals. While the trivalent arsenite inhibited PONase by 33% (plasma) and 46% (HDL), respectively, the pentavalent arsenate inhibited the enzyme by 41 and 34%, respectively. AREase activity was inhibited by 52 and 48% by arsenite, whereas the inhibition amounted to 72 and 67%, respectively by arsenate. The pattern of inhibition in plasma and HDL indicates that arsenite induced a dose-dependent inhibition of PONase whereas arsenate induced a dose-dependent inhibition of AREase. In the VLDL + LDL, arsenate inhibited PONase and AREase while arsenite inhibited PONase. In the hepatic and brain microsomal fractions, only the PONase enzyme was inhibited by the two arsenicals. The inhibition was more pronounced in the hepatic microsomes where a 70% inhibition was observed at the highest dose of pentavalent arsenic. Microsomal cholesterol was increased by the two arsenicals resulting in increased cholesterol/phospholipid ratios. Our findings indicate that decreased PON1 activity observed in arsenic exposure may be an incipient biochemical event in the cardiovascular effects of arsenic. Modulation of PON1 activity by arsenic may also be

  3. Glucose-6-phosphate mediates activation of the carbohydrate responsive binding protein (ChREBP)

    SciTech Connect

    Li, Ming V.; Chen, Weiqin; Harmancey, Romain N.; Nuotio-Antar, Alli M.; Imamura, Minako; Saha, Pradip; Taegtmeyer, Heinrich; Chan, Lawrence

    2010-05-07

    Carbohydrate response element binding protein (ChREBP) is a Mondo family transcription factor that activates a number of glycolytic and lipogenic genes in response to glucose stimulation. We have previously reported that high glucose can activate the transcriptional activity of ChREBP independent of the protein phosphatase 2A (PP2A)-mediated increase in nuclear entry and DNA binding. Here, we found that formation of glucose-6-phosphate (G-6-P) is essential for glucose activation of ChREBP. The glucose response of GAL4-ChREBP is attenuated by D-mannoheptulose, a potent hexokinase inhibitor, as well as over-expression of glucose-6-phosphatase (G6Pase); kinetics of activation of GAL4-ChREBP can be modified by exogenously expressed GCK. Further metabolism of G-6-P through the two major glucose metabolic pathways, glycolysis and pentose-phosphate pathway, is not required for activation of ChREBP; over-expression of glucose-6-phosphate dehydrogenase (G6PD) diminishes, whereas RNAi knockdown of the enzyme enhances, the glucose response of GAL4-ChREBP, respectively. Moreover, the glucose analogue 2-deoxyglucose (2-DG), which is phosphorylated by hexokinase, but not further metabolized, effectively upregulates the transcription activity of ChREBP. In addition, over-expression of phosphofructokinase (PFK) 1 and 2, synergistically diminishes the glucose response of GAL4-ChREBP. These multiple lines of evidence support the conclusion that G-6-P mediates the activation of ChREBP.

  4. Phosphate-solubility and phosphatase activity in Gangetic alluvial soil as influenced by organophosphate insecticide residues.

    PubMed

    Majumder, Shyam Prasad; Das, Amal Chandra

    2016-04-01

    An experiment was conducted under laboratory conditions to investigate the effect of four organophosphate insecticides, viz. monocrotophos, profenophos, quinalphos and triazophos at their field application rates (0.75, 1.0, 0.5 and 0.6 kg a.i.ha(-1), respectively), on the growth and activities of phosphate solubilizing microorganisms in relation to availability of insoluble phosphates in the Gangetic alluvial soil of West Bengal, India. The proliferation of phosphate solubilizing microorganisms was highly induced with profenophos (38.3%), while monocrotophos exerted maximum stimulation (20.8%) towards the solubility of insoluble phosphates in soil. The phosphatase activities of the soil (both acid phosphatase and alkaline phosphatase) were significantly increased due to the incorporation of the insecticides in general, and the augmentation was more pronounced with quinalphos (43.1%) followed by profenophos (27.6%) for acid phosphatase, and with monocrotophos (25.2%) followed by profenophos (16.1%) for alkaline phosphatase activity in soil. The total phosphorus was highly retained by triazophos (19.9%) followed by monocrotophos (16.5%), while incorporation of triazophos and quinalphos manifested greater availability of water soluble phosphorus in soil.

  5. The glucose-6-phosphate transport is not mediated by a glucose-6-phosphate/phosphate exchange in liver microsomes.

    PubMed

    Marcolongo, Paola; Fulceri, Rosella; Giunti, Roberta; Margittai, Eva; Banhegyi, Gabor; Benedetti, Angelo

    2012-09-21

    A phosphate-linked antiporter activity of the glucose-6-phosphate transporter (G6PT) has been recently described in liposomes including the reconstituded transporter protein. We directly investigated the mechanism of glucose-6-phosphate (G6P) transport in rat liver microsomal vesicles. Pre-loading with inorganic phosphate (Pi) did not stimulate G6P or Pi microsomal inward transport. Pi efflux from pre-loaded microsomes could not be enhanced by G6P or Pi addition. Rapid G6P or Pi influx was registered by light-scattering in microsomes not containing G6P or Pi. The G6PT inhibitor, S3483, blocked G6P transport irrespectively of experimental conditions. We conclude that hepatic G6PT functions as an uniporter.

  6. Alkaline Phosphatase Activity : an overlooked player on the phosphate behavior in macrotidal estuaries

    NASA Astrophysics Data System (ADS)

    Delmas, Daniel; Labry, Claire; Youenou, Agnes; Quere, Julien; Auguet, Jean Christophe; Montanie, Helene

    2014-05-01

    The non-conservative behavior of phosphate within the estuarine salinity gradient is essentially assigned to physico-chemical processes, such as desorption at low salinity and to benthic exchanges. Microbial phosphatase activity (APA), generally related to phosphate deficiency, is seldom studied in phosphate rich estuarine waters. In order to address the impact of microbial activity (bacterial abundance, production BSP, APA) on phosphate behavior, we studied these activities on a seasonal basis within the salinity gradient of two macrotidal estuaries presenting different levels of suspended solids. Whatever the season the Charente estuary is characterized by high levels of Suspended Particulate Matter (SPM > 1g.L-1), particularly in the Maximum Turbidity Zone (MTZ) located at the 5-10 psu. In this area characterized by high BSP and APA there is a significant increase of PO4 levels especially during summer. In the Aulne estuary the particle load is significantly lower (1/10) but high BSP and APA are equally recorded. In the highly turbid waters of the Charente estuary, active phytoplankton is virtually absent as pheopigments constitute up to 80% of the total pigments, particularly in the MTZ, therefore APA may essentially have a bacterial origin. In the Aulne estuary attached bacteria are dominant, both in numbers and production, and their distribution along the haline gradient perfectly follows those of APA and phosphate levels. These observations, associated with the very close relationships observed between APA, SPM and BSP, suggest that APA derive mainly from bacterial (attached) origin and operate at the expense of particulate phosphorus and hence contribute to PO4 regeneration, especially in spring and summer. Finally, as APA increased as PO4, whereas the reverse is observed in both fresh and marine waters, an original scheme for APA regulation, related to the large dominance of attached bacteria can be described for the estuarine waters.

  7. Archaeal Inorganic Pyrophosphatase Displays Robust Activity under High-Salt Conditions and in Organic Solvents.

    PubMed

    McMillan, Lana J; Hepowit, Nathaniel L; Maupin-Furlow, Julie A

    2015-11-06

    Soluble inorganic pyrophosphatases (PPAs) that hydrolyze inorganic pyrophosphate (PPi) to orthophosphate (Pi) are commonly used to accelerate and detect biosynthetic reactions that generate PPi as a by-product. Current PPAs are inactivated by high salt concentrations and organic solvents, which limits the extent of their use. Here we report a class A type PPA of the haloarchaeon Haloferax volcanii (HvPPA) that is thermostable and displays robust PPi-hydrolyzing activity under conditions of 25% (vol/vol) organic solvent and salt concentrations from 25 mM to 3 M. HvPPA was purified to homogeneity as a homohexamer by a rapid two-step method and was found to display non-Michaelis-Menten kinetics with a Vmax of 465 U · mg(-1) for PPi hydrolysis (optimal at 42°C and pH 8.5) and Hill coefficients that indicated cooperative binding to PPi and Mg(2+). Similarly to other class A type PPAs, HvPPA was inhibited by sodium fluoride; however, hierarchical clustering and three-dimensional (3D) homology modeling revealed HvPPA to be distinct in structure from characterized PPAs. In particular, HvPPA was highly negative in surface charge, which explained its extreme resistance to organic solvents. To demonstrate that HvPPA could drive thermodynamically unfavorable reactions to completion under conditions of reduced water activity, a novel coupled assay was developed; HvPPA hydrolyzed the PPi by-product generated in 2 M NaCl by UbaA (a "salt-loving" noncanonical E1 enzyme that adenylates ubiquitin-like proteins in the presence of ATP). Overall, we demonstrate HvPPA to be useful for hydrolyzing PPi under conditions of reduced water activity that are a hurdle to current PPA-based technologies.

  8. Archaeal Inorganic Pyrophosphatase Displays Robust Activity under High-Salt Conditions and in Organic Solvents

    PubMed Central

    McMillan, Lana J.; Hepowit, Nathaniel L.

    2015-01-01

    Soluble inorganic pyrophosphatases (PPAs) that hydrolyze inorganic pyrophosphate (PPi) to orthophosphate (Pi) are commonly used to accelerate and detect biosynthetic reactions that generate PPi as a by-product. Current PPAs are inactivated by high salt concentrations and organic solvents, which limits the extent of their use. Here we report a class A type PPA of the haloarchaeon Haloferax volcanii (HvPPA) that is thermostable and displays robust PPi-hydrolyzing activity under conditions of 25% (vol/vol) organic solvent and salt concentrations from 25 mM to 3 M. HvPPA was purified to homogeneity as a homohexamer by a rapid two-step method and was found to display non-Michaelis-Menten kinetics with a Vmax of 465 U · mg−1 for PPi hydrolysis (optimal at 42°C and pH 8.5) and Hill coefficients that indicated cooperative binding to PPi and Mg2+. Similarly to other class A type PPAs, HvPPA was inhibited by sodium fluoride; however, hierarchical clustering and three-dimensional (3D) homology modeling revealed HvPPA to be distinct in structure from characterized PPAs. In particular, HvPPA was highly negative in surface charge, which explained its extreme resistance to organic solvents. To demonstrate that HvPPA could drive thermodynamically unfavorable reactions to completion under conditions of reduced water activity, a novel coupled assay was developed; HvPPA hydrolyzed the PPi by-product generated in 2 M NaCl by UbaA (a “salt-loving” noncanonical E1 enzyme that adenylates ubiquitin-like proteins in the presence of ATP). Overall, we demonstrate HvPPA to be useful for hydrolyzing PPi under conditions of reduced water activity that are a hurdle to current PPA-based technologies. PMID:26546423

  9. Properties of Ce-activated alkali-lutetium double phosphate scintillators

    SciTech Connect

    Wiśniewski, D.; Wojtowicz, A. J.; Boatner, Lynn A

    2010-01-01

    The scintillation properties of Ce-activated alkali-lutetium double phosphate single crystals that vary with the alkali ion type and activation level are summarized and compared. The materials investigated here have been identified as fast and efficient scintillators for the detection of x-ray and radiation, and in case of Li3Lu(PO4)2:Ce, for thermal neutron detection as well.

  10. A nucleophilic catalysis step is involved in the hydrolysis of aryl phosphate monoesters by human CT acylphosphatase.

    PubMed

    Paoli, Paolo; Pazzagli, Luigia; Giannoni, Elisa; Caselli, Anna; Manao, Giampaolo; Camici, Guido; Ramponi, Giampietro

    2003-01-03

    Acylphosphatase, one of the smallest enzymes, is expressed in all organisms. It displays hydrolytic activity on acyl phosphates, nucleoside di- and triphosphates, aryl phosphate monoesters, and polynucleotides, with acyl phosphates being the most specific substrates in vitro. The mechanism of catalysis for human acylphosphatase (the organ-common type isoenzyme) was investigated using both aryl phosphate monoesters and acyl phosphates as substrates. The enzyme is able to catalyze phosphotransfer from p-nitrophenyl phosphate to glycerol (but not from benzoyl phosphate to glycerol), as well as the inorganic phosphate-H(2)18O oxygen exchange reaction in the absence of carboxylic acids or phenols. In short, our findings point to two different catalytic pathways for aryl phosphate monoesters and acyl phosphates. In particular, in the aryl phosphate monoester hydrolysis pathway, an enzyme-phosphate covalent intermediate is formed, whereas the hydrolysis of acyl phosphates seems a more simple process in which the Michaelis complex is attacked directly by a water molecule generating the reaction products. The formation of an enzyme-phosphate covalent complex is consistent with the experiments of isotope exchange and transphosphorylation from substrates to glycerol, as well as with the measurements of the Brønsted free energy relationships using a panel of aryl phosphates with different structures. His-25 involvement in the formation of the enzyme-phosphate covalent complex during the hydrolysis of aryl phosphate monoesters finds significant confirmation in experiments performed with the H25Q mutated enzyme.

  11. Determination of phosphate in natural waters by activation analysis of tungstophosphoric acid

    USGS Publications Warehouse

    Allen, Herbert E.; Hahn, Richard B.

    1969-01-01

    Activation analysis may be used to determine quantitatively traces of phosphate in natural waters. Methods based on the reaction 31P(n,γ)32P are subject to interference by sulfur and chlorine which give rise to 32P through n,p and n,α reactions. If the ratio of phosphorus to sulfur or chlorine is small, as it is in most natural waters, accurate analyses by these methods are difficult to achieve. In the activation analysis method, molybdate and tungstate ions are added to samples containing phosphate ion to form tungstomolybdophosphoric acid. The complex is extracted with 2,6-dimethyl-4-heptanone. After activation of an aliquot of the organic phase for 1 hour at a flux of 1013 neutrons per cm2, per second, the gamma spectrum is essentially that of tungsten-187. The induced activity is proportional to the concentration of phosphate in the sample. A test of the method showed it to give accurate results at concentrations of 4 to at least 200 p.p.b. of phosphorus when an aliquot of 100 μl. was activated. By suitable reagent purification, counting for longer times, and activation of larger aliquots, the detection limit could be lowered several hundredfold.

  12. Inorganic polyphosphate is a potent activator of the mitochondrial permeability transition pore in cardiac myocytes.

    PubMed

    Seidlmayer, Lea K; Gomez-Garcia, Maria R; Blatter, Lothar A; Pavlov, Evgeny; Dedkova, Elena N

    2012-05-01

    Mitochondrial dysfunction caused by excessive Ca2+ accumulation is a major contributor to cardiac cell and tissue damage during myocardial infarction and ischemia-reperfusion injury (IRI). At the molecular level, mitochondrial dysfunction is induced by Ca2+-dependent opening of the mitochondrial permeability transition pore (mPTP) in the inner mitochondrial membrane, which leads to the dissipation of mitochondrial membrane potential (ΔΨm), disruption of adenosine triphosphate production, and ultimately cell death. Although the role of Ca2+ for induction of mPTP opening is established, the exact molecular mechanism of this process is not understood. The aim of the present study was to test the hypothesis that the adverse effect of mitochondrial Ca2+ accumulation is mediated by its interaction with inorganic polyphosphate (polyP), a polymer of orthophosphates linked by phosphoanhydride bonds. We found that cardiac mitochondria contained significant amounts (280±60 pmol/mg of protein) of short-chain polyP with an average length of 25 orthophosphates. To test the role of polyP for mPTP activity, we investigated kinetics of Ca2+ uptake and release, ΔΨm and Ca2+-induced mPTP opening in polyP-depleted mitochondria. polyP depletion was achieved by mitochondria-targeted expression of a polyP-hydrolyzing enzyme. Depletion of polyP in mitochondria of rabbit ventricular myocytes led to significant inhibition of mPTP opening without affecting mitochondrial Ca2+ concentration by itself. This effect was observed when mitochondrial Ca2+ uptake was stimulated by increasing cytosolic [Ca2+] in permeabilized myocytes mimicking mitochondrial Ca2+ overload observed during IRI. Our findings suggest that inorganic polyP is a previously unrecognized major activator of mPTP. We propose that the adverse effect of polyphosphate might be caused by its ability to form stable complexes with Ca2+ and directly contribute to inner mitochondrial membrane permeabilization.

  13. Tuning laccase catalytic activity with phosphate functionalized carbon dots by visible light.

    PubMed

    Li, Hao; Guo, Sijie; Li, Chuanxi; Huang, Hui; Liu, Yang; Kang, Zhenhui

    2015-05-13

    The phosphate functionalized carbon dots (PCDs) with high biocompatibility and low toxicity can be used as efficient additives for the construction of laccase/PCDs hybrids catalyst. A series of experiments indicated that the activity of laccase/PCDs was higher than that of free laccase (increased by 47.7%). When laccase/PCDs hybrids catalyst was irradiated with visible light (laccase/PCDs-Light), its activity was higher than that of laccase/PCDs hybrids without light irradiation (increased by 92.1%). In the present system, the T1 Cu in laccase was combined with the phosphate group on PCDs, which can increase binding capacity of laccase/PCDs hybrids and substrate. Further, the visible light irradiation increased the donating and accepting electronic capability of the laccase/PCDs hybrids, improving their catalytic activity.

  14. Glutamic acid-149 is important for enzymatic activity of yeast inorganic pyrophosphatase.

    PubMed

    Gonzalez, M A; Cooperman, B S

    1986-11-04

    Modification of Saccharomyces cerevisiae inorganic pyrophosphatase (PPase) with 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide is known to lead to a loss of enzymatic activity, the rate of which is decreased in the presence of ligands binding to the active site [Cooperman, B. S., & Chiu, N. Y. (1973) Biochemistry 12, 1676-1682; Heitman, P., & Uhlig, H. J. (1974) Acta Biol. Med. Ger. 32, 565-594]. In this work we show that, when such inactivation is carried out in the presence of [14C]glycine ethyl ester (GEE), GEE is covalently incorporated into PPase, incorporation into the most highly labeled tryptic peptide is site-specific, as evidenced by the reduction of such incorporation in the presence of the active site ligands Zn2+ and Pi, the extent of formation of this specifically labeled peptide correlates with the fractional loss of PPase activity, and the specifically labeled peptide corresponds to residues 145-153 and the position of incorporation within this peptide is Glu-149. The significance of our findings for the location of the active site and for the catalytic mechanism of PPase is briefly considered in the light of the 3-A X-ray crystallographic structure of Arutyunyun and his colleagues [Arutyunyun, E. G., et al. (1981) Dokl. Akad. Nauk SSSR 258, 1481-1485; Kuranova, I. P., et al. (1983) Bioorg. Khim. 9, 1611-1919; Terzyan, S. S., et al. (1984) Bioorg. Khim. 10, 1469-1482].

  15. Effect of lysine to alanine mutations on the phosphate activation and BPTES inhibition of glutaminase.

    PubMed

    McDonald, Charles J; Acheff, Eric; Kennedy, Ryan; Taylor, Lynn; Curthoys, Norman P

    2015-09-01

    The GLS1 gene encodes a mitochondrial glutaminase that is highly expressed in brain, kidney, small intestine and many transformed cells. Recent studies have identified multiple lysine residues in glutaminase that are sites of N-acetylation. Interestingly, these sites are located within either a loop segment that regulates access of glutamine to the active site or the dimer:dimer interface that participates in the phosphate-dependent oligomerization and activation of the enzyme. These two segments also contain the binding sites for bis-2[5-phenylacetamido-1,2,4-thiadiazol-2-yl]ethylsulfide (BPTES), a highly specific and potent uncompetitive inhibitor of this glutaminase. BPTES is also the lead compound for development of novel cancer chemotherapeutic agents. To provide a preliminary assessment of the potential effects of N-acetylation, the corresponding lysine to alanine mutations were constructed in the hGACΔ1 plasmid. The wild type and mutated proteins were purified by Ni(+)-affinity chromatography and their phosphate activation and BPTES inhibition profiles were analyzed. Two of the alanine substitutions in the loop segment (K311A and K328A) and the one in the dimer:dimer interface (K396A) form enzymes that require greater concentrations of phosphate to produce half-maximal activation and exhibit greater sensitivity to BPTES inhibition. By contrast, the K320A mutation results in a glutaminase that exhibits near maximal activity in the absence of phosphate and is not inhibited by BPTES. Thus, lysine N-acetylation may contribute to the acute regulation of glutaminase activity in various tissues and alter the efficacy of BPTES-type inhibitors.

  16. Microbial activities and phosphorus cycling: An application of oxygen isotope ratios in phosphate

    NASA Astrophysics Data System (ADS)

    Stout, Lisa M.; Joshi, Sunendra R.; Kana, Todd M.; Jaisi, Deb P.

    2014-08-01

    Microorganisms carry out biochemical transformations of nutrients that make up their cells. Therefore, understanding how these nutrients are transformed or cycled in natural environments requires knowledge of microbial activity. Commonly used indicators for microbial activity typically include determining microbial respiration by O2/CO2 measurements, cell counts, and measurement of enzyme activities. However, coupled studies on nutrient cycling and microbial activity are not given enough emphasis. Here we apply phosphate oxygen isotope ratios (δ18OP) as a tool for measurement of microbial activity and compare the rate of isotope exchange with methods of measuring microbial activities that are more commonly applied in environmental studies including respiration, dehydrogenase activity, alkaline phosphatase activity, and cell counts. Our results show that different bacteria may have different strategies for P uptake, storage and release, their respiration and consequently expression of DHA and APase activities, but in general the trend of their enzyme activities are comparable. Phosphate δ18OP values correlated well with these other parameters used to measure microbial activity with the strongest linear relationships between δ18OP and CO2 evolution (r = -0.99). Even though the rate of isotope exchange for each microorganism used in this study is different, the rate per unit CO2 respiration showed one general trend, where δ18OP values move towards equilibrium while CO2 is generated. While this suggests that P cycling among microorganisms used in this study can be generalized, further research is needed to determine whether the microorganism-specific isotope exchange trend may occur in natural environments. In summary, phosphate oxygen isotope measurements may offer an alternative for use as a tracer to measure microbial activity in soils, sediments, and many other natural environments.

  17. Self-assembled organic-inorganic hybrid glucoamylase nanoflowers with enhanced activity and stability.

    PubMed

    Nadar, Shamraja S; Gawas, Sarita D; Rathod, Virendra K

    2016-11-01

    An organic-inorganic hybrid glucoamylase nanoflower was prepared in single pot by simple, facile and highly efficient method. The stepwise formation of enzyme-embedded hybrid nanoflowers and influence of experimental parameters viz. pH of solution mixture, enzyme and copper ion concentration on the activity of prepared hybrid nanoflowers were systematically investigated. The self-assembled hybrid glucoamylase nanoflowers were synthesized by mixing aqueous solution of copper sulphate (200mM) with PBS (pH 7.5, 5mM) containing glucoamylase (1mg/mL) in 24h at room temperature. These prepared nanoflowers were further characterized by FT-IR, SEM and XRD. The hybrid nanoflowers exhibited 204% enhanced activity recovery and two folds improvement in thermal stability in terms of half-life (in the range of 50-70°C) with respect to the free form. The hybrid glucoamylase nanoflowers retained 70% residual activity after eight successive cycles indicating their excellent durability. Additionally, the nanoflowers retained up to 91% residual activity upto 25 days of storage. Moreover, the conformational changes occurred in glucoamylase structure after preparing hybrid nanoflowers were evaluated by FT-IR spectroscopy data tools.

  18. Carbonic anhydrase inhibitors. Interaction of isozymes I, II, IV, V, and IX with phosphates, carbamoyl phosphate, and the phosphonate antiviral drug foscarnet.

    PubMed

    Rusconi, Stefano; Innocenti, Alessio; Vullo, Daniela; Mastrolorenzo, Antonio; Scozzafava, Andrea; Supuran, Claudiu T

    2004-12-06

    A detailed inhibition study of five carbonic anhydrase (CA, EC 4.2.1.1) isozymes with inorganic phosphates, carbamoyl phosphate, the antiviral phosphonate foscarnet as well as formate is reported. The cytosolic isozyme hCA I was weakly inhibited by neutral phosphate, strongly inhibited by carbamoyl phosphate (K(I) of 9.4 microM), and activated by hydrogen- and dihydrogenphosphate, foscarnet and formate (best activator foscarnet, K(A)=12 microM). The cytosolic isozyme hCA II was weakly inhibited by all the investigated anions, with carbamoyl phosphate showing a K(I) of 0.31 mM. The membrane-associated isozyme hCA IV was the most sensitive to inhibition by phosphates/phosphonates, showing a K(I) of 84 nM for PO(4)(3-), of 9.8 microM for HPO(4)(2-), and of 9.9 microM for carbamoyl phosphate. Foscarnet was the best inhibitor of this isozyme (K(I) of 0.82 mM) highly abundant in the kidneys, which may explain some of the renal side effects of the drug. The mitochondrial isozyme hCA V was weakly inhibited by all phosphates/phosphonates, except carbamoyl phosphate, which showed a K(I) of 8.5 microM. Thus, CA V cannot be the isozyme involved in the carbamoyl phosphate synthetase I biosynthetic reaction, as hypothesized earlier. Furthermore, the relative resistance of CA V to inhibition by inorganic phosphates suggests an evolutionary adaptation of this mitochondrial isozyme to the presence of high concentrations of such anions in these energy-converting organelles, where high amounts of ATP are produced by ATP synthetase, from ADP and inorganic phosphates. The transmembrane, tumor-associated isozyme hCA IX was on the other hand slightly inhibited by all these anions.

  19. Impact of phosphate concentration on docosahexaenoic acid production and related enzyme activities in fermentation of Schizochytrium sp.

    PubMed

    Ren, Lu-Jing; Feng, Yun; Li, Juan; Qu, Liang; Huang, He

    2013-09-01

    Docosahexaenoic acid (DHA) is an important and widely used infant food additive. In this study, the effects of phosphate concentration on lipid and especially DHA synthesis in the oleaginous fungi Schizochytrium sp. HX-308 have been investigated in batch cultures. The maximum DHA yield (8.9 g/L) and DHA productivity (148.3 mg/L h) in 0.1 g/L KH2PO4 concentration were higher than the DHA yield (6.2 g/L) and DHA productivity (86.1 mg/L h) in 4 g/L KH2PO4 concentration. Furthermore, differences in related enzyme activities (malic enzyme, glucose-6-phosphate dehydrogenase and NAD(+)-isocitrate dehydrogenase) between phosphate-sufficient and phosphate-limitation conditions were assayed. The results showed that the phosphate-limitation condition could maintain higher activities of malic enzyme and glucose-6-phosphate dehydrogenase in addition to lower activity of NAD(+)-isocitrate dehydrogenase. In addition, glucose-6-phosphate dehydrogenase might be the main supplier of NADPH at the early stage of fermentation while malic enzyme might be the provider at the late stage. This information might explain the regulation mechanism of phosphate limitation for lipid production and be useful for further DHA production enhancement.

  20. A potent synthetic inorganic antibiotic with activity against drug-resistant pathogens.

    PubMed

    Hubick, Shelby; Jayaraman, Arumugam; McKeen, Alexander; Reid, Shelby; Alcorn, Jane; Stavrinides, John; Sterenberg, Brian T

    2017-02-06

    The acronymously named "ESKAPE" pathogens represent a group of bacteria that continue to pose a serious threat to human health, not only due to their propensity for repeated emergence, but also due to their ability to "eskape" antibiotic treatment. The evolution of multi-drug resistance in these pathogens alone has greatly outpaced the development of new therapeutics, necessitating an alternative strategy for antibiotic development that considers the evolutionary mechanisms driving antibiotic resistance. In this study, we synthesize a novel inorganic antibiotic, phosphopyricin, which has antibiotic activity against the Gram-positive methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecium (VRE). We show that this potent antibiotic is bactericidal, and exhibits low toxicity in an acute dose assay in mice. As a synthetic compound that does not occur naturally, phosphopyricin would be evolutionarily foreign to microbes, thereby slowing the evolution of resistance. In addition, it loses antibiotic activity upon exposure to light, meaning that the active antibiotic will not accumulate in the general environment where strong selective pressures imposed by antibiotic residuals are known to accelerate resistance. Phosphopyricin represents an innovation in antimicrobials, having a synthetic core, and a photosensitive chemical architecture that would reduce accumulation in the environment.

  1. A potent synthetic inorganic antibiotic with activity against drug-resistant pathogens

    PubMed Central

    Hubick, Shelby; Jayaraman, Arumugam; McKeen, Alexander; Reid, Shelby; Alcorn, Jane; Stavrinides, John; Sterenberg, Brian T.

    2017-01-01

    The acronymously named “ESKAPE” pathogens represent a group of bacteria that continue to pose a serious threat to human health, not only due to their propensity for repeated emergence, but also due to their ability to “eskape” antibiotic treatment12. The evolution of multi-drug resistance in these pathogens alone has greatly outpaced the development of new therapeutics, necessitating an alternative strategy for antibiotic development that considers the evolutionary mechanisms driving antibiotic resistance. In this study, we synthesize a novel inorganic antibiotic, phosphopyricin, which has antibiotic activity against the Gram-positive methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecium (VRE). We show that this potent antibiotic is bactericidal, and exhibits low toxicity in an acute dose assay in mice. As a synthetic compound that does not occur naturally, phosphopyricin would be evolutionarily foreign to microbes, thereby slowing the evolution of resistance. In addition, it loses antibiotic activity upon exposure to light, meaning that the active antibiotic will not accumulate in the general environment where strong selective pressures imposed by antibiotic residuals are known to accelerate resistance. Phosphopyricin represents an innovation in antimicrobials, having a synthetic core, and a photosensitive chemical architecture that would reduce accumulation in the environment. PMID:28165020

  2. Mn0.95I0.02[PO3(OH)] · 2H2O phosphate-iodate, an inorganic analogue of phosphonates

    NASA Astrophysics Data System (ADS)

    Belokoneva, E. L.; Dimitrova, O. V.; Volkov, A. S.

    2015-09-01

    The new Mn0.95I0.02[PO3(OH)] · 2H2O phosphate-iodate (space group Pnam = Pnma, D {2/h 16}) is obtained under hydrothermal conditions. The crystal structure is determined without preliminary knowledge of the chemical formula. The structure consists of layers of MnО6 octahedra connected with PO4 tetrahedra. Water molecules are located between the layers. [IO3]- groups having a typical umbrella-like coordination are statistically implanted in layers of MnО6 octahedra at a distance of 1.2 Å from Mn atoms. Their content in the crystal is minor. The structures of the phosphate-iodate coincides with the structures of phosphonates with consideration for the replacement of one (OH) vertex of the РО4 tetrahedron by the organic methyl radical СН3. In the structures of phosphonates and earlier studied phosphates, identical layers are distinguished and the cause of the existence of two MDO varieties is established based on the analysis within the OD theory. Possible hybrid structures derived from the prototypes under consideration are predicted.

  3. Activation and splitting of carbon dioxide on the surface of an inorganic electride material.

    PubMed

    Toda, Yoshitake; Hirayama, Hiroyuki; Kuganathan, Navaratnarajah; Torrisi, Antonio; Sushko, Peter V; Hosono, Hideo

    2013-01-01

    Activation of carbon dioxide is the most important step in its conversion into valuable chemicals. Surfaces of stable oxide with a low work function may be promising for this purpose. Here we report that the surfaces of the inorganic electride [Ca24Al28O64](4+)(e(-))4 activate and split carbon dioxide at room temperature. This behaviour is attributed to a high concentration of localized electrons in the near-surface region and a corrugation of the surface that can trap oxygen atoms and strained carbon monoxide and carbon dioxide molecules. The [Ca24Al28O64](4+)(e(-))4 surface exposed to carbon dioxide is studied using temperature-programmed desorption, and spectroscopic methods. The results of these measurements, corroborated with ab initio simulations, show that both carbon monoxide and carbon dioxide adsorb on the [Ca24Al28O64](4+)(e(-))4 surface at RT and above and adopt unusual configurations that result in desorption of molecular carbon monoxide and atomic oxygen upon heating.

  4. Enhancing the adsorption of ionic liquids onto activated carbon by the addition of inorganic salts

    PubMed Central

    Neves, Catarina M. S. S.; Lemus, Jesús; Freire, Mara G.; Palomar, Jose; Coutinho, João A. P.

    2014-01-01

    Most ionic liquids (ILs) are either water soluble or present a non-negligible miscibility with water that may cause some harmful effects upon their release into the environment. Among other methods, adsorption of ILs onto activated carbon (AC) has shown to be an effective technique to remove these compounds from aqueous solutions. However, this method has proved to be viable only for hydrophobic ILs rather than for the hydrophilic that, being water soluble, have a larger tendency for contamination. In this context, an alternative approach using the salting-out ability of inorganic salts is here proposed to enhance the adsorption of hydrophilic ILs onto activated carbon. The effect of the concentrations of Na2SO4 on the adsorption of five ILs onto AC was investigated. A wide range of ILs that allow the inspection of the IL cation family (imidazolium- and pyridinium-based) and the anion nature (accounting for its hydrophilicity and fluorination) through the adsorption onto AC was studied. In general, it is shown that the use of Na2SO4 enhances the adsorption of ILs onto AC. In particular, this effect is highly relevant when dealing with hydrophilic ILs that are those that are actually poorly removed by AC. In addition, the COnductor like Screening MOdel for Real Solvents (COSMO-RS) was used aiming at complementing the experimental data obtained. This work contributes with the development of novel methods to remove ILs from water streams aiming at creating “greener” processes. PMID:25516713

  5. Photoluminescence Mechanism and Photocatalytic Activity of Organic-Inorganic Hybrid Materials Formed by Sequential Vapor Infiltration.

    PubMed

    Akyildiz, Halil I; Stano, Kelly L; Roberts, Adam T; Everitt, Henry O; Jur, Jesse S

    2016-05-03

    Organic-inorganic hybrid materials formed by sequential vapor infiltration (SVI) of trimethylaluminum into polyester fibers are demonstrated, and the photoluminescence of the fibers is evaluated using a combined UV-vis and photoluminescence excitation (PLE) spectroscopy approach. The optical activity of the modified fibers depends on infiltration thermal processing conditions and is attributed to the reaction mechanisms taking place at different temperatures. At low temperatures a single excitation band and dual emission bands are observed, while, at high temperatures, two distinct absorption bands and one emission band are observed, suggesting that the physical and chemical structure of the resulting hybrid material depends on the SVI temperature. Along with enhancing the photoluminescence intensity of the PET fibers, the internal quantum efficiency also increased to 5-fold from ∼4-5% to ∼24%. SVI processing also improved the photocatalytic activity of the fibers, as demonstrated by photodeposition of Ag and Au metal particles out of an aqueous metal salt solution onto fiber surfaces via UVA light exposure. Toward applications in flexible electronics, well-defined patterning of the metallic materials is achieved by using light masking and focused laser rastering approaches.

  6. Continuous cellularization of calcium phosphate hybrid scaffolds induced by plasma polymer activation.

    PubMed

    Bergemann, Claudia; Cornelsen, Matthias; Quade, Antje; Laube, Thorsten; Schnabelrauch, Matthias; Rebl, Henrike; Weißmann, Volker; Seitz, Hermann; Nebe, Barbara

    2016-02-01

    The generation of hybrid materials based on β-tricalcium phosphate (TCP) and various biodegradable polymers like poly(l-lactide-co-d,l-lactide) (PLA) represents a common approach to overcoming the disadvantages of pure TCP devices. These disadvantages lie in TCP's mechanical properties, such as brittleness. The positive characteristic of PLA - improvement of compressive strength of calcium phosphate scaffolds - is diametrically opposed to its cell attractiveness. Therefore, the objective of this work was to optimize osteoblast migration and cellularization inside a three-dimensionally (3D) printed, PLA polymer stabilized TCP hybrid scaffold by a plasma polymer process depositing amino groups via allylamine. MG-63 osteoblastic cells inside the 10mm hybrid scaffold were dynamically cultivated for 14days in a 3D model system integrated in a perfusion reactor. The whole TCP/PLA hybrid scaffold was continuously colonized due to plasma polymerized allylamine activation inducing the migration potential of osteoblasts.

  7. MESOPOROUS IRON PHOSPHATE AS AN ACTIVE, SELECTIVE AND RECYCLABLE CATALYST FOR THE SYNTHESIS OF NOPOL BY PRINS CONDENSATION

    EPA Science Inventory


    Mesoporous iron phosphate is found to be a highly active and recyclable heterogeneous catalyst for the selective synthesis of nopol by Prins condensation of ?-pinene and paraformaldehyde in acetonitrile at 80 oC.



  8. The oxygen isotope composition of phosphate released from phytic acid by the activity of wheat and Aspergillus niger phytase

    NASA Astrophysics Data System (ADS)

    von Sperber, C.; Tamburini, F.; Brunner, B.; Bernasconi, S. M.; Frossard, E.

    2015-07-01

    Phosphorus (P) is an essential nutrient for living organisms. Under P-limiting conditions plants and microorganisms can exude extracellular phosphatases that release inorganic phosphate (Pi) from organic phosphorus compounds (Porg). Phytic acid (myo-inositol hexakisphosphate, IP6) is an important form of Porg in many soils. The enzymatic hydrolysis of IP6 by phytase yields available Pi and less phosphorylated inositol derivates as products. The hydrolysis of organic P compounds by phosphatases leaves an isotopic imprint on the oxygen isotope composition (δ18O) of released Pi, which might be used to trace P in the environment. This study aims at determining the effect of phytase on the oxygen isotope composition of released Pi. For this purpose, enzymatic assays with histidine acid phytases from wheat and Aspergillus niger were prepared using IP6, adenosine 5'-monophosphate (AMP) and glycerophosphate (GPO4) as substrates. For a comparison to the δ18O of Pi released by other extracellular enzymes, enzymatic assays with acid phosphatases from potato and wheat germ with IP6 as a substrate were prepared. During the hydrolysis of IP6 by phytase, four of the six Pi were released, and one oxygen atom from water was incorporated into each Pi. This incorporation of oxygen from water into Pi was subject to an apparent inverse isotopic fractionation (ϵ ~ 6 to 10 ‰), which was similar to that imparted by acid phosphatase from potato during the hydrolysis of IP6 (ϵ ~ 7 ‰), where less than three Pi were released. The incorporation of oxygen from water into Pi during the hydrolysis of AMP and GPO4 by phytase yielded a normal isotopic fractionation (ϵ ~ -12 ‰), similar to values reported for acid phosphatases from potato and wheat germ. We attribute this similarity in ϵ to the same amino acid sequence motif (RHGXRXP) at the active site of these enzymes, which leads to similar reaction mechanisms. We suggest that the striking

  9. ATPases and phosphate exchange activities in magnesium chelatase subunits of Rhodobacter sphaeroides.

    PubMed

    Hansson, M; Kannangara, C G

    1997-11-25

    Three separate proteins, BchD, BchH, and BchI, together with ATP, insert magnesium into protoporphyrin IX. An analysis of ATP utilization by the subunits revealed the following: BchH catalyzed ATP hydrolysis at the rate of 0.9 nmol per min per mg of protein. BchI and BchD, tested individually, had no ATPase activity but, when combined, hydrolyzed ATP at the rate of 117.9 nmol/min per mg of protein. Magnesium ions were required for the ATPase activities of both BchH and BchI+D, and these activities were inhibited 50% by 2 mM o-phenanthroline. BchI additionally catalyzed a phosphate exchange reaction from ATP and ADP. We conclude that ATP hydrolysis by BchI+D is required for an activation step in the magnesium chelatase reaction, whereas ATPase activity of BchH and the phosphate exchange activity of BchI participate in subsequent reactions leading to the insertion of Mg2+ into protoporphyrin IX.

  10. Enzymatically synthesized inorganic polymers as morphogenetically active bone scaffolds: application in regenerative medicine.

    PubMed

    Wang, Xiaohong; Schröder, Heinz C; Müller, Werner E G

    2014-01-01

    In recent years a paradigm shift in understanding of human bone formation has occurred that starts to change current concepts in tissue engineering of bone and cartilage. New discoveries revealed that fundamental steps in biomineralization are enzyme driven, not only during hydroxyapatite deposition, but also during initial bioseed formation, involving the transient deposition and subsequent transformation of calcium carbonate to calcium phosphate mineral. The principal enzymes mediating these reactions, carbonic anhydrase and alkaline phosphatase, open novel targets for pharmacological intervention of bone diseases like osteoporosis, by applying compounds acting as potential activators of these enzymes. It is expected that these new findings will give an innovation boost for the development of scaffolds for bone repair and reconstruction, which began with the use of bioinert materials, followed by bioactive materials and now leading to functional regenerative tissue units. These new developments have become possible with the discovery of the morphogenic activity of bioinorganic polymers, biocalcit, bio-polyphosphate and biosilica that are formed by a biogenic, enzymatic mechanism, a driving force along with the development of novel rapid-prototyping three-dimensional (3D) printing methods and bioprinting (3D cell printing) techniques that may allow a fabrication of customized implants for patients suffering in bone diseases in the future.

  11. Applications of inorganic nanoparticles in diabetes

    NASA Astrophysics Data System (ADS)

    Elhabush, Nada Atiya Omar

    Diabetes Mellitus (DM) is an endocrine and metabolic disease that has become a global emergency because of the rapid rise in morbidity and mortality rates worldwide. Since the direct delivery of biomolecules, such as insulin, to treat DM is inefficient and subjected to enzymatic degradation, nanotechnology and nanomedicine research have been devoted to the development of more effective methods to treat DM. Nanoparticles (NP), organic, inorganic, or hybrid, have served as potential carrier for safe and efficient transport for insulin. Additionally, several NP have biological activities that help treat and/or prevent DM and diabetes complications, such as antioxidant, anti-apoptotic, or insulin-mimetic activities. Moreover, physicochemical properties of some NP allow them to be used in diagnostic tools for potential diagnosis or monitoring purposes. This work highlights the applications of inorganic NP such as, gold, selenium, silver, calcium phosphate, zinc oxide, cerium oxide, and iron oxide and in the treatment or diagnosis of DM.

  12. Studies on Anion Promoted Titania.1: Preparation, Characterization, and Catalytic Activity toward Alcohol and Cumene Conversion Reactions of Phosphated Titania.

    PubMed

    Parida; Acharya; Samantaray; Mishra

    1999-09-15

    Phosphate impregnated titania samples with varying amount of phosphate have been prepared by solid-solid kneading as well as aqueous impregnation method. All the samples are characterized by XRD, TG-DTA, and N(2) adsorption-desorption isotherm. Surface area is found to increase with the increase in phosphate content up to 7.5 wt% loading and thereafter decreases. The average pore diameter and crystallite size of titania decreases with the addition of phosphate. However, total acidity (determined by base adsorption method) and the catalytic activity increases with the increase in phosphate content up to 10 wt%. Phosphated samples prepared using phosphoric acid as the source of phosphate exhibit higher acidity compared to the samples prepared using (NH(4))(3)PO(4). However, the sample prepared from (NH(4))(3)PO(4) shows the presence of both acid and basic sites. Though from the cumene conversion study it is understood that phosphated samples contain both Lewis and Brønsted acid sites, the latter predominates over the former. Copyright 1999 Academic Press.

  13. Dietary P regulates phosphate transporter expression, phosphatase activity, and effluent P partitioning in trout culture.

    PubMed

    Coloso, R M; King, K; Fletcher, J W; Weis, P; Werner, A; Ferraris, R P

    2003-08-01

    Phosphate utilization by fish is an important issue because of its critical roles in fish growth and aquatic environmental pollution. High dietary phosphorus (P) levels typically decrease the efficiency of P utilization, thereby increasing the amount of P excreted as metabolic waste in effluents emanating from rainbow trout aquaculture. In mammals, vitamin D3 is a known regulator of P utilization but in fish, its regulatory role is unclear. Moreover, the effects of dietary P and vitamin D3 on expression of enzymatic and transport systems potentially involved in phosphate utilization are little known. We therefore monitored production of effluent P, levels of plasma vitamin D3 metabolites, as well as expression of phosphatases and the sodium phosphate cotransporter (NaPi2) in trout fed semipu diets that varied in dietary P and vitamin D3 levels. Mean soluble P concentrations varied markedly with dietary P but not with vitamin D3, and constituted 40-70% of total effluent P production by trout. Particulate P concentrations accounted for 25-50% of effluent P production, but did not vary with dietary P or vitamin D3. P in settleable wastes accounted for <10% of effluent P. The stronger effect of dietary P on effluent P levels is paralleled by its striking effects on phosphatases and NaPi2. The mRNA abundance of the intestinal and renal sodium phosphate transporters increased in fish fed low dietary P; vitamin D3 had no effect. Low-P diets reduced plasma phosphate concentrations. Intracellular phytase activity increased but brushborder alkaline phosphatase activity decreased in the intestine, pyloric caeca, and gills of trout fed diets containing low dietary P. Vitamin D3 had no effect on enzyme activities. Moreover, plasma concentrations of 25-hydroxyvitamin D3 and of 1,25-dihydroxyvitamin D3 were unaffected by dietary P and vitamin D3 levels. The major regulator of P metabolism, and ultimately of levels of P in the effluent from trout culture, is dietary P.

  14. Loaded Ce-Ag organic-inorganic hybrids and their antibacterial activity.

    PubMed

    Truffault, Laurianne; Rodrigues, Danilo Fernando; Salgado, Hérida Regida Nunes; Santilli, Celso Valentim; Pulcinelli, Sandra Helena

    2016-11-01

    There are requirements for surfaces with antibacterial properties in various technological fields. U-PEO hybrids with antibacterial properties were synthesized by the sol-gel process, incorporating combinations of cerium and silver salts at different silver molar fractions (0, 0.02, 0.05, 0.10, and 1) relative to the total amount of doped cations. The loaded hybrids were characterized by TGA, XRD, and Raman spectroscopy. Release tests were performed using UV-vis spectroscopy, and the antibacterial properties of the hybrids were studied in agar tests and turbidimetry assays. The nanostructural evolution of the hybrids during the release of the antibacterial agents was investigated by in situ SAXS. XRD results showed the presence of the AgCl crystalline phase in the loaded hybrids from a silver molar fraction of 0.05. Raman spectroscopy evidenced the interaction of silver cations with the polymeric part of the hybrid. SAXS results confirmed these interactions and showed that cerium species interacted with both organic and inorganic parts of the hybrids. The loaded U-PEO hybrids were found to release all the incorporated cerium in 1h, while the hybrid containing 100% of silver released only 78% of the incorporated silver. All the loaded hybrids displayed antibacterial activity against the Pseudomonas aeruginosa bacterium. The antibacterial activity was found to increase with silver molar fraction. Due to its high antibacterial activity and low silver molar fraction, the loaded hybrid with silver molar fraction of 0.10 seemed to be a good compromise between efficiency, esthetic transparency, and photostability.

  15. Type I phosphatidylinositol 4-phosphate 5-kinase homo- and heterodimerization determines its membrane localization and activity.

    PubMed

    Lacalle, Rosa Ana; de Karam, Juan C; Martínez-Muñoz, Laura; Artetxe, Ibai; Peregil, Rosa M; Sot, Jesús; Rojas, Ana M; Goñi, Félix M; Mellado, Mario; Mañes, Santos

    2015-06-01

    Type I phosphatidylinositol 4-phosphate 5-kinases (PIP5KIs; α, β, and γ) are a family of isoenzymes that produce phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] using phosphatidylinositol 4-phosphate as substrate. Their structural homology with the class II lipid kinases [type II phosphatidylinositol 5-phosphate 4-kinase (PIP4KII)] suggests that PIP5KI dimerizes, although this has not been formally demonstrated. Neither the hypothetical structural dimerization determinants nor the functional consequences of dimerization have been studied. Here, we used Förster resonance energy transfer, coprecipitation, and ELISA to show that PIP5KIβ forms homo- and heterodimers with PIP5KIγ_i2 in vitro and in live human cells. Dimerization appears to be a general phenomenon for PIP5KI isoenzymes because PIP5KIβ/PIP5KIα heterodimers were also detected by mass spectrometry. Dimerization was independent of actin cytoskeleton remodeling and was also observed using purified proteins. Mutagenesis studies of PIP5KIβ located the dimerization motif at the N terminus, in a region homologous to that implicated in PIP4KII dimerization. PIP5KIβ mutants whose dimerization was impaired showed a severe decrease in PI(4,5)P2 production and plasma membrane delocalization, although their association to lipid monolayers was unaltered. Our results identify dimerization as an integral feature of PIP5K proteins and a central determinant of their enzyme activity.

  16. Influence of plant activity and phosphates on thorium bioavailability in soils from Baotou area, Inner Mongolia.

    PubMed

    Guo, Pengran; Jia, Xiaoyu; Duan, Taicheng; Xu, Jingwei; Chen, Hangting

    2010-09-01

    Harm of thorium to living organisms is governed by its bioavailability. Thorium bioavailability in the soil-plant system of Baotou rare earth industrial area was studied using pot experiments of wheat and single extraction methods. The effects of wheat growth stage and phosphate on thorium bioavailability were also investigated. Based on extractabilities of various extraction methods (CaCl(2), NH(4)NO(3), EDTA, HOAc) and correlation analysis of thorium uptake by wheat plant and extractable thorium, a mixture of 0.02M EDTA+0.5M NH(4)OAc (pH 4.6) was found suitable for evaluation of thorium bioavailability in Baotou soil, which could be predicted quantitatively by multiple regression models. Because of differences of wheat root activities, thorium bioavailability in rhizosphere soil was higher than in bulk soil at tillering stage, but the reverse occurred at jointing stage. Phosphate addition induced the mineralization of soluble thorium by forming stable thorium phosphate compounds, and reduced thorium bioavailability in soil.

  17. Inorganic nanoparticles and the immune system: detection, selective activation and tolerance

    NASA Astrophysics Data System (ADS)

    Bastús, Neus G.; Sánchez-Tilló, Ester; Pujals, Silvia; Comenge, Joan; Giralt, Ernest; Celada, Antonio; Lloberas, Jorge; Puntes, Victor F.

    2012-03-01

    The immune system is the responsible for body integrity and prevention of external invasion. On one side, nanoparticles are no triggers that the immune system is prepared to detect, on the other side it is known that foreign bodies, not only bacteria, viruses and parasites, but also inorganic matter, can cause various pathologies such as silicosis, asbestosis or inflammatory reactions. Therefore, nanoparticles entering the body, after interaction with proteins, will be either recognized as self-agents or detected by the immune system, encompassing immunostimulation or immunosuppression responses. The nature of these interactions seems to be dictated not specially by the composition of the material but by modifications of NP coating (composition, surface charge and structure). Herein, we explore the use of gold nanoparticles as substrates to carry multifunctional ligands to manipulate the immune system in a controlled manner, from undetection to immunostimulation. Murine bone marrow macrophages can be activated with artificial nanometric objects consisting of a gold nanoparticle functionalized with peptides. In the presence of some conjugates, macrophage proliferation was stopped and pro-inflammatory cytokines were induced. The biochemical type of response depended on the type of conjugated peptide and was correlated with the degree of ordering in the peptide coating. These findings help to illustrate the basic requirements involved in medical NP conjugate design to either activate the immune system or hide from it, in order to reach their targets before being removed by phagocytes. Additionally, it opens up the possibility to modulate the immune response in order to suppress unwanted responses resulting from autoimmunity, or allergy or to stimulate protective responses against pathogens.

  18. Effect of chloroquine phosphate and toxic concentrations of lead acetate on Ca2+-ATPase activity in isolates and clones of Plasmodium falciparum.

    PubMed

    Bolaji, O M; Happi, T C; Oduola, A M J; Babafunmi, E A

    2011-12-20

    The basal activity of Ca2+-ATPase in two isolates (NL56, UNC) and two clones (D6, W2) of P.falciparum was assessed. The effects of various concentrations of chloroquine phosphate and toxic concentrations of lead acetate were also evaluated in the clones and strains of P.falciparum. The Ca2+-ATPase activity was measured by monitoring the rate of release of inorganic phosphate from the gamma-position of ATP on spectrophotometer at 820nm wavelength. The various concentrations of chloroquine (3, 6, 9, 12, 18µg/ml) and lead acetate (5, 10, 20, 30, 40µg/ml) on Ca2+-ATPase activity were measured respectively. Chloroquine phosphate inhibited Ca2+-ATPase activity in both the isolates and the cloned strains of P.falciparum in concentration dependent manner. Median Inhibitory concentration of chloroquine (MIC50) estimated from the plot of activity against chloroquine concentration was found to be 2.6mg/ml at pH 7.4 for both the isolates and cloned strains examined. Lead acetate at concentrations 5-20µg/ml inhibited Ca2+-ATPase activity in concentration dependent manner in clone W2 (Chloroquine resistant strain) while the same range of concentrations of lead acetate stimulated the activity of the enzyme in clone D6 (Chloroquine sensitive strain).The inhibitory effect of lead acetate on the enzyme in clone D6 was observed at concentrations above 20µg/ml. The result also suggests that lead ions could modulate and moderate calcium ion homeostasis in P. falciparum via its effect on Ca2+-ATPase activity. Also sufficient influx of lead ions into P. falciparum may transform the biochemical or bioenergetics nature of chloroquine sensitive strain of P. falciparum (D6) to that similar to chloroquine resistant strain (W2). In conclusion, inhibition of Ca2+-ATPase activity of P.falciparum may be part of the mechanism of action of chloroquine in its use as chemotherapy for malaria. The study implies that populations simultaneously exposed to lead pollution and malaria infection may

  19. Glutathion peroxidase and glucose-6-phosphate dehydrogenase activities in bovine blood and liver.

    PubMed

    Abd Ellah, Mahmoud Rushdi; Niishimori, Kazuhiro; Goryo, Masanobu; Okada, Keiji; Yasuda, Jun

    2004-10-01

    A total of 46 cattle, including 25 as control, 16 with glycogen degeneration and 5 with severe fatty degeneration were studied. Whole blood and liver tissue specimens were used to measure glutathione peroxidase (GSH-Px) and Glucose-6-Phosphate Dehydrogenase (G6PD) activities. The present study determined the value of these parameters in diagnosing glycogen and fatty degeneration in cattle from the point of the status of antioxidation and lipid peroxidation. The results showed a significant decrease in hepatic GSH-Px activity and a significant increase in hepatic G6PD activity in cases of fatty degeneration. On the other hand, there were no significant changes in erythrocytic and hepatic GSH-Px and G6PD activities in cases of glycogen degeneration. The results indicated lipoperoxidation process in the liver tissues increased in cases of fatty degeneration. Therefore, supplying animals suffering from fatty liver with sufficient quantities of nutrient antioxidants may be valuable when treatment is considered.

  20. Protein phosphorylation as a mechanism for regulation of spinach leaf sucrose-phosphate synthase activity

    SciTech Connect

    Huber, J.L.A.; Huber, S.C. )

    1989-04-01

    Protein phosphorylation has been identified as a mechanism for the light-dark regulation of spinach sucrose-phosphate synthase (SPS) activity, previously shown to involve some type of covalent modification of the enzyme. The 120 kD subunit of SPS in extracts of light-treated leaves was labeled with {sup 32}P in the presence of ({gamma}-{sup 32}P) ATP. In this in vitro system, {sup 32}P incorporation into light-activated SPS was dependent upon ATP and magnesium concentrations as well as time, and was closely paralleled by inactivation of the enzyme. The soluble protein kinase involved in the interconversion of SPS between activated and deactivated forms may be specific for SPS as it co-purifies with SPS during partial purification of the enzyme. The kinase appears not to be calcium activated and no evidence has been obtained for metabolite control of SPS phosphorylation/inactivation.

  1. EFFECT OF PHOSPHATE ION AND 2,4-DINITROPHENOL ON THE ACTIVITY OF INTACT CELLS OF THIOBACILLUS FERROOXIDANS

    PubMed Central

    Beck, Jay V.; Shafia, Fred M.

    1964-01-01

    Beck, Jay V. (Brigham Young University, Provo, Utah), and Fred M. Shafia. Effect of phosphate ion and 2,4-dinitrophenol on the activity of cell suspensions of Thiobacillus ferrooxidans. J. Bacteriol. 88:850–857. 1964.—The rate of oxidation of ferrous salts or elemental sulfur by aged cell suspensions, phosphate-depleted cells, or 2,4-dinitrophenol (DNP)-treated cells of Thiobacillus ferrooxidans was increased by addition of orthophosphate salts. The effect was found to be transitory, with the rate gradually approaching that observed prior to phosphate ion addition. The total increased oxygen uptake was observed to be roughly proportional to the amount of phosphate salt added. The efficiency of CO2 fixation accompanying oxidation of ferrous salts was found to be about 1.7 μmoles of CO2 fixed per 100 μmoles of O2 absorbed, in contrast to a value of about 8.0 μmoles of CO2 fixed per 100 μmoles of O2 uptake during sulfur oxidation. The rate of oxidation did not affect the CO2 fixation efficiency. Whereas addition of phosphate salts to aged or phosphate-depleted cells increased slightly the already high efficiency of CO2 fixation, it did not affect the complete inhibition of CO2 fixation observed in the presence of 10-5m DNP. The results indicate that the phosphate ion is essential for oxidation of the ferrous ion, and that dinitrophenol and other so-called upcoupling agents interfere with phosphate metabolism. The latter may be a result of action at the site of assimilation of the ferrous ion or it may be an effect on the electron-transport system. In any event, it seems obvious that the phosphate ion is converted into a nonactive form in the presence of dinitrophenol-treated cells, because additional quantities of orthophosphate salts cause an immediate, marked restoration of oxidative activity. PMID:14219046

  2. Conformational and activity changes during guanidine denaturation of D-glyceraldehyde-3-phosphate dehydrogenase.

    PubMed

    Xie, G F; Tsou, C L

    1987-01-05

    Changes in intrinsic protein fluorescence of lobster muscle D-glyceraldehyde-3-phosphate dehydrogenase (D-glyceraldehyde-3-phosphate: NAD+ oxidoreductase (phosphorylating), EC 1.2.1.12) have been compared with inactivation of the enzyme during denaturation in guanidine solutions. The holoenzyme is completely inactivated at guanidine concentrations less than 0.5 M and this is accompanied by a red shift of the emission maximum at 335 nm and a marked decrease in intensity of the intrinsic fluorescence. At 0.5 M guanidine, the inactivation is a slow process, with a first-order rate constant of 2.4 X 10(-3) s-1. A further red shift in the emission maximum and a decrease in intensity occur at guanidine concentrations higher than 1.5 M. The emission peak at 410 nm of the fluorescent NAD derivative introduced at the active site of this enzyme (Tsou, C.L. et al. (1983) Biochem. Soc. Trans. 11, 425-429) shows both a red shift and a marked decrease in intensity at the same guanidine concentration required to bring about the inactivation and the initial changes in the intrinsic fluorescence of the holoenzyme. It appears that treatment by low guanidine concentrations leads to both complete inactivation and perturbation of the active site conformation and that a tryptophan residue is situated at or near the active site.

  3. On the reactive adsorption of ammonia on activated carbons modified by impregnation with inorganic compounds.

    PubMed

    Bandosz, Teresa J; Petit, Camille

    2009-10-15

    Ammonia adsorption was studied under dynamic conditions, at room temperature, on activated carbons of different origins (coal-based, wood-based and coconut-shell-based carbons) before and after their impregnation with various inorganic compounds including metal chlorides, metal oxides and polycations. The role of humidity was evaluated by running tests in both dry and moist conditions. Adsorbents were analyzed before and after exposure to ammonia by thermal analyses, sorption of nitrogen, potentiometric titration, X-ray diffraction and FTIR spectroscopy. Results of breakthrough tests show significant differences in terms of adsorption capacity depending on the parent carbon, the impregnates and the experimental conditions. It is found that surface chemistry governs ammonia adsorption on the impregnated carbons. More precisely, it was demonstrated that a proper combination of the surface pH, the strength, type and amount of functional groups present on the adsorbents' surface is a key point in ammonia uptake. Water can have either positive or negative effects on the performance of adsorbents. It can enhance NH(3) adsorption capacity since it favors ammonia dissolution and thus enables reaction between ammonium ions and carboxylic groups from the carbons' surface. On the other hand, water can also reduce the performance from the strength of adsorption standpoint. It promotes dissolution of ammonia and that ammonia is first removed from the system when the adsorbent bed is purged with air. Ammonia, besides adsorption by van der Waals forces and dissolution in water, is also retained on the surface via reactive mechanisms such as acid-base reactions (Brønsted and Lewis) or complexation. Depending on the materials used and the experimental conditions, 6-47% ammonia adsorbed is strongly retained on the surface even when the bed is purged with air.

  4. On the reactive adsorption of ammonia on activated carbons modified by impregnation with inorganic compounds

    SciTech Connect

    Bandosz, T.J.; Petit, C.

    2009-10-15

    Ammonia adsorption was studied under dynamic conditions, at room temperature, on activated carbons of different origins (coal-based, wood-based and coconut-shell-based carbons) before and after their impregnation with various inorganic compounds including metal chlorides, metal oxides and polycations. The role of humidity was evaluated by running tests in both dry and moist conditions. Adsorbents were analyzed before and after exposure to ammonia by thermal analyses, sorption of nitrogen, potentiometric titration, X-ray diffraction and FTIR spectroscopy. Results of breakthrough tests show significant differences in terms of adsorption capacity depending on the parent carbon, the impregnates and the experimental conditions. It is found that surface chemistry governs ammonia adsorption on the impregnated carbons. More precisely, it was demonstrated that a proper combination of the surface pH, the strength, type and amount of functional groups present on the adsorbents' surface is a key point in ammonia uptake. Water can have either positive or negative effects on the performance of adsorbents. It can enhance NH{sub 3} adsorption capacity since it favors ammonia dissolution and thus enables reaction between ammonium ions and carboxylic groups from the carbons' surface. On the other hand, water can also reduce the performance from the strength of adsorption standpoint. It promotes dissolution of ammonia and that ammonia is first removed from the system when the adsorbent bed is purged with air. Ammonia, besides adsorption by van der Waals forces and dissolution in water, is also retained on the surface via reactive mechanisms such as acid-base reactions (Bronsted and Lewis) or complexation. Depending on the materials used and the experimental conditions, 6-47% ammonia adsorbed is strongly retained on the surface even when the bed is purged with air.

  5. Spontaneous activation of [FeFe]-hydrogenases by an inorganic [2Fe] active site mimic

    PubMed Central

    Esselborn, Julian; Berggren, Gustav; Noth, Jens; Siebel, Judith; Hemschemeier, Anja; Artero, Vincent; Reijerse, Edward; Fontecave, Marc; Lubitz, Wolfgang; Happe, Thomas

    2013-01-01

    Hydrogenases catalyze the formation of hydrogen. The cofactor (H-cluster) of [FeFe]-hydrogenases consists of a [4Fe-4S]-cluster bridged to a unique [2Fe]-subcluster whose biosynthesis in vivo requires hydrogenase-specific maturases. Here we show that a chemical mimic of the [2Fe]-subcluster can reconstitute apo-hydrogenase to full activity, independent of helper proteins. The assembled H-cluster is virtually indistinguishable from the native cofactor. This procedure will be a powerful tool for developing novel artificial H2-producing catalysts. PMID:23934246

  6. Inorganic arsenic represses interleukin-17A expression in human activated Th17 lymphocytes

    SciTech Connect

    Morzadec, Claudie; Macoch, Mélinda; Robineau, Marc; Sparfel, Lydie; Fardel, Olivier; Vernhet, Laurent

    2012-08-01

    Trivalent inorganic arsenic [As(III)] is an efficient anticancer agent used to treat patients suffering from acute promyelocytic leukemia. Recently, experimental studies have clearly demonstrated that this metalloid can also cure lymphoproliferative and/or pro-inflammatory syndromes in different murine models of chronic immune-mediated diseases. T helper (Th) 1 and Th17 lymphocytes play a central role in development of these diseases, in mice and humans, especially by secreting the potent pro-inflammatory cytokine interferon-γ and IL-17A, respectively. As(III) impairs basic functions of human T cells but its ability to modulate secretion of pro-inflammatory cytokines by differentiated Th lymphocytes is unknown. In the present study, we demonstrate that As(III), used at concentrations clinically achievable in plasma of patients, has no effect on the secretion of interferon-γ from Th1 cells but almost totally blocks the expression and the release of IL-17A from human Th17 lymphocytes co-stimulated for five days with anti-CD3 and anti-CD28 antibodies, in the presence of differentiating cytokines. In addition, As(III) specifically reduces mRNA levels of the retinoic-related orphan receptor (ROR)C gene which encodes RORγt, a key transcription factor controlling optimal IL-17 expression in fully differentiated Th17 cells. The metalloid also blocks initial expression of IL-17 gene induced by the co-stimulation, probably in part by impairing activation of the JNK/c-Jun pathway. In conclusion, our results demonstrate that As(III) represses expression of the major pro-inflammatory cytokine IL-17A produced by human Th17 lymphocytes, thus strengthening the idea that As(III) may be useful to treat inflammatory immune-mediated diseases in humans. -- Highlights: ► Arsenic inhibits secretion of IL-17A from human naïve and memory Th17 lymphocytes. ► Arsenic represses early expression of IL-17A gene in human activated T lymphocytes. ► Arsenic interferes with activation of

  7. [Adsorption Characteristics of Nitrate and Phosphate from Aqueous Solution on Zirconium-Hexadecyltrimethylammonium Chloride Modified Activated Carbon].

    PubMed

    Zheng, Wen-jing; Lin, Jian-wei; Zhan, Yan-hui; Wang, Hong

    2015-06-01

    A novel adsorbent material, i.e., zirconium-cationic surfactant modified activated carbon (ZrSMAC) was prepared by loading zirconium hydroxide and hexadecyltrimethylammonium chloride (CTAC) on activated carbon, and was used as an adsorbent for nitrate and phosphate removal from aqueous solution. The adsorption characteristics of nitrate and phosphate on ZrSMAC from aqueous solution were investigated in batch mode. Results showed that the ZrSMAC was effective for nitrate and phosphate removal from aqueous solution. The pseudo-second-order kinetic model fitted both the nitrate and phosphate kinetic experimental data well. The equilibrium isotherm data of nitrate adsorption onto the ZrSMAC were well fitted to the Langmuir, Dubinin-Radushkevich (D-R) and Freundlich isotherm models. The equilibrium isotherm data of phosphate adsorption onto the ZrSMAC could be described by the Langmuir and,D- R isotherm models. According to the Langmuir isotherm model, the maximum nitrate and phosphate adsorption capacities for the ZrSMAC were 7.58 mg x g(-1) and 10.9 mg x g(-1), respectively. High pH value was unfavorable for nitrate and phosphate adsorption onto the ZrSMAC. The presence of Cl-, HCO3- and SO4(2-) in solution reduced the nitrate and phosphate adsorption capacities for the ZrSMAC. The nitrate adsorption capacity for the ZrSMAC was reduced by the presence of coexisting phosphate in solution, and the phosphate adsorption capacity for the ZrSMAC was also reduced by the presence of coexisting nitrate in solution. About 90% of nitrate adsorbed on the ZrSMAC could be desorbed in 1 mol x L(-1) NaCl solution, and about 78% of phosphate adsorbed on the ZrSMAC could be desorbed in 1 mol x L(-1) NaOH solution. The adsorption mechanism of nitrate on the ZrSMAC included the anion exchange interactions and electrostatic attraction, and the adsorption mechanism of phosphate on the ZrSMAC included the ligand exchange interaction, electrostatic attraction and anion exchange interaction.

  8. Lupus anticoagulant activities of murine monoclonal antibodies to liposomal phosphatidylinositol phosphate.

    PubMed Central

    Alving, B M; Banerji, B; Fogler, W E; Alving, C R

    1987-01-01

    Four murine monoclonal antibodies having high levels of activity against phosphatidyl-inositol phosphate (PIP) were tested for lupus anticoagulant activity. The antibodies showed different degrees of potency in a modified partial thromboplastin time test (APTT) that used dilutions of either bovine brain extract (Thrombofax) or liposomes consisting of phosphatidylcholine/phosphatidylserine (PC/PS) as the phospholipid source. The same relative order of anticoagulant potency that was observed in the APTT that used the PC/PS liposomes was maintained when the anti-PIP antibodies were tested for cross-reactivity either by induction of complement-dependent immune damage to liposomes containing PS, or in enzyme-linked immunosorbent assays that used PS, cardiolipin (CL), or phosphatidylinositol (PI) as antigens. The data indicate that monoclonal antibodies to PIP can express anticoagulant activity in a modified APTT that correlates with their different degrees of cross-reactivity against the negatively-charged phospholipids PS, CL, and PI. PMID:2820640

  9. Enhancing the value of commodity polymers: Part 1. Structure-property relationships in composite materials based on maleated polypropylene/inorganic phosphate glasses. Part 2. New value-added applications for polyesters

    NASA Astrophysics Data System (ADS)

    Gupta, Mohit

    The first part of the thesis (Chapters 2 & 3) describes a new class of organic polymer/inorganic glass composite materials with property improvements that are impossible to achieve with classical polymer blends or composites. These materials exhibit good processability, superior mechanical performance, good thermal stability, and have excellent gas barrier properties. Low glass transition temperature phosphate glasses (Pglass) are used as inorganic fillers and slightly maleated polypropylene is used as the organic polymer matrix. The Pglass, which was dispersed as spherical droplets in the unoriented composites can be elongated into high aspect ratio platelets during the biaxial stretching process. Biaxially oriented films exhibited a brick wall type microstructure with highly aligned inorganic platelets in a ductile organic matrix and the oxygen barrier properties are significantly improved due to presence of Pglass platelets as impermeable inclusions. Mechanical properties of the biaxially oriented films showed significant improvements compared to neat polymer due to uniform dispersion of the Pglass platelets. Properly dispersed and aligned platelets have proven to be very effective for increasing the composite modulus. These developed materials therefore show promise to help fulfill the ever increasing demand for new advanced materials for a wide variety of advanced packaging applications because of their gas barrier properties, flexibility, transparency, mechanical strength and performance under humid conditions. The second part of the thesis (Chapters 4 & 5) describes new value-added applications for polyesters. Chapter 4 reports a novel process for the decolorization of green and blue colored PET bottle flakes using hydrogen peroxide. The decolorized flakes were characterized for color, intrinsic viscosity values. Decolorized flakes exhibited color values similar to those of colorless recycled PET and even though IV values decreased, bleached flakes still

  10. Phosphate, inositol and polyphosphates.

    PubMed

    Livermore, Thomas M; Azevedo, Cristina; Kolozsvari, Bernadett; Wilson, Miranda S C; Saiardi, Adolfo

    2016-02-01

    Eukaryotic cells have ubiquitously utilized the myo-inositol backbone to generate a diverse array of signalling molecules. This is achieved by arranging phosphate groups around the six-carbon inositol ring. There is virtually no biological process that does not take advantage of the uniquely variable architecture of phosphorylated inositol. In inositol biology, phosphates are able to form three distinct covalent bonds: phosphoester, phosphodiester and phosphoanhydride bonds, with each providing different properties. The phosphoester bond links phosphate groups to the inositol ring, the variable arrangement of which forms the basis of the signalling capacity of the inositol phosphates. Phosphate groups can also form the structural bridge between myo-inositol and diacylglycerol through the phosphodiester bond. The resulting lipid-bound inositol phosphates, or phosphoinositides, further expand the signalling potential of this family of molecules. Finally, inositol is also notable for its ability to host more phosphates than it has carbons. These unusual organic molecules are commonly referred to as the inositol pyrophosphates (PP-IPs), due to the presence of high-energy phosphoanhydride bonds (pyro- or diphospho-). PP-IPs themselves constitute a varied family of molecules with one or more pyrophosphate moiety/ies located around the inositol. Considering the relationship between phosphate and inositol, it is no surprise that members of the inositol phosphate family also regulate cellular phosphate homoeostasis. Notably, the PP-IPs play a fundamental role in controlling the metabolism of the ancient polymeric form of phosphate, inorganic polyphosphate (polyP). Here we explore the intimate links between phosphate, inositol phosphates and polyP, speculating on the evolution of these relationships.

  11. [Point contacts of T7 RNA polymerase in the promotor complex, as determined with phosphate-activated oligonucleotide derivatives].

    PubMed

    Filippova, S E; Ivanovskaia, M G; Romanova, E A; Tunitskaia, V L; Kochetkov, S N

    2002-01-01

    The contacts between phosphate groups of promoter DNA an Lys or His of T7 RNA polmerase (Pol) in the Pol-promoter complex were studied with single- and double- stranded oligonucleotides, which corresponded to the T7 promoter consensus and contained activated phosphate groups at position +1, +2, or -14 relative to the transcription start. To obtain reactive groups, terminal phosphates were modified with N-oxybenzotriazole (HOBT), and internucleotide phosphates were repalced with a trisubstituted pyrophosphate (TSP). The resulting derivatives produced covalent complexes with T7 Pol. Covalent bonding involved His in the case of TSP at position +1 or HOBT at position +1 or -14, and Lys in the case of TSB at position -14.

  12. Arabidopsis WRKY45 transcription factor activates PHOSPHATE TRANSPORTER1;1 expression in response to phosphate starvation.

    PubMed

    Wang, Hui; Xu, Qian; Kong, You-Han; Chen, Yun; Duan, Jun-Ye; Wu, Wei-Hua; Chen, Yi-Fang

    2014-04-01

    The WRKY transcription factor family has more than 70 members in the Arabidopsis (Arabidopsis thaliana) genome, and some of them are involved in plant responses to biotic and abiotic stresses. This study evaluated the role of WRKY45 in regulating phosphate (Pi) uptake in Arabidopsis. WRKY45 was localized in the nucleus and mainly expressed in roots. During Pi starvation, WRKY45 expression was markedly induced, typically in roots. WRKY45 overexpression in Arabidopsis increased Pi content and uptake, while RNA interference suppression of WRKY45 decreased Pi content and uptake. Furthermore, the WRKY45-overexpressing lines were more sensitive to arsenate, the analog of Pi, compared with wild-type seedlings. These results indicate that WRKY45 positively regulates Arabidopsis Pi uptake. Quantitative real-time polymerase chain reaction and β-glucuronidase staining assays showed that PHOSPHATE TRANSPORTER1;1 (PHT1;1) expression was enhanced in the WRKY45-overexpressing lines and slightly repressed in the WRKY45 RNA interference line. Chromatin immunoprecipitation and electrophoretic mobility shift assay results indicated that WRKY45 can bind to two W-boxes within the PHT1;1 promoter, confirming the role of WRKY45 in directly up-regulating PHT1;1 expression. The pht1;1 mutant showed decreased Pi content and uptake, and overexpression of PHT1;1 resulted in enhanced Pi content and uptake. Furthermore, the PHT1;1-overexpressing line was much more sensitive to arsenate than WRKY45-overexpressing and wild-type seedlings, indicating that PHT1;1 overexpression can enhance Arabidopsis Pi uptake. Moreover, the enhanced Pi uptake and the increased arsenate sensitivity of the WRKY45-overexpressing line was impaired by pht1;1 (35S:WRKY45-18::pht1;1), demonstrating an epistatic genetic regulation between WRKY45 and PHT1;1. Together, our results demonstrate that WRKY45 is involved in Arabidopsis response to Pi starvation by direct up-regulation of PHT1;1 expression.

  13. Phosphate glass core/silica clad fibres with a high concentration of active rare-earth ions

    NASA Astrophysics Data System (ADS)

    Egorova, O. N.; Galagan, B. I.; Denker, B. I.; Sverchkov, S. E.; Semjonov, S. L.

    2016-12-01

    We report a study of silica-clad composite optical fibres having a phosphate glass core doped with active rare-earth elements. The phosphate glass core allows a high concentration of active rare-earth ions to be obtained, and the silica cladding ensures high mechanical strength and facilitates fusion splicing of such fibres to silica fibres. Owing to the high concentration of active rare-earth ions, this type of fibre is potentially attractive for applications where a small cavity length and high lasing efficiency are needed.

  14. Cytocompatibility and osteogenic activity of a novel calcium phosphate silicate bioceramic: Silicocarnotite.

    PubMed

    Duan, Wei; Ning, Congqin; Tang, Tingting

    2013-07-01

    In the present study, the effect of a novel bioceramic, silicon-containing calcium phosphate ceramic (silicocarnotite, Ca5 (PO4 )2 SiO4 , CPS) on attachment, proliferation, and osteogenic differentiation of rat bone marrow-derived mesenchymal stem cells (rBMSC) has been investigated in comparison to hydroxyapatite (HA). The CPS showed a similar cell attachment behavior to HA, while the proliferation of rBMSC on CPS was significantly higher than that on HA, which indicated that CPS had a good cytocompatibility. Moreover, the expression of alkaline phosphatase activity and osteogenic-related genes, including Runx-2, osteopontin (OPN), bone sialoprotein (BSP) and osteocalcin (OC), demonstrated that CPS enhanced the osteogenic differentiation of rBMSC and accelerated the differentiation process. The results suggest that CPS ceramic exhibits a good cytocompatibility and osteogenic activity, which might be used as a potential candidate material for bone tissue engineering.

  15. 3D correlation NMR spectrum between three distinct heteronuclei for the characterization of inorganic samples: Application on sodium alumino-phosphate materials.

    PubMed

    Nagashima, Hiroki; Tricot, Grégory; Trébosc, Julien; Lafon, Olivier; Amoureux, Jean-Paul; Pourpoint, Frédérique

    2017-03-22

    We report here an original NMR sequence allowing the acquisition of 3D correlation NMR spectra between three distinct heteronuclei, among which two are half-integer spin quadrupolar nuclei. Furthermore, as two of them exhibit close Larmor frequency, this experiment was acquired using a standard triple-resonance probe equipped with a commercial frequency splitter. This NMR technique was tested and applied to sodium alumino-phosphate compounds with (31)P as the spin-1/2 nucleus and (23)Na and (27)Al as the close Larmor frequencies isotopes. To the best of our knowledge, such experiment with direct (31)P and indirect (27)Al and (23)Na detection is the first example of 3D NMR experiment in solids involving three distinct heteronuclei. This sequence has first been demonstrated on a mixture of Al(PO3)3 and NaAlP2O7 crystalline phases, for which a selective observation of NaAlP2O7 is possible through the 3D map edition. This 3D correlation experiment is then applied to characterize mixing and phase segregation in a partially devitrified glass that has been proposed as a material for the sequestration of radioactive waste. The (31)P-{(23)Na,(27)Al} 3D experiment conducted on the partially devitrified glass material conclusively demonstrates that the amorphous component of the material does not contain aluminum. The as-synthesized material thus presents a poor resistance against water, which is a severe limitation for its application in the radioactive waste encapsulation domain.

  16. Deletion of hexose-6-phosphate dehydrogenase activates the unfolded protein response pathway and induces skeletal myopathy.

    PubMed

    Lavery, Gareth G; Walker, Elizabeth A; Turan, Nil; Rogoff, Daniela; Ryder, Jeffery W; Shelton, John M; Richardson, James A; Falciani, Francesco; White, Perrin C; Stewart, Paul M; Parker, Keith L; McMillan, Daniel R

    2008-03-28

    Hexose-6-phosphate dehydrogenase (H6PD) is the initial component of a pentose phosphate pathway inside the endoplasmic reticulum (ER) that generates NADPH for ER enzymes. In liver H6PD is required for the 11-oxoreductase activity of 11beta-hydroxysteroid dehydrogenase type 1, which converts inactive 11-oxo-glucocorticoids to their active 11-hydroxyl counterparts; consequently, H6PD null mice are relatively insensitive to glucocorticoids, exhibiting fasting hypoglycemia, increased insulin sensitivity despite elevated circulating levels of corticosterone, and increased basal and insulin-stimulated glucose uptake in muscles normally enriched in type II (fast) fibers, which have increased glycogen content. Here, we show that H6PD null mice develop a severe skeletal myopathy characterized by switching of type II to type I (slow) fibers. Running wheel activity and electrically stimulated force generation in isolated skeletal muscle are both markedly reduced. Affected muscles have normal sarcomeric structure at the electron microscopy level but contain large intrafibrillar membranous vacuoles and abnormal triads indicative of defects in structure and function of the sarcoplasmic reticulum (SR). SR proteins involved in calcium metabolism, including the sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA), calreticulin, and calsequestrin, show dysregulated expression. Microarray analysis and real-time PCR demonstrate overexpression of genes encoding proteins in the unfolded protein response pathway. We propose that the absence of H6PD induces a progressive myopathy by altering the SR redox state, thereby impairing protein folding and activating the unfolded protein response pathway. These studies thus define a novel metabolic pathway that links ER stress to skeletal muscle integrity and function.

  17. Adsorption between TC-stabilized AuNPs and the phosphate group: application of the PTP1B activity assay.

    PubMed

    Lv, Jun; Wang, Xiaonan; Zhang, Yuanyuan; Li, Defeng; Zhang, Juan; Sun, Lizhou

    2015-12-07

    Based on the adsorption between tetracycline (TC) and phosphate groups, a general colorimetric method is explored in this work by using TC-stabilized gold nanoparticles (TC/AuNPs) and 4-aminophenyl phosphate-functionalized Fe3O4 magnetic nanoparticles (APP/MNPs). Taking protein tyrosine phosphatase 1B (PTP1B) as an example, 4-aminophenyl phosphate (APP) can be hydrolyzed into 4-aminophenol (AP) by PTP1B, resulting in the disappearance of the phosphate group on the outer layer of MNPs and the loss of corresponding adsorptive ability. Upon addition of TC/AuNP solution, TC/AuNPs will remain in the supernatant solution after magnetic separation and a high absorbance value can be observed. So PTP1B activity is related to the concentrations of TC/AuNPs in the supernatant solution. In this work, the enzyme activity can be determined at levels as low as 0.0885 U mL(-1) and over a linear detection range as wide as 0.1 U mL(-1) to 0.9 U mL(-1). Moreover, using the proposed method, the inhibition effect of betulinic acid (BA) and sodium orthovanadate (Na3VO4) on PTP1B activity can be tested with IC50 values of 30 μM and 4 μM, respectively. Therefore, a universal platform for the accurate colorimetric analysis of kinase and phosphatase activities can be established through the adsorption between TC and phosphate groups.

  18. Chromium liquid waste inertization in an inorganic alkali activated matrix: leaching and NMR multinuclear approach.

    PubMed

    Ponzoni, Chiara; Lancellotti, Isabella; Barbieri, Luisa; Spinella, Alberto; Saladino, Maria Luisa; Martino, Delia Chillura; Caponetti, Eugenio; Armetta, Francesco; Leonelli, Cristina

    2015-04-09

    A class of inorganic binders, also known as geopolymers, can be obtained by alkali activation of aluminosilicate powders at room temperature. The process is affected by many parameters (curing time, curing temperature, relative humidity etc.) and leads to a resistant matrix usable for inertization of hazardous waste. In this study an industrial liquid waste containing a high amount of chromium (≈ 2.3 wt%) in the form of metalorganic salts is inertized into a metakaolin based geopolymer matrix. One of the innovative aspects is the exploitation of the water contained in the waste for the geopolymerization process. This avoided any drying treatment, a common step in the management of liquid hazardous waste. The evolution of the process--from the precursor dissolution to the final geopolymer matrix hardening--of different geopolymers containing a waste amount ranging from 3 to 20%wt and their capability to inertize chromium cations were studied by: i) the leaching tests, according to the EN 12,457 regulation, at different curing times (15, 28, 90 and 540 days) monitoring releases of chromium ions (Cr(III) and Cr(VI)) and the cations constituting the aluminosilicate matrix (Na, Si, Al); ii) the humidity variation for different curing times (15 and 540 days); iii) SEM characterization at different curing times (28 and 540 days); iv) the trend of the solution conductivity and pH during the leaching test; v) the characterization of the short-range ordering in terms of TOT bonds (where T is Al or Si) by (29)Si and (27)Al solid state magic-angle spinning nuclear magnetic resonance (ss MAS NMR) for geopolymers containing high amounts of waste (10-20%wt). The results show the formation of a stable matrix after only 15 days independently on the waste amount introduced; the longer curing times increase the matrices stabilities and their ability to immobilize chromium cations. The maximum amount of waste that can be inertized is around 10 wt% after a curing time of 28 days.

  19. Synthesis and evaluation of chromogenic and fluorogenic substrates for high-throughput detection of enzymes that hydrolyze inorganic polyphosphate.

    PubMed

    Hebbard, Carleigh F F; Wang, Yan; Baker, Catherine J; Morrissey, James H

    2014-08-11

    Inorganic polyphosphates, linear polymers of orthophosphate, occur naturally throughout biology and have many industrial applications. Their biodegradable nature makes them attractive for a multitude of uses, and it would be important to understand how polyphosphates are turned over enzymatically. Studies of inorganic polyphosphatases are, however, hampered by the lack of high-throughput methods for detecting and quantifying rates of polyphosphate degradation. We now report chromogenic and fluorogenic polyphosphate substrates that permit spectrophotometric monitoring of polyphosphate hydrolysis and allow for high-throughput analyses of both endopolyphosphatase and exopolyphosphatase activities, depending on assay configuration. These substrates contain 4-nitrophenol or 4-methylumbelliferone moieties that are covalently attached to the terminal phosphates of polyphosphate via phosphoester linkages formed during reactions mediated by EDAC (1-ethyl-3-(3-(dimethylamino)propyl)carbodiimide). This report identifies Nudt2 as an inorganic polyphosphatase and also adds to the known coupling chemistry for polyphosphates, permitting facile covalent linkage of alcohols with the terminal phosphates of inorganic polyphosphate.

  20. FTY720 Phosphate Activates Sphingosine-1-Phosphate Receptor 2 and Selectively Couples to Gα12/13/Rho/ROCK to Induce Myofibroblast Contraction.

    PubMed

    Sobel, Katrin; Monnier, Lucile; Menyhart, Katalin; Bolinger, Matthias; Studer, Rolf; Nayler, Oliver; Gatfield, John

    2015-06-01

    FTY720 phosphate (FTY720-P; 2-amino-2-[2-(4-octylphenyl)ethyl]-1,3-propanediol, monodihydrogen phosphate ester) is a nonselective sphingosine-1-phosphate (S1P) receptor agonist thought to be devoid of activity at the S1P2 receptor subtype. However, we have recently shown that FTY720-P displays significant S1P2 receptor agonist activity in recombinant cells and fibroblasts expressing endogenous S1P2 receptors. To elucidate the S1P2-dependent signaling pathways that were activated by FTY720-P, we employed second messenger assays and impedance-based assays in combination with pharmacological and small interfering RNA-based pathway inhibition in recombinant Chinese hamster ovary (CHO)-S1P2 cells as well as human lung myofibroblasts generated in vitro. In CHO-S1P2 cells, FTY720-P did not modulate cAMP or calcium levels. However, reporter-gene assays, impedance-based assays with a selective Rho-associated kinase (ROCK) inhibitor, Gα12/13 knockdown and activated Rho-pull-down assays demonstrated that FTY720-P potently activated Gα12/13/Rho/ROCK signaling. S1P similarly activated Gα12/13/Rho/ROCK signaling via S1P2 receptors, whereas the two selective S1P1 receptor agonists (Z,Z)-5-(3-chloro-4-[(2R)-2,3-dihydroxy-propoxy]-benzylidene)-2-propylimino-3-o-tolyl-thiazolidin-4-one (ponesimond) and 5-[4-phenyl-5-(trifluoromethyl)thiophen-2-yl]-3-[3-(trifluoromethyl)phenyl]1,2,4-oxadiazole (SEW2871) were inactive. In lung myofibroblasts, which mainly expressed the S1P2 receptor subtype, we showed that FTY720-P selectively activated the Gα12/13/Rho/ROCK pathway via the S1P2 receptor. Moreover, the activation of the Gα12/13/Rho/ROCK pathway in myofibroblasts by FTY720-P caused potent myofibroblast contraction similar to that induced by the natural ligand S1P. Thus, complementing second messenger assays with unbiased label-free assays or phenotypic assays in native expression systems can uncover activation of additional pathways, such as Gα12/13/Rho/ROCK signaling.

  1. Effect of chronologic age on induction of cystathionine synthase, uroporphyrinogen I synthase, and glucose-6-phosphate dehydrogenase activities in lymphocytes.

    PubMed Central

    Gartler, S M; Hornung, S K; Motulsky, A G

    1981-01-01

    The activities of cystathionine synthase [L-serine hydro-lyase (adding homocysteine), EC 4.2.1.22], uroporphyrinogen I synthase [porphobilinogen ammonia-lyase (polymerizing), EC 4.3.1.8], and glucose-6-phosphate dehydrogenase (D-glucose-6-phosphate:NADP+ 1-oxidoreductase, EC 1.1.1.49) have been measured in phytohemagglutinin-stimulated lymphocytes of young and old human subjects. A significant decrease in activity with age was observed for cystathionine synthase and uroporphyrinogen I synthase but not for glucose-6-phosphate dehydrogenase. These changes could not be related to declining phytohemagglutinin response with aging. Age-related decreases in activity of some enzymes may be relevant for an understanding of the biology of aging. False assignment of heterozygosity, and even homozygosity, for certain genetic disorders, such as homocystinuria, may result when low enzyme levels are detected in the lymphocytes of older people. PMID:6940198

  2. Dietary phosphorus regulates intestinal transport and plasma concentrations of phosphate in rainbow trout.

    PubMed

    Avila, E M; Tu, H; Basantes, S; Ferraris, R P

    2000-05-01

    Intestinal inorganic phosphate transport and its regulation have not been studied in fish. In this study, we initially characterized the mechanisms of intestinal inorganic phosphate transport in rainbow trout (Oncorhynchus mykiss) then determined the effects of dietary phosphorus concentrations on intestinal inorganic phosphate uptake, plasma inorganic phosphate, and intestinal luminal inorganic phosphate concentrations. In 11-g trout, the saturable mechanism of brushborder inorganic phosphate uptake had a Kt= 1.2 mmol l(-1) and a Vmax = 0.22 nmol mg(-1) min(-1), while the diffusive component had a Kd = 0.012 min(-1). Similar kinetic constants were obtained from 51-g trout, suggesting that development or size had little effect on transport. Tracer inorganic phosphate (1.18 mmol l(-1)) uptake was almost completely inhibited (>95%) by 20 mmol l(-1) unlabeled inorganic phosphate. Inorganic phosphate uptake (0.2 mmol l(-1)) was strongly inhibited (approximately 75% inhibition) by phosphonoformic acid, a competitive inhibitor of mammalian inorganic phosphate transport, as well as by the absence of Na+ (approximately 90% inhibition). Northern blot and reverse transcription-polymerase chain reaction indicated that the intestinal inorganic phosphate transporter in trout is not related to the cloned Na+ inorganic phosphate-II transporter of winter flounder. Intestinal luminal and plasma inorganic phosphate concentrations each increased with dietary P concentrations. Intestinal inorganic phosphate, but not proline, absorption rates decreased with dietary phosphorus concentrations. As in mammals and birds, a Na-dependent inorganic phosphate carrier that is tightly regulated by diet is present in trout small intestine.

  3. A rapid and simple method for estimating sulfate reduction activity and quantifying inorganic sulfides

    USGS Publications Warehouse

    Ulrich, G.A.; Krumholz, L.R.; Suflita, J.M.

    1997-01-01

    A simplified passive extraction procedure for quantifying reduced inorganic sulfur compounds from sediments and water is presented. This method may also be used for the estimation of sulfate reduction rates. Efficient extraction of FeS, FeS(inf2), and S(sup2-) was obtained with this procedure; however, the efficiency for S(sup0) depended on the form that was tested. Passive extraction can be used with samples containing up to 20 mg of reduced sulfur. We demonstrated the utility of this technique in a determination of both sulfate reduction rates and reduced inorganic sulfur pools in marine and freshwater sediments. A side-by-side comparison of the passive extraction method with the established single-step distillation technique yielded comparable results with a fraction of the effort.

  4. Antibacterial and photocatalytic activity of TiO2 and ZnO nanomaterials in phosphate buffer and saline solution.

    PubMed

    Ng, Alan Man Ching; Chan, Charis May Ngor; Guo, Mu Yao; Leung, Yu Hang; Djurišić, Aleksandra B; Hu, Xu; Chan, Wai Kin; Leung, Frederick Chi Chung; Tong, Shuk Yin

    2013-06-01

    We studied antibacterial and photocatalytic activity of anatase TiO2 and ZnO in phosphate buffer and saline solution. We found that the different anions in the suspension medium (chloride and phosphate) significantly affected the following suspension properties: the stability of nanoparticle suspension, the release of metal ions from the nanoparticles, and the production of the reactive oxygen species by the nanoparticles. As a result, antibacterial activity and photocatalytic dye degradation were also affected. However, the effect of the suspension medium was different for ZnO and TiO2. Obtained results are discussed.

  5. Rev1 is a base excision repair enzyme with 5′-deoxyribose phosphate lyase activity

    PubMed Central

    Prasad, Rajendra; Poltoratsky, Vladimir; Hou, Esther W.; Wilson, Samuel H.

    2016-01-01

    Rev1 is a member of the Y-family of DNA polymerases and is known for its deoxycytidyl transferase activity that incorporates dCMP into DNA and its ability to function as a scaffold factor for other Y-family polymerases in translesion bypass events. Rev1 also is involved in mutagenic processes during somatic hypermutation of immunoglobulin genes. In light of the mutation pattern consistent with dCMP insertion observed earlier in mouse fibroblast cells treated with a base excision repair-inducing agent, we questioned whether Rev1 could also be involved in base excision repair (BER). Here, we uncovered a weak 5′-deoxyribose phosphate (5′-dRP) lyase activity in mouse Rev1 and demonstrated the enzyme can mediate BER in vitro. The full-length Rev1 protein and its catalytic core domain are similar in their ability to support BER in vitro. The dRP lyase activity in both of these proteins was confirmed by NaBH4 reduction of the Schiff base intermediate and kinetics studies. Limited proteolysis, mass spectrometry and deletion analysis localized the dRP lyase active site to the C-terminal segment of Rev1's catalytic core domain. These results suggest that Rev1 could serve as a backup polymerase in BER and could potentially contribute to AID-initiated antibody diversification through this activity. PMID:27683219

  6. Diel activity of sucrose phosphate synthase in rice. [Oryza sativa L

    SciTech Connect

    Hussain, M.W.; Bowes, G.; Rowland-Bamford, A.J.; Allen, L.H. )

    1991-05-01

    Rice (Oryza sativa L.) was grown in growth chambers at 28/23C day/night temperatures with 16-h photoperiod at 600 umol m{sup {minus}2} s{sup {minus}1}. Diel sucrose phosphate synthase (SPS) activity, at 21 days after planting, was measured at saturating substrate concentrations. Data suggests that SPS activity increased during illumination to a maximum of 0.8 nmol mg{sup {minus}1} protein min{sup {minus}1} after 5h. Throughout the remainder of the light period there was a slow decline in activity. Upon darkening, activity further declined to 0.4 nmol mg{sup {minus}1} protein min{sup {minus}1}, a basal level that was maintained throughout the night. It appears that rice SPS undergoes light/dark transitions, suggesting there may be two kinetic forms of SPS. Changes in SPS activity will be discussed in relation to kinetic studies, and also CO{sub 2} enrichment of rice during growth.

  7. Potassium ion-activated hydrolysis of p-nitrophenyl phosphate in pancreatic islet-cell membranes.

    PubMed Central

    Lernmark, A; Parman, A; Täljedal, I B

    1977-01-01

    Hydrolysis of p-nitrophenyl phosphate was measured in a fraction enriched in plasma membranes from pancreatic islets of non-inbred ob/ob mice. Hydrolysis was stimulated by K+ (10mM) in the pH range 5--10; a small peak of K+-induced activation was observed between pH7.5 and 8. Both the K+-induced activation and the hydrolysis in the absence of K+ were Mg2+-dependent; maximum activation was obtained with 10mM-K+ plus 5 mM-Mg2+. Rb+ was as effective an activator as K+. Ouabain was inhibitory, the effect being inversely related to the K+ concentration; 0.1--0.2mM-ouabain caused about 50% inhibition in the presence of 1 mM-K+, but had no demonstrable effect in the presence of 4--5mM-K+. The K+-stimulated activity was markedly inhibited by 0.1mM-ATP, 35--140 MM-Na+, or 0.01 mM-p-chloromercuribenzenesulphonic acid. Similarities to Rb+ accumulation suggest that catalysis of univalent cation flow in pancreatic beta-cells may be coupled to a phosphoryl-transfer reaction with ATP as natural substrate or regulator. PMID:20876

  8. Osteogenic activity of cyclodextrin-encapsulated doxycycline in a calcium phosphate PCL and PLGA composite.

    PubMed

    Trajano, V C C; Costa, K J R; Lanza, C R M; Sinisterra, R D; Cortés, M E

    2016-07-01

    Composites of biodegradable polymers and calcium phosphate are bioactive and flexible, and have been proposed for use in tissue engineering and bone regeneration. When associated with the broad-spectrum antibiotic doxycycline (DOX), they could favor antimicrobial action and enhance the action of osteogenic composites. Composites of polycaprolactone (PCL), poly(lactic-co-glycolic acid) (PLGA), and a bioceramic of biphasic calcium phosphate Osteosynt® (BCP) were loaded with DOX encapsulated in β-cyclodextrin (βCD) and were evaluated for effects on osteoblastic cell cultures. The DOX/βCD composite was prepared with a double mixing method. Osteoblast viability was assessed with methyl tetrazolium (MTT) assays after 1day, 7day, and 14days of composite exposure; alkaline phosphatase (AP) activity and collagen production were evaluated after 7days and 14days, and mineral nodule formation after 14days. Composite structures were evaluated by scanning electron microscopy (SEM). Osteoblasts exposed to the composite containing 25μg/mL DOX/βCD had increased cell proliferation (p<0.05) compared to control osteoblast cultures at all experimental time points, reaching a maximum in the second week. AP activity and collagen secretion levels were also elevated in osteoblasts exposed to the DOX/βCD composite (p<0.05 vs. controls) and reached a maximum after 14days. These results were corroborated by Von Kossa test results, which showed strong formation of mineralization nodules during the same time period. SEM of the composite material revealed a surface topography with pore sizes suitable for growing osteoblasts. Together, these results suggest that osteoblasts are viable, proliferative, and osteogenic in the presence of a DOX/βCD-containing BCP ceramic composite.

  9. Impact of organic and inorganic fertilizers application on the phytochemical and antioxidant activity of Kacip Fatimah (Labisia pumila Benth).

    PubMed

    Ibrahim, Mohd Hafiz; Jaafar, Hawa Z E; Karimi, Ehsan; Ghasemzadeh, Ali

    2013-09-05

    A study was conducted to compare secondary metabolites and antioxidant activity of Labisia pumila Benth (Kacip Fatimah) in response to two sources of fertilizer [i.e., organic (chicken dung; 10% N:10% P₂O₅:10% K₂O) and inorganic fertilizer (NPK green; 15% N, 15% P₂O₅, 15% K₂O)] under different N rates of 0, 90, 180 and 270 kg N/ha. The experiment was arranged in a randomized complete block design replicated three times. At the end of 15 weeks, it was observed that the application of organic fertilizer enhanced the production of total phenolics, flavonoids, ascorbic acid, saponin and gluthathione content in L. pumila, compared to the use of inorganic fertilizer. The nitrate content was also reduced under organic fertilization. The application of nitrogen at 90 kg N/ha improved the production of secondary metabolites in Labisia pumila. Higher rates in excess of 90 kg N/ha reduced the level of secondary metabolites and antioxidant activity of this herb. The DPPH and FRAP activity was also highest at 90 kg N/ha. The results indicated that the use of chicken dung can enhance the production of secondary metabolites and improve antioxidant activity of this herb.

  10. Epidermal growth factor-induced cellular invasion requires sphingosine-1-phosphate/sphingosine-1-phosphate 2 receptor-mediated ezrin activation

    PubMed Central

    Orr Gandy, K. Alexa; Adada, Mohamad; Canals, Daniel; Carroll, Brittany; Roddy, Patrick; Hannun, Yusuf A.; Obeid, Lina M.

    2013-01-01

    Ezrin, radixin, and moesin (ERM) proteins link cortical actin to the plasma membrane and coordinate cellular events that require cytoskeletal rearrangement, including cell division, migration, and invasion. While ERM proteins are involved in many important cellular events, the mechanisms regulating their function are not completely understood. Our laboratory previously identified reciprocal roles for the sphingolipids ceramide and sphingosine-1-phosphate (S1P) in the regulation of ERM proteins. We recently showed that ceramide-induced activation of PP1α leads to dephosphorylation and inactivation of ERM proteins, while S1P results in phosphorylation and activation of ERM proteins. Following these findings, we aimed to examine known inducers of the SK/S1P pathway and evaluate their ability to regulate ERM proteins. We examined EGF, a known inducer of the SK/S1P pathway, for its ability to regulate the ERM family of proteins. We found that EGF induces ERM c-terminal threonine phosphorylation via activation of the SK/S1P pathway, as this was prevented by siRNA knockdown or pharmacological inhibition of SK. Using pharmacological, as well as genetic, knockdown approaches, we determined that EGF induces ERM phosphorylation via activation of S1PR2. In addition, EGF led to cell polarization in the form of lamellipodia, and this occurred through a mechanism involving S1PR2-mediated phosphorylation of ezrin T567. EGF-induced cellular invasion was also found to be dependent on S1PR2-induced T567 ezrin phosphorylation, such that S1PR2 antagonist, JTE-013, and expression of a dominant-negative ezrin mutant prevented cellular invasion toward EGF. In this work, a novel mechanism of EGF-stimulated invasion is unveiled, whereby S1P-mediated activation of S1PR2 and phosphorylation of ezrin T567 is required.—Orr Gandy, K. A., Adada, M., Canals, D., Carroll, B., Roddy, P., Hannun, Y. A., Obeid, L. M. Epidermal growth factor-induced cellular invasion requires sphingosine-1-phosphate

  11. Comparison of phosphate estimating methods in the presence of phytic acid for the determination of phytase activity.

    PubMed

    Sanikommu, Suma; Pasupuleti, Mukesh; Vadalkonda, Lakshmipathi

    2014-01-01

    Phosphate released from phytic acid can be used as a measure of phytase activity. However, most of the phosphate estimation methods have not examined the interference or interaction of phytic acid in the assay. In this article, we report the kinetics and influence of unreduced phytic acid on phosphate estimation by three of the often-used methods for phytase estimation, the AOAC, Cooper-Gowing, and Fiske-Subbarow methods. Our results show that the AOAC method is most suitable to estimate the phytase activity in the presence of phytate in the medium. In the Fiske and Subbarow method, we noticed that the time factor plays a role in the interference of the phytic acid; especially the readings taken during the second hour of incubation are influenced by the presence of phytic acid. The method of Cooper and Gowing is labor-intensive and is prone to give error values at higher concentrations.

  12. Antibacterial activity of plastics coated with silver-doped organic-inorganic hybrid coatings prepared by sol-gel processes.

    PubMed

    Marini, M; De Niederhausern, S; Iseppi, R; Bondi, M; Sabia, C; Toselli, M; Pilati, F

    2007-04-01

    Silver-doped organic-inorganic hybrid coatings were prepared starting from tetraethoxysilane- and triethoxysilane-terminated poly(ethylene glycol)-block-polyethylene by the sol-gel process. They were applied as a thin layer (0.6-1.1 microm) to polyethylene (PE) and poly(vinyl chloride) (PVC) films and the antibacterial activity of the coated films was tested against Gram-negative (Escherichia coli ATCC 25922) and Gram-positive (Staphylococcus aureus ATCC 6538) bacteria. The effect of several factors (such as organic-inorganic ratio, type of catalyst, time of post-curing, silver ion concentration, etc.) was investigated. Measurements at different contact times showed a rapid decrease of the viable count for both tested strains. The highest antibacterial activity [more than 6 log reduction within 6 h starting from 106 colony-forming units (cfu) mL-1] was obtained for samples with an organic-inorganic weight ratio of 80:20 and 5 wt % silver salt with respect to the coating. For the coatings prepared by an acid-catalyzed process, a high level of permanence of the antibacterial activity of the coated films was demonstrated by repeatedly washing the samples in warm water or by immersion in physiological saline solution at 37 degrees C for 3 days. The release of silver ions per square meter of coating is very similar to that previously observed for polyamides filled with metallic silver nanoparticles; however, when compared on the basis of Ag content, the concentration of silver ions released from the coating is much higher than that released from 1 mm thick specimens of polyamide (PA) filled with silver nanoparticles. Transparency and good adhesion of the coating to PE and PVC plastic substrates without any previous surface treatment are further interesting features.

  13. Sphingosine-1-phosphate mediates epidermal growth factor-induced muscle satellite cell activation.

    PubMed

    Nagata, Yosuke; Ohashi, Kazuya; Wada, Eiji; Yuasa, Yuki; Shiozuka, Masataka; Nonomura, Yoshiaki; Matsuda, Ryoichi

    2014-08-01

    Skeletal muscle can regenerate repeatedly due to the presence of resident stem cells, called satellite cells. Because satellite cells are usually quiescent, they must be activated before participating in muscle regeneration in response to stimuli such as injury, overloading, and stretch. Although satellite cell activation is a crucial step in muscle regeneration, little is known of the molecular mechanisms controlling this process. Recent work showed that the bioactive lipid sphingosine-1-phosphate (S1P) plays crucial roles in the activation, proliferation, and differentiation of muscle satellite cells. We investigated the role of growth factors in S1P-mediated satellite cell activation. We found that epidermal growth factor (EGF) in combination with insulin induced proliferation of quiescent undifferentiated mouse myoblast C2C12 cells, which are also known as reserve cells, in serum-free conditions. Sphingosine kinase activity increased when reserve cells were stimulated with EGF. Treatment of reserve cells with the D-erythro-N,N-dimethylsphingosine, Sphingosine Kinase Inhibitor, or siRNA duplexes specific for sphingosine kinase 1, suppressed EGF-induced C2C12 activation. We also present the evidence showing the S1P receptor S1P2 is involved in EGF-induced reserve cell activation. Moreover, we demonstrated a combination of insulin and EGF promoted activation of satellite cells on single myofibers in a manner dependent on SPHK and S1P2. Taken together, our observations show that EGF-induced satellite cell activation is mediated by S1P and its receptor.

  14. Residual triose phosphate isomerase activity and color measurements to determine adequate cooking of ground beef patties.

    PubMed

    Sair, A I; Booren, A M; Berry, B W; Smith, D M

    1999-02-01

    The objectives were to (i) compare the use of triose phosphate isomerase (TPI) activity and internal color scores for determination of cooking adequacy of beef patties and (ii) determine the effect of frozen storage and fat content on residual TPI activity in ground beef. Ground beef patties (24.4% fat) were cooked to five temperatures ranging from 60.0 to 82.2 degrees C. TPI activity decreased as beef patty cooking temperature was increased from 60.0 to 71.1 degrees C; however, no difference (P > 0.05) in activity (6.3 U/kg meat) was observed in patties cooked to 71.1 degrees C and above. Degree of doneness color scores, a* values and b* values, of ground beef patties decreased as internal temperature was increased from 60.0 to 71.1 degrees C; however, temperature had no effect on L* values. TPI activity in raw ground beef after five freeze-thaw cycles did not differ from the control. Three freeze-thaw cycles of raw ground beef resulted in a 57.2% decrease in TPI activity after cooking. TPI activity of cooked beef increased during 2 months of frozen storage, but TPI activity in ground beef stored for 3 months or longer did not differ from the unfrozen control. While past research has shown color to be a poor indicator of adequate thermal processing, our results suggest that undercooked ground beef patties could be distinguished from those that had been adequately cooked following U.S. Department of Agriculture guidelines using residual TPI activity as a marker.

  15. Quantification of Galactose-1-Phosphate Uridyltransferase Enzyme Activity by Liquid Chromatography–Tandem Mass Spectrometry

    PubMed Central

    Li, Yijun; Ptolemy, Adam S.; Harmonay, Lauren; Kellogg, Mark; Berry, Gerard T.

    2013-01-01

    Background The diagnosis of galactosemia usually involves the measurement of galactose-1-phosphate uridyltransferase (GALT) activity. Traditional radioactive and fluorescent GALT assays are nonspecific, laborious, and/or lack sufficient analytical sensitivity. We developed a liquid chromatography–tandem mass spectrometry (LC-MS/MS)–based assay for GALT enzyme activity measurement. Method Our assay used stable isotope-labeled α-galactose-1-phosphate ([13C6]-Gal-1-P) as an enzyme substrate. Sample cleanup and separation were achieved by reversed-phase ion-pair chromatography, and the enzymatic product, isotope-labeled uridine diphosphate galactose ([13C6]-UDPGal), was detected by MS/MS at mass transition (571 > 323) and quantified by use of [13C6]-Glu-1-P (265 > 79) as an internal standard. Results The method yielded a mean (SD) GALT enzyme activity of 23.8 (3.8) µmol · (gHgb)−1 · h−1 in erythrocyte extracts from 71 controls. The limit of quantification was 0.04 µmol · (g Hgb)−1 · h−1 (0.2% of normal control value). Intraassay imprecision was determined at 4 different levels (100%, 25%, 5%, and 0.2% of the normal control values), and the CVs were calculated to be 2.1%, 2.5%, 4.6%, and 9.7%, respectively (n = 3). Interassay imprecision CVs were 4.5%, 6.7%, 8.2%, and 13.2% (n = 5), respectively. The assay recoveries at the 4 levels were higher than 90%. The apparent Km of the 2 substrates, Gal-1-P and UDPGlc, were determined to be 0.38 mmol/L and 0.071 mmol/L, respectively. The assay in erythrocytes of 33 patients with classical galactosemia revealed no detectable activity. Conclusions This LC-MS/MS–based assay for GALT enzyme activity will be useful for the diagnosis and study of biochemically heterogeneous patients with galactosemia, especially those with uncommon genotypes and detectable but low residual activities. PMID:20348403

  16. Modulation of arginine decarboxylase activity from Mycobacterium smegmatis. Evidence for pyridoxal-5'-phosphate-mediated conformational changes in the enzyme.

    PubMed

    Balasundaram, D; Tyagi, A K

    1989-08-01

    Arginine decarboxylase (arginine carboxy-lyase, EC 4.1.1.19) from Mycobacterium smegmatis, TMC 1546 has been purified to homogeneity. The enzyme has a molecular mass of 232 kDa and a subunit mass of 58.9 kDa. The enzyme from mycobacteria is totally dependent on pyridoxal 5'-phosphate for its activity at its optimal pH and, unlike that from Escherichia coli, Mg2+ does not play an active role in the enzyme conformation. The enzyme is specific for arginine (Km = 1.6 mM). The holoenzyme is completely resolved in dialysis against hydroxylamine. Reconstitution of the apoenzyme with pyridoxal 5'-phosphate shows sigmoidal binding characteristics at pH 8.4 with a Hill coefficient of 2.77, whereas at pH 6.2 the binding is hyperbolic in nature. The kinetics of reconstitution at pH 8.4 are apparently sigmoidal, indicating the occurrence of two binding types of differing strengths. A low-affinity (Kd = 22.5 microM) binding to apoenzyme at high pyridoxal 5'-phosphate concentrations and a high-affinity (Kd = 3.0 microM) binding to apoenzyme at high pyridoxal 5'-phosphate concentrations. The restoration of full activity occurred in parallel with the tight binding (high affinity) of pyridoxal 5'-phosphate to the apoenzyme. Along with these characteristics, spectral analyses of holoenzyme and apoenzyme at pH 8.4 and pH 6.2 indicate a pH-dependent modulation of coenzyme function. Based on the pH-dependent changes in the polarity of the active-site environment, pyridoxal 5'-phosphate forms different Schiff-base tautomers at pH 8.4 and pH 6.2 with absorption maxima at 415 nm and 333 nm, respectively. These separate forms of Schiff-base confer different catalytic efficiencies to the enzyme.

  17. Short-term hypothermia activates hepatic mitochondrial sn-glycerol-3-phosphate dehydrogenase and thermogenic systems.

    PubMed

    Bobyleva, V; Pazienza, L; Muscatello, U; Kneer, N; Lardy, H

    2000-08-15

    The contribution of the sn-glycerol-3-phosphate (G-3-P) shuttle in the control of energy metabolism is well established. It is also known that its activity may be modulated by hormones involved in thermogenesis, such as thyroid hormones or dehydroepiandrosterone and its metabolites, that act by inducing de novo synthesis of mitochondrial G-3-P dehydrogenase (mGPDH). However, little is known as to the factors that may influence the activity without enzyme induction. In the present study we investigated the possible role of the G-3-P shuttle in the thermogenic response to different hypothermic stresses. It was found that a decrease of body temperature causes the liver rapidly to enhance mGPDH activity and G-3-P-dependent respiration. The enhancement, which does not result from de novo synthesis of enzymes, has the potential of increasing heat production both by decreased ATP synthesis during the oxidation of G-3-P and by activation of the glycolytic pathway.

  18. Generally recognized as safe (GRAS) Lactococcus lactis strains associated with Lippia sidoides Cham. are able to solubilize/mineralize phosphate.

    PubMed

    de Lacerda, Jackeline Rossetti Mateus; da Silva, Thais Freitas; Vollú, Renata Estebanez; Marques, Joana Montezano; Seldin, Lucy

    2016-01-01

    Eight strains isolated from the stems of Lippia sidoides were identified as belonging to Lactococcus lactis, a bacterial species considered as "generally recognized as safe". Their capacity to solubilize/mineralize phosphate was tested in vitro with different inorganic and organic phosphorus (P) sources. All strains were able to solubilize calcium phosphate as an inorganic P source, and the best result was observed with strain 003.41 which solubilized 31 % of this P source. Rock phosphate, a mined rock containing high amounts of phosphate bearing minerals, was solubilized by five strains. When calcium phytate was the organic P source used, the majority of the strains tested showed phosphate mineralization activity. Moreover, all strains were able to solubilize/mineralize phosphate from poultry litter, a complex P source containing inorganic and predominantly organic P. The presence of genes coding for phytase and alkaline phosphatase was searched within the strains studied. However, only gene sequences related to alkaline phosphatase (phoA and phoD) could be detected in the majority of the strains (excepting strain 006.29) with identities varying from 67 to 88 %. These results demonstrate for the first time the potential of L. lactis strains for phosphate solubilization/mineralization activity using a broad spectrum of P sources; therefore, they are of great importance for the future development of more safe bioinoculants with possible beneficial effects for agriculture.

  19. Optimization of the thermophilic anaerobic co-digestion of pig manure, agriculture waste and inorganic additive through specific methanogenic activity.

    PubMed

    Jiménez, J; Cisneros-Ortiz, M E; Guardia-Puebla, Y; Morgan-Sagastume, J M; Noyola, A

    2014-01-01

    The anaerobic co-digestion of three wastes (manure, rice straw and clay residue, an inorganic additive) at different concentration levels and their interactive effects on methanogenic activity were investigated in this work at thermophilic conditions in order to enhance hydrolytic activity and methane production. A central composite design and the response surface methodology were applied for the optimization of specific methanogenic activity (SMA) by assessing their interaction effects with a reduced number of experiments. The results showed a significant interaction among the wastes on the SMA and confirmed that co-digestion enhances methane production. Rice straw apparently did not supply a significant amount of substrate to make a difference in SMA or methane yield. On the other hand, clay residue had a positive effect as an inorganic additive for stimulating the anaerobic process, based on its mineral content and its adsorbent properties for ammonia. Finally, the optimal conditions for achieving a thermophilic SMA value close to 1.4 g CH4-COD/g VSS · d(-1) were 20.3 gVSS/L of manure, 9.8 gVSS/L of rice straw and 3.3 gTSS/L of clay.

  20. Dengue Virus NS1 Protein Modulates Cellular Energy Metabolism by Increasing Glyceraldehyde-3-Phosphate Dehydrogenase Activity

    PubMed Central

    Allonso, Diego; Andrade, Iamara S.; Conde, Jonas N.; Coelho, Diego R.; Rocha, Daniele C. P.; da Silva, Manuela L.; Ventura, Gustavo T.

    2015-01-01

    ABSTRACT Dengue is one of the main public health concerns worldwide. Recent estimates indicate that over 390 million people are infected annually with the dengue virus (DENV), resulting in thousands of deaths. Among the DENV nonstructural proteins, the NS1 protein is the only one whose function during replication is still unknown. NS1 is a 46- to 55-kDa glycoprotein commonly found as both a membrane-associated homodimer and a soluble hexameric barrel-shaped lipoprotein. Despite its role in the pathogenic process, NS1 is essential for proper RNA accumulation and virus production. In the present study, we identified that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) interacts with intracellular NS1. Molecular docking revealed that this interaction occurs through the hydrophobic protrusion of NS1 and the hydrophobic residues located at the opposite side of the catalytic site. Moreover, addition of purified recombinant NS1 enhanced the glycolytic activity of GAPDH in vitro. Interestingly, we observed that DENV infection promoted the relocalization of GAPDH to the perinuclear region, where NS1 is commonly found. Both DENV infection and expression of NS1 itself resulted in increased GAPDH activity. Our findings indicate that the NS1 protein acts to increase glycolytic flux and, consequently, energy production, which is consistent with the recent finding that DENV induces and requires glycolysis for proper replication. This is the first report to propose that NS1 is an important modulator of cellular energy metabolism. The data presented here provide new insights that may be useful for further drug design and the development of alternative antiviral therapies against DENV. IMPORTANCE Dengue represents a serious public health problem worldwide and is caused by infection with dengue virus (DENV). Estimates indicate that half of the global population is at risk of infection, with almost 400 million cases occurring per year. The NS1 glycoprotein is found in both the

  1. Pharmacological evaluation of poly(3-methylthiophene) and its titanium(IV)phosphate nanocomposite: DNA interaction, molecular docking, and cytotoxic activity.

    PubMed

    Baig, Umair; Gondal, M A; Alam, Md Fazle; Wani, Waseem A; Younus, Hina

    2016-11-01

    Cancer and pathogenic microbial diseases have terribly affected human health over a longer period of time. In response to the increasing casualties due to cancer and microbial diseases, unique poly(3-methylthiophene) and poly(3-methylthiophene)-titanium(IV)phosphate composite were prepared via in-situ oxidative chemical polymerization in this work. The poly(3-methylthiophene) and poly(3-methylthiophene)-titanium(IV)phosphate composite were well characterized by Fourier transform infrared spectroscopy and field emission scanning electron microscopy. DNA binding studies by UV-Visible and fluorescence spectroscopic investigations indicated strong binding affinities of poly(3-methylthiophene) and poly(3-methylthiophene)-titanium(IV)phosphate nanocomposite; leading to structural damage of DNA. Poly(3-methylthiophene)-titanium(IV)phosphate nanocomposite showed stronger interactions with DNA as compared to poly(3-methylthiophene) and from dye displacement assay it was confirmed that mode of binding of both the formulations was intercalative. The antimicrobial screening revealed that polymer and its composite displayed stronger antibacterial effects than ampicillin against Bacillus subtilis, Staphylococcus aureus, Pseudomonas aeruginosa and Salmonella typhimurium. Besides, the poly(3-methylthiophene) and poly(3-methylthiophene)-titanium(IV)phosphate nanocomposite showed dose dependent effects towards estrogen receptor positive breast cancer (MCF-7) and estrogen receptor negative breast cancer (MDA-MB-231) cell lines; with poly(3-methylthiophene)-titanium(IV)phosphate nanocomposite showing better activities against both cell lines. In all in-vitro biological investigations, poly(3-methylthiophene)-titanium(IV)phosphate composite showed superior properties to that of the pure poly(3-methylthiophene), which encouraged us to suggest its potential as future therapeutic gear in drug delivery and other allied fields.

  2. Effects of low cell pH and elevated inorganic phosphate on the pCa-force relationship in single muscle fibers at near-physiological temperatures.

    PubMed

    Nelson, Cassandra R; Fitts, Robert H

    2014-04-01

    Intense muscle contraction induces high rates of ATP hydrolysis with resulting increases in Pi, H(+), and ADP, factors thought to induce fatigue by interfering with steps in the cross-bridge cycle. Force inhibition is less at physiological temperatures; thus the role of low pH in fatigue has been questioned. Effects of pH 6.2 and collective effects with 30 mM Pi on the pCa-force relationship were assessed in skinned fast and slow rat skeletal muscle fibers at 15 and 30°C. At 30°C, pH 6.2 + 30 mM Pi significantly depressed peak force in all fiber types, with the greatest effect in type IIx fibers. Across fiber types, Ca(2+) sensitivity was depressed by low pH and low pH + high Pi, with the greater effect at 30°C. For type IIx fibers at 30°C, half-maximal activation (pCa50) was 5.36 at pH 6.2 (no added Pi) and 4.98 at pH 6.2 + 30 mM Pi compared with 6.58 in the control condition (pH 7, no added Pi). At 30°C, n2, reflective of thick filament cooperativity, was unchanged by low cell pH but was depressed from 5.02 to 2.46 in type IIx fibers with pH 6.2 + 30 mM Pi. With acidosis, activation thresholds of all fiber types required higher free Ca(2+) at 15 and 30°C. With the exception of type IIx fibers, the Ca(2+) required to reach activation threshold increased further with added Pi. In conclusion, it is clear that fatigue-inducing effects of low cell pH and elevated Pi at near-physiological temperatures are substantial.

  3. Structural and Chemical Basis for Glucosamine 6-Phosphate Binding and Activation of the glmS Ribozyme

    SciTech Connect

    Cochrane, J.; Lipchock, S; Smith, K; Strobel, S

    2009-01-01

    The glmS ribozyme is the first naturally occurring catalytic RNA that relies on an exogenous, nonnucleotide cofactor for reactivity. From a biochemical perspective, the glmS ribozyme derived from Bacillus anthracis is the best characterized. However, much of the structural work to date has been done on a variant glmS ribozyme, derived from Thermoanaerobacter tengcongensis. Here we present structures of the B. anthracis glmS ribozyme in states before the activating sugar, glucosamine 6-phosphate (GlcN6P), has bound and after the reaction has occurred. These structures show an active site preorganized to bind GlcN6P that retains some affinity for the sugar even after cleavage of the RNA backbone. A structure of an inactive glmS ribozyme with a mutation distal from the ligand-binding pocket highlights a nucleotide critical to the reaction that does not affect GlcN6P binding. Structures of the glmS ribozyme bound to a naturally occurring inhibitor, glucose 6-phosphate (Glc6P), and a nonnatural activating sugar, mannosamine 6-phosphate (MaN6P), reveal a binding mode similar to that of GlcN6P. Kinetic analyses show a pH dependence of ligand binding that is consistent with titration of the cofactor's phosphate group and support a model in which the major determinant of activity is the sugar amine independent of its stereochemical presentation.

  4. Interactions between organic additives and active powders in water-based lithium iron phosphate electrode slurries

    NASA Astrophysics Data System (ADS)

    Li, Chia-Chen; Lin, Yu-Sheng

    2012-12-01

    The interactions of organic additives with active powders are investigated and are found to have great influence on the determination of the mixing process for preparing electrode slurries with good dispersion and electrochemical properties of lithium iron phosphate (LiFePO4) electrodes. Based on the analyses of zeta potential, sedimentation, and rheology, it is shown that LiFePO4 prefers to interact with styrene-butadiene rubber (SBR) relative to other organic additives such as sodium carboxymethyl cellulose (SCMC), and thus shows preferential adsorption by SBR, whereas SBR has much lower efficiency than SCMC in dispersing LiFePO4. Therefore, for SCMC to interact with and disperse LiFePO4 before the interaction of LiFePO4 with SBR, it is suggested to mix SCMC with LiFePO4 prior to the addition of SBR during the slurry preparation process. For the electrode prepared via the suggested process, i.e., the sequenced adding process in which SCMC is mixed with active powders prior to the addition of SBR, a much better electrochemical performance is obtained than that of the one prepared via the process referred as the simultaneous adding process, in which mixing of SCMC and SBR with active powders in simultaneous.

  5. Antibacterial activity evaluation of bioactive glass and biphasic calcium phosphate nanopowders mixtures

    NASA Astrophysics Data System (ADS)

    Nazemi, Zahra; Mehdikhani-Nahrkhalaji, Mehdi; Haghbin-Nazarpak, Masoumeh; Staji, Hamid; Kalani, Mohammad Mehdi

    2016-12-01

    The aim of this work was to evaluate the antibacterial activity of bioactive glass (BG) and biphasic calcium phosphate (BCP) nanopowders mixtures for the first time. 37S BG and BCP (50% HA-50% β-TCP) nanopowders were prepared via sol-gel technique. Characterization techniques such as X-ray diffraction, scanning electron microscopy, transition electron microscopy, and X-ray fluorescent. The antibacterial activity was studied using Escherichia coli and Salmonella typhi as gram-negative, and Staphylococcus aureus as gram-positive bacteria. The antibacterial effect of BG, BCP nanopowders, and their mixtures was evaluated at different concentrations. The 37S BG nanopowders showed minimum bactericidal concentration at 25 mg/ml. At broth concentrations below 300 mg/ml, BCP showed no antibacterial activity. BCP and BG nanopowders mixture (M2) with 60/40 ratio of BCP/BG showed noticeable antibacterial effect. It was concluded that BCP and 37S BG nanopowders mixture could be used as a good candidate for dental and orthopedic applications.

  6. A serum factor that activates the phosphatidylinositol phosphate signaling system in Xenopus oocytes.

    PubMed Central

    Tigyi, G; Dyer, D; Matute, C; Miledi, R

    1990-01-01

    Blood sera from many vertebrate species elicit large oscillatory chloride currents in oocytes from the frog Xenopus laevis. Rabbit serum was active at dilutions as great as one part in 10 million. Intracellularly applied serum was ineffective, and externally applied serum failed to trigger oscillatory currents when the intracellular level of ionized calcium was prevented from rising by loading the oocyte with EGTA. The serum also caused an increase of inositol 1,4,5-trisphosphate in the oocyte. We conclude that serum contains a factor which activates a membrane receptor that is coupled to the phosphatidylinositol second messenger system. The active factor is a protein with an apparent molecular mass of 60-70 kDa in gel permeation chromatography. Although the normal function of the serum factor is still unknown, it may have far-reaching implications, because it acts on the multifunctional phosphatidylinositol phosphate signaling system. Also, because of its great potency the serum factor and Xenopus oocytes are very useful for probing the operation of the phosphatidylinositol system. PMID:1689488

  7. Aldosterone impairs vascular reactivity by decreasing glucose-6-phosphate dehydrogenase activity

    PubMed Central

    Leopold, Jane A.; Dam, Aamir; Maron, Bradley A.; Scribner, Anne W.; Liao, Ronglih; Handy, Diane E.; Stanton, Robert C.; Pitt, Bertram; Loscalzo, Joseph

    2013-01-01

    Hyperaldosteronism is associated with impaired vascular reactivity; however, the mechanism by which aldosterone promotes endothelial dysfunction remains unknown. Glucose-6-phosphate dehydrogenase (G6pd), the principal source of Nadph, modulates vascular function by limiting oxidant stress to preserve bioavailable nitric oxide (NO•). In these studies, we show that aldosterone (10−9-10−7 mol/l) decreases endothelial G6pd expression and activity in vitro resulting in increased oxidant stress and decreased cGMP levels similar to what is observed in G6pd-deficient cells. Aldosterone decreases G6pd expression by protein kinase A activation to increase expression of Crem, which interferes with Creb binding to the G6pd promoter. In vivo, infusion of aldosterone decreases vascular G6pd expression and impairs vascular reactivity. These effects are abrogated by spironolactone or vascular gene transfer of G6pd. These studies demonstrate that aldosterone induces a G6pd-deficient phenotype to impair endothelial function; aldosterone antagonism or gene transfer of G6pd improves vascular reactivity by restoring G6pd activity. PMID:17273168

  8. Sphingosine-1-phosphate mediates epidermal growth factor-induced muscle satellite cell activation

    SciTech Connect

    Nagata, Yosuke Ohashi, Kazuya; Wada, Eiji; Yuasa, Yuki; Shiozuka, Masataka; Nonomura, Yoshiaki; Matsuda, Ryoichi

    2014-08-01

    Skeletal muscle can regenerate repeatedly due to the presence of resident stem cells, called satellite cells. Because satellite cells are usually quiescent, they must be activated before participating in muscle regeneration in response to stimuli such as injury, overloading, and stretch. Although satellite cell activation is a crucial step in muscle regeneration, little is known of the molecular mechanisms controlling this process. Recent work showed that the bioactive lipid sphingosine-1-phosphate (S1P) plays crucial roles in the activation, proliferation, and differentiation of muscle satellite cells. We investigated the role of growth factors in S1P-mediated satellite cell activation. We found that epidermal growth factor (EGF) in combination with insulin induced proliferation of quiescent undifferentiated mouse myoblast C2C12 cells, which are also known as reserve cells, in serum-free conditions. Sphingosine kinase activity increased when reserve cells were stimulated with EGF. Treatment of reserve cells with the D-erythro-N,N-dimethylsphingosine, Sphingosine Kinase Inhibitor, or siRNA duplexes specific for sphingosine kinase 1, suppressed EGF-induced C2C12 activation. We also present the evidence showing the S1P receptor S1P2 is involved in EGF-induced reserve cell activation. Moreover, we demonstrated a combination of insulin and EGF promoted activation of satellite cells on single myofibers in a manner dependent on SPHK and S1P2. Taken together, our observations show that EGF-induced satellite cell activation is mediated by S1P and its receptor. - Highlights: • EGF in combination with insulin induces proliferation of quiescent C2C12 cells. • Sphingosine kinase activity increases when reserve cells are stimulated with EGF. • EGF-induced activation of reserve cells is dependent on sphingosine kinase and ERK. • The S1P receptor S1P2 is involved in EGF-induced reserve cell activation. • EGF-induced reserve cell activation is mediated by S1P and its

  9. Optimizations of particle size and pulp density for solubilization of rock phosphate by a microbial consortium from activated sludge.

    PubMed

    Xiao, Chunqiao; Wu, Xiaoyan; Liu, Tingting; Xu, Guang; Chi, Ruan

    2016-12-29

    Microbial solubilization of rock phosphate is getting more and more attention recently. However, the microorganisms used in previous studies were mostly single or known species, and seldom studies focused on the mixed microorganisms or microbial consortia from natural environments. In this study, a microbial consortium taken from activated sludge was used to solubilize two different mid-low-grade rock phosphates. The results showed that the microbial consortium could effectively solubilize the rock phosphates in National Botanical Research Institute's phosphate growth medium and released soluble phosphorus in the broth. The biomass increased gradually, whereas the pH decreased sharply during the solubilizing process. The maximum phosphorus solubilization was recorded at particle size of 150 µm. Higher or lower than this optimal particle size, the phosphorus solubilization decreased. The phosphorus solubilization gradually decreased with a larger pulp density from 1 to 5%, and the optimal pulp density was 1%. The solubilization level of microbial consortium varied with different rock phosphates. The results revealed that the soluble phosphorus released from high-silicon ore was higher than which from high-magnesium ore. A strong positive correlation between biomass and phosphorus solubilization in the broth was observed from regression analysis results, and the phosphorus solubilization also had a significant negative correlation with pH in the broth.

  10. The desorption and reactivity of butanol adsorbed on lithium iron phosphate (LISICON) activated in a hydrogen plasma

    NASA Astrophysics Data System (ADS)

    Pylinina, A. I.; Mikhalenko, I. I.; Yagodovskaya, T. V.; Yagodovskii, V. D.

    2010-12-01

    The reactivity and desorption of butanol-2 adsorbed on Li3Fe2(PO4)3 not subjected and subjected to treatment in a glow discharge hydrogen plasma were studied under flow conditions with a gas chromatographic analysis of products. X-ray photoelectron spectroscopy data showed that the number of phosphate groups on the surface of the phosphate was two times larger than the stoichiometric number and increased after plasma chemical treatment. The strength of butanol-phosphate bonds also increased, and the selectivity of alcohol decomposition with the formation of an olefin (dehydration) and ketone (dehydrogenation) changed. After plasma treatment, dehydrogenation centers were deactivated. The selectivities of alcohol transformations in the adsorbed state and under vapor phase conditions were different. Ketone was formed from adsorbed alcohol because the activation energies of dehydrogenation were equal for the two reaction variants.

  11. Equilibrium and kinetic studies of adsorption of phosphate onto ZnCl2 activated coir pith carbon.

    PubMed

    Namasivayam, C; Sangeetha, D

    2004-12-15

    Phosphate removal from aqueous solution was investigated using ZnCl(2)-activated carbon developed from coir pith, an agricultural solid waste. Studies were conducted to delineate the effect of contact time, adsorbent dose, phosphate concentration, pH, and temperature. The adsorption equilibrium data followed both Langmuir and Freundlich isotherms. Langmuir adsorption capacity was found to be 5.1 mg/g. Adsorption followed second-order kinetics. The removal was maximum in the pH range 3-10. pH effect and desorption studies showed that adsorption occurred by both ion exchange and chemisorption mechanisms. Adsorption was found to be spontaneous and endothermic. Effect of foreign ions on adsorption shows that perchlorate, sulfate, and selenite decreased the percent removal of phosphate.

  12. Activated platelets release sphingosine 1-phosphate and induce hypersensitivity to noxious heat stimuli in vivo

    PubMed Central

    Weth, Daniela; Benetti, Camilla; Rauch, Caroline; Gstraunthaler, Gerhard; Schmidt, Helmut; Geisslinger, Gerd; Sabbadini, Roger; Proia, Richard L.; Kress, Michaela

    2015-01-01

    At the site of injury activated platelets release various mediators, one of which is sphingosine 1-phosphate (S1P). It was the aim of this study to explore whether activated human platelets had a pronociceptive effect in an in vivo mouse model and whether this effect was based on the release of S1P and subsequent activation of neuronal S1P receptors 1 or 3. Human platelets were prepared in different concentrations (105/μl, 106/μl, 107/μl) and assessed in mice with different genetic backgrounds (WT, S1P1fl/fl, SNS-S1P1−/−, S1P3−/−). Intracutaneous injections of activated human platelets induced a significant, dose-dependent hypersensitivity to noxious thermal stimulation. The degree of heat hypersensitivity correlated with the platelet concentration as well as the platelet S1P content and the amount of S1P released upon platelet activation as measured with LC MS/MS. Despite the significant correlations between S1P and platelet count, no difference in paw withdrawal latency (PWL) was observed in mice with a global null mutation of the S1P3 receptor or a conditional deletion of the S1P1 receptor in nociceptive primary afferents. Furthermore, neutralization of S1P with a selective anti-S1P antibody did not abolish platelet induced heat hypersensitivity. Our results suggest that activated platelets release S1P and induce heat hypersensitivity in vivo. However, the platelet induced heat hypersensitivity was caused by mediators other than S1P. PMID:25954148

  13. Anti-tumor and immunomodulatory activity of iron hepta-tungsten phosphate oxygen clusters complex.

    PubMed

    Zhang, Bisong; Qiu, Jianping; Wu, Changsheng; Li, Yunxia; Liu, Zhenxiang

    2015-12-01

    Polyoxometalates (POMs) have attracted a considerable attention due to their unique structural characteristics, physicochemical properties and biological activities. In this study, iron hepta-tungsten phosphate oxygen clusters complex Na12H[Fe(HPW7O28)2]·44H2O (IHTPO) was synthesized and evaluated for in vitro cytotoxic activities on human hepatoma HepG2, leukemia K562, lung carcinoma A549, and large cell lung cancer NCI-H460 cells, therapeutic efficacies on mice transplantable tumor, and immunomodulatory potentials on the immune response in tumor-bearing mice. IHTPO exhibited lower in vitro cytotoxic activities against four human tumor cell lines, with the IC50 values being higher than 62.5μM (ca. 300μg/ml). IHTPO, however, significantly inhibited the growth of S180 sarcoma transplanted in mice. It was further showed that IHTPO could not only significantly promote splenocytes proliferation, NK cell and CTL activity from splenocytes, but remarkably enhance serum antigen-specific IgG, IgG2a and IgG2b antibody levels in S180-bearing mice. IHTPO also significantly promoted Th1 cytokines IFN-γ and IL-2 production, and up-regulated the mRNA expression levels of IFN-γ, IL-2 and Th1 transcription factors T-bet and STAT-4 in splenocytes from the S180-bearing mice. These results suggested that IHTPO significantly inhibited the growth of mice transplantable tumor, and that its in vivo antitumor activity might be achieved by improving Th1 protective cell-mediated immunity. IHTPO could act as antitumor agent with immunomodulatory activity.

  14. Cellular recovery of glyceraldehyde-3-phosphate dehydrogenase activity and thiol status after exposure to hydroperoxides

    SciTech Connect

    Brodie, A.E.; Reed, D.J. )

    1990-01-01

    The activity of the thiol-dependent enzyme glyceraldehyde-3-phosphate dehydrogenase (GPD), in vertebrate cells, was modulated by a change in the intracellular thiol:disulfide redox status. Human lung carcinoma cells (A549) were incubated with 1-120 mM H2O2, 1-120 mM t-butyl hydroperoxide, 1-6 mM ethacrynic acid, or 0.1-10 mM N-ethylmaleimide for 5 min. Loss of reduced protein thiols, as measured by binding of the thiol reagent iodoacetic acid to GPD, and loss of GPD enzymatic activity occurred in a dose-dependent manner. Incubation of the cells, following oxidative treatment, in saline for 30 min or with 20 mM dithiothreitol (DTT) partially reversed both changes in GPD. The enzymatic recovery of GPD activity was observed either without addition of thiols to the medium or by incubation of a sonicated cell mixture with 2 mM cysteine, cystine, cysteamine, or glutathione (GSH); GSSG had no effect. Treatment of cells with buthionine sulfoximine (BSO) to decrease cellular GSH by varying amounts caused a dose-related increase in sensitivity of GPD activity to inactivation by H2O2 and decreased cellular ability for subsequent recovery. GPD responded in a similar fashion with oxidative treatment of another lung carcinoma cell line (A427) as well as normal lung tissue from human and rat. These findings indicate that the cellular thiol redox status can be important in determining GPD enzymatic activity.

  15. Effect of an organophosphate pesticide, monocrotophos, on phosphate-solubilizing efficiency of soil fungal isolates.

    PubMed

    Jain, Rachna; Garg, Veena; Saxena, Jyoti

    2015-01-01

    Soil is a sink of pesticide residues as well as microorganisms. Fungi are well known for solubilization of inorganic phosphates, and this activity of fungal isolates may be affected by the presence of pesticide residues in the soil. In the present study, five generically different fungal isolates, viz. Aspergillus niger JQ660373, Aspergillus flavus, Penicillium aculeatum JQ660374, Fusarium pallidoroseum and Macrophomina sp., were tested and compared for their phosphate-solubilizing ability in the absence and presence of monocrotophos (500 mg L(-1)). After 168 h of incubation, four times high amount of tricalcium phosphate was solubilized by isolates in the growth medium containing monocrotophos in comparison to control (without monocrotophos). Concurrently, 78 % of the applied monocrotophos was degraded by these fungal isolates. Kinetics of phosphate solubilization shifted from logarithmic to power model in the presence of monocrotophos. Similarly, the phosphatase activity was also found significantly high in the presence of monocrotophos. The combined order of phosphate solubilization as well as monocrotophos degradation was found to be A. niger JQ660373 > P. aculeatum JQ660374 > A. flavus > F. pallidoroseum > Macrophomina sp. On the contrary, phosphate solubilization negatively correlated with the pH of the growth medium. Hence, it could be concluded that these fungal species efficiently solubilize inorganic phosphates and monocrotophos poses a positive effect on their ability and in turn degraded by them. To the best of our knowledge, this is the first report on P solubilization by Macrophomina sp. and F. pallidoroseum.

  16. Abnormal high-energy phosphate molecule metabolism during regional brain activation in patients with bipolar disorder.

    PubMed

    Yuksel, C; Du, F; Ravichandran, C; Goldbach, J R; Thida, T; Lin, P; Dora, B; Gelda, J; O'Connor, L; Sehovic, S; Gruber, S; Ongur, D; Cohen, B M

    2015-09-01

    Converging evidence suggests bioenergetic abnormalities in bipolar disorder (BD). In the brain, phosphocreatine (PCr) acts a reservoir of high-energy phosphate (HEP) bonds, and creatine kinases (CK) catalyze the transfer of HEP from adenosine triphosphate (ATP) to PCr and from PCr back to ATP, at times of increased need. This study examined the activity of this mechanism in BD by measuring the levels of HEP molecules during a stimulus paradigm that increased local energy demand. Twenty-three patients diagnosed with BD-I and 22 healthy controls (HC) were included. Levels of phosphorus metabolites were measured at baseline and during visual stimulation in the occipital lobe using (31)P magnetic resonance spectroscopy at 4T. Changes in metabolite levels showed different patterns between the groups. During stimulation, HC had significant reductions in PCr but not in ATP, as expected. In contrast, BD patients had significant reductions in ATP but not in PCr. In addition, PCr/ATP ratio was lower at baseline in patients, and there was a higher change in this measure during stimulation. This pattern suggests a disease-related failure to replenish ATP from PCr through CK enzyme catalysis during tissue activation. Further studies measuring the CK flux in BD are required to confirm and extend this finding.

  17. Preliminary evidence for biologic activity of toceranib phosphate (Palladia(®)) in solid tumours.

    PubMed

    London, C; Mathie, T; Stingle, N; Clifford, C; Haney, S; Klein, M K; Beaver, L; Vickery, K; Vail, D M; Hershey, B; Ettinger, S; Vaughan, A; Alvarez, F; Hillman, L; Kiselow, M; Thamm, D; Higginbotham, M L; Gauthier, M; Krick, E; Phillips, B; Ladue, T; Jones, P; Bryan, J; Gill, V; Novasad, A; Fulton, L; Carreras, J; McNeill, C; Henry, C; Gillings, S

    2012-09-01

    The purpose of this study was to provide an initial assessment of the potential biologic activity of toceranib phosphate (Palladia®, Pfizer Animal Health, Madison, NJ, USA) in select solid tumours in dogs. Cases in which toceranib was used to treat dogs with apocrine gland anal sac adenocarcinoma (AGASACA), metastatic osteosarcoma (OSA), thyroid carcinoma, head and neck carcinoma and nasal carcinoma were included. Clinical benefit (CB) was observed in 63/85 (74%) dogs including 28/32 AGASACA [8 partial response (PR), 20 stable disease (SD)], 11/23 OSAs (1 PR and 10 SD), 12/15 thyroid carcinomas (4 PR and 8 SD), 7/8 head and neck carcinomas [1 complete response (CR), 5 PR and 1 SD] and 5/7 (1 CR and 4 SD) nasal carcinomas. For dogs experiencing CB, the median dose of toceranib was 2.8 mg kg(-1) , 36/63 (58.7%) were dosed on a Monday/Wednesday/Friday basis and 47/63 (74.6%) were treated 4 months or longer. Although these data provide preliminary evidence that toceranib exhibits CB in dogs with certain solid tumours, future prospective studies are necessary to define its true activity.

  18. Neonatal phosphate nutrition alters in vivo and in vitro satellite cell activity in pigs.

    PubMed

    Alexander, Lindsey S; Seabolt, Brynn S; Rhoads, Robert P; Stahl, Chad H

    2012-06-01

    Satellite cell activity is necessary for postnatal skeletal muscle growth. Severe phosphate (PO(4)) deficiency can alter satellite cell activity, however the role of neonatal PO(4) nutrition on satellite cell biology remains obscure. Twenty-one piglets (1 day of age, 1.8 ± 0.2 kg BW) were pair-fed liquid diets that were either PO(4) adequate (0.9% total P), supra-adequate (1.2% total P) in PO(4) requirement or deficient (0.7% total P) in PO(4) content for 12 days. Body weight was recorded daily and blood samples collected every 6 days. At day 12, pigs were orally dosed with BrdU and 12 h later, satellite cells were isolated. Satellite cells were also cultured in vitro for 7 days to determine if PO(4) nutrition alters their ability to proceed through their myogenic lineage. Dietary PO(4) deficiency resulted in reduced (P < 0.05) sera PO(4) and parathyroid hormone (PTH) concentrations, while supra-adequate dietary PO(4) improved (P < 0.05) feed conversion efficiency as compared to the PO(4) adequate group. In vivo satellite cell proliferation was reduced (P < 0.05) among the PO(4) deficient pigs, and these cells had altered in vitro expression of markers of myogenic progression. Further work to better understand early nutritional programming of satellite cells and the potential benefits of emphasizing early PO(4) nutrition for future lean growth potential is warranted.

  19. Effectiveness of phosphate removal during anaerobic digestion of waste activated sludge by dosing iron(III).

    PubMed

    Cheng, Xiang; Wang, Jue; Chen, Bing; Wang, Yu; Liu, Jiaqi; Liu, Lubo

    2017-05-15

    Phosphate-Fe(II) precipitation induced by Fe(III) reduction during the anaerobic digestion of excess activated sludge was investigated for the removal of phosphorus and its possible recovery. The experiments were conducted with three Fe(III) sources at 35 °C and 55 °C. The results show that ferrihydrite-Fe(III) was effectively reduced during the anaerobic sludge digestion by 63% and 96% under mesophilic and thermophilic conditions, respectively. Whereas FeCl3-Fe(III) was only mesophilically reducible and the reduction of hematite-Fe(III) was unnoticeable at either temperature. Efficient precipitation of vivianite was not observed although high saturation index values, e.g., >14 (activity reduction not considered), had been reached. This reveals the complexity of vivianite precipitation in anaerobic digestion systems; for example, Fe(II) complexation and organic interference could not be ignored. With ferrihydrite amendments at a Fe/TP of 1.5, methane production from sludge digestion was reduced by 35.1% at 35 °C, and was unaffected when the digestion temperature went up to 55 °C. But, acidic FeCl3 severely inhibited the methane production and consequently the sludge biomass degradation.

  20. Inorganic concepts relevant to metal binding, activity, and toxicity in a biological system

    SciTech Connect

    Hoeschele, J.D. . Parke-Davis Pharmaceutical Research Div.); Turner, J.E.; England, M.W. )

    1990-01-01

    The purpose of this paper is to review selected physical and inorganic concepts and factors which might be important in assessing and/or understanding the fact and disposition of a metal system in a biological environment. Hopefully, such inquiries will ultimately permit us to understand, rationalize, and predict differences and trends in biological effects as a function of the basic nature of a metal system and, in optimal cases, serve as input to a system of guidelines for the notion of Chemical Dosimetry.'' The plan of this paper is to first review, in general terms, the basic principles of the Crystal Field Theory (CFT), a unifying theory of bonding in metal complexes. This will provide the necessary theoretical background for the subsequent discussion of selected concepts and factors. 21 refs., 7 figs., 6 tabs.

  1. Reducing the Genetic Redundancy of Arabidopsis PHOSPHATE TRANSPORTER1 Transporters to Study Phosphate Uptake and Signaling1[OPEN

    PubMed Central

    Ayadi, Amal; David, Pascale; Arrighi, Jean-François; Chiarenza, Serge; Thibaud, Marie-Christine; Nussaume, Laurent; Marin, Elena

    2015-01-01

    Arabidopsis (Arabidopsis thaliana) absorbs inorganic phosphate (Pi) from the soil through an active transport process mediated by the nine members of the PHOSPHATE TRANSPORTER1 (PHT1) family. These proteins share a high level of similarity (greater than 61%), with overlapping expression patterns. The resulting genetic and functional redundancy prevents the analysis of their specific roles. To overcome this difficulty, our approach combined several mutations with gene silencing to inactivate multiple members of the PHT1 family, including a cluster of genes localized on chromosome 5 (PHT1;1, PHT1;2, and PHT1;3). Physiological analyses of these lines established that these three genes, along with PHT1;4, are the main contributors to Pi uptake. Furthermore, PHT1;1 plays an important role in translocation from roots to leaves in high phosphate conditions. These genetic tools also revealed that some PHT1 transporters likely exhibit a dual affinity for phosphate, suggesting that their activity is posttranslationally controlled. These lines display significant phosphate deficiency-related phenotypes (e.g. biomass and yield) due to a massive (80%–96%) reduction in phosphate uptake activities. These defects limited the amount of internal Pi pool, inducing compensatory mechanisms triggered by the systemic Pi starvation response. Such reactions have been uncoupled from PHT1 activity, suggesting that systemic Pi sensing is most probably acting downstream of PHT1. PMID:25670816

  2. Inorganic Carbon Turnover caused by Digestion of Carbonate Sands and Metabolic Activity of Holothurians

    SciTech Connect

    Schneider, Kenneth; Silverman, Jacob; Kravitz, Benjamin S.; Rivlin, Tanya; Schneider-Mor, Aya; Barbosa, Sergio; Byrne, Maria; Caldeira, Ken

    2013-11-20

    Recent measurements have shown that holothurians (sea cucumbers) play an important role in the cycling of CaCO3 in tropical coral reef systems through ingestion and processing of carbonate sediment. In this study inorganic additional aspects of carbon turnover were determined in laboratory incubations of Holothuria atra, H. leucospilota and Stichopus herrmanni from One Tree Reef, Great Barrier Reef. The pH values of the gut lumen ranged from 6.1 to 6.7 in animals with empty digestive tracts as opposed to 7.0 to 7.6 when digestive tracts were filled with sediment. Empty gut volume estimates for H. atra and S. herrmanni were 36 ± 4 mL and 151 ± 14 mL, respectively. Based on these measurements it is estimated that these species process 19 ± 2kg and 80 ± 7kg CaCO3 sand yr-1 per individual, respectively. The annual dissolution rates of H. atra and S. herrmanni of 6.5±1.9g and 9.6±1.4g, respectively, suggest that 0.05±0.02% and 0.1±0.02% of the CaCO3 processed through their gut annually is dissolved. During the incubations the CaCO3 dissolution was 0.07±0.01%, 0.04±0.01% and 0.21±0.05% of the fecal casts for H. atra, H. leucospilota and S. herrmanni, respectively. The CaCO3 saturation state for both aragonite and calcite minerals during laboratory incubations decreased markedly due to a greater increase in dissolved inorganic carbon (DIC) relative to total alkalinity (AT) as a result of respiration by the animals. Our results support the hypothesis that deposit feeders such as sea cucumbers play an important ecological role in the coral reef CaCO3 cycle.

  3. Inorganic carbon turnover caused by digestion of carbonate sands and metabolic activity of holothurians

    NASA Astrophysics Data System (ADS)

    Schneider, Kenneth; Silverman, Jacob; Kravitz, Ben; Rivlin, Tanya; Schneider-Mor, Aya; Barbosa, Sergio; Byrne, Maria; Caldeira, Ken

    2013-11-01

    Recent measurements have shown that holothurians (sea cucumbers) may play an important role in the cycling of CaCO3 in tropical coral reef systems through ingestion and processing of carbonate sediment. In this report, we present estimates of inorganic carbon turnover rates determined from laboratory incubations of Holothuria atra, Holothuria leucospilota and Stichopus herrmanni. The pH values of the gut lumen ranged from 7.0 to 7.6 when digestive tracts were filled with sediment compared with 6.1-6.7 in animals with empty digestive tracts. Empty gut volume estimates for H. atra and S. herrmanni were 36 ± 4 mL and 151 ± 14 mL, respectively. Based on these measurements and the density and porosity of carbonate sediments of coral reefs, it is estimated that these species process 19 ± 2 kg and 80 ± 7 kg CaCO3 sand yr-1 per individual, respectively. The annual CaCO3 dissolution rates per H. atra and S. herrmanni individual are estimated to be 6.5 ± 1.9 g and 9.6 ± 1.4 g, respectively, suggesting that 0.05 ± 0.02% and 0.1 ± 0.02% of the CaCO3 processed through their gut annually is dissolved. During incubations the CaCO3 dissolution of the fecal casts was 0.07 ± 0.01%, 0.04 ± 0.01% and 0.21 ± 0.05% for H. atra, H. leucospilota and S. herrmanni, respectively. The CaCO3 saturation state in the incubation seawater decreased markedly due to a greater increase in dissolved inorganic carbon (DIC) relative to total alkalinity (AT) as a result of respiration by the animals. Our results support the hypothesis that deposit feeders such as sea cucumbers play an important ecological role in the coral reef CaCO3 cycle.

  4. Role of the pyridine nitrogen in pyridoxal 5'-phosphate catalysis: activity of three classes of PLP enzymes reconstituted with deazapyridoxal 5'-phosphate.

    PubMed

    Griswold, Wait R; Toney, Michael D

    2011-09-21

    Pyridoxal 5'-phosphate (PLP; vitamin B(6))-catalyzed reactions have been well studied, both on enzymes and in solution, due to the variety of important reactions this cofactor catalyzes in nitrogen metabolism. Three functional groups are central to PLP catalysis: the C4' aldehyde, the O3' phenol, and the N1 pyridine nitrogen. In the literature, the pyridine nitrogen has traditionally been assumed to be protonated in enzyme active sites, with the protonated pyridine ring providing resonance stabilization of carbanionic intermediates. This assumption is certainly correct for some PLP enzymes, but the structures of other active sites are incompatible with protonation of N1, and, consequently, these enzymes are expected to use PLP in the N1-unprotonated form. For example, aspartate aminotransferase protonates the pyridine nitrogen for catalysis of transamination, while both alanine racemase and O-acetylserine sulfhydrylase are expected to maintain N1 in the unprotonated, formally neutral state for catalysis of racemization and β-elimination. Herein, kinetic results for these three enzymes reconstituted with 1-deazapyridoxal 5'-phosphate, an isosteric analogue of PLP lacking the pyridine nitrogen, are compared to those for the PLP enzyme forms. They demonstrate that the pyridine nitrogen is vital to the 1,3-prototropic shift central to transamination, but not to reactions catalyzed by alanine racemase or O-acetylserine sulfhydrylase. Not all PLP enzymes require the electrophilicity of a protonated pyridine ring to enable formation of carbanionic intermediates. It is proposed that modulation of cofactor electrophilicity plays a central role in controlling reaction specificity in PLP enzymes.

  5. Structural definition of the active site and catalytic mechanism of 3,4-dihydroxy-2-butanone-4-phosphate synthase.

    PubMed

    Liao, Der-Ing; Zheng, Ya-Jun; Viitanen, Paul V; Jordan, Douglas B

    2002-02-12

    X-ray crystal structures of L-3,4-dihydroxy-2-butanone-4-phosphate synthase from Magnaporthe grisea are reported for the E-SO(4)(2-), E-SO(4)(2-)-Mg(2+), E-SO(4)(2)(-)-Mn(2+), E-SO(4)(2)(-)-Mn(2+)-glycerol, and E-SO(4)(2)(-)-Zn(2+) complexes with resolutions that extend to 1.55, 0.98, 1.60, 1.16, and 1.00 A, respectively. Active-site residues of the homodimer are fully defined. The structures were used to model the substrate ribulose 5-phosphate in the active site with the phosphate group anchored at the sulfate site and the placement of the ribulose group guided by the glycerol site. The model includes two Mg(2+) cations that bind to the oxygen substituents of the C2, C3, C4, and phosphate groups of the substrate, the side chains of Glu37 and His153, and water molecules. The position of the metal cofactors and the substrate's phosphate group are further stabilized by an extensive hydrogen-bond and salt-bridge network. On the basis of their proximity to the substrate's reaction participants, the imidazole of an Asp99-His136 dyad from one subunit, the side chains of the Asp41, Cys66, and Glu174 residues from the other subunit, and Mg(2+)-activated water molecules are proposed to serve specific roles in the catalytic cycle as general acid-base functionalities. The model suggests that during the 1,2-shift step of the reaction, the substrate's C3 and C4 hydroxyl groups are cis to each other. A cis transition state is calculated to have an activation barrier that is 2 kcal/mol greater than that of the trans transition state in the absence of the enzyme.

  6. Structural definition of the active site and catalytic mechanism of 3,4-dihydroxy-2-butanone 4-phosphate synthase

    SciTech Connect

    Liao, D.-I.; Zheng, Y.-J.; Viitanen, P.V.; Jordan, D.B.

    2010-03-08

    X-ray crystal structures of L-3,4-dihydroxy-2-butanone-4-phosphate synthase from Magnaporthe grisea are reported for the E-SO{sub 4}{sup 2-}, E-{sub 4}{sup 2-}-Mg{sup 2+}, E-SO{sub 4}{sup 2-}-Mn{sup 2+}, E-SO{sub 4}{sup 2-}-Mn{sup 2+}-glycerol, and E-SO{sub 4}{sup 2-}-Zn{sup 2+} complexes with resolutions that extend to 1.55, 0.98, 1.60, 1.16, and 1.00 {angstrom}, respectively. Active-site residues of the homodimer are fully defined. The structures were used to model the substrate ribulose 5-phosphate in the active site with the phosphate group anchored at the sulfate site and the placement of the ribulose group guided by the glycerol site. The model includes two Mg{sup 2+} cations that bind to the oxygen substituents of the C2, C3, C4, and phosphate groups of the substrate, the side chains of Glu37 and His153, and water molecules. The position of the metal cofactors and the substrate's phosphate group are further stabilized by an extensive hydrogen-bond and salt-bridge network. On the basis of their proximity to the substrate's reaction participants, the imidazole of an Asp99-His136 dyad from one subunit, the side chains of the Asp41, Cys66, and Glu174 residues from the other subunit, and Mg{sup 2+}-activated water molecules are proposed to serve specific roles in the catalytic cycle as general acid-base functionalities. The model suggests that during the 1,2-shift step of the reaction, the substrate's C3 and C4 hydroxyl groups are cis to each other. A cis transition state is calculated to have an activation barrier that is 2 kcal/mol greater than that of the trans transition state in the absence of the enzyme.

  7. Sphingosine-1-phosphate lyase downregulation promotes colon carcinogenesis through STAT3-activated microRNAs.

    PubMed

    Degagné, Emilie; Pandurangan, Ashok; Bandhuvula, Padmavathi; Kumar, Ashok; Eltanawy, Abeer; Zhang, Meng; Yoshinaga, Yuko; Nefedov, Mikhail; de Jong, Pieter J; Fong, Loren G; Young, Stephen G; Bittman, Robert; Ahmedi, Yasmin; Saba, Julie D

    2014-12-01

    Growing evidence supports a link between inflammation and cancer; however, mediators of the transition between inflammation and carcinogenesis remain incompletely understood. Sphingosine-1-phosphate (S1P) lyase (SPL) irreversibly degrades the bioactive sphingolipid S1P and is highly expressed in enterocytes but downregulated in colon cancer. Here, we investigated the role of SPL in colitis-associated cancer (CAC). We generated mice with intestinal epithelium-specific Sgpl1 deletion and chemically induced colitis and tumor formation in these animals. Compared with control animals, mice lacking intestinal SPL exhibited greater disease activity, colon shortening, cytokine levels, S1P accumulation, tumors, STAT3 activation, STAT3-activated microRNAs (miRNAs), and suppression of miR-targeted anti-oncogene products. This phenotype was attenuated by STAT3 inhibition. In fibroblasts, silencing SPL promoted tumorigenic transformation through a pathway involving extracellular transport of S1P through S1P transporter spinster homolog 2 (SPNS2), S1P receptor activation, JAK2/STAT3-dependent miR-181b-1 induction, and silencing of miR-181b-1 target cylindromatosis (CYLD). Colon biopsies from patients with inflammatory bowel disease revealed enhanced S1P and STAT3 signaling. In mice with chemical-induced CAC, oral administration of plant-type sphingolipids called sphingadienes increased colonic SPL levels and reduced S1P levels, STAT3 signaling, cytokine levels, and tumorigenesis, indicating that SPL prevents transformation and carcinogenesis. Together, our results suggest that dietary sphingolipids can augment or prevent colon cancer, depending upon whether they are metabolized to S1P or promote S1P metabolism through the actions of SPL.

  8. Sphingosine-1-phosphate lyase downregulation promotes colon carcinogenesis through STAT3-activated microRNAs

    PubMed Central

    Degagné, Emilie; Pandurangan, Ashok; Bandhuvula, Padmavathi; Kumar, Ashok; Eltanawy, Abeer; Zhang, Meng; Yoshinaga, Yuko; Nefedov, Mikhail; de Jong, Pieter J.; Fong, Loren G.; Young, Stephen G.; Bittman, Robert; Ahmedi, Yasmin; Saba, Julie D.

    2014-01-01

    Growing evidence supports a link between inflammation and cancer; however, mediators of the transition between inflammation and carcinogenesis remain incompletely understood. Sphingosine-1-phosphate (S1P) lyase (SPL) irreversibly degrades the bioactive sphingolipid S1P and is highly expressed in enterocytes but downregulated in colon cancer. Here, we investigated the role of SPL in colitis-associated cancer (CAC). We generated mice with intestinal epithelium-specific Sgpl1 deletion and chemically induced colitis and tumor formation in these animals. Compared with control animals, mice lacking intestinal SPL exhibited greater disease activity, colon shortening, cytokine levels, S1P accumulation, tumors, STAT3 activation, STAT3-activated microRNAs (miRNAs), and suppression of miR-targeted anti-oncogene products. This phenotype was attenuated by STAT3 inhibition. In fibroblasts, silencing SPL promoted tumorigenic transformation through a pathway involving extracellular transport of S1P through S1P transporter spinster homolog 2 (SPNS2), S1P receptor activation, JAK2/STAT3-dependent miR-181b-1 induction, and silencing of miR-181b-1 target cylindromatosis (CYLD). Colon biopsies from patients with inflammatory bowel disease revealed enhanced S1P and STAT3 signaling. In mice with chemical-induced CAC, oral administration of plant-type sphingolipids called sphingadienes increased colonic SPL levels and reduced S1P levels, STAT3 signaling, cytokine levels, and tumorigenesis, indicating that SPL prevents transformation and carcinogenesis. Together, our results suggest that dietary sphingolipids can augment or prevent colon cancer, depending upon whether they are metabolized to S1P or promote S1P metabolism through the actions of SPL. PMID:25347472

  9. Multisite inhibition of Pinus pinea isocitrate lyase by phosphate.

    PubMed

    Ranaldi, F; Vanni, P; Giachetti, E

    2000-11-01

    Our results show that the phosphate ion is a nonlinear competitive inhibitor of Pinus pinea isocitrate lyase. In addition, this compound induces a sigmoidal response of the enzyme, which usually exhibits standard Michaelis-Menten kinetics. This peculiar behavior of P. pinea isocitrate lyase could be explained by a dimer (two-site) model, in which phosphate binds cooperatively, but the affinity of the vacant site for substrate (the magnesium-isocitrate complex) remains the same. As a result, the interaction of phosphate with free enzyme produces an inhibitor-enzyme-inhibitor species that is of significant importance in determining reaction rate; a possible regulatory role of the glyoxylate cycle by inorganic phosphate is suggested. The mode of phosphate inhibition is consistent with both the mechanism for magnesium ion activation of P. pinea isocitrate lyase and its site heterogeneity. Our results explain the cooperative effects observed by some authors in kinetic studies of isocitrate lyase carried out in phosphate buffers and also account for the higher K(m) values determined by using such assay systems. Phosphate buffer should be avoided in performing isocitrate lyase kinetics.

  10. Nicotinic acid-adenine dinucleotide phosphate activates the skeletal muscle ryanodine receptor.

    PubMed Central

    Hohenegger, Martin; Suko, Josef; Gscheidlinger, Regina; Drobny, Helmut; Zidar, Andreas

    2002-01-01

    Calcium is a universal second messenger. The temporal and spatial information that is encoded in Ca(2+)-transients drives processes as diverse as neurotransmitter secretion, axonal outgrowth, immune responses and muscle contraction. Ca(2+)-release from intracellular Ca(2+) stores can be triggered by diffusible second messengers like Ins P (3), cyclic ADP-ribose or nicotinic acid-adenine dinucleotide phosphate (NAADP). A target has not yet been identified for the latter messenger. In the present study we show that nanomolar concentrations of NAADP trigger Ca(2+)-release from skeletal muscle sarcoplasmic reticulum. This was due to a direct action on the Ca(2+)-release channel/ryanodine receptor type-1, since in single channel recordings, NAADP increased the open probability of the purified channel protein. The effects of NAADP on Ca(2+)-release and open probability of the ryanodine receptor occurred over a similar concentration range (EC(50) approximately 30 nM) and were specific because (i) they were blocked by Ruthenium Red and ryanodine, (ii) the precursor of NAADP, NADP, was ineffective at equimolar concentrations, (iii) NAADP did not affect the conductance and reversal potential of the ryanodine receptor. Finally, we also detected an ADP-ribosyl cyclase activity in the sarcoplasmic reticulum fraction of skeletal muscle. This enzyme was not only capable of synthesizing cyclic GDP-ribose but also NAADP, with an activity of 0.25 nmol/mg/min. Thus, we conclude that NAADP is generated in the vicinity of type 1 ryanodine receptor and leads to activation of this ion channel. PMID:12102654

  11. Control of glycolytic flux in Zymomonas mobilis by glucose 6-phosphate dehydrogenase activity

    SciTech Connect

    Snoep, J.L. |; Arfman, N.; Yomano, L.P.; Ingram, L.O.; Westerhoff, H.V.; Conway, T.

    1996-07-20

    Alycolytic genes in Zymomonas mobilis are highly expressed and constitute half of the cytoplasmic protein. The first four genes (glf, zwf, edd, glk) in this pathway form an operon encoding a glucose permease, glucose 6-phosphate dehydrogenase (G6-P dehydrogenase), 6-phosphogluconate dehydratase, and glucokinase, respectively. Each gene was overexpressed from a tac promoter to investigate the control of glycolysis during the early stages of batch fermentation when flux (qCO{sub 2}) is highest. Almost half of flux control appears to reside with G6-P dehydrogenase (C{sub G6-P dehydrogenase}{sup J} = 0.4). Although Z. mobilis exhibits one of the highest rates of glycolysis known, recombinants with elevated G6-P dehydrogenase had a 10% to 13% higher glycolytic flux than the native organism. A small increase in flux was also observed for recombinants expressing glf. Results obtained did not allow a critical evaluation of glucokinase and this enzyme may also represent an important control point. 6-Phosphogluconate dehydratase appears to be saturating at native levels. With constructs containing the full operon, growth rate and flux were both reduced, complicating interpretations. However, results obtained were also consistent with G6-P dehydrogenase as a primary site of control. Flux was 17% higher in operon constructs which exhibited a 17% increase in G6-P dehydrogenase specific activity, relative to the average of other operon constructs which contain a frameshift mutation in zwf.

  12. Antibacterial activity and ion release of bonding agent containing amorphous calcium phosphate nanoparticles

    PubMed Central

    Chen, Chen; Weir, Michael D.; Cheng, Lei; Lin, Nancy; Lin-Gibson, Sheng; Chow, Laurence C.; Zhou, Xuedong; Xu, Hockin H. K.

    2015-01-01

    Objectives Recurrent caries at the margins is a primary reason for restoration failure. The objectives of this study were to develop bonding agent with the double benefits of antibacterial and remineralizing capabilities, to investigate the effects of NACP filler level and solution pH on Ca and P ion release from adhesive, and to examine the antibacterial and dentin bond properties. Methods Nanoparticles of amorphous calcium phosphate (NACP) and a quaternary ammonium monomer (dimethylaminododecyl methacrylate, DMADDM) were synthesized. Scotchbond Multi-Purpose (SBMP) primer and adhesive served as control. DMADDM was incorporated into primer and adhesive at 5% by mass. NACP was incorporated into adhesive at filler mass fractions of 10%, 20%, 30% and 40%. A dental plaque microcosm biofilm model was used to test the antibacterial bonding agents. Calcium (Ca) and phosphate (P) ion releases from the cured adhesive samples were measured vs. filler level and solution pH of 7, 5.5 and 4. Results Adding 5% DMADDM and 10–40% NACP into bonding agent, and water-aging for 28 days, did not affect dentin bond strength, compared to SBMP control at 1 day (p > 0.1). Adding DMADDM into bonding agent substantially decreased the biofilm metabolic activity and lactic acid production. Total microorganisms, total streptococci, and mutans streptococci were greatly reduced for bonding agents containing DMADDM. Increasing NACP filler level from 10% to 40% in adhesive increased the Ca and P ion release by an order of magnitude. Decreasing solution pH from 7 to 4 increased the ion release from adhesive by 6–10 folds. Significance Bonding agents containing antibacterial DMADDM and remineralizer NACP were formulated to have Ca and P ion release, which increased with NACP filler level from 10% to 40% in adhesive. NACP adhesive was “smart” and dramatically increased the ion release at cariogenic pH 4, when these ions would be most-needed to inhibit caries. Therefore, bonding agent

  13. Selection of pecan shell-based activated carbons for removal of organic and inorganic impurities from water.

    PubMed

    Niandou, Mohamed A S; Novak, Jeffrey M; Bansode, Rishipal R; Yu, Jianmei; Rehrah, Djaafar; Ahmedna, Mohamed

    2013-01-01

    Activated carbons are a byproduct from pyrolysis and have value as a purifying agent. The effectiveness of activated carbons is dependent on feedstock selection and pyrolysis conditions that modify their surface properties. Therefore, pecan shell-based activated carbons (PSACs) were prepared by soaking shells in 50% (v/v) HPO or 25 to 50% of KOH-NaHCO followed by pyrolysis at 400 to 700°C under a N atmosphere. Physically activated PSACs were produced by pyrolysis at 700°C under N followed by activation with steam or CO at 700 to 900°C. Physicochemical, surface, and adsorption properties of the PSACs were compared with two commercially available activated carbons. The average mass yield of PSACs with respect to the initial mass of the biomass was about 20 and 34% for physically activated and chemically activated carbons, respectively. Acid-activated carbons exhibited higher surface area, higher bulk density, and lower ash content compared with steam- or CO-activated carbons and the two commercial products. Base activation led to the development of biochar with moderate to high surface area with surface charges suitable for adsorption of anionic species. Regardless of the activation method, PSACs had high total surface area ranging from 400 to 1000 m g, better pore size distribution, and more surface charges than commercial samples. Our results also showed that PSACs were effective in removing inorganic contaminants such as Cu and NO as well as organic contaminants such as atrazine and metolachlor. This study showed that pyrolysis conditions and activation had a large influence on the PSAC's surface characteristics, which can limit its effectiveness as a custom sorbent for targeted water contaminants.

  14. Effect of phosphate activating group on oligonucleotide formation on montmorillonite: the regioselective formation of 3',5'-linked oligoadenylates

    NASA Technical Reports Server (NTRS)

    Prabahar, K. J.; Cole, T. D.; Ferris, J. P.

    1994-01-01

    The effects of amine structure on the montmorillonite-catalyzed oligomerization of the 5'-phosphoramidates of adenosine are investigated. 4-Aminopyridine derivatives yielded oligoadenylates as long as dodecamers with a regioselectivity for 3',5'-phosphodiester bond formation averaging 88%. Linear and cyclic oligomers are obtained and no A5'ppA-containing products are detected. Oligomers as long as the hexanucleotide are obtained using 2-aminobenzimidazole as the activating group. A predominance of pA2'pA is detected in the dimer fraction along with cyclic 3',5'-trimer; no A5'ppA-containing oligomers were detected. Little or no oligomer formation was observed when morpholine, piperidine, pyrazole, 1,2,4-triazole, and 2-pyridone are used as phosphate-activating groups. The effects of the structure of the phosphate activating group on the oligomer structure and chain lengths are discussed.

  15. Characterization of cytolytic neutrophil activation in vitro by amorphous hydrated calcium phosphate as a model of biomaterial inflammation.

    PubMed

    Edwards, Felicity C; Taheri, Amir; Dann, Sophie C; Dye, Julian F

    2011-03-01

    Calcium ions are utilized in biomolecular biomaterial design for osteomimetic scaffolds and as divalent cross-linking agents, typically for gelation of alginates, stabilisation of protein structure (e.g., fibrinogen) and enzyme activation (e.g., thrombin). Biological interactions with defined calcium phosphates (e.g., hydroxyapatite) are exploited for osteogenesis, although crystalline calcium phosphates (e.g., calcium pyrophosphate) stimulate inflammation. We found that the calcium concentration used in the manufacture of prototype dermal scaffolds made from fibrin/alginate composite was related to the inflammatory infiltration during in vivo integration. In investigating a cause for this inflammatory response, we have identified and characterized a cytolytic inflammatory effect of amorphous calcium phosphate (CaP) formed in physiological solutions, relevant to biomaterial biocompatibility. Isolated human neutrophils (Nφ) were incubated in phosphate-buffered saline with CaCl(2) ranging 2.5-20 mM total calcium. Nφ activation was assessed by morphology and integrin-β2 (CD18a) expression. Mediator release (Nφ-elastase, IL-8, and TNFα) was measured from both Nφ and whole blood cultures plus CaCl(2). CaP exposure increased CD18a expression over 1 h (maximal at 10 mM calcium/ phosphate) with concurrent phagocytosis, cytolysis, and Nφ-elastase release. CaCl(2) induced expression of IL-8 and TNFα in whole blood cultures. These results suggest that CaP formed from the resorption of calcium-containing biomaterials could induce inflammation and accelerate biomaterial degradation, driving further CaP release. This demonstrates a novel mechanism for biomaterial-induced inflammation. The in vitro system described could aid preclinical evaluation of novel biomaterial inflammatory potential.

  16. Comparison of the adjuvant activity of aluminum hydroxide and calcium phosphate on the antibody response towards Bothrops asper snake venom.

    PubMed

    Olmedo, Hidekel; Herrera, María; Rojas, Leonardo; Villalta, Mauren; Vargas, Mariángela; Leiguez, Elbio; Teixeira, Catarina; Estrada, Ricardo; Gutiérrez, José María; León, Guillermo; Montero, Mavis L

    2014-01-01

    The adjuvanticity of aluminum hydroxide and calcium phosphate on the antibody response in mice towards the venom of the snake Bothrops asper was studied. It was found that, in vitro, most of the venom proteins are similarly adsorbed by both mineral salts, with the exception of some basic phospholipases A2, which are better adsorbed by calcium phosphate. After injection, the adjuvants promoted a slow release of the venom, as judged by the lack of acute toxicity when lethal doses of venom were administered to mice. Leukocyte recruitment induced by the venom was enhanced when it was adsorbed on both mineral salts; however, venom adsorbed on calcium phosphate induced a higher antibody response towards all tested HPLC fractions of the venom. On the other hand, co-precipitation of venom with calcium phosphate was the best strategy for increasing: (1) the capacity of the salt to couple venom proteins in vitro; (2) the venom ability to induce leukocyte recruitment; (3) phagocytosis by macrophages; and (4) a host antibody response. These findings suggest that the chemical nature is not the only one determining factor of the adjuvant activity of mineral salts.

  17. Sphingosine-1-phosphate signaling and biological activities in the cardiovascular system.

    PubMed

    Takuwa, Yoh; Okamoto, Yasuo; Yoshioka, Kazuaki; Takuwa, Noriko

    2008-09-01

    The plasma lysophospholipid mediator sphingosine-1-phosphate (S1P) is produced exclusively by sphingosine kinase (SPHK) 1 and SPHK2 in vivo, and plays diverse biological and pathophysiological roles by acting largely through three members of the G protein-coupled S1P receptors, S1P1, S1P2 and S1P3. S1P1 expressed on endothelial cells mediates embryonic vascular maturation and maintains vascular integrity by contributing to eNOS activation, inhibiting vascular permeability and inducing endothelial cell chemotaxis via Gi-coupled mechanisms. By contrast, S1P2, is expressed in high levels on vascular smooth muscle cells (VSMCs) and certain types of tumor cells, inhibiting Rac and cell migration via a G(12/13)-and Rho-dependent mechanism. In rat neointimal VSMCs, S1P1 is upregulated to mediate local production of platelet-derived growth factor, which is a key player in vascular remodeling. S1P3 expressed on endothelial cells also mediates chemotaxis toward S1P and vasorelaxation via NO production in certain vascular bed, playing protective roles for vascular integrity. S1P3 expressed on VSMCs and cardiac sinoatrial node cells mediates vasopressor and negative chronotropic effect, respectively. In addition, S1P3, together with S1P2 and SPHK1, is suggested to play a protective role against acute myocardial ischemia. However, our recent work indicates that overexpressed SPHK1 is involved in cardiomyocyte degeneration and fibrosis in vivo, in part through S1P activation of the S1P3 signaling. We also demonstrated that exogenously administered S1P accelerates neovascularization and blood flow recovery in ischemic limbs, suggesting its usefulness for angiogenic therapy. These results provide evidence for S1P receptor subtype-specific pharmacological intervention as a novel therapeutic approach to cardiovascular diseases and cancer.

  18. Calcium phosphate/porous silicon biocomposites prepared by cyclic deposition methods: spin coating vs electrochemical activation.

    PubMed

    Hernandez-Montelongo, J; Gallach, D; Naveas, N; Torres-Costa, V; Climent-Font, A; García-Ruiz, J P; Manso-Silvan, M

    2014-01-01

    Porous silicon (PSi) provides an excellent platform for bioengineering applications due to its biocompatibility, biodegradability, and bioresorbability. However, to promote its application as bone engineering scaffold, deposition of calcium phosphate (CaP) ceramics in its hydroxyapatite (HAP) phase is in progress. In that sense, this work focuses on the synthesis of CaP/PSi composites by means of two different techniques for CaP deposition on PSi: Cyclic Spin Coating (CSC) and Cyclic Electrochemical Activation (CEA). Both techniques CSC and CEA consisted on alternate Ca and P deposition steps on PSi. Each technique produced specific morphologies and CaP phases using the same independent Ca and P stem-solutions at neutral pH and at room temperature. The brushite (BRU) phase was favored with the CSC technique and the hydroxyapatite (HAP) phase was better synthesized using the CEA technique. Analyses by elastic backscattering spectroscopy (EBS) on CaP/PSi structures synthesized by CEA supported that, by controlling the CEA parameters, an HAP coating with the required Ca/P atomic ratio of 1.67 can be promoted. Biocompatibility was evaluated by bone-derived progenitor cells, which grew onto CaP/PSi prepared by CSC technique with a long-shaped actin cytoskeleton. The density of adhered cells was higher on CaP/PSi prepared by CEA, where cells presented a normal morphological appearance and active mitosis. These results can be used for the design and optimization of CaP/PSi composites with enhanced biocompatibility for bone-tissue engineering.

  19. Microbial Community Structure of Activated Sludge for Biosolubilization of Two Different Rock Phosphates.

    PubMed

    Xiao, Chunqiao; Wu, Xiaoyan; Liu, Tingting; Xu, Guang; Chi, Ruan

    2016-12-16

    A microbial consortium was directly taken from activated sludge and was used to solubilize rock phosphates (RPs) in a lab-scale bioreactor in this study. Results showed that the microbial consortium could efficiently release soluble phosphorus (P) from the RPs, and during 30-day incubation, it grew well in the bioreactor and reduced the pH of the solutions. The biosolubilization process was also illustrated by the observation of scanning electron microscopy combined with an energy dispersive X-ray spectroscopy (SEM-EDX), which showed an obvious corrosion on the ore surfaces, and most elements were removed from the ore samples. The analysis of microbial community structure by Illumina 16S ribosomal RNA (rRNA) gene and 18S rRNA gene MiSeq sequencing reflected different microbial diversity and richness in the solutions added with different ore samples. A lower richness and diversity of bacteria but a higher richness and diversity of fungi occurred in the solution added with ore sample 1 compared to that of in the solution added with ore sample 2. Alphaproteobacteria and Saccharomycetes were the dominating bacterial and fungal group, respectively, both in the solutions added with ore samples 1 and 2 at the class level. However, their abundances in the solution added with ore sample 1 were obviously lower than that in the solution added with ore sample 2. This study provides new insights into our understanding of the microbial community structure in the biosolubilization of RPs by a microbial consortium directly taken from activated sludge.

  20. New Advances in Nanotechnology-Based Diagnosis and Therapeutics for Breast Cancer: An Assessment of Active-Targeting Inorganic Nanoplatforms.

    PubMed

    Falagan-Lotsch, Priscila; Grzincic, Elissa M; Murphy, Catherine J

    2017-01-18

    Breast cancer is a major cause of suffering and mortality among women. Limitations in the current diagnostic methods and treatment approaches have led to new strategies to positively impact the survival rates and quality of life of breast cancer patients. Nanotechnology offers a real possibility of mitigating breast cancer mortality by early-stage cancer detection and more precise diagnosis as well as more effective treatments with minimal side effects. The current nanoplatforms approved for breast cancer therapeutics are based on passive tumor targeting using organic nanoparticles and have not provided the expected significant improvements in the clinic. In this review, we present the emerging approaches in breast cancer nanomedicine based on active targeting using versatile inorganic nanoplatforms with biomedical relevance, such as gold, silica, and iron oxide nanoparticles, as well as their efficacy in breast cancer imaging, drug and gene delivery, thermal therapy, combinational therapy, and theranostics in preclinical studies. The main challenges for clinical translation and perspectives are discussed.

  1. Effects of pH and inorganic salts on the adsorption of phenol from aqueous systems on activated decolorizing charcoal

    SciTech Connect

    Halhouli, K.A.; Darwish, N.A.; Al-Dhoon, N.M.

    1995-10-01

    An experimental investigation of the effects of pH and three inorganic salts (KCl, KI, and NaCl) on the adsorption isotherms of phenol (from a dilute aqueous solution) on activated charcoal was conducted. Each salt was studied at three different concentrations, i.e., 0.1, 0.01, and 0.005 M. The effect of pH (in the pH range 3 to 11) in the presence of KI, KCl, and NaCl was also investigated. The concentration of phenol in the aqueous systems studied ranged from 10 to 200 ppm. The temperature effect was also studied, and the resulting experimental equilibrium isotherms at 30, 40, and 55{degrees}C are well represented by Freundlich, Langmuir, and Redlich-Paterson isotherms. The relevant parameters for these isotherms are presented.

  2. Lowering effect of firefly squid powder on triacylglycerol content and glucose-6-phosphate dehydrogenase activity in rat liver.

    PubMed

    Takeuchi, Hiroyuki; Morita, Ritsuko; Shirai, Yoko; Nakagawa, Yoshihisa; Terashima, Teruya; Ushikubo, Shun; Matsuo, Tatsuhiro

    2014-01-01

    Effects of dietary firefly squid on serum and liver lipid levels were investigated. Male Wistar rats were fed a diet containing 5% freeze-dried firefly squid or Japanese flying squid for 2 weeks. There was no significant difference in the liver triacylglycerol level between the control and Japanese flying squid groups, but the rats fed the firefly squid diet had a significantly lower liver triacylglycerol content than those fed the control diet. No significant difference was observed in serum triacylglycerol levels between the control and firefly squid groups. The rats fed the firefly squid had a significantly lower activity of liver glucose-6-phosphate dehydrogenase compared to the rats fed the control diet. There was no significant difference in liver fatty acid synthetase activity among the three groups. Hepatic gene expression and lipogenic enzyme activity were investigated; a DNA microarray showed that the significantly enriched gene ontology category of down-regulated genes in the firefly squid group was "lipid metabolic process". The firefly squid group had lower mRNA level of glucose-6-phosphate dehydrogenase compared to the controls. These results suggest that an intake of firefly squid decreases hepatic triacylglycerol in rats, and the reduction of mRNA level and enzyme activity of glucose-6-phosphate dehydrogenase might be related to the mechanisms.

  3. A limitation of the continuous spectrophotometric assay for the measurement of myo-inositol-1-phosphate synthase activity.

    PubMed

    Huang, Xinyi; Hernick, Marcy

    2011-10-15

    Myo-inositol-1-phosphate synthase (MIPS) catalyzes the conversion of glucose-6-phosphate to myo-inositol-1-phosphate. The reaction catalyzed by MIPS is the first step in the biosynthesis of inositol and inositol-containing molecules that serve important roles in both eukaryotes and prokaryotes. Consequently, MIPS is a target for the development of therapeutic agents for the treatment of infectious diseases and bipolar disorder. We recently reported a continuous spectrophotometric method for measuring MIPS activity using a coupled assay that allows the rapid characterization of MIPS in a multiwell plate format. Here we validate the continuous assay as a high-throughput alternative for measuring MIPS activity and report on one limitation of this assay-the inability to examine the effect of divalent metal ions (at high concentrations) on MIPS activity. In addition, we demonstrate that the activity of MIPS from Arabidopsis thaliana is moderately enhanced by the addition Mg(2+) and is not enhanced by other divalent metal ions (Zn(2+) and Mn(2+)), consistent with what has been observed for other eukaryotic MIPS enzymes. Our findings suggest that the continuous assay is better suited for characterizing eukaryotic MIPS enzymes that require monovalent cations as cofactors than for characterizing bacterial or archeal MIPS enzymes that require divalent metal ions as cofactors.

  4. Phosphate binding by cerebral microsomes in relation to adenosine-triphosphatase activity

    PubMed Central

    Rodnight, R.; Hems, D. A.; Lavin, B. E.

    1966-01-01

    1. Microsomes prepared from guinea-pig and ox brain were incubated for periods of a few seconds with low concentrations of Mg-[32P]ATP, the reaction was stopped with trichloroacetic acid and determinations were made of the phosphate bound to the acid-washed, and in some cases solvent-extracted, residue. 2. At 20 μm-ATP, at 37° and in the presence of Na+ ions, 30–50 μμmoles of phosphate/mg. of microsomal protein were bound by the preparation within 1 sec. of starting the reaction; little further change in level occurred until hydrolysis of ATP exceeded 50%, when the bound phosphate began to decline fairly rapidly to the zero-time value. 3. At 20μm-ATP without Na+ ions present or in the presence of K+ ions, the level of bound phosphate increased gradually and did not decline as ATP hydrolysis approached completion. 4. Potassium ions either inhibited the formation of Na+-dependent bound phosphate or, when added during the course of the reaction, rapidly reduced its level. 5. At 200 μm-ATP the bound phosphate formed in the presence of Na+ ions appeared to consist of a mixture of the unstable Na+-dependent type and the stable type requiring only Mg2+ ions for its formation. 6. Non-radioactive ATP added during the course of the reaction at 20 μm-ATP with Na+ions present rapidly discharged virtually all the bound 32P counts; at 200 μm-ATP only a proportion of the label was similarly discharged. The Na+-dependent bound phosphate is therefore turning over, in contrast with that formed in the absence of Na+ions, which proved more stable. 7. The Na+-dependent bound phosphate was not in the form of ATP; experiments with [14C]ATP instead of [32P]ATP showed a small and invariable binding of ATP by the preparation unaffected by Na+ ions or time of incubation. 8. Under the usual conditions employed in this work ouabain stimulated formation of Na+-dependent bound phosphate when Na+ ions were suboptimum and inhibited it when optimum Na+ ions were present. 9. The Na

  5. Arsenic, inorganic

    Integrated Risk Information System (IRIS)

    Arsenic , inorganic ; CASRN 7440 - 38 - 2 Human health assessment information on a chemical substance is included in the IRIS database only after a comprehensive review of toxicity data , as outlined in the IRIS assessment development process . Sections I ( Health Hazard Assessments for Noncarcinoge

  6. Mapping calcium phosphate activated gene networks as a strategy for targeted osteoinduction of human progenitors in vitro and in vivo

    PubMed Central

    Eyckmans, J.; Roberts, S.J.; Bolander, J.; Schrooten, J.; Chen, C.S.; Luyten, F.P.

    2014-01-01

    Although calcium phosphate-containing biomaterials are promising scaffolds for bone regenerative strategies, the osteoinductive capacity of such materials is poorly understood. In this study, we investigated whether endogenous mechanisms of in vivo calcium phosphate-driven, ectopic bone formation could be identified and used to induce enhanced differentiation in vitro of the same progenitor population. To accomplish this, human periosteum derived cells (hPDCs) were seeded on hydroxyapatite/collagen scaffolds (calcium phosphate rich matrix or CPRM), or on decalcified scaffolds (calcium phosphate depleted matrix or CPDM), followed by subcutaneous implantation in nude mice to trigger ectopic bone formation. In this system, osteoblast differentiation occurred in CPRM scaffolds, but not in CPDM scaffolds. Gene expression was assessed by human full-genome microarray at 20 hours after seeding, and 2, 8 and 18 days after implantation. In both matrices, implantation of the cell constructs triggered a similar gene expression cascade, however, gene expression dynamics progressed faster in CPRM scaffolds than in CPDM scaffolds. The difference in gene expression dynamics was associated with differential activation of hub genes and molecular signaling pathways related to calcium signaling (CREB), inflammation (TNFα, NFkB, and IL6) and bone development (TGFβ, β-catenin, BMP, EGF, and ERK signaling). Starting from this set of pathways, a growth factor cocktail was developed that robustly enhanced osteogenesis in vitro and in vivo. Taken together, our data demonstrate that through the identification and subsequent stimulation of genes, proteins and signaling pathways associated with calcium phosphate mediated osteoinduction, a focused approach to develop targeted differentiation protocols in adult progenitor cells can be achieved. PMID:23537666

  7. Comparison of calcium ionophore and receptor-activated inositol phosphate formation in primary glial cell cultures.

    PubMed

    Wigginton, S A; Minneman, K P

    1991-11-13

    The possible role of Ca2+ influx in alpha 1-adrenoceptor-stimulated [3H]inositol phosphate [( 3H]InsP) formation was examined in primary cultures of glial cells from 1-day-old rat brain. The Ca2+ ionophore A23187 caused a concentration- and time-dependent increase in [3H]InsP formation similar in magnitude to that caused by norepinephrine (NE). Responses to A23187 and NE were both completely dependent on extracellular Ca2+, with a similar concentration dependence. However, cadmium was more potent in blocking the response to A23187 than to NE. Lanthanum (1 mM) blocked the response to NE, although cobalt (5 mM) did not. The [3H]InsP response to A23187 was not additive with the response to NE or to the muscarinic agonist carbachol, although responses to NE and carbachol were addictive Both A23187 and ionomycin inhibited the additive stimulation caused by a combination of NE and carbachol, and this inhibition was potentiated by cadmium. Ionomycin stimulated [3H]InsP formation at concentrations lower than those inhibiting receptor-mediated responses, and this stimulation was not additive with responses to NE or carbachol. High-performance liquid chromatography separation showed similar patterns of [3H]InsPs formed in response to both Ca2+ ionophore and receptor agonists. These results raise the possibility that receptor-activated Ca2+ influx may be involved in stimulation of [3H]InsP formation in these cells.

  8. Differential effects of zinc and magnesium ions on mineralization activity of phosphatidylserine calcium phosphate complexes.

    PubMed

    Wu, Licia N Y; Genge, Brian R; Wuthier, Roy E

    2009-07-01

    Mg(2+) and Zn(2+) are present in the mineral of matrix vesicles (MVs) and biological apatites, and are known to influence the onset and progression of mineral formation by amorphous calcium phosphate (ACP) and hydroxyapatite (HAP). However, neither has been studied systematically for its effect on mineral formation by phosphatidylserine-Ca(2+)-Pi complexes (PS-CPLX), an important constituent of the MV nucleation core. Presented here are studies on the effects of increasing levels of Mg(2+) and Zn(2+) on the process of mineral formation, either when present in synthetic cartilage lymph (SCL), or when incorporated during the formation of PS-CPLX. Pure HAP and PS-CPLX proved to be powerful nucleators, but ACP took much longer to induce mineral formation. In SCL, Mg(2+) and Zn(2+) had significantly different inhibitory effects on the onset and amount of mineral formation; HAP and PS-CPLX were less affected than ACP. Mg(2+) and Zn(2+) caused similar reductions in the rate and length of rapid mineral formation, but Zn(2+) was a more potent inhibitor on a molar basis. When incorporated into PS-CPLX, Mg(2+) and Zn(2+) caused significantly different effects than when present in SCL. Even low, subphysiological levels of Mg(2+) altered the inherent structure of PS-CPLX and markedly reduced its ability to induce and propagate mineral formation. Incorporated Zn(2+) caused significantly less effect, low (<20 microM) levels causing almost no inhibition. Levels of Zn(2+) present in MVs do not appear to inhibit their nucleational activity.

  9. Renal threshold phosphate concentration (TmPO4/GFR).

    PubMed Central

    Kruse, K; Kracht, U; Göpfert, G

    1982-01-01

    The ratio of maximum rate of renal tubular reabsorption of phosphate to glomerular filtration rate (TmPO4/GFR) was determined in 546 schoolchildren, aged between 6 and 17.9 years, using the nomogram of Walton and Bijvoet.1 TmPO4/GFR correlated with chronological age in girls and boys and in each remained significantly higher than in adults. TmPO4/GFR in the children correlated neither with fasting serum immunoreactive calcitonin and parathyroid hormone levels nor with the urinary cyclic AMP excretion. The study showed a parallel decrease in TmPO4/GFR, excretion of total hydroxyproline and serum alkaline phosphatase activities after puberty, with a significant relationship of both these indices of bone turnover to TmPO4/GFR values. This indicates that the high renal phosphate threshold of children may be an important factor for bone mineralisation by providing high extracellular inorganic phosphate concentrations during normal growth. PMID:6280622

  10. Alkaline phosphatase revisited: hydrolysis of alkyl phosphates.

    PubMed

    O'Brien, Patrick J; Herschlag, Daniel

    2002-03-05

    Escherichia coli alkaline phosphatase (AP) is the prototypical two metal ion catalyst with two divalent zinc ions bound approximately 4 A apart in the active site. Studies spanning half a century have elucidated many structural and mechanistic features of this enzyme, rendering it an attractive model for investigating the potent catalytic power of bimetallic centers. Unfortunately, fundamental mechanistic features have been obscured by limitations with the standard assays. These assays generate concentrations of inorganic phosphate (P(i)) in excess of its inhibition constant (K(i) approximately 1 muM). This tight binding by P(i) has affected the majority of published kinetic constants. Furthermore, binding limits k(cat)/K(m) for reaction of p-nitrophenyl phosphate, the most commonly employed substrate. We describe a sensitive (32)P-based assay for hydrolysis of alkyl phosphates that avoids the complication of product inhibition. We have revisited basic mechanistic features of AP with these alkyl phosphate substrates. The results suggest that the chemical step for phosphorylation of the enzyme limits k(cat)/K(m). The pH-rate profile and additional results suggest that the serine nucleophile is active in its anionic form and has a pK(a) of < or = 5.5 in the free enzyme. An inactivating pK(a) of 8.0 is observed for binding of both substrates and inhibitors, and we suggest that this corresponds to ionization of a zinc-coordinated water molecule. Counter to previous suggestions, inorganic phosphate dianion appears to bind to the highly charged AP active site at least as strongly as the trianion. The dependence of k(cat)/K(m) on the pK(a) of the leaving group follows a Brønsted correlation with a slope of beta(lg) = -0.85 +/- 0.1, differing substantially from the previously reported value of -0.2 obtained from data with a less sensitive assay. This steep leaving group dependence is consistent with a largely dissociative transition state for AP-catalyzed hydrolysis of

  11. Sphingosine 1-phosphate receptor activation enhances BMP-2-induced osteoblast differentiation

    SciTech Connect

    Sato, Chieri; Iwasaki, Tsuyoshi; Kitano, Sachie; Tsunemi, Sachi; Sano, Hajime

    2012-06-22

    Highlights: Black-Right-Pointing-Pointer We investigated the role of S1P signaling for osteoblast differentiation. Black-Right-Pointing-Pointer Both S1P and FTY enhanced BMP-2-stimulated osteoblast differentiation by C2C12 cells. Black-Right-Pointing-Pointer S1P signaling enhanced BMP-2-stimulated Smad and ERK phosphorylation by C2C12 cells. Black-Right-Pointing-Pointer MEK/ERK signaling is a pathway underlying S1P signaling for osteoblast differentiation. -- Abstract: We previously demonstrated that sphingosine 1-phosphate (S1P) receptor-mediated signaling induced proliferation and prostaglandin productions by synovial cells from rheumatoid arthritis (RA) patients. In the present study we investigated the role of S1P receptor-mediated signaling for osteoblast differentiation. We investigated osteoblast differentiation using C2C12 myoblasts, a cell line derived from murine satellite cells. Osteoblast differentiation was induced by the treatment of bone morphogenic protein (BMP)-2 in the presence or absence of either S1P or FTY720 (FTY), a high-affinity agonist of S1P receptors. Osteoblast differentiation was determined by osteoblast-specific transcription factor, Runx2 mRNA expression, alkaline phosphatase (ALP) activity and osteocalcin production by the cells. Smad1/5/8 and extracellular signal-regulated kinase (ERK) 1/2 phosphorylation was examined by Western blotting. Osteocalcin production by C2C12 cells were determined by ELISA. Runx2 expression and ALP activity by BMP-2-stimulated C2C12 cells were enhanced by addition of either S1P or FTY. Both S1P and FTY enhanced BMP-2-induced ERK1/2 and Smad1/5/8 phosphorylation. The effect of FTY was stronger than that of S1P. S1P receptor-mediated signaling on osteoblast differentiation was inhibited by addition of mitogen-activated protein kinase/ERK kinase (MEK) 1/2 inhibitor, indicating that the S1P receptor-mediated MEK1/2-ERK1/2 signaling pathway enhanced BMP-2-Smad signaling. These results indicate that S1P

  12. Microbial Populations Associated with Phosphate-Mediated Vadose Zone Sequestration of Strontium and Uranium

    NASA Astrophysics Data System (ADS)

    Wu, C. H.; Chou, J.; Fujita, Y.; Bill, M.; Brodie, E. L.; Andersen, G. L.; Hazen, T. C.; Conrad, M. S.

    2007-12-01

    Significant quantities of metals and radionuclides are contained in thick unsaturated zones at several contaminated sites in the western US. In many cases, this contamination has migrated to underlying groundwater, sometimes decades after being released into the subsurface. Because of the prohibitive costs associated with physically removing the contamination, an attractive remedy to this problem is to develop methods for long-term in situ stabilization of the contamination in the vadose zone. Our research focuses on developing a method of introducing gaseous compounds to stimulate precipitation of stable phosphate mineral phases in the vadose zone to immobilize soluble contaminants thus minimizing further transport to groundwater. Preliminary studies have demonstrated that biological precipitation of phosphate minerals can be stimulated under unsaturated conditions by injection of triethyl phosphate (TEP) gas. Microorganisms hydrolyze TEP, releasing inorganic phosphate, catalyzing the precipitation of metals and radionuclide-containing phosphate minerals. Our initial results demonstrate that a mixed culture of aerobic microorganisms from vadose zone sediments, enriched with TEP, produce significantly higher concentrations of inorganic phosphate than the no TEP control. A high-density microarray (PhyloChip) capable of detecting up to 9,000 prokaryotic taxa will be used to identify the microbial community composition of the enriched culture. In addition, the metabolically active organisms will be investigated through extraction and hybridization of ribosomal RNA. Organisms capable of hydrolyzing TEP to inorganic phosphate will be further characterized to determine the requirements for aerobic microbially-mediated radionuclide immobilization. The chemical and isotopic compositions of the reactants and products will be measured to enable in situ monitoring of microbial TEP utilization. The result of these studies will be the basis for unsaturated column experiments

  13. Extracellular phosphates enhance activities of voltage-gated proton channels and production of reactive oxygen species in murine osteoclast-like cells.

    PubMed

    Li, Guangshuai; Miura, Katsuyuki; Kuno, Miyuki

    2017-02-01

    Osteoclasts are highly differentiated bone-resorbing cells and play a significant role in bone remodelling. In the resorption pit, inorganic phosphate (Pi) concentrations increase because of degradation of hydroxyapatite. We studied effects of extracellular Pi on voltage-gated H(+) channels in osteoclast-like cells derived from a macrophage cell line (RAW264). Extracellular Pi (1.25-20 mM) increased the H(+) channel currents dose dependently and reversibly. The Pi-induced increases were attenuated by removal of extracellular Na(+) and by phosphonoformic acid, a blocker of Na(+)-dependent Pi transporters. Pi increased the maximal conductance, decreased activation time constant, increased deactivation time constant, and shifted the conductance-voltage relationship to more negative voltages. The most marked change was enhanced gating which was mainly caused by elevation of intracellular Pi levels. The Pi-induced enhanced gating was partially inhibited by protein kinase C (PKC) inhibitors, GF109203X and staurosporine, indicating that PKC-mediated phosphorylation was involved in part. The increase in the maximal conductance was mainly due to accompanying decrease in intracellular pH. These effects of Pi were not affected by intracellular Mg(2+), bafilomycin A1 (V-ATPase inhibitor) and removal of intracellular ATP. Extracellular Pi also upregulated reactive oxygen species (ROS). Diphenyleneiodonium chloride, an inhibitor of NADPH oxidases, decreased ROS production and partially attenuated the enhanced gating. In the cells during later passages where osteoclastogenesis declined, H(+) channel activities and ROS production were both modest. These results suggest that, in osteoclasts, ambient Pi is a common enhancer for H(+) channels and ROS production and that potentiation of H(+) channels may help ROS production.

  14. Bifunctional sucrose phosphate synthase/phosphatase is involved in the sucrose biosynthesis by Methylobacillus flagellatus KT.

    PubMed

    But, Sergey Y; Khmelenina, Valentina N; Reshetnikov, Alexander S; Trotsenko, Yuri A

    2013-10-01

    The aerobic obligate methylotroph Methylobacillus flagellatus KT was shown to synthesize sucrose in the presence of 0.5-2% NaCl in the growth medium. In the genome of this bacterium, an open reading frame (ORF) encoding a predicted 84-kD polypeptide homologous to the plant and cyanobacterial sucrose phosphate synthases (SPSs) was found. Using heterologous expression of the putative sps gene in Escherichia coli, followed by affinity chromatography, pure recombinant protein SPS-His6 was obtained. The enzyme catalyzed two reactions: conversion of fructose 6-phosphate and UDP-glucose into sucrose 6-phosphate and hydrolysis of sucrose 6-phosphate to sucrose. The bifunctional sucrose phosphate synthase/phosphatase (SPS/SPP) was a 340 kDa homotetrameric Mg(2+) -dependent enzyme activated by fructose 1,6-phosphate2 and ATP but inhibited by glucose 6-phosphate, fructose 1-phosphate, AMP and inorganic phosphate. The amino acid sequence of the protein had a C-terminal domain homologous to SPPs. This correlated with the absence of the spp gene in the M. flagellatus chromosome. The ORFs homologous to the M. flagellatus SPS were found in the genomes of another obligate methylotroph Methylovorus glucosetrophus as well as the lithoautotrophic bacteria Acidithiobacillus ferrooxidans, Nitrosomonas europaea and Nitrosospira multiformis whose genomes lacked the spp genes. Thus, data extending the knowledge of biochemical properties of bacterial SPSs have been obtained.

  15. The study of redox-active inorganic substituents of cellulase enzyme

    SciTech Connect

    Not Available

    1992-01-01

    Hexaammineruthemium(III) chloride enhances the catalytic activity of Trichoderma reesei cellobiohydrolase I (CBHI) by as much as 45 percent over a 24 hr period. The mechanism involved could be related to the redox activity and reduction of O[sub 2] by RU(II) complexes. Since the addition of ascorbic acid is not required for the enhancement of CBHI activity, we speculate that the cellobiose generated by the enzyme activity may serve as the reducing agent for the formation of RU(II) species.

  16. Rapid and direct estimation of active biomass on granular activated carbon through adenosine tri-phosphate (ATP) determination.

    PubMed

    Velten, Silvana; Hammes, Frederik; Boller, Markus; Egli, Thomas

    2007-05-01

    Granular activated carbon (GAC) filtration is used during drinking water treatment for the removal of micropollutants such as taste and odour compounds, halogenated hydrocarbons, pesticides and pharmaceuticals. In addition, the active microbial biomass established on GAC is responsible for the removal of biodegradable dissolved organic carbon compounds present in water or formed during oxidation (e.g., ozonation and chlorination) processes. In order to conduct correct kinetic evaluations of DOC removal during drinking water treatment, and to assess the state and performance of full-scale GAC filter installations, an accurate and sensitive method for active biomass determination on GAC is required. We have developed a straight-forward method based on direct measurement of the total adenosine tri-phosphate (ATP) content of a GAC sample and other support media. In this method, we have combined flow-cytometric absolute cell counting and ATP analysis to derive case-specific ATP/cell conversion values. In this study, we present the detailed standardisation of the ATP method. An uncertainty assessment has shown that heterogeneous colonisation of the GAC particles makes the largest contribution to the combined standard uncertainty of the method. The method was applied for the investigation of biofilm formation during the start-up period of a GAC pilot-scale plant treating Lake Zurich water. A rapid increase in the biomass of up to 1.1 x 10(10)cells/g GAC dry weight (DW) within the first 33 days was observed, followed by a slight decrease to an average steady-state concentration of 7.9 x 10(9)cells/g GAC DW. It was shown that the method can be used to determine the biomass attached to the GAC for both stable and developing biofilms.

  17. Synthesis and Catalytic Activity of Ruthenium-Indenylidene Complexes for Olefin Metathesis: Microscale Experiments for the Undergraduate Inorganic or Organometallic Laboratories

    ERIC Educational Resources Information Center

    Pappenfus, Ted M.; Hermanson, David L.; Ekerholm, Daniel P.; Lilliquist, Stacie L.; Mekoli, Megan L.

    2007-01-01

    A series of experiments for undergraduate laboratory courses (e.g., inorganic, organometallic or advanced organic) have been developed. These experiments focus on understanding the design and catalytic activity of ruthenium-indenylidene complexes for olefin metathesis. Included in the experiments are the syntheses of two ruthenium-indenylidene…

  18. Parallel determination of enzyme activities and in vivo fluxes in Brassica napus embryos grown on organic or inorganic nitrogen source.

    PubMed

    Junker, Björn H; Lonien, Joachim; Heady, Lindsey E; Rogers, Alistair; Schwender, Jörg

    2007-01-01

    After the completion of the genomic sequencing of model organisms, numerous post-genomic studies, integrating transcriptome and metabolome data, are aimed at developing a more complete understanding of cell physiology. Here, we measure in vivo metabolic fluxes by steady state labeling, and in parallel, the activity of enzymes in central metabolism in cultured developing embryos of Brassica napus. Embryos were grown on either the amino acids glutamine and alanine as an organic nitrogen source, or on ammonium nitrate as an inorganic nitrogen source. The type of nitrogen made available to developing embryos caused substantial differences in fluxes associated with the tricarboxylic acid cycle, including flux reversion. The changes observed in enzyme activity were not consistent with our estimates of metabolic flux. Furthermore, most extractable enzyme activities are in large surplus relative to the requirements for the observed in vivo fluxes. The results demonstrate that in this model system the metabolic response of central metabolism to changes in environmental conditions can be achieved largely without regulatory reprogramming of the enzyme machinery.

  19. Influence of triethyl phosphate on phosphatase activity in shooting range soil: Isolation of a zinc-resistant bacterium with an acid phosphatase.

    PubMed

    Story, Sandra; Brigmon, Robin L

    2017-03-01

    Phosphatase-mediated hydrolysis of organic phosphate may be a viable means of stabilizing heavy metals via precipitation as a metal phosphate in bioremediation applications. We investigated the effect of triethyl phosphate (TEP) on soil microbial-phosphatase activity in a heavy-metal contaminated soil. Gaseous TEP has been used at subsurface sites for bioremediation of organic contaminants but not applied in heavy-metal contaminated areas. Little is known about how TEP affects microbial activity in soils and it is postulated that TEP can serve as a phosphate source in nutrient-poor groundwater and soil/sediments. Over a 3-week period, TEP amendment to microcosms containing heavy-metal contaminated soil resulted in increased activity of soil acid-phosphatase and repression of alkaline phosphatase, indicating a stimulatory effect on the microbial population. A soil-free enrichment of microorganisms adapted to heavy-metal and acidic conditions was derived from the TEP-amended soil microcosms using TEP as the sole phosphate source and the selected microbial consortium maintained a high acid-phosphatase activity with repression of alkaline phosphatase. Addition of 5mM zinc to soil-free microcosms had little effect on acid phosphatase but inhibited alkaline phosphatase. One bacterial member from the consortium, identified as Burkholderia cepacia sp., expressed an acid-phosphatase activity uninhibited by high concentrations of zinc and produced a soluble, indigo pigment under phosphate limitation. The pigment was produced in a phosphate-free medium and was not produced in the presence of TEP or phosphate ion, indicative of purple acid-phosphatase types that are pressed by bioavailable phosphate. These results demonstrate that TEP amendment was bioavailable and increased overall phosphatase activity in both soil and soil-free microcosms supporting the possibility of positive outcomes in bioremediation applications.

  20. Roles of phosphate recognition in inositol 1,3,4,5,6-pentakisphosphate 2-kinase (IPK1) substrate binding and activation.

    PubMed

    Gosein, Varin; Miller, Gregory J

    2013-09-13

    Inositol phosphate kinases (IPKs) sequentially phosphorylate inositol phosphates (IPs) to yield a group of small signaling molecules involved in diverse cellular processes. IPK1 (inositol 1,3,4,5,6-pentakisphosphate 2-kinase) phosphorylates inositol 1,3,4,5,6-pentakisphosphate to inositol 1,2,3,4,5,6-hexakisphosphate; however, the mechanism of IP recognition employed by IPK1 is currently unresolved. We demonstrated previously that IPK1 possesses an unstable N-terminal lobe in the absence of IP, which led us to propose that the phosphate profile of the IP was linked to stabilization of IPK1. Here, we describe a systematic study to determine the roles of the 1-, 3-, 5-, and 6-phosphate groups of inositol 1,3,4,5,6-pentakisphosphate in IP binding and IPK1 activation. The 5- and 6-phosphate groups were the most important for IP binding to IPK1, and the 1- and 3-phosphate groups were more important for IPK1 activation than the others. Moreover, we demonstrate that there are three critical residues (Arg-130, Lys-170, and Lys-411) necessary for IPK1 activity. Arg-130 is the only substrate-binding N-terminal lobe residue that can render IPK1 inactive; its 1-phosphate is critical for full IPK1 activity and for stabilization of the active conformation of IPK1. Taken together, our results support the model for recognition of the IP substrate by IPK1 in which (i) the 4-, 5-, and 6-phosphates are initially recognized by the C-terminal lobe, and subsequently, (ii) the interaction between the 1-phosphate and Arg-130 stabilizes the N-terminal lobe and activates IPK1. This model of IP recognition, believed to be unique among IPKs, could be exploited for selective inhibition of IPK1 in future studies that investigate the role of higher IPs.

  1. In vitro evaluation of the antibacterial and osteogenic activity promoted by selenium-doped calcium phosphate coatings.

    PubMed

    Rodríguez-Valencia, C; Freixeiro, P; Serra, J; Ferreirós, C M; González, P; López-Álvarez, M

    2017-02-24

    Selenium is an essential trace element present in 25 selenoenzymes, playing critical roles in a variety of physiological processes, such as anti-oxidative defense and the modulation of cell proliferation and differentiation. This paper characterizes selenium-doped calcium phosphate coatings and evaluates their effects on the osteogenic activity, the proliferation of osteosarcoma cells and biofilm formation. To do so, the structure and elemental composition of the obtained coatings were analyzed, in addition to their thicknesses, and they were compared to pure calcium phosphate coatings. Moreover, the dose-effect ratio of two coatings with the lower (0.6 at%) and the higher (2.7 at%) selenium content was studied in terms of osteogenic, anti-biofilm and cancerous anti-proliferative properties. The results showed the incorporation of selenium in the form of selenite groups into the hydroxyapatite structure, with a similar crystalline pattern to the latter and increased roughness of the coatings. The calcium phosphate coatings with 2.7 at% of selenium resulted in significant osteogenic activity (p < 0.01) of healthy pre-osteoblasts (MC3T3-E1) over long periods of incubation, a significant anti-proliferative effect (p < 0.01) on cancerous osteoblasts (MG63) in a preliminary study, and anti-biofilm properties (p < 0.01) against Staphylococcus epidermidis and Staphylococcus aureus bacterial strains, which are responsible for most infections after orthopedic surgeries.

  2. Inorganic nitrite attenuates NADPH oxidase-derived superoxide generation in activated macrophages via a nitric oxide-dependent mechanism.

    PubMed

    Yang, Ting; Peleli, Maria; Zollbrecht, Christa; Giulietti, Alessia; Terrando, Niccolo; Lundberg, Jon O; Weitzberg, Eddie; Carlström, Mattias

    2015-06-01

    Oxidative stress contributes to the pathogenesis of many disorders, including diabetes and cardiovascular disease. Immune cells are major sources of superoxide (O2(∙-)) as part of the innate host defense system, but exaggerated and sustained O2(∙-) generation may lead to progressive inflammation and organ injuries. Previous studies have proven organ-protective effects of inorganic nitrite, a precursor of nitric oxide (NO), in conditions manifested by oxidative stress and inflammation. However, the mechanisms are still not clear. This study aimed at investigating the potential role of nitrite in modulating NADPH oxidase (NOX) activity in immune cells. Mice peritoneal macrophages or human monocytes were activated by lipopolysaccharide (LPS), with or without coincubation with nitrite. O2(∙-) and peroxynitrite (ONOO(-)) formation were detected by lucigenin-based chemiluminescence and fluorescence techniques, respectively. The intracellular NO production was measured by DAF-FM DA fluorescence. NOX isoforms and inducible NO synthase (iNOS) expression were detected by qPCR. LPS increased both O2(∙-) and ONOO(-) production in macrophages, which was significantly reduced by nitrite (10µmol/L). Mechanistically, the effects of nitrite are (1) linked to increased NO generation, (2) similar to that observed with the NO donor DETA-NONOate, and (3) can be abolished by the NO scavenger carboxy-PTIO or by the xanthine oxidase (XO) inhibitor febuxostat. Nox2 expression was increased in activated macrophages, but was not influenced by nitrite. However, nitrite attenuated LPS-induced upregulation of iNOS expression. Similar to that observed in mice macrophages, nitrite also reduced O2(∙-) generation in LPS-activated human monocytes. In conclusion, XO-mediated reduction of nitrite attenuates NOX activity in activated macrophages, which may modulate the inflammatory response.

  3. Apyrase Functions in Plant Phosphate Nutrition and Mobilizes Phosphate from Extracellular ATP1

    PubMed Central

    Thomas, Collin; Sun, Yu; Naus, Katie; Lloyd, Alan; Roux, Stanley

    1999-01-01

    ATP, which is present in the extracellular matrix of multicellular organisms and in the extracellular fluid of unicellular organisms, has been shown to function as a signaling molecule in animals. The concentration of extracellular ATP (xATP) is known to be functionally modulated in part by ectoapyrases, membrane-associated proteins that cleave the γ- and β-phosphates on xATP. We present data showing a previously unreported (to our knowledge) linkage between apyrase and phosphate transport. An apyrase from pea (Pisum sativum) complements a yeast (Saccharomyces cerevisiae) phosphate-transport mutant and significantly increases the amount of phosphate taken up by transgenic plants overexpressing the gene. The transgenic plants show enhanced growth and augmented phosphate transport when the additional phosphate is supplied as inorganic phosphate or as ATP. When scavenging phosphate from xATP, apyrase mobilizes the γ-phosphate without promoting the transport of the purine or the ribose. PMID:9952450

  4. Inorganic polymers from laterite using activation with phosphoric acid and alkaline sodium silicate solution: Mechanical and microstructural properties

    SciTech Connect

    Lassinantti Gualtieri, Magdalena

    2015-01-15

    Geopolymers from laterite, an iron-rich soil available in developing countries, have great potential as building materials. In this work, laterite from Togo (Africa) was used to prepare geopolymers using both phosphoric acid and alkaline sodium silicate solution. Microstructural properties were investigated by scanning electron microscopy, X-ray powder diffraction and mercury porosimetry, whereas thermal properties were evaluated by thermal analyses. The local environment of iron was studied by X-ray Absorption Spectroscopy (XANES region). The mechanical properties were determined. Modulus of Rupture and Young's modulus fell in the ranges 3.3–4.5 MPa and 12–33 GPa, respectively, rendering the materials good candidates for construction purposes. Heating above 900 °C results in weight-gain, presumably due to iron redox reactions. X-ray Absorption Spectroscopy data evidence changes in the chemical and structural environments of iron following thermal treatment of geopolymers. These changes indicate interaction between the geopolymer structure and iron during heating, possibly leading to redox properties. -- Highlights: •Geopolymerization of laterite is promising for fabrication of building materials. •Both phosphoric acid and alkaline sodium silicate solution can be used for activation. •Thermally activated redox properties of the inorganic polymers were observed.

  5. Improving spatio-temporal resolution of infrared images to detect thermal activity of defect at the surface of inorganic glass

    NASA Astrophysics Data System (ADS)

    Corvec, Guillaume; Robin, Eric; Le Cam, Jean-Benoît; Sangleboeuf, Jean-Christophe; Lucas, Pierre

    2016-07-01

    This paper proposes a noise suppression methodology to improve the spatio-temporal resolution of infrared images. The methodology is divided in two steps. The first one consists in removing the noise from the temporal signal at each pixel. Three basic temporal filters are considered for this purpose: average filter, cost function minimization (FIT) and short time Fast Fourier Transform approach (STFFT). But while this step effectively reduces the temporal signal noise at each pixel, the infrared images may still appear noisy. This is due to a random distribution of a residual offset value of pixels signal. Hence in the second step, the residual offset is identified by considering thermal images for which no mechanical loading is applied. In this case, the temperature variation field is homogeneous and the value of temperature variation at each pixel is theoretically equal to zero. The method is first tested on synthetic images built from infrared computer-generated images combined with experimental noise. The results demonstrate that this approach permits to keep the spatial resolution of infrared images equal to 1 pixel. The methodology is then applied to characterize thermal activity of a defect at the surface of inorganic glass submitted to cyclic mechanical loading. The three basic temporal filters are quantitatively compared and contrasted. Results obtained demonstrate that, contrarily to a basic spatio-temporal approach, the denoising method proposed is suitable to characterize low thermal activity combined to strong spatial gradients induced by cyclic heterogeneous deformations.

  6. Cerium-activated rare-earth orthophosphate and double-phosphate scintillators for x-and gamma-ray detection

    SciTech Connect

    Boatner, Lynn A; Keefer, Lara A; Farmer, James Matthew; Wisniewski, D.; Wojtowicz, A. J.

    2004-01-01

    When activated with an appropriate rare-earth ion (e.g., Ce or Nd), rare-earth orthophosphates of the form REPO4 (where RE = a rare-earth cation) and alkali rare-earth double phosphates of the form A{sub 3}RE(PO{sub 4}){sub 2} (where A = K, Rb, or Cs) are characterized by light yields and decay times that make these materials of interest for radiation-detection applications. Crystals of the compound Rb{sub 3}Lu(PO{sub 4}){sub 2} when activated with {approx}0.1 mol % Ce exhibit a light yield that is {approx}250% that of BGO with a decay time on the order of {approx}40 nsec. The cerium-activated rare-earth orthophosphate LuPO{sub 4}:Ce is also characterized by a high light yield and a relatively fast decay time of {approx}25 nsec. Additionally, the rare-earth orthophosphates are extremely chemically, physically, and thermally durable hosts that recover easily from radiation damage effects. The properties of the rare-earth orthophosphates and double phosphates that pertain to their use as X- and gamma-ray detectors are reviewed. This review includes information related to the use of Nd-doped LuPO{sub 4} as a scintillator with a sufficiently energetic, short-wavelength output ({lambda} = 90 nm) so that it can be used in conjunction with appropriately activated proportional counters. Information is presented on the details of the synthesis, structure, and luminescence properties of lanthanide double phosphates that, when activated with cerium, are efficient scintillators with output wavelengths that are sufficiently long to be well matched to the response of silicon photodiode detectors.

  7. Effects on operant learning and brain acetylcholine esterase activity in rats following chronic inorganic arsenic intake.

    PubMed

    Nagaraja, T N; Desiraju, T

    1994-05-01

    1. Very young and adult Wistar rats were given As5+, 5 mg arsenic kg-1 body weight day-1 (sodium arsenate). 2. Operant learning was tested in a Skinner box at the end of exposure and, in the case of developing animals, also after a recovery period. 3. Acetylcholine esterase (AChE) activity was estimated in discrete brain regions of these animals. 4. The animals exposed to arsenic took longer to acquire the learned behaviour and to extinguish the operant. AChE activity was inhibited in some regions of the brain.

  8. The zinc spark is an inorganic signature of human egg activation

    PubMed Central

    Duncan, Francesca E.; Que, Emily L.; Zhang, Nan; Feinberg, Eve C.; O’Halloran, Thomas V.; Woodruff, Teresa K.

    2016-01-01

    Egg activation refers to events required for transition of a gamete into an embryo, including establishment of the polyspermy block, completion of meiosis, entry into mitosis, selective recruitment and degradation of maternal mRNA, and pronuclear development. Here we show that zinc fluxes accompany human egg activation. We monitored calcium and zinc dynamics in individual human eggs using selective fluorophores following activation with calcium-ionomycin, ionomycin, or hPLCζ cRNA microinjection. These egg activation methods, as expected, induced rises in intracellular calcium levels and also triggered the coordinated release of zinc into the extracellular space in a prominent “zinc spark.” The ability of the gamete to mount a zinc spark response was meiotic-stage dependent. Moreover, chelation of intracellular zinc alone was sufficient to induce cell cycle resumption and transition of a meiotic cell into a mitotic one. Together, these results demonstrate critical functions for zinc dynamics and establish the zinc spark as an extracellular marker of early human development. PMID:27113677

  9. Similar healthy osteoclast and osteoblast activity on nanocrystalline hydroxyapatite and nanoparticles of tri-calcium phosphate compared to natural bone.

    PubMed

    MacMillan, Adam K; Lamberti, Francis V; Moulton, Julia N; Geilich, Benjamin M; Webster, Thomas J

    2014-01-01

    While there have been numerous studies to determine osteoblast (bone forming cell) functions on nanocrystalline compared to micron crystalline ceramics, there have been few studies which have examined osteoclast activity (including tartrate-resistant acid phosphatase, formation of resorption pits, size of resorption pits, and receptor activator of nuclear factor κB [RANK]). This is despite the fact that osteoclasts are an important part of maintaining healthy bone since they resorb bone during the bone remodeling process. Moreover, while it is now well documented that bone formation is enhanced on nanoceramics compared to micron ceramics, some have pondered whether osteoblast functions (such as osteoprotegerin and RANK ligand [RANKL]) are normal (ie, non-diseased) on such materials compared to natural bone. For these reasons, the objective of the present in vitro study was to determine various functions of osteoclasts and osteoblasts on nanocrystalline and micron crystalline hydroxyapatite as well as tri-calcium phosphate materials and compare such results to cortical and cancellous bone. Results showed for the first time similar osteoclast activity (including tartrate-resistant acid phosphatase, formation of resorption pits, size of resorption pits, and RANK) and osteoblast activity (osteoprotegerin and RANKL) on nanocrystalline hydroxyapatite compared to natural bone, whereas osteoclast and osteoblast functions on micron crystalline versions of these ceramics were much different than natural bone. In this manner, this study provides additional evidence that nanocrystalline calcium phosphates can serve as suitable synthetic analogs to natural bone to improve numerous orthopedic applications. It also provides the first data of healthy osteoclast and osteoblast functions on nanocrystalline calcium phosphates compared to natural bone.

  10. The influence of metakaolin substitution by slag in alkali-activated inorganic binders for civil engineering

    NASA Astrophysics Data System (ADS)

    Kadlec, J.; Rieger, D.; Kovářík, T.; Novotný, P.; Franče, P.; Pola, M.

    2017-02-01

    In this study the effect of metakaolin replacement by milled blast furnace slag in alkali-activated geopolymeric binder was investigated in accordance to their rheological and mechanical properties. It was demonstrated that slag addition into the metakaolin binder can improve mechanical properties of final products. Our investigation was focused on broad interval of metakaolin substitution in the range from 100 to 40 volume per cents of metakaolin so that the volume content of solids in final binder was maintained constant. Prepared binders were activated by alkaline solution of potassium silicate with silicate module of 1.61. The particle size analyses were performed for determination of particle size distribution. The rheological properties were determined in accordance to flow properties by measurements on Ford viscosity cup and by oscillatory measurements of hardening process. For the investigation of hardening process, the strain controlled small amplitude oscillatory rheometry was used in plane-plate geometry. For determination of applied mechanical properties were binders filled by ceramic grog in the granularity range 0-1 mm. The filling was maintained constant at 275 volume per cents in accordance to ratio of solids in dry binder. The mechanical properties were investigated after 1, 7 and 28 days and microstructure was documented by scanning electron microscopy. The results indicate that slag addition have beneficial effect not only on mechanical properties of hardened binder but also on flow properties of fresh geopolymer paste and subsequent hardening kinetics of alkali-activated binders.

  11. Nanocomposites of polymers with layered inorganic nanofillers: Antimicrobial activity, thermo-mechanical properties, morphology, and dispersion

    NASA Astrophysics Data System (ADS)

    Songtipya, Ponusa

    In the first part of the thesis, polyethylene/layered silicate nanocomposites that exhibit an antimicrobial activity were synthesized and studied. Their antimicrobial activity was designed to originate from non-leaching, novel cationic modifiers---amine-based surfactants---used as the organic-modification of the fillers. Specifically, PE/organically-modified montmorillonite ( mmt) nanocomposites were prepared via melt-processing, and simultaneous dispersion and antimicrobial activity was designed by proper choice of the fillers' organic modification. The antimicrobial activity was measured against three micotoxinogen fungal strains (Penicillium roqueforti and claviforme, and Fusarium graminearum ). Various mmt-based organofillers, which only differ in the type or amount of their organic modification, were used to exemplify how these surfactants can be designed to render antifungal activity to the fillers themselves and the respective nanocomposites. A comparative discussion of the growth of fungi on unfilled PE and nanocomposite PE films is used to demonstrate how the antimicrobial efficacy is dictated by the surfactant chemistry and, further, how the nanocomposites' inhibitory activity compares to that of the organo-fillers and the surfactants. An attempt to improve the thermomechanical reinforcement of PE/mmt nanocomposites while maintaining their antimicrobial activity, was also carried out by combining two different organically modified montmorillonites. However, a uniform microscopic dispersion could not be achieved through this approach. In the second part of this thesis, a number of fundamental studies relating to structure-property relations in nanocomposites were carried out, towards unveiling strategies that can concurrently optimize selected properties of polymers by the addition of nanofillers. Specifically, the dispersion-crystallinity-reinforcement relations in HDPE/mmt nanocomposites was investigated. The influence of a functional HDPE compatibilizer

  12. Modulation of hexa-acyl pyrophosphate lipid A population under Escherichia coli phosphate (Pho) regulon activation.

    PubMed

    Lamarche, Martin G; Kim, Sang-Hyun; Crépin, Sébastien; Mourez, Michael; Bertrand, Nicolas; Bishop, Russell E; Dubreuil, J Daniel; Harel, Josée

    2008-08-01

    Environmental phosphate is an important signal for microorganism gene regulation, and it has recently been shown to trigger some key bacterial virulence mechanisms. In many bacteria, the Pho regulon is the major circuit involved in adaptation to phosphate limitation. The Pho regulon is controlled jointly by the two-component regulatory system PhoR/PhoB and by the phosphate-specific transport (Pst) system, which both belong to the Pho regulon. We showed that a pst mutation results in virulence attenuation in extraintestinal pathogenic Escherichia coli (ExPEC) strains. Our results indicate that the bacterial cell surface of the pst mutants is altered. In this study, we show that pst mutants of ExPEC strains display an increased sensitivity to different cationic antimicrobial peptides and vancomycin. Remarkably, the hexa-acylated 1-pyrophosphate form of lipid A is significantly less abundant in pst mutants. Among differentially expressed genes in the pst mutant, lpxT coding for an enzyme that transfers a phosphoryl group to lipid A, forming the 1-diphosphate species, was found to be downregulated. Our results strongly suggest that the Pho regulon is involved in lipid A modifications, which could contribute to bacterial surface perturbations. Since the Pho regulon and the Pst system are conserved in many bacteria, such a lipid A modification mechanism could be widely distributed among gram-negative bacterial species.

  13. Acid inactivation of and incorporation of phosphate into alkaline phosphatase from Escherichia coli

    PubMed Central

    Pigretti, M. M.; Milstein, C.

    1965-01-01

    1. Alkaline phosphatase of Escherichia coli undergoes below pH 6·0 a reversible acid inactivation that has been studied and related to the extent of uptake of inorganic phosphate occurring below pH 6·0. 2. The rate of inactivation is rapid in the first few minutes but later it decreases markedly. Temperature, pH, composition of buffer and other factors have an important effect on the inactivation. 3. About 60% of the activity lost at pH values above 3·5 is rapidly recovered when the enzyme is taken back to pH 8·0, independently (within certain limits) of the extent of the inactivation. 4. Phosphate and Zn2+, although very good protectors of the inactivation by acid, are not by themselves able to reverse the acid inactivation. 5. Inorganic phosphate seems not to be incorporated into the acid-inactivated enzyme. 6. Incorporation of more than one mole of phosphate/mole of enzyme has been obtained, but the phosphate residues seem to be incorporated to serine residues with a common sequence, suggesting two identical active serine residues/molecule of active enzyme. ImagesFig. 3.Fig. 4. PMID:14342215

  14. Effects of inorganic nanoparticles on viability and catabolic activities of Agrobacterium sp. PH-08 during biodegradation of dibenzofuran.

    PubMed

    Le, Thao Thanh; Murugesan, Kumarasamy; Kim, Eun-Ju; Chang, Yoon-Seok

    2014-09-01

    This study investigated the cytotoxicity, genotoxicity, and growth inhibition effects of four different inorganic nanoparticles (NPs) such as aluminum (nAl), iron (nFe), nickel (nNi), and zinc (nZn) on a dibenzofuran (DF) degrading bacterium Agrobacterium sp. PH-08. NP (0-1,000 mg L(-1)) -treated bacterial cells were assessed for cytotoxicity, genotoxicity, growth and biodegradation activities at biochemical and molecular levels. In an aqueous system, the bacterial cells treated with nAl, nZn and nNi at 500 mg L(-1) showed significant reduction in cell viability (30-93.6 %, p < 0.05), while nFe had no significant inhibition on bacterial cell viability. In the presence of nAl, nZn and nNi, the cells exhibited elevated levels of reactive oxygen species (ROS), DNA damage and cell death. Furthermore, NP exposure showed significant (p < 0.05) impairment in DF and catechol biodegradation activities. The reduction in DF biodegradation was ranged about 71.7-91.6 % with single NPs treatments while reached up to 96.3 % with a mixture of NPs. Molecular and biochemical investigations also clearly revealed that NP exposure drastically affected the catechol-2,3-dioxygenase activities and its gene (c23o) expression. However, no significant inhibition was observed in nFe treatment. The bacterial extracellular polymeric materials and by-products from DF degradation can be assumed as key factors in diminishing the toxic effects of NPs, especially for nFe. This study clearly demonstrates the impact of single and mixed NPs on the microbial catabolism of xenobiotic-degrading bacteria at biochemical and molecular levels. This is the first study on estimating the impact of mixed NPs on microbial biodegradation.

  15. Templated, layered manganese phosphate

    DOEpatents

    Thoma, Steven G.; Bonhomme, Francois R.

    2004-08-17

    A new crystalline maganese phosphate composition having an empirical formula: O). The compound was determined to crystallize in the trigonal space group P-3c1 with a=8.8706(4) .ANG., c=26.1580(2) .ANG., and V (volume)=1783 .ANG..sup.3. The structure consists of sheets of corner sharing Mn(II)O.sub.4 and PO.sub.4 tetrahedra with layers of (H.sub.3 NCH.sub.2 CH.sub.2).sub.3 N and water molecules in-between. The pronated (H.sub.3 NCH.sub.2 CH.sub.2).sub.3 N molecules provide charge balancing for the inorganic sheets. A network of hydrogen bonds between water molecules and the inorganic sheets holds the structure together.

  16. Role of Acetyl-Phosphate in Activation of the Rrp2-RpoN-RpoS Pathway in Borrelia burgdorferi

    PubMed Central

    Xu, Haijun; Caimano, Melissa J.; Lin, Tao; He, Ming; Radolf, Justin D.; Norris, Steven J.; Gheradini, Frank; Wolfe, Alan J.; Yang, X. Frank

    2010-01-01

    Borrelia burgdorferi, the Lyme disease spirochete, dramatically alters its transcriptome and proteome as it cycles between the arthropod vector and mammalian host. During this enzootic cycle, a novel regulatory network, the Rrp2-RpoN-RpoS pathway (also known as the σ54–σS sigma factor cascade), plays a central role in modulating the differential expression of more than 10% of all B. burgdorferi genes, including the major virulence genes ospA and ospC. However, the mechanism(s) by which the upstream activator and response regulator Rrp2 is activated remains unclear. Here, we show that none of the histidine kinases present in the B. burgdorferi genome are required for the activation of Rrp2. Instead, we present biochemical and genetic evidence that supports the hypothesis that activation of the Rrp2-RpoN-RpoS pathway occurs via the small, high-energy, phosphoryl-donor acetyl phosphate (acetyl∼P), the intermediate of the Ack-Pta (acetate kinase-phosphate acetyltransferase) pathway that converts acetate to acetyl-CoA. Supplementation of the growth medium with acetate induced activation of the Rrp2-RpoN-RpoS pathway in a dose-dependent manner. Conversely, the overexpression of Pta virtually abolished acetate-induced activation of this pathway, suggesting that acetate works through acetyl∼P. Overexpression of Pta also greatly inhibited temperature and cell density-induced activation of RpoS and OspC, suggesting that these environmental cues affect the Rrp2-RpoN-RpoS pathway by influencing acetyl∼P. Finally, overexpression of Pta partially reduced infectivity of B. burgdorferi in mice. Taken together, these findings suggest that acetyl∼P is one of the key activating molecule for the activation of the Rrp2-RpoN-RpoS pathway and support the emerging concept that acetyl∼P can serve as a global signal in bacterial pathogenesis. PMID:20862323

  17. Antimicrobial effects of silver zeolite, silver zirconium phosphate silicate and silver zirconium phosphate against oral microorganisms

    PubMed Central

    Saengmee-anupharb, Sirikamon; Srikhirin, Toemsak; Thaweboon, Boonyanit; Thaweboon, Sroisiri; Amornsakchai, Taweechai; Dechkunakorn, Surachai; Suddhasthira, Theeralaksna; Kamaguchi, Arihide

    2013-01-01

    Objective To evaluate the antimicrobial activities of silver inorganic materials, including silver zeolite (AgZ), silver zirconium phosphate silicate (AgZrPSi) and silver zirconium phosphate (AgZrP), against oral microorganisms. In line with this objective, the morphology and structure of each type of silver based powders were also investigated. Methods The antimicrobial activities of AgZ, AgZrPSi and AgZrP were tested against Streptococcus mutans, Lactobacillus casei, Candida albicans and Staphylococcus aureus using disk diffusion assay as a screening test. The minimum inhibitory concentration (MIC) and minimum lethal concentration (MLC) were determined using the modified membrane method. Scanning electron microscope and X-ray diffraction were used to investigate the morphology and structure of these silver materials. Results All forms of silver inorganic materials could inhibit the growth of all test microorganisms. The MIC of AgZ, AgZrPSi and AgZrP was 10.0 g/L whereas MLC ranged between 10.0–60.0 g/L. In terms of morphology and structure, AgZrPSi and AgZrP had smaller sized particles (1.5–3.0 µm) and more uniformly shaped than AgZ. Conclusions Silver inorganic materials in the form of AgZ, AgZrPSi and AgZrP had antimicrobial effects against all test oral microorganisms and those activities may be influenced by the crystal structure of carriers. These results suggest that these silver materials may be useful metals applied to oral hygiene products to provide antimicrobial activity against oral infection. PMID:23570016

  18. Evolutionary conservation of a phosphate transporter in the arbuscular mycorrhizal symbiosis

    PubMed Central

    Karandashov, Vladimir; Nagy, Réka; Wegmüller, Sarah; Amrhein, Nikolaus; Bucher, Marcel

    2004-01-01

    Arbuscular mycorrhizae are ancient symbioses that are thought to have originated >400 million years ago in the roots of plants, pioneering the colonization of terrestrial habitats. In these associations, a key process is the transfer of phosphorus as inorganic phosphate to the host plant across the fungus–plant interface. Mycorrhiza-specific phosphate transporter genes and their regulation are conserved in phylogenetically distant plant species, and they are activated selectively by fungal species from the phylum Glomeromycota. The potato phosphate transporter gene StPT3 is expressed in a temporally defined manner in root cells harboring various mycorrhizal structures, including thick-coiled hyphae. The results highlight the role of different symbiotic structures in phosphorus transfer, and they indicate that cell–cell contact between the symbiotic partners is required to induce phosphate transport. PMID:15075387

  19. Assessment of salivary calcium, phosphate, magnesium, pH, and flow rate in healthy subjects, periodontitis, and dental caries

    PubMed Central

    Rajesh, K. S.; Zareena; Hegde, Shashikanth; Arun Kumar, M. S.

    2015-01-01

    Aim: This study was conducted to estimate and compare inorganic salivary calcium, phosphate, magnesium, salivary flow rate, and pH of unstimulated saliva and oral hygiene status of healthy subjects, subjects with periodontitis and dental caries, and to correlate salivary calcium level with number of intact teeth. Materials and Methods: The study population consisted of 48 systemically healthy subjects in the age group of 18-55 years, which was further divided into three groups: healthy, periodontitis, and dental caries. Oral hygiene index-simplified, probing pocket depth, clinical attachment level, the number of intact teeth, and active carious lesions were recorded. Estimation of inorganic salivary calcium, phosphate, and magnesium was performed spectrophotometrically using Vitros 5.1 FS. Statistical analysis was performed using the one-way analysis of variance test at 5% significance level. Results: There was a statistically significant increase in inorganic salivary calcium, phosphate, pH, flow rate, and poor oral hygiene status in periodontitis group compared to dental caries and healthy group. Conclusion: Subjects with increased inorganic salivary calcium, phosphate, pH, flow rate, and poor oral hygiene are at a higher risk of developing periodontitis. Since there is increased remineralization potential, these subjects have more number of intact teeth compared to the dental caries group. PMID:26681848

  20. Toxicological effects of selective herbicides on plant growth promoting activities of phosphate solubilizing Klebsiella sp. strain PS19.

    PubMed

    Ahemad, Munees; Saghir Khan, Md

    2011-02-01

    This study examines the effect of four herbicides, quizalafop-p-ethyl, clodinafop, metribuzin and glyphosate, on plant growth promoting activities like phosphate solubilization, siderophores, indole acetic acid, exo-polysaccharides, hydrogen cyanide and ammonia production by herbicide tolerant Klebsiella sp. strain PS19. The strain was isolated from mustard rhizosphere. The selected herbicides were applied two to three times at the recommended rates. Klebsiella sp. strain PS19 tolerated a concentration of 1600 μg/ml each of quizalafop-p-ethyl and clodinafop, and 3200 and 2800 μg/ml of metribuzin and glyphosate, respectively. The activities of Klebsiella sp. strain PS19 observed under in vitro environment were persistent in the presence of all herbicides at lower rates. The plant growth promoting activities even-though decreased regularly, but was not lost completely, as the concentration of each herbicide was increased from the recommended to three times of higher doses. Among all herbicides, quizalafop-p-ethyl, generally, showed maximum toxicity to plant growth promoting activities of Klebsiella sp. strain PS19. As an example, 40, 80 and 120 μg/l of quizalafop-p-ethyl added to liquid culture Pikovskaya medium, decreased phosphate solubilizing activity of strain PS19 by 93, 95 and 97%, respectively over untreated control. The study revealed that the higher rates of herbicides though decreased the plant growth promoting activity but it did not completely inhibit the metabolic activities of strain PS19. The herbicide tolerance together with growth promoting activities observed under herbicide stress suggests that Klebsiella sp. strain PS19 could be used as bacterial preparation for facilitating the growth and yields of crops even in soils polluted with herbicides.

  1. [Determination of content and specific activity of radioactive elements in phosphate slag by spectral and energy spectrum analysis].

    PubMed

    Bao, P Y

    2001-10-01

    Phosphate slag is the slag discarded after phosphate ore is smelted. The content and specific activity of radioactive elements in slag must be determined accurately for environmental protection and comprehensive utilization. This paper discusses how IR spectrum and X-ray diffraction method are used to study its components. The main phase composition is glassy slag. The samples are decomposed with HF-HNO3-HClO4. After anion-exchange separation, arsenazo III is used to determine the content of uranium and thorium in slag. The average content of U is 32.11 micrograms.g-1 and 8.5 micrograms.g-1 for Th. gamma spectrum is used to determine the specific activity of radioactive nuclear elements in it. The specific activities of 226Ra, 232Th and 40K are 112 +/- 2.0% Bq.kg-1, 18.8 +/- 6.4% Bq.kg-1 and 77.6 +/- 3.7% Bq.kg-1, respectively. The values are calculated as follow: M(Ra) = 0.6 Bq.kg-1, M gamma = 0.5 Bq.kg-1. The results show that the values are below 1.0 Bq.kg-1, which is stipulated by national GB6566-86 standard for radioactivity of building materials. The slag is therefore can be utilized to produce slag cement. This provides theoretical basis for the treatment of it.

  2. Isolation, identification, and synthesis of 2-carboxyarabinitol 1-phosphate, a diurnal regulator of ribulase-bisphosphate carboxylase activity

    SciTech Connect

    Berry, J.A.; Lorimer, G.H.; Pierce, J.; Seemann, J.R.; Meek, J.; Freas, S.

    1987-02-01

    The diurnal change in activity of ribulose 1,5-bisphosphate (Rbu-1,5-P/sub 2/) carboxylase (3-phospho-D-glycerate carboxy-lyase (dimerizing); EC 4.1.1.39) of leaves of Phaseolus vulgaris is regulated (in part) by mechanisms that control the level of an endogenous inhibitor that binds tightly to the activated (carbamoylated) form of Rbu-1,5-P/sub 2/ carboxylase. This inhibitor was extracted from leaves and copurified with the Rbu-1,5-P/sub 2/ carboxylase of the leaves. Further purification by ion-exchange chromatography, adsorption to purified Rbu-1,5-P/sub 2/ carboxylase, barium precipitation, and HPLC separation yielded a phosphorylated compound that was a strong inhibitor of Rbu-1,5-P/sub 2/ carboxylase. The compound was analyzed by GC/MS, /sup 13/C NMR, and /sup 1/H NMR and shown to be 2-carboxyarabinitol 1-phosphate ((2-C-phosphohydroxymethyl)-D-ribonic acid). The structure of the isolated compound differs from the Rbu-1,5-P/sub 2/ carboxylase transition-state analogue 2-carboxyarabinitol 1,5-bisphosphate only by the lack of the C-5 phosphate group. This difference results in a higher binding constant for the monophosphate compared with the bisphosphate. The less tightly bound compound acts in a light-dependent, reversible regulation of Rbu-1,5-P/sub 2/ carboxylase activity in vivo.

  3. Removal of organic and inorganic sulfur compounds by ozone and granular activated carbon

    SciTech Connect

    Shepherd, B.; Ball, G.W.

    1996-11-01

    Most groundwater supplies in the western U.S. are relatively low in dissolved organic matter, are generally free of bacteria, and are platable to their consumers. In areas of western Nevada, certain groundwaters are near active geothermal areas, which can produce sulfurous types of tastes and odors (T&Os) in the water. Other water quality characteristics can consist of either relatively low or highly mineralized waters, variations in pH, and temperatures ranging from those slightly above normal groundwaters to pressurized steam. Watersource Consulting Engineers (WCE) and Shepherd Laboratories (SL) conducted an engineering study of a high-capacity well for a local northwestern Nevada utility. WCE`s original task had been to design pumping and storage facilities for the well, in addition to evaluating basic treatment. Originally, WCE anticipated designing facilities to remove hydrogen sulfide (H{sub 2}S) and reduce color, primarily with chlorination and aeration. SL was requested to evaluate existing water quality and eventually conduct bench-scale testing of several treatment processes. As the study proceeded, the original goals were modified when it became evident that water quality conditions required more extensive evaluation. The study was done in several stages, reflecting the information gained during each stage. The final recommended design criteria included treatment for improving water quality relative to T&O, color, total organic carbon (TOC), and, to a limited extent, fluoride. The water quality goals adopted by the utility encompassed primary maximum contaminant levels (MCLs) for regulatory compliance and secondary MCLs for aesthetically pleasing water. The treatment processes evaluated and recommended in this study were designed primarily to improve the aesthetic qualities of color, taste, and odor. Fluoride reduction was evaluated but was not included in the final design requirements, except for the overall reduction provided by the recommended process.

  4. Coarse and Fine Control and Annual Changes of Sucrose-Phosphate Synthase in Norway Spruce Needles.

    PubMed Central

    Loewe, A.; Einig, W.; Hampp, R.

    1996-01-01

    Annual changes of activity of sucrose-phosphate synthase (SPS) from spruce (Picea abies [L.] Karst.) needles were studied with respect to three regulatory levels: metabolic fine control, covalent modification (phosphorylation), and protein amount. Glucose-6-phosphate served as an allosteric activator of spruce SPS by shifting the Michaelis constant for the substrate fructose-6-phosphate from 4.2 to 0.59 mM, whereas inorganic phosphate competitively inhibited this activation. The affinity for the other substrate, UDP-glucose, was unaffected. Incubation of the crude extract with ATP resulted in a time- and concentration-dependent decrease of the maximal velocity of SPS. This inactivation was sensitive to staurosporine, a potent protein kinase inhibitor, indicating the participation of a protein kinase. Probing SPS protein with heterologous antibodies showed that the subunit of spruce SPS is an approximately 139-kD protein and that changes in the extractable activity during the course of a year were correlated with the amount of SPS protein. High SPS activities in winter were paralleled by increased levels of the activator glucose-6-phosphate and the substrate fructose-6-phosphate, indicating a high capacity for sucrose synthesis that may be necessary to maintain photosynthetic CO2 fixation in cold-hardened spruce needles. PMID:12226418

  5. Expression of FGF23 is correlated with serum phosphate level in isolated fibrous dysplasia.

    PubMed

    Kobayashi, Keisuke; Imanishi, Yasuo; Koshiyama, Hiroyuki; Miyauchi, Akimitsu; Wakasa, Kenichi; Kawata, Takehisa; Goto, Hitoshi; Ohashi, Hirotsugu; Koyano, Hajime M; Mochizuki, Ryuichi; Miki, Takami; Inaba, Masaaki; Nishizawa, Yoshiki

    2006-04-11

    Fibrous dysplasia (FD) patients sometimes suffer from concomitant hypophosphatemic rickets/osteomalacia, resulting from renal phosphate wasting. It was recently reported that FD tissue in the patients with McCune-Albright syndrome (MAS) expressed fibroblast growth factor-23 (FGF-23), which is now known to be as a pathogenic phosphaturic factor in patients with oncogenic osteomalacia and X-linked hypophosphatemic rickets. Since it remains controversial whether serum phosphate levels are influenced by FGF23 expressions in FD tissue, isolated FD patients without MAS syndrome were examined for the relationship between FGF23 expressions, circulating levels of FGF-23 and phosphate to negate the effects of MAS-associated endocrine abnormalities on serum phosphate. Eighteen paraffin embedded FD tissues and 2 frozen tissues were obtained for the study. Sixteen of 18 isolated FD tissues were successfully analyzed GNAS gene, which exhibited activated mutations observed in MAS. Eight of 16 FD tissues, which exhibited GNAS mutations, revealed positive staining for FGF-23. These evidence indicate that postzygotic activated mutations of GNAS is necessary for the FD tissue formation by mosaic distribution of mutated osteogenic cell lineage, but is not sufficient to elevate FGF23 expression causing generalized osteomalacia with severe renal phosphate wasting. The expression level of FGF23 in isolated FD tissue with hypophosphatemic osteomalacia determined by real-time PCR was abundant close to the levels in OOM tumors. Osteoblasts/osteocytes in woven bone were predominant source of circulating FGF-23 in FD tissues by immunohistochemistry. A negative correlation of the intensity of FGF-23 staining with serum inorganic phosphate levels indicated that the expression of FGF23 in focal FD tissues could be a prominent determinant of serum phosphate levels in isolated FD patient. These data provide novel insights into the regulatory mechanism of serum inorganic phosphate levels in

  6. Ceramide 1-Phosphate Increases P-Glycoprotein Transport Activity at the Blood-Brain Barrier via Prostaglandin E2 Signaling.

    PubMed

    Mesev, Emily V; Miller, David S; Cannon, Ronald E

    2017-04-01

    P-glycoprotein, an ATP-driven efflux pump, regulates permeability of the blood-brain barrier (BBB). Sphingolipids, endogenous to brain tissue, influence inflammatory responses and cell survival in vitro. Our laboratory has previously shown that sphingolipid signaling by sphingosine 1-phosphate decreases basal P-glycoprotein transport activity. Here, we investigated the potential for another sphingolipid, ceramide 1-phosphate (C1P), to modulate efflux pumps at the BBB. Using confocal microscopy and measuring luminal accumulation of fluorescent substrates, we assessed the transport activity of several efflux pumps in isolated rat brain capillaries. C1P treatment induced P-glycoprotein transport activity in brain capillaries rapidly and reversibly. In contrast, C1P did not affect transport activity of two other major efflux transporters, multidrug resistance protein 2 and breast cancer resistance protein. C1P induced P-glycoprotein transport activity without changing transporter protein expression. Inhibition of the key signaling components in the cyclooxygenase-2 (COX-2)/prostaglandin E2 signaling cascade (phospholipase A2, COX-2, multidrug resistance protein 4, and G-protein-coupled prostaglandin E2 receptors 1 and 2), abolished P-glycoprotein induction by C1P. We show that COX-2 and prostaglandin E2 are required for C1P-mediated increases in P-glycoprotein activity independent of transporter protein expression. This work describes how C1P activates a signaling cascade to dynamically regulate P-glycoprotein transport at the BBB and offers potential clinical targets to modulate neuroprotection and drug delivery to the CNS.

  7. Ceramide 1-Phosphate Increases P-Glycoprotein Transport Activity at the Blood-Brain Barrier via Prostaglandin E2 Signaling

    PubMed Central

    Mesev, Emily V.; Miller, David S.

    2017-01-01

    P-glycoprotein, an ATP-driven efflux pump, regulates permeability of the blood-brain barrier (BBB). Sphingolipids, endogenous to brain tissue, influence inflammatory responses and cell survival in vitro. Our laboratory has previously shown that sphingolipid signaling by sphingosine 1-phosphate decreases basal P-glycoprotein transport activity. Here, we investigated the potential for another sphingolipid, ceramide 1-phosphate (C1P), to modulate efflux pumps at the BBB. Using confocal microscopy and measuring luminal accumulation of fluorescent substrates, we assessed the transport activity of several efflux pumps in isolated rat brain capillaries. C1P treatment induced P-glycoprotein transport activity in brain capillaries rapidly and reversibly. In contrast, C1P did not affect transport activity of two other major efflux transporters, multidrug resistance protein 2 and breast cancer resistance protein. C1P induced P-glycoprotein transport activity without changing transporter protein expression. Inhibition of the key signaling components in the cyclooxygenase-2 (COX-2)/prostaglandin E2 signaling cascade (phospholipase A2, COX-2, multidrug resistance protein 4, and G-protein–coupled prostaglandin E2 receptors 1 and 2), abolished P-glycoprotein induction by C1P. We show that COX-2 and prostaglandin E2 are required for C1P-mediated increases in P-glycoprotein activity independent of transporter protein expression. This work describes how C1P activates a signaling cascade to dynamically regulate P-glycoprotein transport at the BBB and offers potential clinical targets to modulate neuroprotection and drug delivery to the CNS. PMID:28119480

  8. Enriched inorganic compounds in diesel exhaust particles induce mitogen-activated protein kinase activation, cytoskeleton instability, and cytotoxicity in human bronchial epithelial cells.

    PubMed

    Seriani, Robson; Junqueira, Mara S; Carvalho-Sousa, Claudia E; Arruda, Alessandra C T; Martinez, Diana; Alencar, Adriano M; Garippo, Ana L; Brito, Jôse Mara; Martins, Milton A; Saldiva, Paulo H N; Negri, Elnara M; Mauad, Thais; Macchione, Mariangela

    2015-04-01

    This study assessed the effects of the diesel exhaust particles on ERK and JNK MAPKs activation, cell rheology (viscoelasticity), and cytotoxicity in bronchial epithelial airway cells (BEAS-2B). Crude DEP and DEP after extraction with hexane (DEP/HEX) were utilized. The partial reduction of some DEP/HEX organics increased the biodisponibility of many metallic elements. JNK and ERK were activated simultaneously by crude DEP with no alterations in viscoelasticity of the cells. Mitochondrial activity, however, revealed a decrease through the MTT assay. DEP/HEX treatment increased viscoelasticity and cytotoxicity (membrane damage), and also activated JNK. Our data suggest that the greater bioavailability of metals could be involved in JNK activation and, consequently, in the reduction of fiber coherence and increase in the viscoelasticity and cytotoxicity of BEAS cells. The adverse findings detected after exposure to crude DEP and to DEP/HEX reflect the toxic potential of diesel compounds. Considering the fact that the cells of the respiratory epithelium are the first line of defense between the body and the environment, our data contribute to a better understanding of the pathways leading to respiratory cell injury and provide evidence for the onset of or worsening of respiratory diseases caused by inorganic compounds present in DEP.

  9. Artefactual nanoparticle activation of the inflammasome platform: in vitro evidence with a nano-formed calcium phosphate

    PubMed Central

    Pele, Laetitia; Haas, Carolin T; Hewitt, Rachel; Faria, Nuno; Brown, Andy; Powell, Jonathan

    2015-01-01

    Aim To determine whether in vitro experimental conditions dictate cellular activation of the inflammasome by apatitic calcium phosphate nanoparticles. Material & methods The responses of blood-derived primary human cells to in situ-formed apatite were investigated under different experimental conditions to assess the effect of aseptic culture, cell rest and duration of particle exposure. Cell death and particle uptake were assessed, while IL-1β and caspase 1 responses, with and without lipopolysaccharide prestimulation, were evaluated as markers of inflammasome activation. Results Under carefully addressed experimental conditions, apatitic nanoparticles did not induce cell death or engage the inflammasome platform, although both could be triggered through artefacts of experimentation. Conclusion In vitro studies often predict that engineered nanoparticles, such as synthetic apatite, are candidates for inflammasome activation and, hence, are toxic. However, the experimental setting must be very carefully considered as it may promote false-positive outcomes. PMID:24991724

  10. Activity of tedizolid phosphate (TR-701) in murine models of infection with penicillin-resistant and penicillin-sensitive Streptococcus pneumoniae.

    PubMed

    Choi, Sunghak; Im, Weonbin; Bartizal, Ken

    2012-09-01

    The in vitro activity of tedizolid (previously known as torezolid, TR-700) against penicillin-resistant Streptococcus pneumoniae (PRSP) clinical isolates and the in vivo efficacy of tedizolid phosphate (torezolid phosphate, TR-701) in murine models of PRSP systemic infection and penicillin-susceptible S. pneumoniae (PSSP) pneumonia were examined using linezolid as a comparator. The MIC(90) against 28 PRSP isolates was 0.25 μg/ml for tedizolid, whereas it was 1 μg/ml for linezolid. In mice infected systemically with a lethal inoculum of PRSP 1 h prior to a single administration of either antimicrobial, oral tedizolid phosphate was equipotent to linezolid (1 isolate) to 2-fold more potent than linezolid (3 isolates) for survival at day 7, with tedizolid phosphate 50% effective dose (ED(50)) values ranging from 3.19 to 11.53 mg/kg of body weight/day. In the PSSP pneumonia model, the ED(50) for survival at day 15 was 2.80 mg/kg/day for oral tedizolid phosphate, whereas it was 8.09 mg/kg/day for oral linezolid following 48 h of treatment with either agent. At equivalent doses (10 mg/kg once daily tedizolid phosphate or 5 mg/kg twice daily linezolid), pneumococcal titers in the lungs at 52 h postinfection were approximately 3 orders of magnitude lower with tedizolid phosphate treatment than with linezolid treatment or no treatment. Lung histopathology showed less inflammatory cell invasion into alveolar spaces in mice treated with tedizolid phosphate than in untreated or linezolid-treated mice. These results demonstrate that tedizolid phosphate is effective in murine models of PRSP systemic infection and PSSP pneumonia.

  11. Inositol phosphates induce DAPI fluorescence shift.

    PubMed

    Kolozsvari, Bernadett; Parisi, Federica; Saiardi, Adolfo

    2014-06-15

    The polymer inorganic polyP (polyphosphate) and inositol phosphates, such as IP6 (inositol hexakisphosphate; also known as phytic acid), share many biophysical features. These similarities must be attributed to the phosphate groups present in these molecules. Given the ability of polyP to modify the excitation-emission spectra of DAPI we decided to investigate whether inositol phosphates possess the same property. We discovered that DAPI-IP6 complexes emit at approximately 550 nm when excited with light of wavelength 410-420 nm. IP5 (inositol pentakisphosphate) is also able to induce a similar shift in DAPI fluorescence. Conversely, IP3 (inositol trisphosphate) and IP4 (inositol tetrakisphosphate) are unable to shift DAPI fluorescence. We have employed this newly discovered feature of DAPI to study the enzymatic activity of the inositol polyphosphate multikinase and to monitor phytase phosphatase reactions. Finally, we used DAPI-IP6 fluorescence to determine the amount of IP6 in plant seeds. Using an IP6 standard curve this straight-forward analysis revealed that among the samples tested, borlotti beans possess the highest level of IP6 (9.4 mg/g of dry mass), whereas the Indian urad bean the lowest (3.2 mg/g of dry mass). The newly identified fluorescence properties of the DAPI-IP5 and DAPI-IP6 complexes allow the levels and enzymatic conversion of these two important messengers to be rapidly and reliably monitored.

  12. Conditional iron and pH-dependent activity of a non-enzymatic glycolysis and pentose phosphate pathway.

    PubMed

    Keller, Markus A; Zylstra, Andre; Castro, Cecilia; Turchyn, Alexandra V; Griffin, Julian L; Ralser, Markus

    2016-01-01

    Little is known about the evolutionary origins of metabolism. However, key biochemical reactions of glycolysis and the pentose phosphate pathway (PPP), ancient metabolic pathways central to the metabolic network, have non-enzymatic pendants that occur in a prebiotically plausible reaction milieu reconstituted to contain Archean sediment metal components. These non-enzymatic reactions could have given rise to the origin of glycolysis and the PPP during early evolution. Using nuclear magnetic resonance spectroscopy and high-content metabolomics that allowed us to measure several thousand reaction mixtures, we experimentally address the chemical logic of a metabolism-like network constituted from these non-enzymatic reactions. Fe(II), the dominant transition metal component of Archean oceanic sediments, has binding affinity toward metabolic sugar phosphates and drives metabolism-like reactivity acting as both catalyst and cosubstrate. Iron and pH dependencies determine a metabolism-like network topology and comediate reaction rates over several orders of magnitude so that the network adopts conditional activity. Alkaline pH triggered the activity of the non-enzymatic PPP pendant, whereas gentle acidic or neutral conditions favored non-enzymatic glycolytic reactions. Fe(II)-sensitive glycolytic and PPP-like reactions thus form a chemical network mimicking structural features of extant carbon metabolism, including topology, pH dependency, and conditional reactivity. Chemical networks that obtain structure and catalysis on the basis of transition metals found in Archean sediments are hence plausible direct precursors of cellular metabolic networks.

  13. Zinc-phosphate nanoparticles with reversibly attached TNF-α analogs: an interesting concept for potential use in active immunotherapy

    NASA Astrophysics Data System (ADS)

    Hribar, Gorazd; Žnidaršič, Andrej; Bele, Marjan; Maver, Uroš; Caserman, Simon; Gaberšček, Miran; Gaberc-Porekar, Vladka

    2011-07-01

    The authors' intention was to prepare nanometer-sized zinc-phosphate nanoparticles that would be capable of binding histidine-rich TNF-α analogs onto their surface via a coordinative bond. Zinc-phosphate nanoparticles with a size of around 60 nm were prepared by a wet precipitation method and characterized using SEM, EDX, XRD, and DLS. First, BSA was bound as a testing protein, afterward two TNF-α analogs with decreased activity were bound to the described nanoparticles. The efficiency of binding and the existence of coordinative bond were confirmed with SDS-PAGE analysis. During binding, particle storage, and release experiments, the prepared TNF-α analogs retained their biological activity—hence the epitopes necessary for formation of antibodies stayed intact. The particle size did not change within a period of 2 weeks. No significant agglomeration was observed, the particles could be quickly dispersed in ultrasound. The present nanoparticles and the general approach of coordinative binding are widely applicable for natural and engineered histidine-rich proteins. The nanoparticles bearing appropriate TNF-α analogs could also be potentially used for active immunotherapy to tackle the chronic inflammatory diseases associated with pathogenically elevated levels of TNF-α.

  14. Conditional iron and pH-dependent activity of a non-enzymatic glycolysis and pentose phosphate pathway

    PubMed Central

    Keller, Markus A.; Zylstra, Andre; Castro, Cecilia; Turchyn, Alexandra V.; Griffin, Julian L.; Ralser, Markus

    2016-01-01

    Little is known about the evolutionary origins of metabolism. However, key biochemical reactions of glycolysis and the pentose phosphate pathway (PPP), ancient metabolic pathways central to the metabolic network, have non-enzymatic pendants that occur in a prebiotically plausible reaction milieu reconstituted to contain Archean sediment metal components. These non-enzymatic reactions could have given rise to the origin of glycolysis and the PPP during early evolution. Using nuclear magnetic resonance spectroscopy and high-content metabolomics that allowed us to measure several thousand reaction mixtures, we experimentally address the chemical logic of a metabolism-like network constituted from these non-enzymatic reactions. Fe(II), the dominant transition metal component of Archean oceanic sediments, has binding affinity toward metabolic sugar phosphates and drives metabolism-like reactivity acting as both catalyst and cosubstrate. Iron and pH dependencies determine a metabolism-like network topology and comediate reaction rates over several orders of magnitude so that the network adopts conditional activity. Alkaline pH triggered the activity of the non-enzymatic PPP pendant, whereas gentle acidic or neutral conditions favored non-enzymatic glycolytic reactions. Fe(II)-sensitive glycolytic and PPP-like reactions thus form a chemical network mimicking structural features of extant carbon metabolism, including topology, pH dependency, and conditional reactivity. Chemical networks that obtain structure and catalysis on the basis of transition metals found in Archean sediments are hence plausible direct precursors of cellular metabolic networks. PMID:26824074

  15. Periplasmic protein related to the sn-glycerol-3-phosphate transport system of Escherichia coli.

    PubMed Central

    Silhavy, T J; Hartig-Beecken, I; Boos, W

    1976-01-01

    Two-dimensional gel electrophoresis of shock fluids of Escherichia coli K-12 revealed the presence of a periplasmic protein related to sn-glycerol-3-phosphate transport (GLPT) that is under the regulation of glpR, the regulatory gene of the glp regulon. Mutants selected for their resistance to phosphonomycin and found to be defective in sn-glycerol-3-phosphate transport either did not produce GLPT or produced it in reduced amounts. Other mutations exhibited no apparent effect of GLPT. Transductions of glpT+ nalA phage P1 into these mutants and selection for growth on sn-glycerol-3-phosphate revealed a 50% cotransduction frequency to nalA. Reversion of mutants taht did not produce GLPT to growth on sn-glycerol-3-phosphate resulted in strains that produce GLPT. This suggests a close relationship of GLPT to the glpT gene and to sn-glycerol-3-phosphate transport. Attempts to demonstrate binding activity of GLPT in crude shock fluid towards sn-glycerol-3-phosphate have failed so far. However, all shock fluids, independent of their GLPT content, exhibited an enzymatic activity that hydrolyzes under the conditions of the binding assay, 30 to 60% of the sn-glycerol-3-phosphate to glycerol and inorganic orthophosphate. Images PMID:770459

  16. A plant proton-pumping inorganic pyrophosphatase functionally complements the vacuolar ATPase transport activity and confers bafilomycin resistance in yeast.

    PubMed

    Pérez-Castiñeira, José R; Hernández, Agustín; Drake, Rocío; Serrano, Aurelio

    2011-07-15

    V-ATPases (vacuolar H+-ATPases) are a specific class of multi-subunit pumps that play an essential role in the generation of proton gradients across eukaryotic endomembranes. Another simpler proton pump that co-localizes with the V-ATPase occurs in plants and many protists: the single-subunit H+-PPase [H+-translocating PPase (inorganic pyrophosphatase)]. Little is known about the relative contribution of these two proteins to the acidification of intracellular compartments. In the present study, we show that the expression of a chimaeric derivative of the Arabidopsis thaliana H+-PPase AVP1, which is preferentially targeted to internal membranes of yeast, alleviates the phenotypes associated with V-ATPase deficiency. Phenotypic complementation was achieved both with a yeast strain with its V-ATPase specifically inhibited by bafilomycin A1 and with a vma1-null mutant lacking a catalytic V-ATPase subunit. Cell staining with vital fluorescent dyes showed that AVP1 recovered vacuole acidification and normalized the endocytic pathway of the vma mutant. Biochemical and immunochemical studies further demonstrated that a significant fraction of heterologous H+-PPase is located at the vacuolar membrane. These results raise the question of the occurrence of distinct proton pumps in certain single-membrane organelles, such as plant vacuoles, by proving yeast V-ATPase activity dispensability and the capability of H+-PPase to generate, by itself, physiologically suitable internal pH gradients. Also, they suggest new ways of engineering macrolide drug tolerance and outline an experimental system for testing alternative roles for fungal and animal V-ATPases, other than the mere acidification of subcellular organelles.

  17. Characterization of the human and mouse sphingosine 1-phosphate receptor, S1P5 (Edg-8): structure-activity relationship of sphingosine1-phosphate receptors.

    PubMed

    Im, D S; Clemens, J; Macdonald, T L; Lynch, K R

    2001-11-20

    Five G protein-coupled receptors (S1P(1)/Edg-1, S1P(3)/Edg-3, S1P(2)/Edg-5, S1P(4)/Edg-6, and S1P(5)/Edg-8) for the intercellular lipid mediator sphingosine 1-phosphate have been cloned and characterized. We found human and mouse sequences closely related to rat S1P(5) (97% identical amino acids) and report now the characterization of the human and mouse S1P(5) gene products as encoding sphingosine 1-phosphate receptors. When HEK293T cells were cotransfected with S1P(5) and G protein DNAs, prepared membranes showed sphingosine 1-phosphate concentration-dependent increases in [gamma-(35)S]GTP binding (EC(50) = 12.7 nM). The lipid mediator inhibited forskolin-driven rises in cAMP by greater than 80% after introduction of the mouse or human S1P(5) DNAs into rat hepatoma RH7777 cells (IC(50) = 0.22 nM). This response is blocked fully by prior treatment of cultures with pertussis toxin, thus implicating signaling through G(i/o)alpha proteins. Northern blot analysis showed high expression of human S1P(5) mRNA in spleen, corpus collosum, peripheral blood leukocytes, placenta, lung, aorta, and fetal tissues. Mouse S1P(5) mRNA is also expressed in spleen and brain. Finally, we found that one enantiomer of a sphingosine 1-phosphate analogue wherein the 3-hydroxyl and 4,5-olefin are replaced by an amide functionality shows some selectivity as an agonist S1P(1) and S1P(3) vs S1P(2) and S1P(5).

  18. Sphingosine-1-phosphate enhances satellite cell activation in dystrophic muscles through a S1PR2/STAT3 signaling pathway.

    PubMed

    Loh, Kenneth C; Leong, Weng-In; Carlson, Morgan E; Oskouian, Babak; Kumar, Ashok; Fyrst, Henrik; Zhang, Meng; Proia, Richard L; Hoffman, Eric P; Saba, Julie D

    2012-01-01

    Sphingosine-1-phosphate (S1P) activates a widely expressed family of G protein-coupled receptors, serves as a muscle trophic factor and activates muscle stem cells called satellite cells (SCs) through unknown mechanisms. Here we show that muscle injury induces dynamic changes in S1P signaling and metabolism in vivo. These changes include early and profound induction of the gene encoding the S1P biosynthetic enzyme SphK1, followed by induction of the catabolic enzyme sphingosine phosphate lyase (SPL) 3 days later. These changes correlate with a transient increase in circulating S1P levels after muscle injury. We show a specific requirement for SphK1 to support efficient muscle regeneration and SC proliferation and differentiation. Mdx mice, which serve as a model for muscular dystrophy (MD), were found to be S1P-deficient and exhibited muscle SPL upregulation, suggesting that S1P catabolism is enhanced in dystrophic muscle. Pharmacological SPL inhibition increased muscle S1P levels, improved mdx muscle regeneration and enhanced SC proliferation via S1P receptor 2 (S1PR2)-dependent inhibition of Rac1, thereby activating Signal Transducer and Activator of Transcription 3 (STAT3), a central player in inflammatory signaling. STAT3 activation resulted in p21 and p27 downregulation in a S1PR2-dependent fashion in myoblasts. Our findings suggest that S1P promotes SC progression through the cell cycle by repression of cell cycle inhibitors via S1PR2/STAT3-dependent signaling and that SPL inhibition may provide a therapeutic strategy for MD.

  19. Ionophore A23187 induces a refractory state in thrombin-activated release of inositol phosphates.

    PubMed Central

    Moscat, G; Moreno, F; Iglesias, S; Garcia-Barreno, P; Municio, A M

    1986-01-01

    The phosphatidylinositol cycle has been proposed to be involved in the regulation of platelet functionality through the control of cytoplasmic Ca2+ levels. However, the requirements of phospholipase C for Ca2+ has not yet been elucidated in intact platelets. The primary purpose of the present study was to investigate the Ca2+ requirements of this enzyme in platelets from miniature swine by taking advantage of the permeabilizing properties of the ionophore A23187. Our results strongly suggest that the treatment of platelets with A23187 induces a refractory state in thrombin-stimulated release of inositol phosphates while 5-hydroxytryptamine (serotonin)-secretory capacity in response to thrombin remained constant. This refractory state seems to be dependent on some cytochalasin-inhibitable cytoskeletal phenomena. PMID:3099773

  20. Science Update: Inorganic Chemistry.

    ERIC Educational Resources Information Center

    Rawls, Rebecca

    1981-01-01

    Describes areas of inorganic chemistry which have changed dramatically in the past year or two, including photochemistry, electrochemistry, organometallic complexes, inorganic reaction theory, and solid state chemistry. (DS)

  1. EFFECT OF ACTIVE ACCUMULATION OF CALCIUM AND PHOSPHATE IONS ON THE STRUCTURE OF RAT LIVER MITOCHONDRIA.

    PubMed

    GREENAWALT, J W; ROSSI, C S; LEHNINGER, A L

    1964-10-01

    Rat liver mitochondria allowed to accumulate maximal amounts of Ca(++) and HPO(4) (=) ions from the suspending medium in vitro during respiration have a considerably higher specific gravity than normal mitochondria and may be easily separated from the latter by isopycnic centrifugation in density gradients of sucrose or cesium chloride. When the mitochondria are allowed to accumulate less than maximal amounts of Ca(++) and HPO(4) (=) from the medium, they have intermediate specific gravities which are roughly proportional to their content of calcium phosphate. Maximally "loaded" mitochondria are relatively homogeneous with respect to specific gravity. Correlated biochemical and electron microscopic studies show that Ca(++)-loaded mitochondria contain numerous dense granules, of which some 85 per cent are over 500 A in diameter. These granules are electron-opaque not only following fixation and staining with heavy metal reagents, but also following fixation with formaldehyde, demonstrating that the characteristic granules in Ca(++)-loaded mitochondria have intrinsic electron-opacity. The dense granules are almost always located within the inner compartment of the mitochondria and not in the space between the inner and outer membranes. They are frequently located at or near the cristae and they often show electron-transparent "cores." Such granules appear to be made up of clusters of smaller dense particles, but preliminary x-ray diffraction analysis and electron diffraction studies have revealed no evidence of crystallinity in the deposits. The electron-opaque granules decrease in number when the Ca(++)-loaded mitochondria are incubated with 2,4-dinitrophenol; simultaneously there is discharge of Ca(++) and phosphate from the mitochondria into the medium.

  2. Biomineralization of U(VI) phosphate promoted by microbially-mediated phytate hydrolysis in contaminated soils

    NASA Astrophysics Data System (ADS)

    Salome, Kathleen R.; Beazley, Melanie J.; Webb, Samuel M.; Sobecky, Patricia A.; Taillefert, Martial

    2017-01-01

    The bioreduction of uranium may immobilize a significant fraction of this toxic contaminant in reduced environments at circumneutral pH. In oxic and low pH environments, however, the low solubility of U(VI)-phosphate minerals also makes them good candidates for the immobilization of U(VI) in the solid phase. As inorganic phosphate is generally scarce in soils, the biomineralization of U(VI)-phosphate minerals via microbially-mediated organophosphate hydrolysis may represent the main immobilization process of uranium in these environments. In this study, contaminated sediments were incubated aerobically in two pH conditions to examine whether phytate, a naturally-occurring and abundant organophosphate in soils, could represent a potential phosphorous source to promote U(VI)-phosphate biomineralization by natural microbial communities. While phytate hydrolysis was not evident at pH 7.0, nearly complete hydrolysis was observed both with and without electron donor at pH 5.5, suggesting indigenous microorganisms express acidic phytases in these sediments. While the rate of hydrolysis of phytate generally increased in the presence of uranium, the net rate of inorganic phosphate production in solution was decreased and inositol phosphate intermediates were generated in contrast to similar incubations conducted without uranium. These findings suggest uranium stress enhanced the phytate-metabolism of the microbial community, while simultaneously inhibiting phosphatase production and/or activity by the indigenous population. Finally, phytate hydrolysis drastically decreased uranium solubility, likely due to formation of ternary sorption complexes, U(VI)-phytate precipitates, and U(VI)-phosphate minerals. Overall, the results of this study provide evidence for the ability of natural microbial communities to liberate phosphate from phytate in acidic sediments, possibly as a detoxification mechanism, and demonstrate the potential utility of phytate-promoted uranium

  3. Biologically-synthesized inorganic nanomaterials

    NASA Astrophysics Data System (ADS)

    Kramer, Ryan M.; Stone, Morley O.; Naik, Rajesh R.

    2004-06-01

    A hallmark of biological systems is their ability to self-assemble. This self-assembly can occur on the molecular, macromolecular and mesoscale. In this work, we have chosen to exploit biology's ability to self-assemble by incorporating additional functionality within the final structure. Our research efforts have been directed at not only understanding how biological organisms control nucleation and growth of inorganic materials, but also how this activity can be controlled in vitro. In previous work, we have demonstrated how peptides can be selected from a combinatorial library that possesses catalytic activity with respect to inorganic nucleation and deposition. We have engineered some of these peptide sequences into self-assembling protein structures. The goal of the project was to create an organic/inorganic hybrid that retained the "memory" properties of the organic, but possessed the superior optical and electronic properties of the inorganic.

  4. Autosomal factors with correlated effects on the activities of the glucose 6-phosphate and 6-phosphogluconate dehydrogenases in Drosophila melanogaster.

    PubMed

    Laurie-Ahlberg, C C; Williamson, J H; Cochrane, B J; Wilton, A N; Chasalow, F I

    1981-09-01

    Isogenic lines, in which chromosomes sampled from natural populations of C. melanogaster are substituted into a common genetic background, were used to detect and partially characterize autosomal factors that affect the activities of the two pentose phosphate pathway enzymes, glucose 6-phosphate dehydrogenase (G6PD) and 6-phosphogluconate dehydrogenase (6PGD). The chromosome 3 effects on G6PD and 6PGD are clearly correlated; the chromosome 2 effects, which are not so great, also appear to be correlated, but the evidence in this case is not so strong. Examination of activity variation of ten other enzymes revealed that G6PD and 6PGD are not the only pair of enzymes showing a high positive correlation, but it is among the highest in both sets of lines. In addition, there was some evidence that the factor(s) affecting G6PD and 6PGD may also affect two other metabolically related enzymes, transaldolase and phosphoglucose isomerase.--Rocket immunoelectrophoresis was used to estimate specific CRM levels for three of the enzymes studied: G6PD, 6PGD and ME. This experiment shows that a large part of the activity variation is accounted for by variation in CRM level (especially for chromosome 3 lines), but there remains a significant fraction of the genetic component of activity variation that is not explained by CRM level.--These results suggest that the autosomal factors are modifiers involved in regulation of the expression of the X-linked structural genes for G6PD and 6PGD, but a role in determining part of the enzymes' primary structure cannot be excluded with the present evidence.

  5. Autosomal Factors with Correlated Effects on the Activities of the Glucose 6-Phosphate and 6-Phosphogluconate Dehydrogenases in DROSOPHILA MELANOGASTER

    PubMed Central

    Laurie-Ahlberg, C. C.; Williamson, J. H.; Cochrane, B. J.; Wilton, A. N.; Chasalow, F. I.

    1981-01-01

    Isogenic lines, in which chromosomes sampled from natural populations of D. melanogaster are substituted into a common genetic background, were used to detect and partially characterize autosomal factors that affect the activities of the two pentose phosphate pathway enzymes, glucose 6-phosphate dehydrogenase (G6PD) and 6-phosphogluconate dehydrogenase (6PGD). The chromosome 3 effects on G6PD and 6PGD are clearly correlated; the chromosome 2 effects, which are not so great, also appear to be correlated, but the evidence in this case is not so strong. Examination of activity variation of ten other enzymes revealed that G6PD and 6PGD are not the only pair of enzymes showing a high positive correlation, but it is among the highest in both sets of lines. In addition, there was some evidence that the factor(s) affecting G6PD and 6PGD may also affect two other metabolically related enzymes, transaldolase and phosphoglucose isomerase.—Rocket immunoelectrophoresis was used to estimate specific CRM levels for three of the enzymes studied: G6PD, 6PGD and ME. This experiment shows that a large part of the activity variation is accounted for by variation in CRM level (especially for chromosome 3 lines), but there remains a significant fraction of the genetic component of activity variation that is not explained by CRM level.—These results suggest that the autosomal factors are modifiers involved in regulation of the expression of the X-linked structural genes for G6PD and 6PGD, but a role in determining part of the enzymes' primary structure cannot be excluded with the present evidence. PMID:6804300

  6. Effects of Sphingosine-1-Phosphate on Pacemaker Activity of Interstitial Cells of Cajal from Mouse Small Intestine

    PubMed Central

    Kim, Young Dae; Han, Kyoung Taek; Lee, Jun; Park, Chan Guk; Kim, Man Yoo; Shahi, Pawan Kumar; Zuo, Dong Chuan; Choi, Seok; Jun, Jae Yeoul

    2013-01-01

    Interstitial cells of Cajal (ICC) are the pacemaker cells that generate the rhythmic oscillation responsible for the production of slow waves in gastrointestinal smooth muscle. Spingolipids are known to present in digestive system and are responsible for multiple important physiological and pathological processes. In this study, we are interested in the action of sphingosine 1-phosphate (S1P) on ICC. S1P depolarized the membrane and increased tonic inward pacemaker currents. FTY720 phosphate (FTY720P, an S1P1,3,4,5 agonist) and SEW 2871 (an S1P1 agonist) had no effects on pacemaker activity. Suramin (an S1P3 antagonist) did not block the S1P-induced action on pacemaker currents. However, JTE-013 (an S1P2 antagonist) blocked the S1P-induced action. RT-PCR revealed the presence of the S1P2 in ICC. Calphostin C (a protein kinase C inhibitor), NS-398 (a cyclooxygenase-2 inhibitor), PD 98059 (a p42/44 inhibitor), or SB 203580 (a p38 inhibitor) had no effects on S1P-induced action. However, c-jun NH2-terminal kinase (JNK) inhibitor II suppressed S1P-induced action. External Ca2+-free solution or thapsigargin (a Ca2+-ATPase inhibitor of endoplasmic reticulum) suppressed action of S1P on ICC. In recording of intracellular Ca2+ ([Ca2+]i) concentration using fluo-4/AM S1P increased intensity of spontaneous [Ca2+]i oscillations in ICC. These results suggest that S1P can modulate pacemaker activity of ICC through S1P2 via regulation of external and internal Ca2+ and mitogen-activated protein kinase activation. PMID:23307289

  7. Time and Demand are Two Critical Dimensions of Immunometabolism: The Process of Macrophage Activation and the Pentose Phosphate Pathway

    PubMed Central

    Nagy, Csörsz; Haschemi, Arvand

    2015-01-01

    A process is a function of time; in immunometabolism, this is reflected by the stepwise adaptation of metabolism to sustain the bio-energetic demand of an immune-response in its various states and shades. This perspective article starts by presenting an early attempt to investigate the physiology of inflammation, in order to illustrate one of the basic concepts of immunometabolism, wherein an adapted metabolism of infiltrating immune cells affects tissue function and inflammation. We then focus on the process of macrophage activation and aim to delineate the factor time within the current molecular context of metabolic-rewiring important for adapting primary carbohydrate metabolism. In the last section, we will provide information on how the pentose phosphate pathway may be of importance to provide both nucleotide precursors and redox-equivalents, and speculate how carbon-scrambling events in the non-oxidative pentose phosphate pathway might be regulated within cells by demand. We conclude that the adapted metabolism of inflammation is specific in respect to the effector-function and appears as a well-orchestrated event, dynamic by nature, and based on a functional interplay of signaling- and metabolic-pathways. PMID:25904920

  8. Chloroplast Activity and 3′phosphadenosine 5′phosphate Signaling Regulate Programmed Cell Death in Arabidopsis1

    PubMed Central

    Mazubert, Christelle; Prunier, Florence; Chan, Kai Xun; Pogson, Barry James; Krieger-Liszkay, Anja; Delarue, Marianne; Benhamed, Moussa; Bergounioux, Catherine; Raynaud, Cécile

    2016-01-01

    Programmed cell death (PCD) is a crucial process both for plant development and responses to biotic and abiotic stress. There is accumulating evidence that chloroplasts may play a central role during plant PCD as for mitochondria in animal cells, but it is still unclear whether they participate in PCD onset, execution, or both. To tackle this question, we have analyzed the contribution of chloroplast function to the cell death phenotype of the myoinositol phosphate synthase1 (mips1) mutant that forms spontaneous lesions in a light-dependent manner. We show that photosynthetically active chloroplasts are required for PCD to occur in mips1, but this process is independent of the redox state of the chloroplast. Systematic genetic analyses with retrograde signaling mutants reveal that 3′-phosphoadenosine 5′-phosphate, a chloroplast retrograde signal that modulates nuclear gene expression in response to stress, can inhibit cell death and compromises plant innate immunity via inhibition of the RNA-processing 5′-3′ exoribonucleases. Our results provide evidence for the role of chloroplast-derived signal and RNA metabolism in the control of cell death and biotic stress response. PMID:26747283

  9. In Vitro Antiviral Activity of 6-Substituted 9-β-d-Ribofuranosylpurine 3′,5′-Cyclic Phosphates

    PubMed Central

    Sidwell, Robert W.; Huffman, John H.; Allen, Lois B.; Meyer, Rich B.; Shuman, Dennis A.; Simon, Lionel N.; Robins, Roland K.

    1974-01-01

    A series of twelve recently synthesized 6-substituted derivatives of 9-β-d-ribofuranosylpurine 3′,5′-cyclic phosphate (RPcMP) were evaluated for in vitro antiviral activity, using inhibition of viral cytopathogenic effect as the primary parameter for evaluation. Inhibition of the development of intra- and extracellular virus titer was used as a secondary criterion with certain viruses. Five derivatives were considered to have significant antiviral activity. 6-Hydroxylamino-RPcMP was active against type 1 herpes simplex, cytomegalo-, and vaccinia viruses. 6-Thio-RPcMP was inhibitory to types 1 and 2 herpes simplex, cytomegalo-, vaccinia, and type 3 parainfluenza viruses. The 6-methylthio derivative was active against types 1 and 2 herpes simplex, cytomegalo-, and vaccinia viruses, and types 1A, 2, 8, and 13 rhinoviruses; alteration of this 6-substitution to 6-ethylthio or to 6-benzylthio weakened the herpes- and vaccinia virus activity of the compound, but each continued to have significant antirhinovirus activity. The effect of time of addition of 6-methylthio-RPcMP to type 1 herpes simplex virus-infected cells was determined; the compound was most active when added prior to the virus. Early removal of the compound from the infected cells markedly reduced its antiviral effectiveness. PMID:15825420

  10. Increased mitochondrial glycerol-3-phosphate acyltransferase protein and enzyme activity in rat epididymal fat upon cessation of wheel running.

    PubMed

    Kump, David S; Laye, Matthew J; Booth, Frank W

    2006-03-01

    Triacylglycerol synthesis in rat epididymal fat overshoots sedentary levels at 10, 29, and 53 h of physical inactivity after 21 days of wheel running. The purposes of the present study were to determine 1) whether this effect is also observed after an acute bout of physical activity and 2) what enzymatic changes might contribute to this effect. We show that more than one bout of physical activity, such as that which occurs with 21 days of wheel running, is necessary for palmitic acid incorporation into triacylglyceride (triglyceride synthesis) to overshoot sedentary values, which suggests that pretranslational mechanisms may be responsible for this overshoot effect. Ten hours after 21 days of wheel running, activity of the mitochondrial glycerol-3-phosphate acyltransferase-1 (mtGPAT1) isoform, a key regulator of triacylglycerol synthesis, overshot sedentary values by 48% and remained higher than sedentary values at 29 and 53 h of reduced physical activity. The overshoot in mtGPAT1 activity was accompanied by an increase in mtGPAT protein level. Cyclic AMP response element-binding protein-binding protein level was higher in sedentary 29 h after 21 days of wheel running. AMP kinase-alpha Thr(172) phosphorylation was increased immediately after treadmill running, but decreased to sedentary values by 5 h after activity. Casein kinase-2alpha protein level and activity were unchanged. We conclude that an increase in mtGPAT protein might contribute to the overshoot in triacylglycerol synthesis.

  11. Glutamine-dependent carbamoyl-phosphate synthetase and other enzyme activities related to the pyrimidine pathway in spleen of Squalus acanthias (spiny dogfish).

    PubMed

    Anderson, P M

    1989-07-15

    The first two steps of urea synthesis in liver of marine elasmobranchs involve formation of glutamine from ammonia and of carbamoyl phosphate from glutamine, catalysed by glutamine synthetase and carbamoyl-phosphate synthetase, respectively [Anderson & Casey (1984) J. Biol. Chem. 259, 456-462]; both of these enzymes are localized exclusively in the mitochondrial matrix. The objective of this study was to establish the enzymology of carbamoyl phosphate formation and utilization for pyrimidine nucleotide biosynthesis in Squalus acanthias (spiny dogfish), a representative elasmobranch. Aspartate carbamoyltransferase could not be detected in liver of dogfish. Spleen extracts, however, had glutamine-dependent carbamoyl-phosphate synthetase, aspartate carbamoyltransferase, dihydro-orotase, and glutamine synthetase activities, all localized in the cytosol; dihydro-orotate dehydrogenase, orotate phosphoribosyltransferase, and orotidine-5'-decarboxylase activities were also present. Except for glutamine synthetase, the levels of all activities were very low. The carbamoyl-phosphate synthetase activity is inhibited by UTP and is activated by 5-phosphoribosyl 1-pyrophosphate. The first three enzyme activities of the pyrimidine pathway were eluted in distinctly different positions during gel filtration chromatography under a number of different conditions; although complete proteolysis of inter-domain regions of a multifunctional complex during extraction cannot be excluded, the evidence suggests that in dogfish, in contrast to mammalian species, these three enzymes of the pyrimidine pathway exist as individual polypeptide chains. These results: (1) establish that dogfish express two different glutamine-dependent carbamoyl-phosphate synthetase activities, (2) confirm the report [Smith, Ritter & Campbell (1987) J. Biol. Chem. 262, 198-202] that dogfish express two different glutamine synthetases, and (3) provide indirect evidence that glutamine may not be available in liver for

  12. Adenine derivatives as phosphate-activating groups for the regioselective formation of 3',5'-linked oligoadenylates on montmorillonite: possible phosphate-activating groups for the prebiotic synthesis of RNA

    NASA Technical Reports Server (NTRS)

    Prabahar, K. J.; Ferris, J. P.

    1997-01-01

    Methyladenine and adenine N-phosphoryl derivatives of adenosine 5'-monophosphate (5'-AMP) and uridine 5'-monophosphate (5'-UMP) are synthesized, and their structures are elucidated. The oligomerization reactions of the adenine derivatives of 5'-phosphoramidates of adenosine on montmorillonite are investigated. 1-Methyladenine and 3-methyladenine derivatives on montmorillonite yielded oligoadenylates as long as undecamer, and the 2-methyladenine and adenine derivatives on montmorillonite yielded oligomers up to hexamers and pentamers, respectively. The 1-methyladenine derivative yielded linear, cyclic, and A5'ppA-derived oligonucleotides with a regioselectivity for the 3',5'-phosphodiester linkages averaging 84%. The effect of pKa and amine structure of phosphate-activating groups on the montmorillonite-catalyzed oligomerization of the 5'-phosphoramidate of adenosine are discussed. The binding and reaction of methyladenine and adenine N-phosphoryl derivatives of adenosine are described.

  13. Synthesis of unsaturated phosphatidylinositol 4-phosphates and the effects of substrate unsaturation on SopB phosphatase activity.

    PubMed

    Furse, Samuel; Mak, LokHang; Tate, Edward W; Templer, Richard H; Ces, Oscar; Woscholski, Rüdiger; Gaffney, Piers R J

    2015-02-21

    In this paper evidence is presented that the fatty acid component of an inositide substrate affects the kinetic parameters of the lipid phosphatase Salmonella Outer Protein B (SopB). A succinct route was used to prepare the naturally occurring enantiomer of phosphatidylinositol 4-phosphate (PI-4-P) with saturated, as well as singly, triply and quadruply unsaturated, fatty acid esters, in four stages: (1) The enantiomers of 2,3:5,6-O-dicyclohexylidene-myo-inositol were resolved by crystallisation of their di(acetylmandelate) diastereoisomers. (2) The resulting diol was phosphorylated regio-selectively exclusively on the 1-O using the new reagent tri(2-cyanoethyl)phosphite. (3) With the 4-OH still unprotected, the glyceride was coupled using phosphate tri-ester methodology. (4) A final phosphorylation of the 4-O, followed by global deprotection under basic then acidic conditions, provided PI-4-P bearing a range of sn-1-stearoyl, sn-2-stearoyl, -oleoyl, -γ-linolenoyl and arachidonoyl, glycerides. Enzymological studies showed that the introduction of cis-unsaturated bonds has a measurable influence on the activity (relative Vmax) of SopB. Mono-unsaturated PI-4-P exhibited a five-fold higher activity, with a two-fold higher KM, over the saturated substrate, when presented in DOPC vesicles. Poly-unsaturated PI-4-P showed little further change with respect to the singly unsaturated species. This result, coupled with our previous report that saturated PI-4-P has much higher stored curvature elastic stress than PI, supports the hypothesis that the activity of inositide phosphatase SopB has a physical role in vivo.

  14. Influence of moderate pre-oxidation treatment on the physical, chemical and phosphate adsorption properties of iron-containing activated carbon.

    PubMed

    Wang, Zhengfang; Shi, Mo; Li, Jihua; Zheng, Zheng

    2014-03-01

    A novel adsorbent based on iron oxide dispersed over activated carbon (AC) were prepared, and used for phosphate removal from aqueous solutions. The influence of pre-oxidation treatment on the physical, chemical and phosphate adsorption properties of iron-containing AC were determined. Two series of ACs, non-oxidized and oxidized carbon modified by iron (denoted as AC-Fe and AC/O-Fe), resulted in a maximum impregnated iron of 4.03% and 7.56%, respectively. AC/O-Fe showed 34.0%-46.6% higher phosphate removal efficiency than the AC-Fe did. This was first attributed to the moderate pre-oxidation of raw AC by nitric acid, achieved by dosing Fe(II) after a pre-oxidation, to obtain higher iron loading, which is favorable for phosphate adsorption. Additionally, the in-situ formed active site on the surface of carbon, which was derived from the oxidation of Fe(II) by nitric acid dominated the remarkably high efficiency with respect to the removal of phosphate. The activation energy for adsorption was calculated to be 10.53 and 18.88 kJ/mol for AC-Fe and AC/O-Fe, respectively. The results showed that the surface mass transfer and intra-particle diffusion were simultaneously occurring during the process and contribute to the adsorption mechanism.

  15. Pumpable/injectable phosphate-bonded ceramics

    DOEpatents

    Singh, Dileep; Wagh, Arun S.; Perry, Lamar; Jeong, Seung-Young

    2001-01-01

    A pumpable ceramic composition is provided comprising an inorganic oxide, potassium phosphate, and an oxide coating material. Also provided is a method for preparing pumpable ceramic-based waste forms comprising selecting inorganic oxides based on solubility, surface area and morphology criteria; mixing the selected oxides with phosphate solution and waste to form a first mixture; combining an additive to the first mixture to create a second mixture; adding water to the second mixture to create a reactive mixture; homogenizing the reactive mixture; and allowing the reactive mixture to cure.

  16. Genome-Wide Transcriptional Analysis of the Phosphate Starvation Stimulon of Bacillus subtilis†

    PubMed Central

    Allenby, Nicholas E. E.; O'Connor, Nicola; Prágai, Zoltán; Ward, Alan C.; Wipat, Anil; Harwood, Colin R.

    2005-01-01

    Bacillus subtilis responds to phosphate starvation stress by inducing the PhoP and SigB regulons. While the PhoP regulon provides a specific response to phosphate starvation stress, maximizing the acquisition of phosphate (Pi) from the environment and reducing the cellular requirement for this essential nutrient, the SigB regulon provides nonspecific resistance to stress by protecting essential cellular components, such as DNA and membranes. We have characterized the phosphate starvation stress response of B. subtilis at a genome-wide level using DNA macroarrays. A combination of outlier and cluster analyses identified putative new members of the PhoP regulon, namely, yfkN (2′,3′ cyclic nucleotide 2′-phosphodiesterase), yurI (RNase), yjdB (unknown), and vpr (extracellular serine protease). YurI is thought to be responsible for the nonspecific degradation of RNA, while the activity of YfkN on various nucleotide phosphates suggests that it could act on substrates liberated by YurI, which produces 3′ or 5′ phosphoribonucleotides. The putative new PhoP regulon members are either known or predicted to be secreted and are likely to be important for the recovery of inorganic phosphate from a variety of organic sources of phosphate in the environment. PMID:16291680

  17. Modifying Polymer Flocculants for the Removal of Inorganic Phophate from Water

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Due to strong hydrogen bonding interactions, thiourea has been shown to have a high affinity for anions such as inorganic phosphate. The interaction between phosphate and thiourea has been used to develop technologies that can detect and even remove phosphate from water. This research investigates t...

  18. Synthesis, characterization and antimicrobial activity of the micro/nano structured biogenic silver doped calcium phosphate

    NASA Astrophysics Data System (ADS)

    Supraja, N.; Prasad, T. N. V. K. V.; David, Ernest

    2016-01-01

    Scale formation in PVC pipelines reduces the water flow efficiency and enhances microbial contamination. A bio-based composite material comprising of silver doped calcium phosphate (Cp-Ag) was synthesized using a simple technique (photo catalysis) and herein, we report for the first time on preparation and evaluation of the antimicrobial efficacy of silver doped calcite extracted from the scale in drinking water pipe lines. Five concentrations of silver doped calcite materials viz,5, 10, 15, 20 and 25 ppm were prepared using chemical ammonia mediated synthetic method. The material Cp-Ag was characterized by using the techniques UV-Visible spectroscopy, X-ray diffraction, Fourier transform infrared spectroscopy, Scanning electron microscopy, Raman spectroscopy, Thermo gravimetric analysis, X-ray photo electron spectroscopy (XPS), Nuclear magnetic resonance spectrometer and X-ray flouresence microscopy (XRF). Typical rhombohedral structure of the silver doped calcite was observed. XRF and XPS studies confirmed the presence of both calcium and silver in the composite material (Cp-Ag). The silver doped calcite material exhibited enhanced inhibition against Escherichia coli and staphylococcus aureus (Kirby-Bauer discs diffusion assay) which is also dependent on the concentration of the Cp-Ag material.

  19. Rheb protein binds CAD (carbamoyl-phosphate synthetase 2, aspartate transcarbamoylase, and dihydroorotase) protein in a GTP- and effector domain-dependent manner and influences its cellular localization and carbamoyl-phosphate synthetase (CPSase) activity.

    PubMed

    Sato, Tatsuhiro; Akasu, Hitomi; Shimono, Wataru; Matsu, Chisa; Fujiwara, Yuki; Shibagaki, Yoshio; Heard, Jeffrey J; Tamanoi, Fuyuhiko; Hattori, Seisuke

    2015-01-09

    Rheb small GTPases, which consist of Rheb1 and Rheb2 (also known as RhebL1) in mammalian cells, are unique members of the Ras superfamily and play central roles in regulating protein synthesis and cell growth by activating mTOR. To gain further insight into the function of Rheb, we carried out a search for Rheb-binding proteins and found that Rheb binds to CAD protein (carbamoyl-phosphate synthetase 2, aspartate transcarbamoylase, and dihydroorotase), a multifunctional enzyme required for the de novo synthesis of pyrimidine nucleotides. CAD binding is more pronounced with Rheb2 than with Rheb1. Rheb binds CAD in a GTP- and effector domain-dependent manner. The region of CAD where Rheb binds is located at the C-terminal region of the carbamoyl-phosphate synthetase domain and not in the dihydroorotase and aspartate transcarbamoylase domains. Rheb stimulated carbamoyl-phosphate synthetase activity of CAD in vitro. In addition, an elevated level of intracellular UTP pyrimidine nucleotide was observed in Tsc2-deficient cells, which was attenuated by knocking down of Rheb. Immunostaining analysis showed that expression of Rheb leads to increased accumulation of CAD on lysosomes. Both a farnesyltransferase inhibitor that blocks membrane association of Rheb and knockdown of Rheb mislocalized CAD. These results establish CAD as a downstream effector of Rheb and suggest a possible role of Rheb in regulating de novo pyrimidine nucleotide synthesis.

  20. Expression, purification, and characterization of cold-adapted inorganic pyrophosphatase from psychrophilic Shewanella sp. AS-11.

    PubMed

    Ginting, Elvy Like; Iwasaki, Syouhei; Maeganeku, Chihiro; Motoshima, Hiroyuki; Watanabe, Keiichi

    2014-01-01

    In the presence of divalent cations, inorganic pyrophosphatase is activated to hydrolyze inorganic pyrophosphate to inorganic phosphate. Here, we clone, express, purify, and characterize inorganic pyrophosphatase from the psychrophilic Shewanella sp. AS-11 (Sh-PPase). The recombinant Sh-PPase was expressed in Escherichia coli BL21 (DE3) at 20°C using pET16b as an expression vector and purified from the cell extracts by a combination of ammonium sulfate fractionation and anion-exchange chromatography. Sh-PPase was found to be a family II PPase with a subunit molecular mass of 34 kD that preferentially utilizes Mn²⁺ over Mg²⁺ ions for activity. The functional characteristics of Sh-PPase, such as activity, temperature dependency, and thermal inactivation, were greatly influenced by manganese ions. Manganese ion activation increased the enzyme's activity at low temperatures; therefore, it was required to gain the cold-adapted characteristics of Sh-PPase.

  1. Regulation of inorganic sulfate activation in filamentous fungi. Allosteric inhibition of ATP sulfurylase by 3'-phosphoadenosine-5'-phosphosulfate.

    PubMed

    Renosto, F; Martin, R L; Wailes, L M; Daley, L A; Segel, I H

    1990-06-25

    ATP sulfurylases from Penicillium chrysogenum, Penicillium duponti, Aspergillus nidulans, and Neurospora crassa are strongly inhibited by 3'-phosphoadenosine-5'-phosphosulfate (PAPS), the product of the second (adenosine-5'-phosphosulfate kinase-catalyzed) reaction in the two-step activation of inorganic sulfate. The v versus [PAPS] plots are sigmoidal. At physiological concentrations of MgATP (0.17-3 mM) and SO4(2-) (0.4-10 mM), the [I]0.5 for PAPS inhibition of the P. chrysogenum enzyme is 35-200 microM; [I]0.9 is 68-310 microM. In the presence of PAPS, the [S]0.5 values for both substrates are increased and the v versus [MgATP] and v versus [SO4(2-)] or [MoO4(2-)] plots are sigmoidal. Fluorosulfonate (FSO3-) and thiosulfate (S2O3(2-] (non-reactive sulfate analogs) inhibit the enzyme at subsaturating substrate concentrations in the absence of PAPS, but low concentrations of the analogs activate the enzyme when PAPS is present. Thus, PAPS behaves as an allosteric inhibitor of ATP sulfurylase. In contrast, adenosine-5'-phosphosulfate (APS = product Q), the immediate product of the SO4(2-)-dependent reaction, is a linear inhibitor of the P. chrysogenum enzyme, competitive with both MgATP and MoO4(2-) (Kiq = 36-73 nM). FSO3- or S2O3(2-) does not activate the enzyme in the presence of APS. The effect of PAPS on fungal ATP sulfurylase is very similar to that observed when a single highly reactive cysteinyl SH group/subunit (SH-1) is covalently modified (Renosto, F., Martin, R. L., and Segel, I. H. (1987) J. Biol. Chem. 262, 16279-16288). The results suggest that in vitro SH-1 modification induces a conformational change in the enzyme that mimics the change induced in vivo by the reversible binding of PAPS. No evidence was obtained to suggest that PAPS covalently modifies SH-1. ATP sulfurylases from rat liver (Yu, M., Martin, R. L., Jain, S., Chen, L. T., and Segel, I. H. (1989) Arch. Biochem. Biophys. 269, 156-174), spinach leaf, cabbage leaf, and Saccharomyces

  2. The marine sponge-derived inorganic polymers, biosilica and polyphosphate, as morphogenetically active matrices/scaffolds for the differentiation of human multipotent stromal cells: potential application in 3D printing and distraction osteogenesis.

    PubMed

    Wang, Xiaohong; Schröder, Heinz C; Grebenjuk, Vladislav; Diehl-Seifert, Bärbel; Mailänder, Volker; Steffen, Renate; Schloßmacher, Ute; Müller, Werner E G

    2014-02-21

    The two marine inorganic polymers, biosilica (BS), enzymatically synthesized from ortho-silicate, and polyphosphate (polyP), a likewise enzymatically synthesized polymer consisting of 10 to >100 phosphate residues linked by high-energy phosphoanhydride bonds, have previously been shown to display a morphogenetic effect on osteoblasts. In the present study, the effect of these polymers on the differential differentiation of human multipotent stromal cells (hMSC), mesenchymal stem cells, that had been encapsulated into beads of the biocompatible plant polymer alginate, was studied. The differentiation of the hMSCs in the alginate beads was directed either to the osteogenic cell lineage by exposure to an osteogenic medium (mineralization activation cocktail; differentiation into osteoblasts) or to the chondrogenic cell lineage by incubating in chondrocyte differentiation medium (triggering chondrocyte maturation). Both biosilica and polyP, applied as Ca²⁺ salts, were found to induce an increased mineralization in osteogenic cells; these inorganic polymers display also morphogenetic potential. The effects were substantiated by gene expression studies, which revealed that biosilica and polyP strongly and significantly increase the expression of bone morphogenetic protein 2 (BMP-2) and alkaline phosphatase (ALP) in osteogenic cells, which was significantly more pronounced in osteogenic versus chondrogenic cells. A differential effect of the two polymers was seen on the expression of the two collagen types, I and II. While collagen Type I is highly expressed in osteogenic cells, but not in chondrogenic cells after exposure to biosilica or polyP, the upregulation of the steady-state level of collagen Type II transcripts in chondrogenic cells is comparably stronger than in osteogenic cells. It is concluded that the two polymers, biosilica and polyP, are morphogenetically active additives for the otherwise biologically inert alginate polymer. It is proposed that alginate

  3. The Marine Sponge-Derived Inorganic Polymers, Biosilica and Polyphosphate, as Morphogenetically Active Matrices/Scaffolds for the Differentiation of Human Multipotent Stromal Cells: Potential Application in 3D Printing and Distraction Osteogenesis

    PubMed Central

    Wang, Xiaohong; Schröder, Heinz C.; Grebenjuk, Vladislav; Diehl-Seifert, Bärbel; Mailänder, Volker; Steffen, Renate; Schloßmacher, Ute; Müller, Werner E. G.

    2014-01-01

    The two marine inorganic polymers, biosilica (BS), enzymatically synthesized from ortho-silicate, and polyphosphate (polyP), a likewise enzymatically synthesized polymer consisting of 10 to >100 phosphate residues linked by high-energy phosphoanhydride bonds, have previously been shown to display a morphogenetic effect on osteoblasts. In the present study, the effect of these polymers on the differential differentiation of human multipotent stromal cells (hMSC), mesenchymal stem cells, that had been encapsulated into beads of the biocompatible plant polymer alginate, was studied. The differentiation of the hMSCs in the alginate beads was directed either to the osteogenic cell lineage by exposure to an osteogenic medium (mineralization activation cocktail; differentiation into osteoblasts) or to the chondrogenic cell lineage by incubating in chondrocyte differentiation medium (triggering chondrocyte maturation). Both biosilica and polyP, applied as Ca2+ salts, were found to induce an increased mineralization in osteogenic cells; these inorganic polymers display also morphogenetic potential. The effects were substantiated by gene expression studies, which revealed that biosilica and polyP strongly and significantly increase the expression of bone morphogenetic protein 2 (BMP-2) and alkaline phosphatase (ALP) in osteogenic cells, which was significantly more pronounced in osteogenic versus chondrogenic cells. A differential effect of the two polymers was seen on the expression of the two collagen types, I and II. While collagen Type I is highly expressed in osteogenic cells, but not in chondrogenic cells after exposure to biosilica or polyP, the upregulation of the steady-state level of collagen Type II transcripts in chondrogenic cells is comparably stronger than in osteogenic cells. It is concluded that the two polymers, biosilica and polyP, are morphogenetically active additives for the otherwise biologically inert alginate polymer. It is proposed that alginate

  4. Kinetic analysis of Escherichia coli 2-C-methyl-D-erythritol-4-phosphate cytidyltransferase, wild type and mutants, reveals roles of active site amino acids.

    PubMed

    Richard, Stéphane B; Lillo, Antonietta M; Tetzlaff, Charles N; Bowman, Marianne E; Noel, Joseph P; Cane, David E

    2004-09-28

    Escherichia coli 2-C-methyl-D-erythritol-4-phosphate cytidyltransferase (YgbP or IspD) catalyzes the conversion of 2-C-methyl-D-erythritol 4-phosphate (MEP) and cytidine triphosphate (CTP) to 4-diphosphocytidyl-2-C-methylerythritol (CDPME). Pulse chase experiments established that the reaction involves an ordered sequential mechanism with mandatory initial binding of CTP. On the basis of analysis of the previously reported crystal structures of apo-YgbP as well as YgbP complexed with both CTP.Mg(2+) and CDPME.Mg(2+) [Richard, S. B., Bowman, M. E., Kwiatkowski, W., Kang, I., Chow, C., Lillo, A. M., Cane, D. E., and Noel, J. P. (2001) Nat. Struct. Biol. 8, 641-648], a group of active site residues were selected for site-directed mutagenesis and steady-state kinetic analysis. Both Lys27 and Lys213 were shown to be essential to catalytic activity, consistent with their proposed role in stabilization of a pentacoordinate phosphate transition state resulting from in-line attack of the MEP phosphate on the alpha-phosphate of CTP. In addition, Thr140, Arg109, Asp106, and Thr165 were all shown to play critical roles in the binding and proper orientation of the MEP substrate.

  5. Phosphate salts

    MedlinePlus

    ... sodium if you have heart disease. Fluid retention (edema): Avoid using phosphate salts that contain sodium if ... heart failure, or other conditions that can cause edema. High levels of calcium in the blood (hypercalcemia): ...

  6. Alkaline phosphatase activity and its relationship to inorganic phosphorus in the transition zone of the North-western African upwelling system

    NASA Astrophysics Data System (ADS)

    Sebastián, Marta; Arístegui, Javier; Montero, María F.; Escanez, Jose; Xavier Niell, F.

    2004-08-01

    The enzymatic activity of alkaline phosphatase (APA) was studied in the transition zone between the African upwelling system and the open ocean waters of the Canary Islands region. This region is recurrently dominated by the presence of upwelling filaments that may transport nutrient-enriched waters out into the open ocean before nutrients become exhausted by plankton. Turnover rates by APA were generally low in the whole region, but detectable in all the measurements carried out. On average, turnover rates were higher in the upwelling stations, and APA in those waters seemed to be mainly generated by heterotrophic bacteria to supply easily assimilable organic C. APA outside the upwelling area showed an inverse hyperbolic relationship with increasing phosphate, suggesting the presence of both constitutive and Pi-inducible APA. In these offshore waters, a threshold of 0.1 μM of phosphate could be defined for the regulatory function of Pi on APA. Thus, APA in nutrient-poor waters seemed to be induced to compensate for Pi-deficiency. Turnover rates in the filaments showed basal (probably constitutive) levels, whereas they increased in the surrounding waters, where phosphate concentration presumably did not satisfy plankton P-demands. The fertilising effect of the filaments and associated cyclonic eddies extended to at least 175 km offshore, where basal alkaline phosphatase activities were still found. The magnitude of this effect depends probably on the intensity of upwelling events and the degree of recirculation of filament water back to the coastal jet.

  7. Phosphate Activation via Reduced Oxidation State Phosphorus (P). Mild Routes to Condensed-P Energy Currency Molecules

    PubMed Central

    Kee, Terence P.; Bryant, David E.; Herschy, Barry; Marriott, Katie E. R.; Cosgrove, Nichola E.; Pasek, Matthew A.; Atlas, Zachary D.; Cousins, Claire R.

    2013-01-01

    The emergence of mechanisms for phosphorylating organic and inorganic molecules is a key step en route to the earliest living systems. At the heart of all contemporary biochemical systems reside reactive phosphorus (P) molecules (such as adenosine triphosphate, ATP) as energy currency molecules to drive endergonic metabolic processes and it has been proposed that a predecessor of such molecules could have been pyrophosphate [P2O74−; PPi(V)]. Arguably the most geologically plausible route to PPi(V) is dehydration of orthophosphate, Pi(V), normally a highly endergonic process in the absence of mechanisms for activating Pi(V). One possible solution to this problem recognizes the presence of reactive-P containing mineral phases, such as schreibersite [(Fe,Ni)3P] within meteorites whose abundance on the early Earth would likely have been significant during a putative Hadean-Archean heavy bombardment. Here, we propose that the reduced oxidation state P-oxyacid, H-phosphite [HPO32−; Pi(III)] could have activated Pi(V) towards condensation via the intermediacy of the condensed oxyacid pyrophosphite [H2P2O52−; PPi(III)]. We provide geologically plausible provenance for PPi(III) along with evidence of its ability to activate Pi(V) towards PPi(V) formation under mild conditions (80 °C) in water. PMID:25369812

  8. Phosphate Activation via Reduced Oxidation State Phosphorus (P). Mild Routes to Condensed-P Energy Currency Molecules.

    PubMed

    Kee, Terence P; Bryant, David E; Herschy, Barry; Marriott, Katie E R; Cosgrove, Nichola E; Pasek, Matthew A; Atlas, Zachary D; Cousins, Claire R

    2013-07-19

    The emergence of mechanisms for phosphorylating organic and inorganic molecules is a key step en route to the earliest living systems. At the heart of all contemporary biochemical systems reside reactive phosphorus (P) molecules (such as adenosine triphosphate, ATP) as energy currency molecules to drive endergonic metabolic processes and it has been proposed that a predecessor of such molecules could have been pyrophosphate [P2O74-; PPi(V)]. Arguably the most geologically plausible route to PPi(V) is dehydration of orthophosphate, Pi(V), normally a highly endergonic process in the absence of mechanisms for activating Pi(V). One possible solution to this problem recognizes the presence of reactive-P containing mineral phases, such as schreibersite [(Fe,Ni)3P] within meteorites whose abundance on the early Earth would likely have been significant during a putative Hadean-Archean heavy bombardment. Here, we propose that the reduced oxidation state P-oxyacid, H-phosphite [HPO32-; Pi(III)] could have activated Pi(V) towards condensation via the intermediacy of the condensed oxyacid pyrophosphite [H2P2O52-; PPi(III)]. We provide geologically plausible provenance for PPi(III) along with evidence of its ability to activate Pi(V) towards PPi(V) formation under mild conditions (80 °C) in water.

  9. Red light generation through the lead boro-telluro-phosphate glasses activated by Eu3+ ions

    NASA Astrophysics Data System (ADS)

    Selvi, S.; Marimuthu, K.; Suriya Murthy, N.; Muralidharan, G.

    2016-09-01

    Lead boro-telluro-phosphate glasses containing 0.05 to 2.0 wt% of Eu3+ ions were prepared through melt quenching technique. Structural characteristics of title glasses were identified through XRD, FTIR and Raman studies. The optical properties of the prepared glasses were studied using UV-Vis-NIR absorption and photoluminescence spectra. From the resultant spectra, we have obtained the bonding parameters (δ), nephelauxetic ratio (β), direct and indirect band gaps and Urbach energy (ΔE) values. A deep red luminescence due to 5D0 → 7F2 transition of Eu3+ ions could be observed for the title glasses. The local site symmetry around the Eu3+ ions and the degree of Eu3+-O2- covalence were assessed from the luminescence intensity ratio of 5D0 → 7F2/5D0 → 7F1 transitions. Judd-Ofelt intensity parameters, calculated from the luminescence spectra, were used to estimate the radiative parameters like transition probability (A), branching ratio (βexp, βcal) and stimulated emission cross-section (σPE) concerning the 5D0 → 7FJ (J = 0, 1, 2, 3 and 4) transitions. The important laser parameters, gain bandwidth and optical gain are also estimated. The decay curves associated with the transition from 5D0 state was found to be single-exponential at all Eu3+ ion concentrations. CIE colour coordinates and colour purity of the prepared glasses were estimated from the CIE chromaticity diagram.

  10. Effect of feeding and of DDT on the activity of hepatic glucose 6- phosphate dehydrogenase in two salmonids

    USGS Publications Warehouse

    Buhler, Donald R.; Benville, P.

    1969-01-01

    The specific activity of liver glucose 6-phosphate dehydrogenase in yearling rainbow trout remained unchanged when the fish were starved for periods as long as 8 weeks and when starved animals were fed diets of various compositions. Injection of insulin concurrently with refeeding also failed to alter the specific activity of the enzyme in trout. The absence of a dietary or insulin influence on the teleost enzyme system is to be contrasted with studies in mammals in which the activity of hepatic glucose 6-P dehydrogenase was markedly stimulated after refeeding starved animals or injection of insulin.Ingestion of the pesticide DDT by juvenile coho salmon or adult rainbow trout also had no effect on the specific activity of liver glucose 6-P dehydrogenase and DDT failed to inhibit the rainbow trout enzyme in vitro. These results also differ considerably from those found in higher animals.These results suggest that the glucose 6-P dehydrogenase enzyme in teleosts may be under a different type of regulatory control from that found in mammals.

  11. Regulation of Enzyme Activities in Drosophila: Genetic Variation Affecting Induction of Glucose 6-Phosphate and 6-Phosphogluconate Dehydrogenases in Larvae

    PubMed Central

    Cochrane, Bruce J.; Lucchesi, John C.; Laurie-Ahlberg, C. C.

    1983-01-01

    The genetic basis of modulation by dietary sucrose of the enzyme activities glucose 6-phosphate dehydrogenase (G6PD) and 6-phosphogluconate dehydrogenase (6PGD) activities in third instar larvae of Drosophila melanogaster was investigated, using isogenic lines derived from wild populations. Considerable genetically determined variation in response was detected among lines that differed only in their third chromosome constitution. Comparison of crossreacting material between a responding and a nonresponding line showed that the G6PD activity variation is due to changes in G6PD protein level. These differences in responses are localized in the fat body, with 300 m m sucrose in the diet resulting in a sixfold stimulation of G6PD activity and a fourfold one of 6PGD in the line showing the strongest response. In this tissue, the responses of the two enzymes are closely correlated with one another. Using recombinant lines, we obtained data that suggested the existence of more than one gene on chromosome III involved in the regulation of G6PD in the fat body, and at least one of these genes affects the level of 6PGD as well. PMID:6416921

  12. Lowered dietary phosphate increases oral bioavailability of arsenate in mice

    EPA Science Inventory

    Arsenate (iAsv), an inorganic oxyanionic species, has physicochemical properties similar to inorganic phosphate (iP). There is evidence that iAsv competes with iP for transmembrane carriers that mediate iP uptake. Thus, it is possible that altered dietary intake of iP could modif...

  13. Novel mode of inhibition by D-tagatose 6-phosphate through a Heyns rearrangement in the active site of transaldolase B variants.

    PubMed

    Stellmacher, Lena; Sandalova, Tatyana; Schneider, Sarah; Schneider, Gunter; Sprenger, Georg A; Samland, Anne K

    2016-04-01

    Transaldolase B (TalB) and D-fructose-6-phosphate aldolase A (FSAA) from Escherichia coli are C-C bond-forming enzymes. Using kinetic inhibition studies and mass spectrometry, it is shown that enzyme variants of FSAA and TalB that exhibit D-fructose-6-phosphate aldolase activity are inhibited covalently and irreversibly by D-tagatose 6-phosphate (D-T6P), whereas no inhibition was observed for wild-type transaldolase B from E. coli. The crystal structure of the variant TalB(F178Y) with bound sugar phosphate was solved to a resolution of 1.46 Å and revealed a novel mode of covalent inhibition. The sugar is bound covalently via its C2 atom to the ℇ-NH2 group of the active-site residue Lys132. It is neither bound in the open-chain form nor as the closed-ring form of D-T6P, but has been converted to β-D-galactofuranose 6-phosphate (D-G6P), a five-membered ring structure. The furanose ring of the covalent adduct is formed via a Heyns rearrangement and subsequent hemiacetal formation. This reaction is facilitated by Tyr178, which is proposed to act as acid-base catalyst. The crystal structure of the inhibitor complex is compared with the structure of the Schiff-base intermediate of TalB(E96Q) formed with the substrate D-fructose 6-phosphate determined to a resolution of 2.20 Å. This comparison highlights the differences in stereochemistry at the C4 atom of the ligand as an essential determinant for the formation of the inhibitor adduct in the active site of the enzyme.

  14. Localization of nicotinamide adenine dinucleotide phosphate-diaphorase activity in electrosensory and electromotor systems of a gymnotiform teleost, Apteronotus leptorhynchus.

    PubMed

    Turner, R W; Moroz, L L

    1995-05-29

    The distribution of nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) activity was determined in electrosensory and electromotor systems of the weakly electric gymnotiform teleost Apteronotus leptorhynchus as an indicator of putative nitric oxide synthase-containing cells. NADPH-d activity was detected in electroreceptors and in afferent nerves of both ampullary and type I and type II tuberous organs. All cell bodies within the anterior lateral line nerve ganglion were positive for NADPH-d activity, as were the primary afferent axons and termination fields in the medullary electrosensory lateral line lobe. In the corpus cerebelli and valvula cerebelli, NADPH-d label was present in Purkinje cell somata, mossy fiber synaptic glomeruli, granule cells, and parallel fibers. In the midbrain, NADPH-d activity was apparent in layer VIIIB of the torus semicircularis dorsalis and in electrosensory laminae of the optic tectum. NADPH-d was particularly associated with diencephalic electrosensory and electromotor nuclei, including the prepacemaker nucleus, the nucleus subelectrosensorius, and the central posterior nucleus of the thalamus. Intense NADPH-d activity was present in pacemaker and relay cells of the medullary pacemaker nucleus but was absent from a novel class of smaller cells in this structure. Relay cell axons and spinal electromotor neurons and their axons within the electric organ were positive for NADPH-d activity. These results indicate that putative nitric oxide synthase-containing neurons in Apteronotus are localized preferentially to electrosensory and electromotor structures, suggesting a role for nitric oxide in determining the activity of cells involved in detecting or generating weakly electric fields.

  15. Modification of TiO2 Nanoparticles with Oleyl Phosphate via Phase Transfer in the Toluene-Water System and Application of Modified Nanoparticles to Cyclo-Olefin-Polymer-Based Organic-Inorganic Hybrid Films Exhibiting High Refractive Indices.

    PubMed

    Takahashi, Shiori; Hotta, Shuhei; Watanabe, Akira; Idota, Naokazu; Matsukawa, Kimihiro; Sugahara, Yoshiyuki

    2017-01-18

    Oleyl-phosphate-modified TiO2 nanoparticles (OP_TiO2) were prepared via phase transfer from an aqueous phase containing dispersed TiO2 nanoparticles to a toluene phase containing oleyl phosphate (OP, a mixture of monoester and diester), and employed for the preparation of OP_TiO2/cyclo-olefin polymer (COP) hybrid films with high-refractive indices. The modification of TiO2 by OP was essentially completed by reaction at room temperature for 8 h, and essentially all the TiO2 nanoparticles in the aqueous phase were transferred to the toluene phase. The infrared and solid-state (13)C cross-polarization and magic-angle spinning (CP/MAS) NMR spectrum of OP_TiO2 showed the presence of oleyl groups originating from oleyl phosphate. The solid-state (31)P MAS NMR spectrum of OP_TiO2 exhibited new signals at -1.4, 2.1, and 4.8 ppm, indicating the formation of Ti-O-P bonds. CHN and inductively coupled plasma analyses revealed that the major species bound to the TiO2 surface was tridentate CH3(CH2)7CH═CH(CH2)8P(OTi)3. These results clearly indicate that the surfaces of the TiO2 nanoparticles were modified by OP moieties via phase transfer. OP_TiO2/COP hybrid films exhibited excellent optical transparency up to 19.1 vol % TiO2 loading, and the light transmittance of the hybrid films with 19.1 vol % TiO2 loading was 99.8% at 633 nm. The refractive index of these hybrid films rose to 1.83.

  16. Probing the location and function of the conserved histidine residue of phosphoglucose isomerase by using an active site directed inhibitor N-bromoacetylethanolamine phosphate.

    PubMed Central

    Meng, M.; Chane, T. L.; Sun, Y. J.; Hsiao, C. D.

    1999-01-01

    Phosphoglucose isomerase (EC 5.3.1.9) catalyzes the interconversion of D-glucopyranose-6-phosphate and D-fructofuranose-6-phosphate by promoting an intrahydrogen transfer between C1 and C2. A conserved histidine exists throughout all phosphoglucose isomerases and was hypothesized to be the base catalyzing the isomerization reaction. In the present study, this conserved histidine, His311, of the enzyme from Bacillus stearothermophilus was subjected to mutational analysis, and the mutational effect on the inactivation kinetics by N-bromoacetylethanolamine phosphate was investigated. The substitution of His311 with alanine, asparagine, or glutamine resulted in the decrease of activity, in k(cat)/K(M), by a factor of 10(3), indicating the importance of this residue. N-bromoacetylethanolamine phosphate inactivated irreversibly the activity of wild-type phosphoglucose isomerase; however, His311 --> Ala became resistant to this inhibitor, indicating that His311 is located in the active site and is responsible for the inactivation of the enzyme by this active site-directed inhibitor. The pKa of His311 was estimated to be 6.31 according to the pH dependence of the inactivation. The proximity of this value with the pKa value of 6.35, determined from the pH dependence of k(cat)/K(M), supports a role of His311 as a general base in the catalysis. PMID:10595547

  17. Evaluating the Effects of Tri-Butyl Phosphate and Normal Paraffin Hydrocarbon in Simulated Low-Activity Waste Solution on Ion Exchange

    SciTech Connect

    Adu-Wusu, K.

    2003-05-13

    Ultrafiltration and ion exchange are among the pretreatment processes selected for the WTP at the Hanford Site. This study is the second part of a two-part study on Evaluating the Effects of Tri-Butyl Phosphate and Normal Paraffin Hydrocarbon in Simulated Low-Activity Waste Solution on Ultrafiltration and Ion Exchange.

  18. Effects of delayed delivery of dexamethasone-21-phosphate via subcutaneous microdialysis implants on macrophage activation in rats.

    PubMed

    Keeler, Geoffrey D; Durdik, Jeannine M; Stenken, Julie A

    2015-09-01

    Macrophage activation is of interest in the biomaterials field since macrophages with an M(Dex) characteristic phenotype, i.e., CD68(+)CD163(+), are believed to result in improved integration of the biomaterial as well as improved tissue remodeling and increased biomaterial longevity. To facilitate delivery of a macrophage modulator, dexamethasone-21-phosphate (Dex), microdialysis probes were subcutaneously implanted in male Sprague-Dawley rats. Dex localized delivery was delayed to the third day post implantation as a means to alter macrophage activation state at an implant site. To better elucidate the molecular mechanisms associated with M(Dex) macrophage activation, CCL2 was quantified in dialysates, gene expression ratios were determined from excised tissue surrounding the implant, histological analyses, and immunohistochemical analyses (CD68, CD163) were performed. Delayed Dex infusion resulted in the up-regulation of IL-6 at the transcript level in the tissue in contact with the microdialysis probe and decreased CCL2 concentrations collected in dialysates. Histological analyses showed increased cellular density as compared to controls in response to delayed Dex infusion. Dex delayed infusion resulted in an increased percentage of CD68(+)CD163(+), M(Dex), macrophages in the tissue surrounding the microdialysis probe as compared to probes that served as controls.

  19. Application of capillary enzyme micro-reactor in enzyme activity and inhibitors studies of glucose-6-phosphate dehydrogenase.

    PubMed

    Camara, Mohamed Amara; Tian, Miaomiao; Guo, Liping; Yang, Li

    2015-05-15

    In this study, we present an on-line measurement of enzyme activity and inhibition of Glucose-6-phosphate dehydrogenase (G6PDH) enzyme using capillary electrophoresis based immobilized enzyme micro-reactor (CE-based IMER). The IMER was prepared using a two-step protocol based on electrostatic assembly. The micro-reactor exhibited good stability and reproducibility for on-line assay of G6PDH enzyme. Both the activity as well as the inhibition of the G6PDH enzyme by six inhibitors, including three metals (Cu(2+), Pb(2+), Cd(2+)), vancomycin, urea and KMnO4, were investigated using on-line assay of the CE-based IMERs. The enzyme activity and inhibition kinetic constants were measured using the IMERs which were found to be consistent with those using traditional off-line enzyme assays. The kinetic mechanism of each inhibitor was also determined. The present study demonstrates the feasibility of using CE-based IMERs for rapid and efficient on-line assay of G6PDH, an important enzyme in the pentosephosphate pathway of human metabolism.

  20. Structural and computational dissection of the catalytic mechanism of the inorganic pyrophosphatase from Mycobacterium tuberculosis.

    PubMed

    Pratt, Andrew C; Dewage, Sajeewa W; Pang, Allan H; Biswas, Tapan; Barnard-Britson, Sandra; Cisneros, G Andrés; Tsodikov, Oleg V

    2015-10-01

    Family I inorganic pyrophosphatases (PPiases) are ubiquitous enzymes that are critical for phosphate metabolism in all domains of life. The detailed catalytic mechanism of these enzymes, including the identity of the general base, is not fully understood. We determined a series of crystal structures of the PPiase from Mycobacterium tuberculosis (Mtb PPiase) bound to catalytic metals, inorganic pyrophosphate (PPi; the reaction substrate) and to one or two inorganic phosphate ions (Pi; the reaction product), ranging in resolution from 1.85 to 3.30Å. These structures represent a set of major kinetic intermediates in the catalytic turnover pathway for this enzyme and suggest an order of association and dissociation of the divalent metals, the substrate and the two products during the catalytic turnover. The active site of Mtb PPiase exhibits significant structural differences from the well characterized Escherichia coli PPiase in the vicinity of the bound PPi substrate. Prompted by these differences, quantum mechanics/molecular mechanics (QM/MM) analysis yielded an atomic description of the hydrolysis step for Mtb PPiase and, unexpectedly, indicated that Asp89, rather than Asp54 that was proposed for E. coli PPiase, can abstract a proton from a water molecule to activate it for a nucleophilic attack on the PPi substrate. Mutagenesis studies of the key Asp residues of Mtb PPiase supported this mechanism. This combination of structural and computational analyses clarifies our understanding of the mechanism of family I PPiases and has potential utility for rational development of drugs targeting this enzyme.

  1. Phosphate-independent utilization of phosphonoacetic acid as sole phosphorus source by a psychrophilic strain of Geomyces pannorum P15.

    PubMed

    Klimek-Ochab, Magdalena

    2014-09-01

    A psychrophilic fungal strain of Geomyces pannorum P15 was screened for its ability to utilize a range of synthetic and natural organophosphonate compounds as the sole source of phosphorus, nitrogen, or carbon. Only phosphonoacetic acid served as a phosphorus source for microbial growth in phosphate-independent manner. Substrate metabolism did not lead to extracellular release of inorganic phosphate. No phosphonate metabolizing enzyme activity was detectable in cell-free extracts prepared from Geomyces biomass pregrown on 2 mmol/L phosphonoacetic acid.

  2. The development of geotextiles incorporating slow-release phosphate beads for the maintenance of oil degrading bacteria in permeable pavements.

    PubMed

    Spicer, G E; Lynch, D E; Coupe, S J

    2006-01-01

    The development of a self-fertilising geotextile mat designed to provide a sustained slow-release of required inorganic nutrients for the growth of oil degrading microorganisms in porous pavement systems (PPS) is reported. The system comprises a geotextile spun from polymer fibres containing spherical phosphated polymer beads that release phosphate upon contact with water at a desirable level for microbial growth. Initial results using model PPS have shown that the self-fertilising geotextile system works extremely effectively as increased microbial activity has been observed throughout the experiment, illustrating that the oil-degrading bacteria can effectively utilise this polymer composite as a suitable nutrient source.

  3. Phosphate control in dialysis

    PubMed Central

    Cupisti, Adamasco; Gallieni, Maurizio; Rizzo, Maria Antonietta; Caria, Stefania; Meola, Mario; Bolasco, Piergiorgio

    2013-01-01

    Prevention and correction of hyperphosphatemia is a major goal of chronic kidney disease–mineral and bone disorder (CKD–MBD) management, achievable through avoidance of a positive phosphate balance. To this aim, optimal dialysis removal, careful use of phosphate binders, and dietary phosphate control are needed to optimize the control of phosphate balance in well-nourished patients on a standard three-times-a-week hemodialysis schedule. Using a mixed diffusive–convective hemodialysis tecniques, and increasing the number and/or the duration of dialysis tecniques are all measures able to enhance phosphorus (P) mass removal through dialysis. However, dialytic removal does not equal the high P intake linked to the high dietary protein requirement of dialysis patients; hence, the use of intestinal P binders is mandatory to reduce P net intestinal absorption. Unfortunately, even a large dose of P binders is able to bind approximately 200–300 mg of P on a daily basis, so it is evident that their efficacy is limited in the case of an uncontrolled dietary P load. Hence, limitation of dietary P intake is needed to reach the goal of neutral phosphate balance in dialysis, coupled to an adequate protein intake. To this aim, patients should be informed and educated to avoid foods that are naturally rich in phosphate and also processed food with P-containing preservatives. In addition, patients should preferentially choose food with a low P-to-protein ratio. For example, patients could choose egg white or protein from a vegetable source. Finally, boiling should be the preferred cooking procedure, because it induces food demineralization, including phosphate loss. The integrated approach outlined in this article should be actively adapted as a therapeutic alliance by clinicians, dieticians, and patients for an effective control of phosphate balance in dialysis patients. PMID:24133374

  4. Phosphate control in dialysis.

    PubMed

    Cupisti, Adamasco; Gallieni, Maurizio; Rizzo, Maria Antonietta; Caria, Stefania; Meola, Mario; Bolasco, Piergiorgio

    2013-10-04

    Prevention and correction of hyperphosphatemia is a major goal of chronic kidney disease-mineral and bone disorder (CKD-MBD) management, achievable through avoidance of a positive phosphate balance. To this aim, optimal dialysis removal, careful use of phosphate binders, and dietary phosphate control are needed to optimize the control of phosphate balance in well-nourished patients on a standard three-times-a-week hemodialysis schedule. Using a mixed diffusive-convective hemodialysis tecniques, and increasing the number and/or the duration of dialysis tecniques are all measures able to enhance phosphorus (P) mass removal through dialysis. However, dialytic removal does not equal the high P intake linked to the high dietary protein requirement of dialysis patients; hence, the use of intestinal P binders is mandatory to reduce P net intestinal absorption. Unfortunately, even a large dose of P binders is able to bind approximately 200-300 mg of P on a daily basis, so it is evident that their efficacy is limited in the case of an uncontrolled dietary P load. Hence, limitation of dietary P intake is needed to reach the goal of neutral phosphate balance in dialysis, coupled to an adequate protein intake. To this aim, patients should be informed and educated to avoid foods that are naturally rich in phosphate and also processed food with P-containing preservatives. In addition, patients should preferentially choose food with a low P-to-protein ratio. For example, patients could choose egg white or protein from a vegetable source. Finally, boiling should be the preferred cooking procedure, because it induces food demineralization, including phosphate loss. The integrated approach outlined in this article should be actively adapted as a therapeutic alliance by clinicians, dieticians, and patients for an effective control of phosphate balance in dialysis patients.

  5. Phosphate Sorption in Water by Several cationic Polyer flocculants

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Although inorganic phosphate is an essential plant nutrient, elevated levels in surface waters may lead to adverse effects in the environment. These effects are attributed to runoff from rain or irrigation events that may cause the sorbed phosphate to move away from the application sites and move i...

  6. Soil phosphate stable oxygen isotopes across rainfall and bedrock gradients.

    PubMed

    Angert, Alon; Weiner, Tal; Mazeh, Shunit; Sternberg, Marcelo

    2012-02-21

    The stable oxygen isotope compositions of soil phosphate (δ(18)O(p)) were suggested recently to be a tracer of phosphorus cycling in soils and plants. Here we present a survey of bioavailable (resin-extractable or resin-P) inorganic phosphate δ(18)O(p) across natural and experimental rainfall gradients, and across soil formed on sedimentary and igneous bedrock. In addition, we analyzed the soil HCl-extractable inorganic δ(18)O(p), which mainly represents calcium-bound inorganic phosphate. The resin-P values were in the range 14.5-21.2‰. A similar range, 15.6-21.3‰, was found for the HCl-extractable inorganic δ(18)O(p), with the exception of samples from a soil of igneous origin that show lower values, 8.2-10.9‰, which indicate that a large fraction of the inorganic phosphate in this soil is still in the form of a primary mineral. The available-P δ(18)O(p) values are considerably higher than the values we calculated for extracellular hydrolysis of organic phosphate, based on the known fractionation from lab experiments. However, these values are close to the values expected for enzymatic-mediated phosphate equilibration with soil-water. The possible processes that can explain this observation are (1) extracellular equilibration of the inorganic phosphate in the soil; (2) fractionations in the soil are different than the ones measured at the lab; (3) effect of fractionation during uptake; and (4) a flux of intercellular-equilibrated inorganic phosphate from the soil microbiota, which is considerably larger than the flux of hydrolyzed organic-P.

  7. Distinguishing Biotic from Abiotic Phosphate Oxygen Isotopic Signatures

    NASA Astrophysics Data System (ADS)

    Blake, R.; Moyer, C.; Colman, A.; Liang, Y.; Dogru, D.

    2006-05-01

    On earth, phosphate has a strong biological oxygen isotope signature due to its concentration and intense cycling by living organisms as an essential nutrient. Phosphate does not undergo oxygen isotope exchange with water at low temperature without enzymatic catalysis, making the oxygen isotope ratio (18O/16O) of phosphate, δ18OP, an attractive biosignature in the search for early and extraterrestrial life. Recent laboratory and field studies have demonstrated that the δ18OP value of dissolved inorganic phosphate (PO4) records specific microbial activity and enzymatic reaction pathways in both laboratory cultures and natural waters/sediments (Blake et al., 2005; Colman et al 2005; Liang and Blake, 2005). Phosphate oxygen isotope biosignatures may be distinguished from abiotic signatures by: (1) evaluating the degree of temperature-dependent PO4-water oxygen isotope exchange in aqueous systems and deviation from equilibrium; and (2) evolution from an abiotic P reservoir signature towards a biotic P reservoir signature. Important abiotic processes potentially affecting phosphate δ18OP values include dissolution/precipitation, adsorption/desorption, recrystallization of PO4 mineral phases, diagenesis and metamorphism. For most of these processes, the recording, retention and alteration of δ18OP biosignatures have not been evaluated. Deep-sea hydrothermal vent fields are an ideal system in which to study the preservation and alteration of δ18OP biosignatures, as well as potential look-alikes produced by heat-promoted PO4 -water oxygen isotope exchange. Results from recent studies of δ18OP biosignatures in hydrothermal deposits near 9 and 21 degrees N. EPR and at Loihi seamount will be presented.

  8. Alkaline Phosphatase Revisited:  Hydrolysis of Alkyl Phosphates (†).

    PubMed

    O'Brien, Patrick J; Herschlag, Daniel

    2002-03-05

    Escherichia coli alkaline phosphatase (AP) is the prototypical two metal ion catalyst with two divalent zinc ions bound ∼4 Å apart in the active site. Studies spanning half a century have elucidated many structural and mechanistic features of this enzyme, rendering it an attractive model for investigating the potent catalytic power of bimetallic centers. Unfortunately, fundamental mechanistic features have been obscured by limitations with the standard assays. These assays generate concentrations of inorganic phosphate (Pi) in excess of its inhibition constant (K i ≈ 1 μM). This tight binding by Pi has affected the majority of published kinetic constants. Furthermore, binding limits k cat/K m for reaction of p-nitrophenyl phosphate, the most commonly employed substrate. We describe a sensitive (32)P-based assay for hydrolysis of alkyl phosphates that avoids the complication of product inhibition. We have revisited basic mechanistic features of AP with these alkyl phosphate substrates. The results suggest that the chemical step for phosphorylation of the enzyme limits k cat/K m. The pH-rate profile and additional results suggest that the serine nucleophile is active in its anionic form and has a pK a of ≤5.5 in the free enzyme. An inactivating pK a of 8.0 is observed for binding of both substrates and inhibitors, and we suggest that this corresponds to ionization of a zinc-coordinated water molecule. Counter to previous suggestions, inorganic phosphate dianion appears to bind to the highly charged AP active site at least as strongly as the trianion. The dependence of k cat/K m on the pK a of the leaving group follows a Brønsted correlation with a slope of βlg = -0.85 ± 0.1, differing substantially from the previously reported value of -0.2 obtained from data with a less sensitive assay. This steep leaving group dependence is consistent with a largely dissociative transition state for AP-catalyzed hydrolysis of phosphate monoesters. The new (32)P

  9. Sphingosine 1-phosphate is a ligand for peroxisome proliferator-activated receptor-γ that regulates neoangiogenesis.

    PubMed

    Parham, Kate A; Zebol, Julia R; Tooley, Katie L; Sun, Wai Y; Moldenhauer, Lachlan M; Cockshell, Michaelia P; Gliddon, Briony L; Moretti, Paul A; Tigyi, Gabor; Pitson, Stuart M; Bonder, Claudine S

    2015-09-01

    Sphingosine 1-phosphate (S1P) is a bioactive lipid that can function both extracellularly and intracellularly to mediate a variety of cellular processes. Using lipid affinity matrices and a radiolabeled lipid binding assay, we reveal that S1P directly interacts with the transcription factor peroxisome proliferator-activated receptor (PPAR)γ. Herein, we show that S1P treatment of human endothelial cells (ECs) activated a luciferase-tagged PPARγ-specific gene reporter by ∼12-fold, independent of the S1P receptors. More specifically, in silico docking, gene reporter, and binding assays revealed that His323 of the PPARγ ligand binding domain is important for binding to S1P. PPARγ functions when associated with coregulatory proteins, and herein we identify that peroxisome proliferator-activated receptor-γ coactivator 1 (PGC1)β binds to PPARγ in ECs and their progenitors (nonadherent endothelial forming cells) and that the formation of this PPARγ:PGC1β complex is increased in response to S1P. ECs treated with S1P selectively regulated known PPARγ target genes with PGC1β and plasminogen-activated inhibitor-1 being increased, no change to adipocyte fatty acid binding protein 2 and suppression of CD36. S1P-induced in vitro tube formation was significantly attenuated in the presence of the PPARγ antagonist GW9662, and in vivo application of GW9662 also reduced vascular development in Matrigel plugs. Interestingly, activation of PPARγ by the synthetic ligand troglitazone also reduced tube formation in vitro and in vivo. To support this, Sphk1(-/-)Sphk2(+/-) mice, with low circulating S1P levels, demonstrated a similar reduction in vascular development. Taken together, our data reveal that the transcription factor, PPARγ, is a bona fide intracellular target for S1P and thus suggest that the S1P:PPARγ:PGC1β complex may be a useful target to manipulate neovascularization.

  10. The level of glucose-6-phosphate dehydrogenase activity strongly influences xylose fermentation and inhibitor sensitivity in recombinant Saccharomyces cerevisiae strains.

    PubMed

    Jeppsson, Marie; Johansson, Björn; Jensen, Peter Ruhdal; Hahn-Hägerdal, Bärbel; Gorwa-Grauslund, Marie F

    2003-11-01

    Disruption of the ZWF1 gene encoding glucose-6-phosphate dehydrogenase (G6PDH) has been shown to reduce the xylitol yield and the xylose consumption in the xylose-utilizing recombinant Saccharomyces cerevisiae strain TMB3255. In the present investigation we have studied the influence of different production levels of G6PDH on xylose fermentation. We used a synthetic promoter library and the copper-regulated CUP1 promoter to generate G6PDH-activities between 0% and 179% of the wild-type level. G6PDH-activities of 1% and 6% of the wild-type level resulted in 2.8- and 5.1-fold increase in specific xylose consumption, respectively, compared with the ZWF1-disrupted strain. Both strains exhibited decreased xylitol yields (0.13 and 0.19 g/g xylose) and enhanced ethanol yields (0.36 and 0.34 g/g xylose) compared with the control strain TMB3001 (0.29 g xylitol/g xylose, 0.31 g ethanol/g xylose). Cytoplasmic transhydrogenase (TH) from Azotobacter vinelandii has previously been shown to transfer NADPH and NAD(+) into NADP(+) and NADH, and TH-overproduction resulted in lower xylitol yield and enhanced glycerol yield during xylose utilization. Strains with low G6PDH-activity grew slower in a lignocellulose hydrolysate than the strain with wild-type G6PDH-activity, which suggested that the availability of intracellular NADPH correlated with tolerance towards lignocellulose-derived inhibitors. Low G6PDH-activity strains were also more sensitive to H(2)O(2) than the control strain TMB3001.

  11. Organic materials as templates for the formation of mesoporous inorganic materials and ordered inorganic nanocomposites

    NASA Astrophysics Data System (ADS)

    Ziegler, Christopher R.

    Hierarchically structured inorganic materials are everywhere in nature. From unicellular aquatic algae such as diatoms to the bones and/or cartilage that comprise the skeletal systems of vertebrates. Complex mechanisms involving site-specific chemistries and precision kinetics are responsible for the formation of such structures. In the synthetic realm, reproduction of even the most basic hierarchical structure effortlessly produced in nature is difficult. However, through the utilization of self-assembling structures or "templates", such as polymers or amphiphilic surfactants, combined with some favorable interaction between a chosen inorganic, the potential exists to imprint an inorganic material with a morphology dictated via synthetic molecular self-assembly. In doing so, a very basic hierarchical structure is formed on the angstrom and nanometer scales. The work presented herein utilizes the self-assembly of either surfactants or block copolymers with the desired inorganic or inorganic precursor to form templated inorganic structures. Specifically, mesoporous silica spheres and copolymer directed calcium phosphate-polymer composites were formed through the co-assembly of an organic template and a precursor to form the desired mesostructured inorganic. For the case of the mesoporous silica spheres, a silica precursor was mixed with cetyltrimethylammonium bromide and cysteamine, a highly effective biomimetic catalyst for the conversion of alkoxysilanes to silica. Through charge-based interactions between anionic silica species and the micelle-forming cationic surfactant, ordered silica structures resulted. The incorporation of a novel, effective catalyst was found to form highly condensed silica spheres for potential application as catalyst supports or an encapsulation media. Ordered calcium phosphate-polymer composites were formed using two routes. Both routes take advantage of hydrogen bonding and ionic interactions between the calcium and phosphate precursors

  12. Efficient production of 2-deoxyribose 5-phosphate from glucose and acetaldehyde by coupling of the alcoholic fermentation system of Baker's yeast and deoxyriboaldolase-expressing Escherichia coli.

    PubMed

    Horinouchi, Nobuyuki; Ogawa, Jun; Kawano, Takako; Sakai, Takafumi; Saito, Kyota; Matsumoto, Seiichiro; Sasaki, Mie; Mikami, Yoichi; Shimizu, Sakayu

    2006-06-01

    2-Deoxyribose 5-phosphate production through coupling of the alcoholic fermentation system of baker's yeast and deoxyriboaldolase-expressing Escherichia coli was investigated. In this process, baker's yeast generates fructose 1,6-diphosphate from glucose and inorganic phosphate, and then the E. coli convert the fructose 1,6-diphosphate into 2-deoxyribose 5-phosphate via D-glyceraldehyde 3-phosphate. Under the optimized conditions with toluene-treated yeast cells, 356 mM (121 g/l) fructose 1,6-diphosphate was produced from 1,111 mM glucose and 750 mM potassium phosphate buffer (pH 6.4) with a catalytic amount of AMP, and the reaction supernatant containing the fructose 1,6-diphosphate was used directly as substrate for 2-deoxyribose 5-phosphate production with the E. coli cells. With 178 mM enzymatically prepared fructose 1,6-diphosphate and 400 mM acetaldehyde as substrates, 246 mM (52.6 g/l) 2-deoxyribose 5-phosphate was produced. The molar yield of 2-deoxyribose 5-phosphate as to glucose through the total two step reaction was 22.1%. The 2-deoxyribose 5-phosphate produced was converted to 2-deoxyribose with a molar yield of 85% through endogenous or exogenous phosphatase activity.

  13. Mild water stress of Phaseolus vulgaris plants leads to reduced starch synthesis and extractable sucrose phosphate synthase activity

    SciTech Connect

    Vassey, T.L.; Sharkey, T.D. )

    1989-04-01

    Mild water stress, on the order of {minus}1.0 megapascals xylem water potential, can reduce the rate of photosynthesis and eliminate the inhibition of photosynthesis caused by O{sub 2} in water-stress-sensitive plants such as Phaseolus vulgaris. To investigate the lack of O{sub 2} inhibition of photosynthesis, we measured stromal and cytosolic fructose-1,6-bisphosphatase, sucrose phosphate synthase, and partitioning of newly fixed carbon between starch and sucrose before, during, and after mild water stress. The extractable activity of the fructose bisphosphatases was unaffected by mild water stress. The extractable activity of SPS was inhibited by more than 60% in plants stressed to water potentials of {minus}0.9 megapascals. Water stress caused a decline in the starch/sucrose partitioning ratio indicating that starch synthesis was inhibited more than sucrose synthesis. We conclude that the reduced rate of photosynthesis during water stress is caused by stomatal closure, and that the restriction of CO{sub 2} supply caused by stomatal closure leads to a reduction in the capacity for both starch and sucrose synthesis. This causes the reduced O{sub 2} inhibition and abrupt CO{sub 2} saturation of photosynthesis.

  14. The Effect of Covalently Immobilized FGF-2 on Biphasic Calcium Phosphate Bone Substitute on Enhanced Biological Compatibility and Activity.

    PubMed

    Moon, Kyung-Suk; Choi, Eun-Joo; Oh, Seunghan; Kim, Sungtae

    2015-01-01

    The purpose of this research was to covalently graft fibroblast growth factor 2 (FGF-2) onto biphasic calcium phosphate (BCP) via a bifunctional cross-linker technique and to estimate the optimal dose of FGF-2 resulting in the best osteogenic differentiation of human mesenchymal stem cells (hMSCs). SEM observation revealed that the surface of the 100 ng FGF-2 coated BCP was completely covered with the nanoparticles expected to be from the silane coupling agent. XRD, FT-IR, and XPS analysis showed that silane treatment, bifunctional cross-linker coating, and FGF-2 covalent grafts were conducted successfully without deforming the crystalline structure of BCP. An MTT assay demonstrated that FGF-2 coated BCP had good biocompatibility, regardless of the concentration of FGF-2, after 24 or 48 h of incubation. An alkaline phosphatase (ALP) activity assay (14 days of incubation) and the ALP gene expression level of real-time PCR analysis (7 days of incubation) revealed that 50, 100, and 200 ng FGF-2 coated BCP induced the highest activities among all experimental groups and control group (P < 0.05). Thus, low concentrations of FGF-2 facilitated excellent osteogenesis and were effective at enhancing osteogenic potential. Also, the bifunctional cross-linker technique is expected to be a more feasible way to induce osteogenic differentiation while minimizing the risk of FGF-2 overdose.

  15. Low red blood cell glutathione reductase and pyridoxine phosphate oxidase activities not related to dietary riboflavin: selection by malaria?

    PubMed

    Anderson, B B; Giuberti, M; Perry, G M; Salsini, G; Casadio, I; Vullo, C

    1993-05-01

    This study was designed to confirm that low dietary riboflavin does not contribute to the flavin-deficient red blood cells commonly found in subjects in Ferrara Province, northern Italy. In this area it is primarily an inherited characteristic believed to have been selected for by malaria, which was endemic from the 12th century. In parallel with assessment of daily riboflavin intake (DRI), flavin adenine dinucleotide-dependent glutathione reductase (EGR) and flavin mononucleotide-dependent pyridoxine phosphate oxidase (PPO) were measured in beta-thalassemic heterozygotes, their normal relatives, and normal spouses (representative of the normal population). In all of these groups there is a high incidence of deficiency of these flavin enzymes. We found that the majority had an adequate riboflavin intake and there was no significant correlation of EGR and PPO activities with DRI. Thus, interpretation of low EGR activity is discussed with reference to studies of EGR done to detect nutritional riboflavin deficiency in countries where there is malnutrition and endemic malaria.

  16. Change of crystallinity and mineral composition of fly ash with mechanical and chemical activation for the improvement of phosphate uptake.

    PubMed

    Liang, Zhu; He, XiaoJia; Ni, JinRen

    2010-10-01

    A detailed investigation of the development of the mineral composition and crystallinity of fly ash (FA) activated by an integrated process, as well as the relation between that development and phosphate uptake (PU) in solution, was conducted. This process, consisting of pretreatment (mechanical milling), alkali fusion (sodium hydroxide) at 550 °C and acid attack (3 mol L⁻¹ sulfuric acid), exhibited a remarkable activation effect. One-hour grinding could enhance PU from 0.67 to 1.66 mg PO₄³⁻-P g⁻¹ FA, and then under the optimum chemical conditions with the ratio of NaOH to FA of 0.5 g g⁻¹ and that of H₂SO₄ to FA of 3 mL g⁻¹, the PU was further improved to 7.14 mg g⁻¹. Results also indicated that the PU performance was closely linked with the crystallinity and mineral composition of FA, that is, the treated material with the lowest crystallinity and least crystal phase could achieve the highest PU. For the purpose of destroying original minerals in raw FA farthest and avoiding the production of new crystals, therefore, control on the ratio of NaOH to FA and that of acid to FA was very important during the chemical treatment.

  17. Organic-Inorganic Composites Toward Biomaterial Application.

    PubMed

    Miyazaki, Toshiki; Sugawara-Narutaki, Ayae; Ohtsuki, Chikara

    2015-01-01

    Bioactive ceramics are known to exhibit specific biological affinities and are able to show direct integration with surrounding bone when implanted in bony defects. However, their inadequate mechanical properties, such as low fracture toughness and high Young's modulus in comparison to natural bone, limit their clinical application. Bone is a kind of organic-inorganic composite where apatite nanocrystals are precipitated onto collagen fibre networks. Thus, one way to address these problems is to mimic the natural composition of bone by using bioactive ceramics via material designs based on organic-inorganic composites. In this chapter, the current research on the development of the various organic-inorganic composites designed for biomaterial applications has been reviewed. Various compounds such as calcium phosphate, calcium sulphate and calcium carbonate can be used for the inorganic phases to design composites with the desired mechanical and biological properties of bone. Not only classical mechanical mixing but also coating of the inorganic phase in aqueous conditions is available for the fabrication of such composites. Organic modifications using various polymers enable the control of the crystalline structure of the calcium carbonate in the composites. These approaches on the fabrication of organic-inorganic composites provide important options for biomedical materials with novel functions.

  18. High affinity of acid phosphatase encoded by PHO3 gene in Saccharomyces cerevisiae for thiamin phosphates.

    PubMed

    Nosaka, K

    1990-02-09

    The enzymatic properties of acid phosphatase (orthophosphoric-monoester phosphohydrolase, EC 3.1.3.2) encoded by PHO3 gene in Saccharomyces cerevisiae, which is repressed by thiamin and has thiamin-binding activity at pH 5.0, were investigated to study physiological functions. The following results led to the conclusion that thiamin-repressible acid phosphatase physiologically catalyzes the hydrolysis of thiamin phosphates in the periplasmic space of S. cerevisiae, thus participating in utilization of the thiamin moiety of the phosphates by yeast cells: (a) thiamin-repressible acid phosphatase showed Km values of 1.6 and 1.7 microM at pH 5.0 for thiamin monophosphate and thiamin pyrophosphate, respectively. These Km values were 2-3 orders of magnitude lower than those (0.61 and 1.7 mM) for p-nitrophenyl phosphate; (b) thiamin exerted remarkable competitive inhibition in the hydrolysis of thiamin monophosphate (Ki 2.2 microM at pH 5.0), whereas the activity for p-nitrophenyl phosphate was slightly affected by thiamin; (c) the inhibitory effect of inorganic phosphate, which does not repress the thiamin-repressible enzyme, on the hydrolysis of thiamin monophosphate was much smaller than that of p-nitrophenyl phosphate. Moreover, the modification of thiamin-repressible acid phosphatase of S. cerevisiae with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide resulted in the complete loss of thiamin-binding activity and the Km value of the modified enzyme for thiamin monophosphate increased nearly to the value of the native enzyme for p-nitrophenyl phosphate. These results also indicate that the high affinity of the thiamin-repressible acid phosphatase for thiamin phosphates is due to the thiamin-binding properties of this enzyme.

  19. Phytosphingosine-phosphate is a signal for AtMPK6 activation and Arabidopsis response to chilling.

    PubMed

    Dutilleul, Christelle; Benhassaine-Kesri, Ghouziel; Demandre, Chantal; Rézé, Nathalie; Launay, Alban; Pelletier, Sandra; Renou, Jean-Pierre; Zachowski, Alain; Baudouin, Emmanuel; Guillas, Isabelle

    2012-04-01

    • Long-chain bases (LCBs) are pleiotropic sphingolipidic signals in eukaryotes. We investigated the source and function of phytosphingosine-1-phosphate (PHS-P), a phospho-LCB rapidly and transiently formed in Arabidopsis thaliana on chilling. • PHS-P was analysed by thin-layer chromatography following in vivo metabolic radiolabelling. Pharmacological and genetic approaches were used to identify the sphingosine kinase isoforms involved in cold-responsive PHS-P synthesis. Gene expression, mitogen-activated protein kinase activation and growth phenotypes of three LCB kinase mutants (lcbk1, sphk1 and lcbk2) were studied following cold exposure. • Chilling provoked the rapid and transient formation of PHS-P in Arabidopsis cultured cells and plantlets. Cold-evoked PHS-P synthesis was reduced by LCB kinase inhibitors and abolished in the LCB kinase lcbk2 mutant, but not in lcbk1 and sphk1 mutants. lcbk2 presented a constitutive AtMPK6 activation at 22°C. AtMPK6 activation was also triggered by PHS-P treatment independently of PHS/PHS-P balance. lcbk2 mutants grew comparably with wild-type plants at 22 and 4°C, but exhibited a higher root growth at 12°C, correlated with an altered expression of the cold-responsive DELLA gene RGL3. • Together, our data indicate a function for LCBK2 in planta. Furthermore, they connect PHS-P formation with plant response to cold, expanding the field of LCB signalling in plants.

  20. Remote Monitoring, Inorganic Monitoring

    EPA Science Inventory

    This chapter provides an overview of applicability, amenability, and operating parameter ranges for various inorganic parameters:this chapter will also provide a compilation of existing and new online technologies for determining inorganic compounds in water samples. A wide vari...

  1. Intracellular activity of tedizolid phosphate and ACH-702 versus Mycobacterium tuberculosis infected macrophages

    PubMed Central

    2014-01-01

    Background Due to the emergency of multidrug-resistant strains of Mycobacterium tuberculosis, is necessary the evaluation of new compounds. Findings Tedizolid, a novel oxazolidinone, and ACH-702, a new isothiazoloquinolone, were tested against M. tuberculosis infected THP-1 macrophages. These two compounds significantly decreased the number of intracellular mycobacteria at 0.25X, 1X, 4X and 16X the MIC value. The drugs were tested either in nanoparticules or in free solution. Conclusion Tedizolid and ACH-702 have a good intracellular killing activity comparable to that of rifampin or moxifloxacin. PMID:24708819

  2. Autophagy and endosomal trafficking inhibition by Vibrio cholerae MARTX toxin phosphatidylinositol-3-phosphate-specific phospholipase A1 activity

    PubMed Central

    Agarwal, Shivani; Kim, Hyunjin; Chan, Robin B.; Agarwal, Shivangi; Williamson, Rebecca; Cho, Wonhwa; Paolo, Gilbert D.; Satchell, Karla J. F.

    2015-01-01

    Vibrio cholerae, responsible for acute gastroenteritis secretes a large multifunctional-autoprocessing repeat-in-toxin (MARTX) toxin linked to evasion of host immune system, facilitating colonization of small intestine. Unlike other effector domains of the multifunctional toxin that target cytoskeleton, the function of alpha-beta hydrolase (ABH) remained elusive. This study demonstrates that ABH is an esterase/lipase with catalytic Ser–His–Asp triad. ABH binds with high affinity to phosphatidylinositol-3-phosphate (PtdIns3P) and cleaves the fatty acid in PtdIns3P at the sn1 position in vitro making it the first PtdIns3P-specific phospholipase A1 (PLA1). Expression of ABH in vivo reduces intracellular PtdIns3P levels and its PtdIns3P-specific PLA1 activity blocks endosomal and autophagic pathways. In accordance with recent studies acknowledging the potential of extracellular pathogens to evade or exploit autophagy to prevent their clearance and facilitate survival, this is the first report highlighting the role of ABH in inhibiting autophagy and endosomal trafficking induced by extracellular V. cholerae. PMID:26498860

  3. Activation of mitochondria-mediated apoptotic pathway in tri-ortho-cresyl phosphate-induced delayed neuropathy.

    PubMed

    Zou, Chaoshuang; Kou, Ruirui; Gao, Yuan; Xie, Keqin; Song, Fuyong

    2013-06-01

    Previous studies suggest that abnormal neurons death has been implicated in organophosphate-induced delayed neuropathy (OPIDN). However, the precise mechanism of neuronal death in OPIDN remains largely unknown. In this study, adult hens were treated with a dosage of 750 mg/kg tri-ortho-cresyl phosphate (TOCP) by gavage, and then sacrificed on the time-points of 1, 5, 10, and 21 days after dosing TOCP, respectively. The apoptotic change of spinal cord neurons induced by TOCP was examined, and the role of mitochondria-mediated apoptosis of neurons during OPIDN was investigated. TUNEL assays showed that apoptotic neurons in hen spinal cords began to appear on day 5 following TOCP exposure. Immunohistochemistry and western blot analysis revealed a translocation of cytochrome C from mitochondria to cytoplasm after dosing TOCP. Moreover, the level of Bcl-2, Bcl-xl, Pro-caspase3 and Pro-caspase9 in hen spinal cord was significantly decreased, whereas that of Bax and cleaved-PARP was significantly elevated. Taken together, these findings indicate that the administration of TOCP can induce neuron apoptosis in hen spinal cords, which might be mediated by the activation of mitochondrial apoptotic pathway.

  4. Wnt5a Increases the Glycolytic Rate and the Activity of the Pentose Phosphate Pathway in Cortical Neurons

    PubMed Central

    Cisternas, Pedro; Salazar, Paulina; Silva-Álvarez, Carmen; Barros, L. Felipe

    2016-01-01

    In the last few years, several reports have proposed that Wnt signaling is a general metabolic regulator, suggesting a role for this pathway in the control of metabolic flux. Wnt signaling is critical for several neuronal functions, but little is known about the correlation between this pathway and energy metabolism. The brain has a high demand for glucose, which is mainly used for energy production. Neurons use energy for highly specific processes that require a high energy level, such as maintaining the electrical potential and synthesizing neurotransmitters. Moreover, an important metabolic impairment has been described in all neurodegenerative disorders. Despite the key role of glucose metabolism in the brain, little is known about the cellular pathways involved in regulating this process. We report here that Wnt5a induces an increase in glucose uptake and glycolytic rate and an increase in the activity of the pentose phosphate pathway; the effects of Wnt5a require the intracellular generation of nitric oxide. Our data suggest that Wnt signaling stimulates neuronal glucose metabolism, an effect that could be important for the reported neuroprotective role of Wnt signaling in neurodegenerative disorders. PMID:27688915

  5. Orbital fluid shear stress promotes osteoblast metabolism, proliferation and alkaline phosphates activity in vitro.

    PubMed

    Aisha, M D; Nor-Ashikin, M N K; Sharaniza, A B R; Nawawi, H; Froemming, G R A

    2015-09-10

    Prolonged disuse of the musculoskeletal system is associated with reduced mechanical loading and lack of anabolic stimulus. As a form of mechanical signal, the multidirectional orbital fluid shear stress transmits anabolic signal to bone forming cells in promoting cell differentiation, metabolism and proliferation. Signals are channeled through the cytoskeleton framework, directly modifying gene and protein expression. For that reason, we aimed to study the organization of Normal Human Osteoblast (NHOst) cytoskeleton with regards to orbital fluid shear (OFS) stress. Of special interest were the consequences of cytoskeletal reorganization on NHOst metabolism, proliferation, and osteogenic functional markers. Cells stimulated at 250 RPM in a shaking incubator resulted in the rearrangement of actin and tubulin fibers after 72 h. Orbital shear stress increased NHOst mitochondrial metabolism and proliferation, simultaneously preventing apoptosis. The ratio of RANKL/OPG was reduced, suggesting that orbital shear stress has the potential to inhibit osteoclastogenesis and osteoclast activity. Increase in ALP activity and OCN protein production suggests that stimulation retained osteoblast function. Shear stress possibly generated through actin seemed to hold an anabolic response as osteoblast metabolism and functional markers were enhanced. We hypothesize that by applying orbital shear stress with suitable magnitude and duration as a non-drug anabolic treatment can help improve bone regeneration in prolonged disuse cases.

  6. Distance between two active-site lysines of ribulosebis-phosphate carboxylase/oxygenase

    SciTech Connect

    Lee, E.H.; Hartman, F.C.

    1986-05-01

    Previous affinity labeling studies and comparative sequence analyses have identified two different lysines at the active site of the title enzyme (Lys-166 and Lys-329 in the Rhodospirillum rubrum enzyme and Lys-175 and Lys-334 in the spinach enzyme). Because the two lysines are mutually exclusive to various reagents, they appear to be in proximity. To challenge this postulate, the authors have explored the reactions of the R. rubrum enzyme (a homodimer) with chemical cross-linking agents. 4,4'-Diisothiocyano-2,2'-disulfonate stilbene, which spans 12 A, rapidly inactivates the enzyme with protection afforded by the competitive inhibitor 2-carboxyribitol-1,5-bisphosphate. The inactivated enzyme was subjected to gel filtration in the presence of urea to remove material arising from intersubunit or intermolecular cross-linking. The monomeric fraction was digested with trypsin; inspection of the digest by HPLC revealed that over-half of the incorporated reagent was associated with a single peptide. This peptide was purified by successive ion-exchange chromatography and gel filtration. The amino acid composition and sequence of the purified peptide demonstrated that it is comprised of two chains, encompassing position 149-168 and 314-337 of the original protein subunit and connected by a cross-link between Lys-166 and Lys-329. Thus, the two active-site lysines can be juxtaposed only 12 A apart.

  7. Characterization of the Proteomic Profiles of the Brown Tide Alga Aureoumbra lagunensis under Phosphate- and Nitrogen-Limiting Conditions and of Its Phosphate Limitation-Specific Protein with Alkaline Phosphatase Activity

    PubMed Central

    Sun, Ming-Ming; Sun, Jin; Qiu, Jian-Wen; Jing, Hongmei

    2012-01-01

    The persistent bloom of the brown tide alga Aureoumbra lagunensis has been reported in coastal embayments along southern Texas, but the molecular mechanisms that sustain such algal bloom are unknown. We compared the proteome and physiological parameters of A. lagunensis grown in phosphate (P)-depleted, P- and nitrogen (N)-depleted, and nutrient-replete cultures. For the proteomic analysis, samples from three conditions were subjected to two-dimensional electrophoresis and tandem mass spectrometry analysis. Because of the paucity of genomic resources in this species, a de novo cross-species protein search was used to identify the differentially expressed proteins, which revealed their involvement in several key biological processes, such as chlorophyll synthesis, antioxidative protection, and protein degradation, suggesting that A. lagunensis may adopt intracellular nutrient compensation, extracellular organic nutrient regeneration, and damage protection to thrive in P-depleted environments. A highly abundant P limitation-specific protein, tentatively identified as a putative alkaline phosphatase, was further characterized by enzyme activity assay on nondenaturing gel and confocal microscopy, which confirmed that this protein has alkaline phosphatase activity, is a cytoplasmic protein, and is closely associated with the cell membrane. The abundance, location, and functional expression of this alkaline phosphatase all indicate the importance of organic P utilization for A. lagunensis under P limitation and the possible role of this alkaline phosphatase in regenerating phosphate from extra- or intracellular organic phosphorus. PMID:22247172

  8. Orbital fluid shear stress promotes osteoblast metabolism, proliferation and alkaline phosphates activity in vitro

    SciTech Connect

    Aisha, M.D.; Nor-Ashikin, M.N.K.; Sharaniza, A.B.R.; Nawawi, H.; Froemming, G.R.A.

    2015-09-10

    Prolonged disuse of the musculoskeletal system is associated with reduced mechanical loading and lack of anabolic stimulus. As a form of mechanical signal, the multidirectional orbital fluid shear stress transmits anabolic signal to bone forming cells in promoting cell differentiation, metabolism and proliferation. Signals are channeled through the cytoskeleton framework, directly modifying gene and protein expression. For that reason, we aimed to study the organization of Normal Human Osteoblast (NHOst) cytoskeleton with regards to orbital fluid shear (OFS) stress. Of special interest were the consequences of cytoskeletal reorganization on NHOst metabolism, proliferation, and osteogenic functional markers. Cells stimulated at 250 RPM in a shaking incubator resulted in the rearrangement of actin and tubulin fibers after 72 h. Orbital shear stress increased NHOst mitochondrial metabolism and proliferation, simultaneously preventing apoptosis. The ratio of RANKL/OPG was reduced, suggesting that orbital shear stress has the potential to inhibit osteoclastogenesis and osteoclast activity. Increase in ALP activity and OCN protein production suggests that stimulation retained osteoblast function. Shear stress possibly generated through actin seemed to hold an anabolic response as osteoblast metabolism and functional markers were enhanced. We hypothesize that by applying orbital shear stress with suitable magnitude and duration as a non-drug anabolic treatment can help improve bone regeneration in prolonged disuse cases. - Highlights: • OFS stress transmits anabolic signals to osteoblasts. • Actin and tubulin fibers are rearranged under OFS stress. • OFS stress increases mitochondrial metabolism and proliferation. • Reduced RANKL/OPG ratio in response to OFS inhibits osteoclastogenesis. • OFS stress prevents apoptosis and stimulates ALP and OCN.

  9. Conformational Changes in Orotidine 5’-Monophosphate Decarboxylase: A Structure-Based Explanation for How the 5’-Phosphate Group Activates the Enzyme†

    PubMed Central

    Desai, Bijoy J.; Wood, McKay; Fedorov, Alexander A.; Fedorov, Elena V.; Goryanova, Bogdana; Amyes, Tina L.; Richard, John P.; Almo, Steven C.; Gerlt, John A.

    2012-01-01

    The binding of a ligand to orotidine 5’-monophosphate decarboxylase (OMPDC) is accompanied by a conformational change from an open, inactive conformation (Eo) to a closed, active conformation (Ec). As the substrate traverses the reaction coordinate to form the stabilized vinyl carbanion/carbene intermediate, interactions are enforced that destabilize the carboxylate group of the substrate as well as stabilize the intermediate (in the Ec•S‡ complex). Focusing on the OMPDC from Methanothermobacter thermautotrophicus, the “remote” 5’-phosphate group of the substrate activates the enzyme 2.4 × 108-fold; the activation is equivalently described by an intrinsic binding energy (IBE) of 11.4 kcal/mol. We studied residues in the activation that 1) directly contact the 5’-phosphate group; 2) participate in a hydrophobic cluster near the base of the active site loop that sequesters the bound substrate from solvent; and 3) form hydrogen-bonding interactions across the interface between the “mobile” and “fixed” half-barrel domains of the (β/α8-barrel structure. Our data support a model in which the IBE provided by the 5’-phosphate group is used to enable interactions both near the N-terminus of the active site loop and across the domain interface that stabilize both the Ec•S and Ec•S‡ complexes relative to the Eo•S complex. The conclusion that the IBE of the 5’-phosphate group provides stabilization of both the Ec•S and Ec•S‡ complexes, not just the Ec•S‡ complex, is central to understanding the structural origins of enzymatic catalysis as well as the requirements for the de novo design of enzymes that catalyze novel reactions. PMID:23030629

  10. Phosphate Mines, Jordan

    NASA Technical Reports Server (NTRS)

    2008-01-01

    Jordan's leading industry and export commodities are phosphate and potash, ranked in the top three in the world. These are used to make fertilizer. The Jordan Phosphate Mines Company is the sole producer, having started operations in 1935. In addition to mining activities, the company produces phosphoric acid (for fertilizers, detergents, pharmaceuticals), diammonium phosphate (for fertilizer), sulphuric acid (many uses), and aluminum fluoride (a catalyst to make aluminum and magnesium).

    The image covers an area of 27.5 x 49.4 km, was acquired on September 17, 2005, and is located near 30.8 degrees north latitude, 36.1 degrees east longitude.

    The U.S. science team is located at NASA's Jet Propulsion Laboratory, Pasadena, Calif. The Terra mission is part of NASA's Science Mission Directorate.

  11. Structural transformations in the a-tricalcium phosphate powders after mechanical activation and subsequent heat treatment

    NASA Astrophysics Data System (ADS)

    Shamray, V. F.; Karpihin, A. E.; Fedotov, A. Y.; Komlev, V. S.; Barinov, S. M.; Sirotinkin, V. P.

    2016-04-01

    X-ray diffraction analysis of the structural phase evolution in the a-TCP powder subjected to milling in the planetary mill in butanol, and followed by annealing at 1300 °C was performed. After milling, there was a systematic smoothing of the X-ray diffraction pattern of α-TCP. Significant changes in lattice constants and atomic parameters were not observed. The average particle size was reduced from 10 to 2 micrometers. The decrease of the size of coherently scattering domains CSD (250 to 80 nm) makes the main contribution to the X-ray lines broadening. The value of microstrain changes from 1 *10-6 to 2*10-4. After annealing at 1300 °C for 2 hours the phase with apatite structure was found. The content of the apatite phase increases with increasing of time of mechanical activation. Possible mechanisms of deformation of α-TCP particles and structural transformations α-TCP → HA are considered.

  12. Digital database of mining-related features at selected historic and active phosphate mines, Bannock, Bear Lake, Bingham, and Caribou counties, Idaho

    USGS Publications Warehouse

    Causey, J. Douglas; Moyle, Phillip R.

    2001-01-01

    This report provides a description of data and processes used to produce a spatial database that delineates mining-related features in areas of historic and active phosphate mining in the core of the southeastern Idaho phosphate resource area. The data have varying degrees of accuracy and attribution detail. Classification of areas by type of mining-related activity at active mines is generally detailed; however, the spatial coverage does not differentiate mining-related surface disturbance features at many of the closed or inactive mines. Nineteen phosphate mine sites are included in the study. A total of 5,728 hc (14,154 ac), or more than 57 km2 (22 mi2), of phosphate mining-related surface disturbance are documented in the spatial coverage of the core of the southeast Idaho phosphate resource area. The study includes 4 active phosphate mines—Dry Valley, Enoch Valley, Rasmussen Ridge, and Smoky Canyon—and 15 historic phosphate mines—Ballard, Champ, Conda, Diamond Gulch, Gay, Georgetown Canyon, Henry, Home Canyon, Lanes Creek, Maybe Canyon, Mountain Fuel, Trail Canyon, Rattlesnake Canyon, Waterloo, and Wooley Valley. Spatial data on the inactive historic mines is relatively up-to-date; however, spatially described areas for active mines are based on digital maps prepared in early 1999. The inactive Gay mine has the largest total area of disturbance: 1,917 hc (4,736 ac) or about 19 km2 (7.4 mi2). It encompasses over three times the disturbance area of the next largest mine, the Conda mine with 607 hc (1,504 ac), and it is nearly four times the area of the Smoky Canyon mine, the largest of the active mines with 497 hc (1,228 ac). The wide range of phosphate mining-related surface disturbance features (approximately 80) were reduced to 13 types or features used in this study—adit and pit, backfilled mine pit, facilities, mine pit, ore stockpile, railroad, road, sediment catchment, tailings or tailings pond, topsoil stockpile, water reservoir, and disturbed

  13. Insulin activation of vacuolar protein sorting 34 mediates localized phosphatidylinositol 3-phosphate production at lamellipodia and activation of mTOR/S6K1.

    PubMed

    Hirsch, Dianne S; Shen, Yi; Dokmanovic, Milos; Yu, Joyce; Mohan, Nishant; Elzarrad, Mohammed Khair; Wu, Wen Jin

    2014-06-01

    The class III phosphatidylinositol 3-kinase, VPS34, phosphorylates the D3 hydroxyl of inositol generating phosphatidylinositol 3-phosphate (ptdins(3)p). Initial studies suggested that ptdins(3)p solely functioned as a component of vesicular and endosomal membranes and that VPS34 did not function in signal transduction. However, VPS34 has recently been shown to be required for insulin-mediated activation of S6 kinase 1 (S6K1). Whether VPS34 activity is directly regulated by insulin is unclear. It is also not known whether VPS34 activity can be spatially restricted in response to extracellular stimuli. Data presented here demonstrate that in response to insulin, VPS34 is activated and translocated to lamellipodia where it produces ptdins(3)p. The localized production of ptdins(3)p is dependent on Src phosphorylation of VPS34. In cells expressing VPS34 with mutations at Y231 or Y310, which are Src-phosphorylation sites, insulin-stimulated VPS34 translocation to the plasma membrane and lamellipodia formation are blocked. mTOR also colocalizes with VPS34 and ptdins(3)p at lamellipodia following insulin-stimulation. In cells expressing the VPS34-Y231F mutant, which blocks lamellipodia formation, mTOR localization at the plasma membrane and insulin-mediated S6K1 activation are reduced. This suggests that mTOR localization at lamellipodia is important for full activation of S6K1 induced by insulin. These data demonstrate that insulin can spatially regulate VPS34 activity through Src-mediated tyrosine phosphorylation and that this membrane localized activity contributes to lamellipodia formation and activation of mTOR/S6K1signaling.

  14. NELL-1 increases pre-osteoblast mineralization using both phosphate transporter Pit1 and Pit2

    SciTech Connect

    Cowan, Catherine M.; Zhang, Xinli; James, Aaron W.; Mari Kim, T.; Sun, Nichole; Wu, Benjamin; Ting, Kang; Soo, Chia

    2012-06-08

    Highlights: Black-Right-Pointing-Pointer NELL-1 accelerates extracellular matrix mineralization in MC3T3-E1 pre-osteoblasts. Black-Right-Pointing-Pointer NELL-1 significantly increases intracellular inorganic phosphate levels. Black-Right-Pointing-Pointer NELL-1 positively regulates osteogenesis but not proliferation in MC3T3-E1 cells. Black-Right-Pointing-Pointer NELL-1 regulates inorganic phosphate transporter activity. -- Abstract: NELL-1 is a potent osteoinductive molecule that enhances bone formation in multiple animal models through currently unidentified pathways. In the present manuscript, we hypothesized that NELL-1 may regulate osteogenic differentiation accompanied by alteration of inorganic phosphate (Pi) entry into the osteoblast via sodium dependent phosphate (NaPi) transporters. To determine this, MC3T3-E1 pre-osteoblasts were cultured in the presence of recombinant human (rh)NELL-1 or rhBMP-2. Analysis was performed for intracellular Pi levels through malachite green staining, Pit-1 and Pit-2 expression, and forced upregulation of Pit-1 and Pit-2. Results showed rhNELL-1 to increase MC3T3-E1 matrix mineralization and Pi influx associated with activation of both Pit-1 and Pit-2 channels, with significantly increased Pit-2 production. In contrast, Pi transport elicited by rhBMP-2 showed to be associated with increased Pit-1 production only. Next, neutralizing antibodies against Pit-1 and Pit-2 completely abrogated the Pi influx effect of rhNELL-1, suggesting rhNELL-1 is dependent on both transporters. These results identify one potential mechanism of action for rhNELL-1 induced osteogenesis and highlight a fundamental difference between NELL-1 and BMP-2 signaling.

  15. Inhibition of phosphate uptake in corn roots by aluminum-fluoride complexes.

    PubMed

    Façanha, Arnoldo Rocha; Okorokova-Façanha, Anna L

    2002-08-01

    F forms stable complexes with Al at conditions found in the soil. Fluoroaluminate complexes (AlF(x)) have been widely described as effective analogs of inorganic phosphate (Pi) in Pi-binding sites of several proteins. In this work, we explored the possibility that the phytotoxicity of AlF(x) reflects their activity as Pi analogs. For this purpose, (32)P-labeled phosphate uptake by excised roots and plasma membrane H(+)-ATPase activity were investigated in an Al-tolerant variety of maize (Zea mays L. var. dwarf hybrid), either treated or not with AlF(x). In vitro, AlF(x) competitively inhibited the rate of root phosphate uptake as well as the H(+)-ATPase activity. Conversely, pretreatment of seedlings with AlF(x) in vivo promoted no effect on the H(+)-ATPase activity, whereas a biphasic effect on Pi uptake by roots was observed. Although the initial rate of phosphate uptake by roots was inhibited by AlF(x) pretreatment, this situation changed over the following minutes as the rate of uptake increased and a pronounced stimulation in subsequent (32)Pi uptake was observed. This kinetic behavior suggests a reversible and competitive inhibition of the phosphate transporter by fluoroaluminates. The stimulation of root (32)Pi uptake induced by AlF(x) pretreatment was tentatively interpreted as a phosphate starvation response. This report places AlF(3) and AlF(4)(-) among Al-phytotoxic species and suggests a mechanism of action where the accumulation of Pi-mimicking fluoroaluminates in the soil may affect the phosphate absorption by plants. The biochemical, physiological, and environmental significance of these findings is discussed.

  16. Inhibition of Phosphate Uptake in Corn Roots by Aluminum-Fluoride Complexes1

    PubMed Central

    Façanha, Arnoldo Rocha; Okorokova-Façanha, Anna L.

    2002-01-01

    F forms stable complexes with Al at conditions found in the soil. Fluoroaluminate complexes (AlFx) have been widely described as effective analogs of inorganic phosphate (Pi) in Pi-binding sites of several proteins. In this work, we explored the possibility that the phytotoxicity of AlFx reflects their activity as Pi analogs. For this purpose, 32P-labeled phosphate uptake by excised roots and plasma membrane H+-ATPase activity were investigated in an Al-tolerant variety of maize (Zea mays L. var. dwarf hybrid), either treated or not with AlFx. In vitro, AlFx competitively inhibited the rate of root phosphate uptake as well as the H+-ATPase activity. Conversely, pretreatment of seedlings with AlFx in vivo promoted no effect on the H+-ATPase activity, whereas a biphasic effect on Pi uptake by roots was observed. Although the initial rate of phosphate uptake by roots was inhibited by AlFx pretreatment, this situation changed over the following minutes as the rate of uptake increased and a pronounced stimulation in subsequent 32Pi uptake was observed. This kinetic behavior suggests a reversible and competitive inhibition of the phosphate transporter by fluoroaluminates. The stimulation of root 32Pi uptake induced by AlFx pretreatment was tentatively interpreted as a phosphate starvation response. This report places AlF3 and AlF4− among Al-phytotoxic species and suggests a mechanism of action where the accumulation of Pi-mimicking fluoroaluminates in the soil may affect the phosphate absorption by plants. The biochemical, physiological, and environmental significance of these findings is discussed. PMID:12177489

  17. Enhanced enzymatic activity of glycerol-3-phosphate dehydrogenase from the cryophilic