Secretion of acid phosphatase by axenic Entamoeba histolytica NIH-200 and properties of the extracellular enzyme.

PubMed

Agrawal, A; Pandey, V C; Kumar, S; Sagar, P

1989-01-01

Entamoeba histolytica (NIH-200) secreted large amounts of acid phosphatase in its external environment when grown axenically in modified TPS-II medium. Fractionation by DEAE-cellulose chromatography of the precipitate obtained from the cell-free medium at 60% ammonium sulfate saturation yielded 3 distinct peaks of enzyme activity. The enzyme in all the peaks showed resistance to tartrate but was inhibited by fluoride, cupric chloride, ethylene diamine-tetra acetic acid, ammonium molybdate and cysteine; however, enzyme associated with different peaks differed in its polyacrylamide gel electrophoretic profiles and behavior towards concanavalin A.

21 CFR 582.1077 - Potassium acid tartrate.

Code of Federal Regulations, 2010 CFR

2010-04-01

... Additives § 582.1077 Potassium acid tartrate. (a) Product. Potassium acid tartrate. (b) Conditions of use. This substance is generally recognized as safe when used in accordance with good manufacturing or...

Increased amount of phosphorylated proinflammatory osteopontin in rheumatoid arthritis synovia is associated to decreased tartrate-resistant acid phosphatase 5B/5A ratio.

PubMed

Luukkonen, Jani; Pascual, Laia Mira; Patlaka, Christina; Lång, Pernilla; Turunen, Sanna; Halleen, Jussi; Nousiainen, Tomi; Valkealahti, Maarit; Tuukkanen, Juha; Andersson, Göran; Lehenkari, Petri

2017-01-01

Osteopontin (OPN) is an immunoregulatory protein which production increases in both rheumatoid arthritis (RA) and osteoarthritis (OA). Phosphorylated osteopontin (Phospho-OPN) is known to increase macrophage and osteoclast activation, this process is controlled by extracellular tartrate-resistant acid phosphatase (TRAcP), also a biomarker for RA. Here, we evaluated the phosphorylation status of OPN in RA and OA synovia, as well as its correlation with TRAcP isoforms. Synovial tissue and fluid were obtained from 24 RA (14 seropositive and 10 seronegative) and 24 OA patients. Western blotting was used to analyze the extent of OPN phosphorylation. TRAcP isoforms were measured in synovial fluid using ELISA; immunohistochemistry assessed the distribution of OPN and TRAcP expressing cells in the synovial tissue, especially distinguishing between the TRAcP isoforms. Full-length OPN was more phosphorylated in RA than in OA (p<0.05). The thrombin cleaved C-terminal end of OPN was also more phosphorylated in RA (p<0.05). RA patients had a lower concentration of TRAcP 5B and higher concentration of less active 5A in their synovial fluid compared to OA patients. The TRAcP 5B/5A ratio was decreased in RA and correlated negatively with the amount of phospho-OPN (p<0.05). TRAcP positive cells for both isoforms were found all along the synovial lining; OPN antibody staining was localized in the extracellular matrix. Our data suggests that in RA the synovial fluid contains insufficient amounts of TRAcP 5B which increase levels of the proinflammatory phospho-OPN. This may lead to increased macrophage and osteoclast activation, resulting in the increased local inflammation and bone resorption present in RA joints.

Tartrate-resistant acid phosphatase (TRAP/ACP5) promotes metastasis-related properties via TGFβ2/TβR and CD44 in MDA-MB-231 breast cancer cells.

PubMed

Reithmeier, Anja; Panizza, Elena; Krumpel, Michael; Orre, Lukas M; Branca, Rui M M; Lehtiö, Janne; Ek-Rylander, Barbro; Andersson, Göran

2017-09-15

Tartrate-resistant acid phosphatase (TRAP/ACP5), a metalloenzyme that is characteristic for its expression in activated osteoclasts and in macrophages, has recently gained considerable focus as a driver of metastasis and was associated with clinically relevant parameters of cancer progression and cancer aggressiveness. MDA-MB-231 breast cancer cells with different TRAP expression levels (overexpression and knockdown) were generated and characterized for protein expression and activity levels. Functional cell experiments, such as proliferation, migration and invasion assays were performed as well as global phosphoproteomic and proteomic analysis was conducted to connect molecular perturbations to the phenotypic changes. We identified an association between metastasis-related properties of TRAP-overexpressing MDA-MB-231 breast cancer cells and a TRAP-dependent regulation of Transforming growth factor (TGFβ) pathway proteins and Cluster of differentiation 44 (CD44). Overexpression of TRAP increased anchorage-independent and anchorage-dependent cell growth and proliferation, induced a more elongated cellular morphology and promoted cell migration and invasion. Migration was increased in the presence of the extracellular matrix (ECM) proteins osteopontin and fibronectin and the basement membrane proteins collagen IV and laminin I. TRAP-induced properties were reverted upon shRNA-mediated knockdown of TRAP or treatment with the small molecule TRAP inhibitor 5-PNA. Global phosphoproteomics and proteomics analyses identified possible substrates of TRAP phosphatase activity or signaling intermediates and outlined a TRAP-dependent regulation of proteins involved in cell adhesion and ECM organization. Upregulation of TGFβ isoform 2 (TGFβ2), TGFβ receptor type 1 (TβR1) and Mothers against decapentaplegic homolog 2 (SMAD2), as well as increased intracellular phosphorylation of CD44 were identified upon TRAP perturbation. Functional antibody-mediated blocking and chemical

[Alkaline phosphatase in Amoeba proteus].

PubMed

Sopina, V A

2005-01-01

In free-living Amoeba proteus (strain B), 3 phosphatase were found after disc-electrophoresis of 10 microg of protein in PAGE and using 1-naphthyl phosphate as a substrate a pH 9.0. These phosphatases differed in their electrophoretic mobilities - "slow" (1-3 bands), "middle" (one band) and "fast" (one band). In addition to 1-naphthyl phosphate, "slow" phosphatases were able to hydrolyse 2-naphthyl phosphate and p-nitrophenyl phosphate. They were slightly activated by Mg2+, completely inhibited by 3 chelators (EDTA, EGTA and 1,10-phenanthroline), L-cysteine, sodium dodecyl sulfate and Fe2+, Zn2+ and Mn2+ (50 mM), considerably inactivated by orthovanadate, molybdate, phosphatase inhibitor cocktail 1, p-nitrophenyl phosphate, Na2HPO4, DL-dithiothreitol and urea and partly inhibited by H2O2, DL-phenylalanine, 2-mercaptoethanol, phosphatase inhibitor cocktail 2 and Ca2+. Imidazole, L-(+)-tartrate, okadaic acid, NaF and sulfhydryl reagents -p-(hydroxy-mercuri)benzoate and N-ethylmaleimide - had no influence on the activity of "slow" phosphatases. "Middle" and "fast" phosphatases, in contrast to "slow" ones, were not inactivated by 3 chelators. The "middle" phosphatase differed from the "fast" one by smaller resistance to urea, Ca2+, Mn2+, phosphates and H2O2 and greater resistance to dithiothreitol and L-(+)-tartrate. In addition, the "fast" phosphatase was inhibited by L-cysteine but the "middle" one was activated by it. Of 5 tested ions (Mg2+, Cu2+, Mn2+, Ca2+ and Zn2+), only Zn2+ reactivated "slow" phosphatases after their inactivation by EDTA treatment. The reactivation of apoenzyme was only partial (about 35 %). Thus, among phosphatases found in amoebae at pH 9.0, only "slow" ones are Zn-metalloenzymes and may be considered as alkaline phosphatases (EC 3.1.3.1). It still remains uncertain, to which particular phosphatase class "middle" and "fast" phosphatases (pH 9.0) may belong.

Studies on the enzymes of Sarcocystis suicanis: purification and characterization of an acid phosphatase.

PubMed

Farooqui, A A; Adams, D D; Hanson, W L; Prestwood, A K

1987-08-01

Percoll density gradient centrifugation was used for isolating large quantities of bradyzoites of Sarcocystis suicanis, which were used for enzymatic analysis. Crude extracts of bradyzoites contained activities suggestive of several acid hydrolases. Levels of acid and alkaline phosphatase were higher than those of beta-N-acetylhexosaminidase and beta-galactosidase. Acid phosphatase was purified 156-fold with an overall recovery of 54% using DEAE-Sepharose 4B and Sephadex G-200 chromatography. The partially purified enzyme was not a glycoprotein and had a molecular weight of approximately 170,000. The enzyme was markedly inhibited by Cu++, Hg++, and iodoacetamide, suggesting the presence of a sulfhydryl group. Sodium tartrate caused strong inhibition of the enzyme. The acid phosphatase of S. suicanis appears to be a unique enzyme that cannot be classified under high or low molecular weight acid phosphatases of widely diverse origin.

Predictors of osteoclast activity in patients with sickle cell disease

PubMed Central

Nouraie, Mehdi; Cheng, Kevin; Niu, Xiaomei; Moore-King, Evadne; Fadojutimi-Akinsi, Margaret F.; Minniti, Caterina P.; Sable, Craig; Rana, Sohail; Dham, Niti; Campbell, Andrew; Ensing, Gregory; Kato, Gregory J.; Gladwin, Mark T.; Castro, Oswaldo L.; Gordeuk, Victor R.

2011-01-01

Background Bone changes are common in sickle cell disease, but the pathogenesis is not fully understood. Tartrate-resistant acid phosphatase (TRACP) type 5b is produced by bone-resorbing osteoclasts. In other forms of hemolytic anemia, increased iron stores are associated with osteoporosis. We hypothesized that transfusional iron overload would be associated with increased osteoclast activity in patients with sickle cell disease. Design and Methods We examined tartrate-resistant acid phosphatase 5b concentrations in patients with sickle cell disease and normal controls of similar age and sex distribution at steady state. Serum tartrate-resistant acid phosphatase 5b concentration was measured using an immunocapture enzyme assay and plasma concentrations of other cytokines were assayed using the Bio-Plex suspension array system. Tricuspid regurgitation velocity, an indirect measure of systolic pulmonary artery pressure, was determined by echocardiography. Results Tartrate-resistant acid phosphatase 5b concentrations were higher in 58 adults with sickle cell disease than in 22 controls (medians of 4.4 versus 2.4 U/L, respectively; P=0.0001). Among the patients with sickle cell disease, tartrate-resistant acid phosphatase 5b independently correlated with blood urea nitrogen (standardized beta=0.40, P=0.003), interleukin-8 (standardized beta=0.30, P=0.020), and chemokine C-C motif ligand 5 (standardized beta=−0.28, P=0.031) concentrations, but not with serum ferritin concentration. Frequent blood transfusions (>10 units in life time) were not associated with higher tartrate-resistant acid phosphatase 5b levels in multivariate analysis. There were strong correlations among tartrate-resistant acid phosphatase 5b, alkaline phosphatase and tricuspid regurgitation velocity (r>0.35, P<0.001). Conclusions Patients with sickle cell disease have increased osteoclast activity as reflected by serum tartrate-resistant acid phosphatase 5b concentrations. Our results may support a

The Leishmania donovani histidine acid ecto-phosphatase LdMAcP: insight into its structure and function.

PubMed

Papadaki, Amalia; Politou, Anastasia S; Smirlis, Despina; Kotini, Maria P; Kourou, Konstadina; Papamarcaki, Thomais; Boleti, Haralabia

2015-05-01

Acid ecto-phosphatase activity has been implicated in Leishmania donovani promastigote virulence. In the present study, we report data contributing to the molecular/structural and functional characterization of the L. donovani LdMAcP (L. donovani membrane acid phosphatase), member of the histidine acid phosphatase (HAcP) family. LdMAcP is membrane-anchored and shares high sequence identity with the major secreted L. donovani acid phosphatases (LdSAcPs). Sequence comparison of the LdMAcP orthologues in Leishmania sp. revealed strain polymorphism and species specificity for the L. donovani complex, responsible for visceral leishmaniasis (Khala azar), proposing thus a potential value of LdMAcP as an epidemiological or diagnostic tool. The extracellular orientation of the LdMAcP catalytic domain was confirmed in L. donovani promastigotes, wild-type (wt) and transgenic overexpressing a recombinant LdMAcP-mRFP1 (monomeric RFP1) chimera, as well as in transiently transfected mammalian cells expressing rLdMAcP-His. For the first time it is demonstrated in the present study that LdMAcP confers tartrate resistant acid ecto-phosphatase activity in live L. donovani promastigotes. The latter confirmed the long sought molecular identity of at least one enzyme contributing to this activity. Interestingly, the L. donovani rLdMAcP-mRFP1 promastigotes generated in this study, showed significantly higher infectivity and virulence indexes than control parasites in the infection of J774 mouse macrophages highlighting thereby a role for LdMAcP in the parasite's virulence.

Phosphatase-Resistant Analogues of Lysophosphatidic Acid

PubMed Central

Prestwich, Glenn D.; Gajewiak, Joanna; Zhang, Honglu; Xu, Xiaoyu; Yang, Guanghui; Serban, Monica

2008-01-01

Isoform-selective agonists and antagonists of the lysophosphatidic acid (LPA) G protein coupled receptors (GPCRs) have important potential applications in cell biology and therapy. LPA GPCRs regulate cancer cell proliferation, invasion, angiogenesis, and also biochemical resistance to chemotherapy- and radiotherapy-induced apoptosis. LPA and its analogues also are feedback inhibitors of the enzyme lysophospholipase D (lysoPLD, a.k.a., autotaxin, ATX), a central regulator of invasion and metastasis. For cancer therapy, the optimal therapeutic profile would be a metabolically stabilized, pan-LPA receptor antagonist that also inhibited lysoPLD. For protection of gastrointestinal mucosa and lymphocytes, LPA agonists would be desirable to minimize or reverse radiation or chemical-induced injury. Analogues of lysophosphatidic acid (LPA) that are chemically modified to be less susceptible to phospholipases and phosphatases show activity as long-lived receptor-specific agonists and antagonists for LPA receptors, as well as inhibitors for the lysoPLD activity of ATX. PMID:18454946

An acid phosphatase in the plasma membranes of human astrocytoma showing marked specificity toward phosphotyrosine protein.

PubMed

Leis, J F; Kaplan, N O

1982-11-01

The plasma membrane from the human tumor astrocytoma contains an active acid phosphatase activity based on hydrolysis of p-nitrophenyl phosphate. Other acid phosphatase substrates--beta-glycerophosphate, O-phosphorylcholine, and 5'-AMP--are not hydrolyzed significantly. The phosphatase activity is tartrate insensitive and is stimulated by Triton X-100 and EDTA. Of the three known phosphoamino acids, only free O-phosphotyrosine is hydrolyzed by the membrane phosphatase activity. Other acid phosphatases tested from potato, wheat germ, milk, and bovine prostate did not show this degree of specificity. The plasma membrane activity also dephosphorylated phosphotyrosine histone at a much greater rate than did the other acid phosphatases. pH profiles for free O-phosphotyrosine and phosphotyrosine histone showed a shift toward physiological pH, indicating possible physiological significance. Phosphotyrosine histone dephosphorylation activity was nearly 10 times greater than that seen for phosphoserine histone dephosphorylation, and Km values were much lower for phosphotyrosine histone dephosphorylation (0.5 microM vs. 10 microM). Fluoride and zinc significantly inhibited phosphoserine histone dephosphorylation. Vanadate, on the other hand, was a potent inhibitor of phosphotyrosine histone dephosphorylation (50% inhibition at 0.5 microM) but not of phosphoserine histone. ATP stimulated phosphotyrosine histone dephosphorylation (160-250%) but inhibited phosphoserine histone dephosphorylation (95%). These results suggest the existence of a highly specific phosphotyrosine protein phosphatase activity associated with the plasma membrane of human astrocytoma.

[Phosphatase activity in Amoeba proteus at low pH].

PubMed

Sopina, V A

2009-01-01

In free-living Amoeba proteus (strain B), three forms of tartrate-sensitive phosphatase were revealed using PAGE of the supernatant of ameba homogenates obtained with 1% Triton X-100 or distilled water and subsequent staining of gels with 2-naphthyl phosphate as substrate (pH 4.0). The form with the highest mobility in the ameba supernatant was sensitive to all tested phosphatase activity modulators. Two other forms with the lower mobilities were completely or significantly inactivated not only by sodium L-(+)-tartrate, but also by L-(+)-tartaric acid, sodium orthovanadate, ammonium molybdate, EDTA, EGTA, o-phospho-L-tyrosine, DL-dithiotreitol, H2O2, 2-mercaptoethanol, and ions of heavy metals - Fe2+, Fe3+, and Cu2+. Based on results of inhibitory analysis, lysosome location in the ameba cell, and wide substrate specificity of these two forms, it has been concluded that they belong to nonspecific acid phosphomonoesterases (AcP, EC 3.1.3.2). This AcP is suggested to have both phosphomonoesterase and phosphotyrosyl-protein phosphatase activitis. Two ecto-phosphatases were revealed in the culture medium, in which amebas were cultivated. One of them was inhibited by the same reagents as the ameba tartrate-sensitive AcP and seems to be the AcP released into the culture medium in the process of exocytosis of the content of food vacuoles. In the culture medium, apart from this AcP, another phosphatase was revealed, which was not inhibited by any tested inhibitors of AcP and alkaline phosphatase. It cannot be ruled out that this phosphatase belong to the ecto-ATPases found in many protists; however, its ability to hydrolyze ATP has not yet been proven.

[The effects of oxygen partial pressure changes on the osteometric markers of the bone tissue in rats].

PubMed

Berezovs'kyĭ, V Ia; Zamors'ka, T M; Ianko, R V

2013-01-01

Our purpose was to investigate the oxygen partial pressure changes on the osteometric and biochemical markers of bone tissue in rats. It was shown that breathing of altered gas mixture did not change the mass, general length, sagittal diameter and density thigh-bones in 12-month Wistar male-rats. The dosed normobaric hypoxia increased the activity of alkaline phosphatase and decreased the activity of tartrate-resistant acid phosphatase. At the same time normobaric hyperoxia with 40 and 90% oxygen conversely decreased the activity of alkaline phosphatase and increased the activity of tartrate-resistant acid phosphatase.

A novel 35 kDa frog liver acid metallophosphatase.

PubMed

Szalewicz, A; Radomska, B; Strzelczyk, B; Kubicz, A

1999-04-12

The lower molecular weight (35 kDa) acid phosphatase from the frog (Rana esculenta) liver is a glycometalloenzyme susceptible to activation by reducing agents and displaying tartrate and fluoride resistance. Metal chelators (EDTA, 1,10-phenanthroline) inactivate the enzyme reversibly in a time- and temperature-dependent manner. The apoenzyme is reactivated by divalent transition metal cations, i. e. cobalt, zinc, ferrous, manganese, cadmium and nickel to 130%, 75%, 63%, 62%, 55% and 34% of the original activity, respectively. Magnesium, calcium, cupric and ferric ions were shown to be ineffective in this process. Metal analysis by the emission spectrometry method (inductively coupled plasma-atomic emission spectrometry) revealed the presence of zinc, iron and magnesium. The time course of the apoenzyme reactivation, the stabilization effect and the relatively high resistance to oxidizing conditions indicate that the zinc ion is crucial for the enzyme activity. The presence of iron was additionally confirmed by the visible absorption spectrum of the enzyme with a shoulder at 417 nm and by the electron paramagnetic resonance line of high spin iron(III) with geff of 2.4. The active center containing only zinc or both zinc and iron ions is proposed. The frog liver lower molecular weight acid phosphatase is a novel metallophosphatase of lower vertebrate origin, distinct from the mammalian tartrate-resistant, purple acid phosphatases.

Isolation, Cloning, and Expression of an Acid Phosphatase Containing Phosphotyrosyl Phosphatase Activity from Prevotella intermedia

PubMed Central

Chen, Xiaochi; Ansai, Toshihiro; Awano, Shuji; Iida, Toshiya; Barik, Sailen; Takehara, Tadamichi

1999-01-01

A novel acid phosphatase containing phosphotyrosyl phosphatase (PTPase) activity, designated PiACP, from Prevotella intermedia ATCC 25611, an anaerobe implicated in progressive periodontal disease, has been purified and characterized. PiACP, a monomer with an apparent molecular mass of 30 kDa, did not require divalent metal cations for activity and was sensitive to orthovanadate but highly resistant to okadaic acid. The enzyme exhibited substantial activity against tyrosine phosphate-containing peptides derived from the epidermal growth factor receptor. On the basis of N-terminal and internal amino acid sequences of purified PiACP, the gene coding for PiACP was isolated and sequenced. The PiACP gene consisted of 792 bp and coded for a basic protein with an Mr of 29,164. The deduced amino acid sequence exhibited striking similarity (25 to 64%) to those of members of class A bacterial acid phosphatases, including PhoC of Morganella morganii, and involved a conserved phosphatase sequence motif that is shared among several lipid phosphatases and the mammalian glucose-6-phosphatases. The highly conservative motif HCXAGXXR in the active domain of PTPase was not found in PiACP. Mutagenesis of recombinant PiACP showed that His-170 and His-209 were essential for activity. Thus, the class A bacterial acid phosphatases including PiACP may function as atypical PTPases, the biological functions of which remain to be determined. PMID:10559178

[Number, activity and thermostability of the electrophoretic forms of acid phosphatase in Amoeba proteus, cultured at different temperatures].

PubMed

Sopina, V A

2001-01-01

In free-living amoebae (Amoeba proteus, strain B), cultured at 10 and 25 degrees C, we compared the number, activity, and thermostability of separate electromorphs of Triton-soluble acid phosphatase (AcP) revealed by disc-electrophoresis in polyacrylamide gel using 2-naphthyl phosphate (pH 4.0) as a substrate. No differences in the number of AcP electromorphs and their mobility were observed at both these temperatures. The total activity of AcP electromorphas per unit of cellular protein and their total thermostability were lower in amoebae acclimated to 10 degrees C than to 25 degrees C. The above decrease may be a consequence of a simultaneous decrease in the activity and thermostability of two tartrate-sensitive electromorphs, both being of lysosomal nature. The total activity and thermostability of tartrate-resistant AcP electromorphs did not differ in amoebae acclimated to the two above temperatures. In amoebae cultured at 10 degrees C the fall of activity and thermostability of lysosomal AcP correlates with the decrease in their primary cell thermoresistance and phagocytic activity. The obtained results confirm the earlier conclusion (Vysotskaya et al., 1994) that lysosomes may be involved in acclimation of electrothermal animals to changing environmental temperatures.

Influence of triethyl phosphate on phosphatase activity in shooting range soil: Isolation of a zinc-resistant bacterium with an acid phosphatase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Story, Sandra; Brigmon, Robin L.

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 soilmore » 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 5 mM 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. Finally, 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.« less

Influence of triethyl phosphate on phosphatase activity in shooting range soil: Isolation of a zinc-resistant bacterium with an acid phosphatase

DOE PAGES

Story, Sandra; Brigmon, Robin L.

2016-12-19

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 soilmore » 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 5 mM 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. Finally, 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.« less

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. Copyright © 2016. Published by Elsevier Inc.

Iron overload causes osteoporosis in thalassemia major patients through interaction with transient receptor potential vanilloid type 1 (TRPV1) channels

PubMed Central

Rossi, Francesca; Perrotta, Silverio; Bellini, Giulia; Luongo, Livio; Tortora, Chiara; Siniscalco, Dario; Francese, Matteo; Torella, Marco; Nobili, Bruno; Di Marzo, Vincenzo; Maione, Sabatino

2014-01-01

The pathogenesis of bone resorption in β-thalassemia major is multifactorial and our understanding of the underlying molecular and cellular mechanisms remains incomplete. Considering the emerging importance of the endocannabinoid/endovanilloid system in bone metabolism, it may be instructive to examine a potential role for this system in the development of osteoporosis in patients with β-thalassemia major and its relationship with iron overload and iron chelation therapy. This study demonstrates that, in thalassemic-derived osteoclasts, tartrate-resistant acid phosphatase expression inversely correlates with femoral and lumbar bone mineral density, and directly correlates with ferritin levels and liver iron concentration. The vanilloid agonist resiniferatoxin dramatically reduces cathepsin K levels and osteoclast numbers in vitro, without affecting tartrate-resistant acid phosphatase expression. The iron chelators deferoxamine, deferiprone and deferasirox decrease both tartrate-resistant acid phosphatase and cathepsin K expression, as well as osteoclast activity. Taken together, these data show that transient receptor potential vanilloid type 1 activation/desensitization influences tartrate-resistant acid phosphatase expression and activity, and this effect is dependent on iron, suggesting a pivotal role for iron overload in the dysregulation of bone metabolism in patients with thalassemia major. Our applied pharmacology provides evidence for the potential of iron chelators to abrogate these effects by reducing osteoclast activity. Whether iron chelation therapy is capable of restoring bone health in humans requires further study, but the potential to provide dual benefits for patients with β-thalassemia major –preventing iron-overload and alleviating associated osteoporotic changes – is exciting. PMID:25216685

21 CFR 184.1804 - Sodium potassium tartrate.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Sodium potassium tartrate. 184.1804 Section 184... Listing of Specific Substances Affirmed as GRAS § 184.1804 Sodium potassium tartrate. (a) Sodium potassium tartrate (C4H4KNaO6·4H2O, CAS Reg. No. 304-59-6) is the sodium potassium salt of l−(+)−tartaric acid and is...

21 CFR 184.1804 - Sodium potassium tartrate.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Sodium potassium tartrate. 184.1804 Section 184... as GRAS § 184.1804 Sodium potassium tartrate. (a) Sodium potassium tartrate (C4H4KNaO6·4H2O, CAS Reg. No. 304-59-6) is the sodium potassium salt of l−(+)−tartaric acid and is also called the Rochelle...

21 CFR 184.1804 - Sodium potassium tartrate.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Sodium potassium tartrate. 184.1804 Section 184... Listing of Specific Substances Affirmed as GRAS § 184.1804 Sodium potassium tartrate. (a) Sodium potassium tartrate (C4H4KNaO6·4H2O, CAS Reg. No. 304-59-6) is the sodium potassium salt of l−(+)−tartaric acid and is...

21 CFR 184.1804 - Sodium potassium tartrate.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Sodium potassium tartrate. 184.1804 Section 184... Listing of Specific Substances Affirmed as GRAS § 184.1804 Sodium potassium tartrate. (a) Sodium potassium tartrate (C4H4KNaO6·4H2O, CAS Reg. No. 304-59-6) is the sodium potassium salt of l−(+)−tartaric acid and is...

Structural and kinetic properties of a novel purple acid phosphatase from phosphate-starved tomato (Lycopersicon esculentum) cell cultures.

PubMed Central

Bozzo, Gale G; Raghothama, Kashchandra G; Plaxton, William C

2004-01-01

An intracellular acid phosphatase (IAP) from P(i)-starved (-P(i)) tomato ( Lycopersicon esculentum ) suspension cells has been purified to homogeneity. IAP is a purple acid phosphatase (PAP), as the purified protein was violet in colour (lambda(max)=546 nm) and was insensitive to L-tartrate. PAGE, periodic acid-Schiff staining and peptide mapping demonstrated that the enzyme exists as a 142 kDa heterodimer composed of an equivalent ratio of glycosylated and structurally dissimilar 63 (alpha-subunit) and 57 kDa (beta-subunit) polypeptides. However, the nine N-terminal amino acids of the alpha- and beta-subunits were identical, exhibiting similarity to the deduced N-terminal portions of several putative plant PAPs. Quantification of immunoblots probed with rabbit anti-(tomato acid phosphatase) immune serum revealed that the 4-fold increase in IAP activity due to P(i)-deprivation was correlated with similar increases in the amount of antigenic IAP alpha- and beta-subunits. IAP displayed optimal activity at pH 5.1, was activated 150% by 10 mM Mg(2+), but was potently inhibited by Zn(2+), Cu(2+), Fe(3+), molybdate, vanadate, fluoride and P(i). Although IAP demonstrated broad substrate selectivity, its specificity constant ( V (max)/ K (m)) with phosphoenolpyruvate was >250% greater than that obtained with any other substrate. IAP exhibited significant peroxidase activity, which was optimal at pH 9.0 and insensitive to Mg(2+) or molybdate. This IAP is proposed to scavenge P(i) from intracellular phosphate esters in -P(i) tomato. A possible secondary IAP role in the metabolism of reactive oxygen species is discussed. IAP properties are compared with those of two extracellular PAP isoenzymes that are secreted into the medium of -P(i) tomato cells [Bozzo, Raghothama and Plaxton (2002) Eur. J. Biochem. 269, 6278-6286]. PMID:14521509

In vitro studies on the translocation of acid phosphatase into the endoplasmic reticulum of the yeast Saccharomyces cerevisiae.

PubMed

Krebs, H O; Hoffschulte, H K; Müller, M

1989-05-01

We demonstrate here the in vitro translocation of yeast acid phosphatase into rough endoplasmic reticulum. The precursor of the repressible acid phosphatase from Saccharomyces cerevisiae encoded by the PHO5 gene, was synthesized in a yeast lysate programmed with in vitro transcribed PHO5 mRNA. In the presence of yeast rough microsomes up to 16% of the acid phosphatase synthesized was found to be translocated into the microsomes, as judged by proteinase resistance, and fully core-glycosylated. The translocation efficiency however, decreased to 3% if yeast rough microsomes were added after synthesis of acid phosphatase had been terminated. When a wheat-germ extract was used for in vitro synthesis, the precursor of acid phosphatase was translocated into canine pancreatic rough microsomes and thereby core-glycosylated in a signal-recognition-particle-dependent manner. Replacing canine with yeast rough microsomes in the wheat-germ translation system, however, resulted in a significant decrease in the ability to translocate and glycosylate the precursor. Translocation and glycosylation were partially restored by a high-salt extract prepared from yeast ribosomes. The results presented here suggest that yeast-specific factors are needed to translocate and glycosylate acid phosphatase efficiently in vitro.

Suppression of DS1 Phosphatidic Acid Phosphatase Confirms Resistance to Ralstonia solanacearum in Nicotiana benthamiana

PubMed Central

Nakano, Masahito; Nishihara, Masahiro; Yoshioka, Hirofumi; Takahashi, Hirotaka; Sawasaki, Tatsuya; Ohnishi, Kouhei; Hikichi, Yasufumi; Kiba, Akinori

2013-01-01

Nicotiana benthamiana is susceptible to Ralstonia solanacearum. To analyze molecular mechanisms for disease susceptibility, we screened a gene-silenced plant showing resistance to R. solanacearum, designated as DS1 (Disease suppression 1). The deduced amino acid sequence of DS1 cDNA encoded a phosphatidic acid phosphatase (PAP) 2. DS1 expression was induced by infection with a virulent strain of R. solanacearum in an hrp-gene-dependent manner. DS1 rescued growth defects of the temperature-sensitive ∆lpp1∆dpp1∆pah1 mutant yeast. Recombinant DS1 protein showed Mg2+-independent PAP activity. DS1 plants showed reduced PAP activity and increased phosphatidic acid (PA) content. After inoculation with R. solanacearum, DS1 plants showed accelerated cell death, over-accumulation of reactive oxygen species (ROS), and hyper-induction of PR-4 expression. In contrast, DS1-overexpressing tobacco plants showed reduced PA content, greater susceptibility to R. solanacearum, and reduced ROS production and PR-4 expression. The DS1 phenotype was partially compromised in the plants in which both DS1 and NbCoi1 or DS1 and NbrbohB were silenced. These results show that DS1 PAP may affect plant immune responses related to ROS and JA cascades via regulation of PA levels. Suppression of DS1 function or DS1 expression could rapidly activate plant defenses to achieve effective resistance against Ralstonia solanacearum. PMID:24073238

Moraxella catarrhalis synthesizes an autotransporter that is an acid phosphatase.

PubMed

Hoopman, Todd C; Wang, Wei; Brautigam, Chad A; Sedillo, Jennifer L; Reilly, Thomas J; Hansen, Eric J

2008-02-01

Moraxella catarrhalis O35E was shown to synthesize a 105-kDa protein that has similarity to both acid phosphatases and autotransporters. The N-terminal portion of the M. catarrhalis acid phosphatase A (MapA) was most similar (the BLAST probability score was 10(-10)) to bacterial class A nonspecific acid phosphatases. The central region of the MapA protein had similarity to passenger domains of other autotransporter proteins, whereas the C-terminal portion of MapA resembled the translocation domain of conventional autotransporters. Cloning and expression of the M. catarrhalis mapA gene in Escherichia coli confirmed the presence of acid phosphatase activity in the MapA protein. The MapA protein was shown to be localized to the outer membrane of M. catarrhalis and was not detected either in the soluble cytoplasmic fraction from disrupted M. catarrhalis cells or in the spent culture supernatant fluid from M. catarrhalis. Use of the predicted MapA translocation domain in a fusion construct with the passenger domain from another predicted M. catarrhalis autotransporter confirmed the translocation ability of this MapA domain. Inactivation of the mapA gene in M. catarrhalis strain O35E reduced the acid phosphatase activity expressed by this organism, and this mutation could be complemented in trans with the wild-type mapA gene. Nucleotide sequence analysis of the mapA gene from six M. catarrhalis strains showed that this protein was highly conserved among strains of this pathogen. Site-directed mutagenesis of a critical histidine residue (H233A) in the predicted active site of the acid phosphatase domain in MapA eliminated acid phosphatase activity in the recombinant MapA protein. This is the first description of an autotransporter protein that expresses acid phosphatase activity.

Human Prostatic Acid Phosphatase: Structure, Function and Regulation

PubMed Central

Muniyan, Sakthivel; Chaturvedi, Nagendra K.; Dwyer, Jennifer G.; LaGrange, Chad A.; Chaney, William G.; Lin, Ming-Fong

2013-01-01

Human prostatic acid phosphatase (PAcP) is a 100 kDa glycoprotein composed of two subunits. Recent advances demonstrate that cellular PAcP (cPAcP) functions as a protein tyrosine phosphatase by dephosphorylating ErbB-2/Neu/HER-2 at the phosphotyrosine residues in prostate cancer (PCa) cells, which results in reduced tumorigenicity. Further, the interaction of cPAcP and ErbB-2 regulates androgen sensitivity of PCa cells. Knockdown of cPAcP expression allows androgen-sensitive PCa cells to develop the castration-resistant phenotype, where cells proliferate under an androgen-reduced condition. Thus, cPAcP has a significant influence on PCa cell growth. Interestingly, promoter analysis suggests that PAcP expression can be regulated by NF-κB, via a novel binding sequence in an androgen-independent manner. Further understanding of PAcP function and regulation of expression will have a significant impact on understanding PCa progression and therapy. PMID:23698773

Effects of multivalent cations on cell wall-associated acid phosphatase activity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tu, S.I.; Brouillette, J.N.; Nagahashi, G.

1988-09-01

Primary cell walls, free from cytoplasmic contamination were prepared from corn (Zea mays L.) roots and potato (Solanum tuberosum) tubers. After EDTA treatment, the bound acid phosphatase activities were measured in the presence of various multivalent cations. Under the conditions of minimized Donnan effect and at pH 4.2, the bound enzyme activity of potato tuber cell walls (PCW) was stimulated by Cu/sup 2 +/, Mg/sup 2 +/, Za/sup 2 +/, and Mn/sup 2 +/; unaffected by Ba/sup 2 +/, Cd/sup 2 +/, and Pb/sup 2 +/; and inhibited by Al/sup 3 +/. The bound acid phosphatase of PCW was stimulatedmore » by a low concentration but inhibited by a higher concentration of Hg/sup 2 +/. On the other hand, in the case of corn root cells walls (CCW), only inhibition of the bound acid phosphatase by Al/sup 3 +/ and Hg/sup 2 +/ was observed. Kinetic analyses revealed that PCW acid phosphatase exhibited a negative cooperativity under all employed experimental conditions except in the presence of Mg/sup 2 +/. In contrast, CCW acid phosphatase showed no cooperative behavior. The presence of Ca/sup 2 +/ significantly reduced the effects of Hg/sup 2 +/ or Al/sup 3 +/, but not Mg/sup 2 +/, to the bound cell wall acid phosphatases. The salt solubilized (free) acid phosphatases from both PCW and CCW were not affected by the presence of tested cations except for Hg/sup 2 +/ or Al/sup 3 +/ which caused a Ca/sup 2 +/-insensitive inhibition of the enzymes. The induced stimulation or inhibition of bound acid phosphatases was quantitatively related to cation binding in the cell wall structure.« less

Vanadate monomers and dimers both inhibit the human prostatic acid phosphatase.

PubMed

Crans, D C; Simone, C M; Saha, A K; Glew, R H

1989-11-30

A combination of enzyme kinetics and 51V NMR spectroscopy was used to identify the species of vanadate that inhibits acid phosphatases. Monomeric vanadate was shown to inhibit wheat germ and potato acid phosphatases. At pH 5.5, the vanadate dimer inhibits the human prostatic acid phosphatase whereas at pH 7.0 it is the vanadate monomer that inhibits this enzyme. The pH-dependent shift in the affinity of the prostatic phosphatase for vanadate is presumably due to deprotonation of an amino acid side chain in or near the binding site resulting in a conformational change in the protein. pH may be a subtle effector of the insulin-like vanadate activity in biological systems and may explain some of the differences in selectivity observed with the protein phosphatases.

Protein phosphatase AP2C1 negatively regulates basal resistance and defense responses to Pseudomonas syringae.

PubMed

Shubchynskyy, Volodymyr; Boniecka, Justyna; Schweighofer, Alois; Simulis, Justinas; Kvederaviciute, Kotryna; Stumpe, Michael; Mauch, Felix; Balazadeh, Salma; Mueller-Roeber, Bernd; Boutrot, Freddy; Zipfel, Cyril; Meskiene, Irute

2017-02-01

Mitogen-activated protein kinases (MAPKs) mediate plant immune responses to pathogenic bacteria. However, less is known about the cell autonomous negative regulatory mechanism controlling basal plant immunity. We report the biological role of Arabidopsis thaliana MAPK phosphatase AP2C1 as a negative regulator of plant basal resistance and defense responses to Pseudomonas syringae. AP2C2, a closely related MAPK phosphatase, also negatively controls plant resistance. Loss of AP2C1 leads to enhanced pathogen-induced MAPK activities, increased callose deposition in response to pathogen-associated molecular patterns or to P. syringae pv. tomato (Pto) DC3000, and enhanced resistance to bacterial infection with Pto. We also reveal the impact of AP2C1 on the global transcriptional reprogramming of transcription factors during Pto infection. Importantly, ap2c1 plants show salicylic acid-independent transcriptional reprogramming of several defense genes and enhanced ethylene production in response to Pto. This study pinpoints the specificity of MAPK regulation by the different MAPK phosphatases AP2C1 and MKP1, which control the same MAPK substrates, nevertheless leading to different downstream events. We suggest that precise and specific control of defined MAPKs by MAPK phosphatases during plant challenge with pathogenic bacteria can strongly influence plant resistance. © The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Acid phosphatase role in chickpea/maize intercropping.

PubMed

Li, S M; Li, L; Zhang, F S; Tang, C

2004-08-01

Organic P comprises 30-80 % of the total P in most agricultural soils. It has been proven that chickpea facilitates P uptake from an organic P source by intercropped wheat. In this study, acid phosphatase excreted from chickpea roots is quantified and the contribution of acid phosphatase to the facilitation of P uptake by intercropped maize receiving phytate is examined. For the first experiment using hydroponics, maize (Zea mays 'Zhongdan No. 2') and chickpea (Cicer arietinum 'Sona') were grown in either the same or separate containers, and P was supplied as phytate, KH2PO4 at 0.25 mmol P L(-1), or not at all. The second experiment involved soil culture with three types of root separation between the two species: (1) plastic sheet, (2) nylon mesh, and (3) no barrier. Maize plants were grown in one compartment and chickpea in the other. Phosphorus was supplied as phytate, Ca(H2PO4)2 at 50 mg P kg(-1), or no P added. In the hydroponics study, the total P uptake by intercropped maize supplied with phytate was 2.1-fold greater than when it was grown as a monoculture. In the soil experiment, when supplied with phytate, total P uptake by maize with mesh barrier and without root barrier was 2.2 and 1.5 times, respectively, as much as that with solid barrier. In both experiments, roots of both maize and chickpea supplied with phytate and no P secreted more acid phosphatase than those with KH2PO4 or Ca(H2PO4)2. However, average acid phosphatase activity of chickpea roots supplied with phytate was 2-3-fold as much as maize. Soil acid phosphatase activity in the rhizosphere of chickpea was also significantly higher than maize regardless of P sources. Chickpea can mobilize organic P in both hydroponic and soil cultures, leading to an interspecific facilitation in utilization of organic P in maize/chickpea intercropping.

"Prostatic acid phosphatase?" A comparison of acid phosphatase activities in epithelial cells, granulocytes, monocytes, lymphocytes, and platelets purified by velocity sedimentation in isokinetic gradients of Ficoll in tissue culture medium.

PubMed Central

Helms, S. R.; Brattain, M. G.; Pretlow, T. G.; Kreisberg, J. I.

1977-01-01

Numerous investigators have found several substrates and inhibitors to be particularly suited for the demonstration of acid phosphatase of prostatic origin. There has been much controversy over the specificity or lack of specificity of several substrates and inhibitors. We have investigated acid phosphatase activities obtained from several kinds of purified cells. None of the substrates or inhibitors which we studied permitted us to discriminate "prostatic" acid phosphatase from acid phosphatase activities obtained from other kinds of cells. PMID:560800

In situ synthesis of di-n-butyl l-tartrate-boric acid complex chiral selector and its application in chiral microemulsion electrokinetic chromatography.

PubMed

Hu, Shaoqiang; Chen, Yonglei; Zhu, Huadong; Zhu, Jinhua; Yan, Na; Chen, Xingguo

2009-11-06

A novel procedure for in situ assembling a complex chiral selector, di-n-butyl l-tartrate-boric acid complex, by the reaction of di-n-butyl l-tartrate with boric acid in a running buffer was reported and its application in the enantioseparation of beta-blockers and structural related compounds by chiral microemulsion electrokinetic chromatography (MEEKC) has been demonstrated. In order to achieve a good enantioseparation, the effect of dibutyl l-tartrate and sodium tetraborate concentration, surfactant identity and concentration, cosurfactant, buffer pH and composition, organic modifiers, as well as applied voltage and capillary length were investigated. Ten pairs of enantiomers that could not be separated with only dibutyl l-tartrate, obtained good chiral separation using the complex chiral selector; among them, seven pairs could be baseline resolved under optimized experimental conditions. The fixation of chiral centers by the formation of five-membered rings, and being oppositely charged with basic analytes were thought to be the key factors giving the complex chiral selector a superior chiral recognition capability. The effect of the molecular structure of analytes on enantioseparation was discussed in terms of molecular interaction.

Lysophosphatidic acids are new substrates for the phosphatase domain of soluble epoxide hydrolase.

PubMed

Oguro, Ami; Imaoka, Susumu

2012-03-01

Soluble epoxide hydrolase (sEH) is a bifunctional enzyme that has a C-terminus epoxide hydrolase domain and an N-terminus phosphatase domain. The endogenous substrates of epoxide hydrolase are known to be epoxyeicosatrienoic acids, but the endogenous substrates of the phosphatase activity are not well understood. In this study, to explore the substrates of sEH, we investigated the inhibition of the phosphatase activity of sEH toward 4-methylumbelliferyl phosphate by using lecithin and its hydrolyzed products. Although lecithin itself did not inhibit the phosphatase activity, the hydrolyzed lecithin significantly inhibited it, suggesting that lysophospholipid or fatty acid can inhibit it. Next, we investigated the inhibition of phosphatase activity by lysophosphatidyl choline, palmitoyl lysophosphatidic acid, monopalmitoyl glycerol, and palmitic acid. Palmitoyl lysophosphatidic acid and fatty acid efficiently inhibited phosphatase activity, suggesting that lysophosphatidic acids (LPAs) are substrates for the phosphatase activity of sEH. As expected, palmitoyl, stearoyl, oleoyl, and arachidonoyl LPAs were efficiently dephosphorylated by sEH (Km, 3-7 μM; Vmax, 150-193 nmol/min/mg). These results suggest that LPAs are substrates of sEH, which may regulate physiological functions of cells via their metabolism.

Prostatic acid phosphatase is an ectonucleotidase and suppresses pain by generating adenosine

PubMed Central

Zylka, Mark J.; Sowa, Nathaniel A.; Taylor-Blake, Bonnie; Twomey, Margaret A.; Herrala, Annakaisa; Voikar, Vootele; Vihko, Pirkko

2008-01-01

SUMMARY Thiamine monophosphatase (TMPase, also known as Fluoride-Resistant Acid Phosphatase) is a classic histochemical marker of small-diameter dorsal root ganglia neurons. The molecular identity of TMPase is currently unknown. We found that TMPase is identical to the transmembrane isoform of Prostatic Acid Phosphatase (PAP), an enzyme with unknown molecular and physiological functions. We then found that PAP knockout mice have normal acute pain sensitivity but enhanced sensitivity in chronic inflammatory and neuropathic pain models. In gain-of-function studies, intraspinal injection of PAP protein has potent anti-nociceptive, anti-hyperalgesic and anti-allodynic effects that last longer than the opioid analgesic morphine. PAP suppresses pain by functioning as an ecto-5’-nucleotidase. Specifically, PAP dephosphorylates extracellular adenosine monophosphate (AMP) to adenosine and activates A1-adenosine receptors in dorsal spinal cord. Our studies reveal molecular and physiological functions for PAP in purine nucleotide metabolism and nociception and suggest a novel use for PAP in the treatment of chronic pain. PMID:18940592

N-Glycosylation Improves the Pepsin Resistance of Histidine Acid Phosphatase Phytases by Enhancing Their Stability at Acidic pHs and Reducing Pepsin's Accessibility to Its Cleavage Sites

PubMed Central

Niu, Canfang; Luo, Huiying; Shi, Pengjun; Huang, Huoqing; Wang, Yaru; Yang, Peilong

2015-01-01

N-Glycosylation can modulate enzyme structure and function. In this study, we identified two pepsin-resistant histidine acid phosphatase (HAP) phytases from Yersinia kristensenii (YkAPPA) and Yersinia rohdei (YrAPPA), each having an N-glycosylation motif, and one pepsin-sensitive HAP phytase from Yersinia enterocolitica (YeAPPA) that lacked an N-glycosylation site. Site-directed mutagenesis was employed to construct mutants by altering the N-glycosylation status of each enzyme, and the mutant and wild-type enzymes were expressed in Pichia pastoris for biochemical characterization. Compared with those of the N-glycosylation site deletion mutants and N-deglycosylated enzymes, all N-glycosylated counterparts exhibited enhanced pepsin resistance. Introduction of the N-glycosylation site into YeAPPA as YkAPPA and YrAPPA conferred pepsin resistance, shifted the pH optimum (0.5 and 1.5 pH units downward, respectively) and improved stability at acidic pH (83.2 and 98.8% residual activities at pH 2.0 for 1 h). Replacing the pepsin cleavage sites L197 and L396 in the immediate vicinity of the N-glycosylation motifs of YkAPPA and YrAPPA with V promoted their resistance to pepsin digestion when produced in Escherichia coli but had no effect on the pepsin resistance of N-glycosylated enzymes produced in P. pastoris. Thus, N-glycosylation may improve pepsin resistance by enhancing the stability at acidic pH and reducing pepsin's accessibility to peptic cleavage sites. This study provides a strategy, namely, the manipulation of N-glycosylation, for improvement of phytase properties for use in animal feed. PMID:26637601

Omega 3 Fatty Acids Reduce Bone Resorption While Promoting Bone Generation in Rat Apical Periodontitis.

PubMed

Azuma, Mariane Maffei; Gomes-Filho, João Eduardo; Ervolino, Edilson; Pipa, Camila Barbosa; Cardoso, Carolina de Barros Morais; Andrada, Ana Cristina; Kawai, Toshihisa; Cintra, Luciano Tavares Angelo

2017-06-01

This study evaluated the effects of the dietary supplement omega 3 polyunsaturated fatty acids (ω-3 PUFAs) on pulp exposure-induced apical periodontitis (AP) in rats. Twenty-eight male rats were divided into groups: control untreated rats (C), control rats treated with ω-3 PUFAs alone (C-O), rats with pulp exposure-induced AP, and rats with pulp exposure-induced AP treated with ω-3 PUFAs (AP-O). The ω-3 PUFAs were administered orally, once a day, for 15 days before pulp exposure and, subsequently, 30 days after pulp exposure. Rats were killed 30 days after pulp exposure, and jaws were subjected to histologic and immunohistochemical analyses. Immunohistochemical analyses were performed to detect tartrate-resistant acid phosphatase-positive osteoclasts and osteocalcin-positive osteoblasts on the bone surface of periapical area. Results were statistically evaluated by using analysis of variance and Tukey honestly significant difference, and P < .05 was considered statistically significant. The bone resorption lesion was significantly larger in the AP group compared with AP-O, C, and C-O groups (P < .05). The level of inflammatory cell infiltration was significantly elevated, and the number of tartrate-resistant acid phosphatase-positive osteoclasts was significantly higher in the periapical lesions of the AP group compared with AP-O, C, and C-O groups (P < .05). The number of osteocalcin-positive osteoblasts was significantly increased in the AP-O group compared with the AP group (P > .05). Supplementation with ω-3 PUFAs not only suppresses bone resorption but also promotes new bone formation in the periapical area of rats with AP in conjunction with downregulation of inflammatory cell infiltration into the lesion. Copyright © 2017 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Partial purification and kinetic characterization of acid phosphatase from garlic seedling.

PubMed

Yenigün, Begüm; Güvenilir, Yüksel

2003-01-01

The objective of this study was to obtain purer acid phosphatases than produced by prior art by operating under conditions that improve the final product. The study features are the use of a mild nonionic detergent, 40-80% saturation with (NH4)2SOm4, maintained at low temperature to remove impurity, and the use of chromatografic columns to concentrate the acid phosphatase and remove non-acid phosphatase proteins with lower or higher molecular weights. Acid phosphatase was isolated and purified from garlic seedlings by a streamline method without the use of proteolytic and lipolytic enzymes, butanol, or other organic solvents. Grown garlic seedlings of 10- 15 cm height were homogenized with 0.1 M acetate buffer containing 0.1 M NaCl and 0.1% Triton X-100. After homogenization, the supernatant was filtered with paper filters. Filtrated supernatant was cooled to 4 degrees C, followed by a threestep fractionation of the proteins with ammonium sulfate. The crude enzyme was isolated as a green precipitate that was dissolved in a small amount of 0.1 M acetate buffer containing 0.1 M NaCl and 0.1% Triton X-100. Garlic seedling acid phosphatase was purified with ion-exchange chromatography (DEAE cellulose). The column was equilibrated with 0.1 M acetate buffer. Acid phosphatase was purified 40-fold from the starting material. The specific activity of the pure enzyme was 168 U/mg. A variety of stability and activity profiles were determined for the purified garlic seedling acid phosphatase: optimum pH, optimum temperature, pH stability, temperature stability, thermal inactivation, substrate specificity, effect of enzyme concentration, effect of substrate concentration, activation energy, and effect of inhibitor and activator. The molecular mass of acid phosphatase was estimated to be 58 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The optimum pH was 5.7 and the optimum temperature was 50 degrees C. The enzyme was stable at pH 4.0-10.0 and 40-60 degrees C

Yeast Acid Phosphatases and Phytases: Production, Characterization and Commercial Prospects

NASA Astrophysics Data System (ADS)

Kaur, Parvinder; Satyanarayana, T.

The element phosphorus is critical to all life forms as it forms the basic component of nucleic acids and ATP and has a number of indispensable biochemical roles. Unlike C or N, the biogeochemical cycling of phosphorus is very slow, and thus making it the growth-limiting element in most soils and aquatic systems. Phosphohydrolases (e.g. acid phosphatases and phytases) are enzymes that break the C-O-P ester bonds and provide available inorganic phosphorus from various inassimilable organic forms of phosphorus like phytates. These enzymes are of significant value in effectively combating phosphorus pollution. Although phytases and acid phosphatases are produced by various plants, animals and micro organisms, microbial sources are more promising for the production on a commercial scale. Yeasts being the simplest eukaryotes are ideal candidates for phytase and phos-phatase research due to their mostly non-pathogenic and GRAS status. They have not, however, been utilized to their full potential. This chapter focuses attention on the present state of knowledge on the production, characterization and potential commercial prospects of yeast phytases and acid phosphatases.

Diversity of multinucleated giant cells by microstructures of hydroxyapatite and plasma components in extraskeletal implantation model.

PubMed

Morishita, Kota; Tatsukawa, Eri; Shibata, Yasuaki; Suehiro, Fumio; Kamitakahara, Masanobu; Yokoi, Taishi; Ioku, Koji; Umeda, Masahiro; Nishimura, Masahiro; Ikeda, Tohru

2016-07-15

Foreign body giant cells (FBGCs) and osteoclasts are multinucleated giant cells (MNGCs), both of which are formed by the fusion of macrophage-derived mononuclear cells. Osteoclasts are distinct from FBGCs due to their bone resorption ability; however, not only morphological, but also functional similarities may exist between these cells. The characterization and diversity of FBGCs that appear in an in vivo foreign body reaction currently remain incomplete. In the present study, we investigated an in vivo foreign body reaction using an extraskeletal implantation model of hydroxyapatite (HA) with different microstructures. The implantation of HA granules in rat subcutaneous tissue induced a foreign body reaction that was accompanied by various MNGCs. HA granules composed of rod-shaped particles predominantly induced cathepsin K (CTSK)-positive FBGCs, whereas HA granules composed of globular-shaped particles predominantly induced CTSK-negative FBGCs. Plasma, which was used as the binder of ceramic granules, stimulated the induction of CTSK-positive FBGCs more strongly than purified fibrin. Furthermore, the implantation of HA composed of rod-shaped particles with plasma induced tartrate-resistant acid phosphatase (TRAP)-positive MNGCs in contrast to HA composed of globular-shaped particles with purified fibrin, which predominantly induced CTSK-negative and TRAP-negative typical FBGCs. These results suggest that CTSK-positive, TRAP-positive, and CTSK- and TRAP-negative MNGCs are induced in this subcutaneous implantation model in a manner that is dependent on the microstructure of HA and presence or absence of plasma. We attempted to elucidate the mechanisms responsible for the foreign body reaction induced by the implantation of hydroxyapatite granules with different microstructures in rat subcutaneous tissue with or without plasma components as the binder of ceramic granules. By analyzing the expression of two reliable osteoclast markers, we detected tartrate-resistant

ISOZYMES OF ACID PHOSPHATASE IN NORMAL AND CALMETTE-GUÉRIN BACILLUS-INDUCED RABBIT ALVEOLAR MACROPHAGES

PubMed Central

Axline, S. G.

1968-01-01

The acid phosphatase activity of normal alveolar and BCG-induced alveolar macrophages has been examined. Five electrophoretically distinct forms of acid phosphatase have been identified in both normal and BCG-induced macrophages. The acid phosphatases can be divided into two major categories. One category, containing four distinct forms, is readily solubilized after repeated freezing and thawing or mechanical disruption The second category, containing one form, is firmly bound to the lysosomal membrane and can be solubilized by treatment of the lysosomal fraction with Triton X-100. The Triton-extractable acid phosphatase and the predominant aqueous soluble acid phosphatase have been shown to differ in the degree of membrane binding, in solubility, in net charge, and in molecular weight. The two pre-dominant phosphatases possess identical pH optimum and do not differ in response to enzyme inhibitors. BCG stimulation has been shown to result in a nearly twofold increase in acid phosphatase activity. A nearly proportionate increase in the major acid phosphatase forms has been observed. PMID:4878908

Zinc-ion-dependent acid phosphatase exhibits magnesium-ion-dependent myo-inositol-1-phosphatase activity.

PubMed

Fujimoto, S; Okano, I; Tanaka, Y; Sumida, Y; Tsuda, J; Kawakami, N; Shimohama, S

1996-06-01

We have purified bovine brain Zn(2+)-dependent acid phosphatase (Zn(2+)-APase), which requires Zn2+ ions to hydrolyze the substrate p-nitrophenyl phosphate (pNPP) in an acidic environment. The substrate specificity and metal requirement of Zn(2+)-APase at a physiological pH was also studied. The enzyme exhibited hydrolytic activity on myo-inositol-1- and -2-monophosphates, 2'-adenosine monophosphate, 2'-guanosine monophosphate, and the alpha- and beta-glycerophosphates, glucose-1-phosphate, and fructose-6-phosphate in 50 mM Tris-HCl buffer (pH 7.4) in the presence of Mg2+ ions, but not on pNPP and phosphotyrosine. Zn2+, Mn2+ and Co2+ ions were less effective for activation. Among the above substrates, myo-inositol-1-phosphate was the most susceptible to hydrolysis by the enzyme in the presence of 3 mM Mg2+ ions. The enzyme exhibited an optimum pH at around 8 for myo-inositol-1-phosphate in the presence of 3 mM Mg2+ ions. The Mg(2+)-dependent myo-inositol-1-phosphatase activity of the enzyme was significantly inhibited by Li+ ions. The Zn(2+)-dependent p-nitrophenyl phosphatase activity and Mg(2+)-dependent myo-inositol-1-phosphatase activity of the purified enzyme fraction exhibited similar behavior on Sephadex G-100 and Mono Q colomns. These findings suggest that Zn(2+)-APase also exhibits Mg(2+)-dependent myo-inositol-1-phosphatase activity under physiological conditions.

Lysophosphatidic acids are new substrates for the phosphatase domain of soluble epoxide hydrolase[S

PubMed Central

Oguro, Ami; Imaoka, Susumu

2012-01-01

Soluble epoxide hydrolase (sEH) is a bifunctional enzyme that has a C-terminus epoxide hydrolase domain and an N-terminus phosphatase domain. The endogenous substrates of epoxide hydrolase are known to be epoxyeicosatrienoic acids, but the endogenous substrates of the phosphatase activity are not well understood. In this study, to explore the substrates of sEH, we investigated the inhibition of the phosphatase activity of sEH toward 4-methylumbelliferyl phosphate by using lecithin and its hydrolyzed products. Although lecithin itself did not inhibit the phosphatase activity, the hydrolyzed lecithin significantly inhibited it, suggesting that lysophospholipid or fatty acid can inhibit it. Next, we investigated the inhibition of phosphatase activity by lysophosphatidyl choline, palmitoyl lysophosphatidic acid, monopalmitoyl glycerol, and palmitic acid. Palmitoyl lysophosphatidic acid and fatty acid efficiently inhibited phosphatase activity, suggesting that lysophosphatidic acids (LPAs) are substrates for the phosphatase activity of sEH. As expected, palmitoyl, stearoyl, oleoyl, and arachidonoyl LPAs were efficiently dephosphorylated by sEH (Km, 3–7 μM; Vmax, 150–193 nmol/min/mg). These results suggest that LPAs are substrates of sEH, which may regulate physiological functions of cells via their metabolism. PMID:22217705

Effect of molecular structure of tartrates on chiral recognition of tartrate-boric acid complex chiral selectors in chiral microemulsion electrokinetic chromatography.

PubMed

Hu, Shao-Qiang; Chen, Yong-Lei; Zhu, Hua-Dong; Shi, Hai-Jun; Yan, Na; Chen, Xing-Guo

2010-08-20

Eight l-tartrates and a d-tartrate with different alcohol moieties were used as chiral oils to prepare chiral microemulsions, which were utilized in conjunction with borate buffer to separate the enantiomers of beta-blockers or structurally related compounds by the chiral microemulsion electrokinetic chromatography (MEEKC) method. Among them, six were found to have a relatively good chiral separation performance and their chiral recognition effect in terms of both enantioselectivity and resolution increases linearly with the number of carbon atoms in the alkyl group of alcohol moiety. The tartrates containing alkyl groups of different structures but the same number of carbon atoms, i.e. one of straight chain and one of branched chain, provide similar enantioseparations. The trend was elucidated according to the changes in the difference of the steric matching between the molecules of two enantiomers and chiral selector. Furthermore, it was demonstrated for the first time that a water insoluble solid compound, di-i-butyl l-tartrate (mp. 73.5 degrees C), can be used as an oil to prepare a stable microemulsion to be used in the chiral MEEKC successfully. And a critical effect of the microemulsion for chiral separation, which has never been reported before, was found in this experiment, namely providing a hydrophobic environment to strengthen the interactions between the chiral selector and enantiomers. Copyright 2010 Elsevier B.V. All rights reserved.

Effects of prey, pitcher age, and microbes on acid phosphatase activity in fluid from pitchers of Sarracenia purpurea (Sarraceniaceae).

PubMed

Luciano, Carl S; Newell, Sandra J

2017-01-01

Carnivory in pitcher plants generally involves digestion of prey, by the plant itself, by symbionts, or both. While symbionts appear to be important in the digestion of prey in Sarracenia purpurea, the importance of pitcher-derived enzymes is less well documented. Our goal was to reduce microbial numbers in pitcher fluid in order to measure the acid phosphatase activity attributable to the pitchers themselves. Preliminary experiments indicated that various antibiotics were minimally effective at reducing microbial populations and that antibiotic-resistant microbes were easily cultured from pitcher fluid. Consequently, we measured the abundance of culturable microbes in every sample taken for the measurement of acid phosphatase activity. Pitchers fed with one sterilized ant had higher levels of acid phosphatase activity than unfed pitchers. Older pitchers were more responsive to feeding than young pitchers. Pitchers with high levels of microbes (on Day 5) had higher acid phosphatase activity than pitchers with low levels of microbes. However, fed pitchers were not more likely to have higher microbe levels and microbe levels were not related to pitcher age. When fluid samples from inside the pitcher were compared to appropriate controls incubated outside the pitcher, acid phosphatase activity was higher inside the pitcher. Results from the feeding experiments are consistent with a primary role of microbes in the digestion of prey in pitchers of S. purpurea. However, the relationship between pitcher age and enzyme activity is not a function of microbes in the pitcher fluid and may depend on enzymes produced by the plant. Our methods would not detect microbes embedded on the inner surface of the pitcher; and if they survived the alcohol rinse and antibiotics, we cannot rule out microbes as the source of the relationship between pitcher age and acid phosphatase activity.

Effects of prey, pitcher age, and microbes on acid phosphatase activity in fluid from pitchers of Sarracenia purpurea (Sarraceniaceae)

PubMed Central

Newell, Sandra J.

2017-01-01

Carnivory in pitcher plants generally involves digestion of prey, by the plant itself, by symbionts, or both. While symbionts appear to be important in the digestion of prey in Sarracenia purpurea, the importance of pitcher-derived enzymes is less well documented. Our goal was to reduce microbial numbers in pitcher fluid in order to measure the acid phosphatase activity attributable to the pitchers themselves. Preliminary experiments indicated that various antibiotics were minimally effective at reducing microbial populations and that antibiotic-resistant microbes were easily cultured from pitcher fluid. Consequently, we measured the abundance of culturable microbes in every sample taken for the measurement of acid phosphatase activity. Pitchers fed with one sterilized ant had higher levels of acid phosphatase activity than unfed pitchers. Older pitchers were more responsive to feeding than young pitchers. Pitchers with high levels of microbes (on Day 5) had higher acid phosphatase activity than pitchers with low levels of microbes. However, fed pitchers were not more likely to have higher microbe levels and microbe levels were not related to pitcher age. When fluid samples from inside the pitcher were compared to appropriate controls incubated outside the pitcher, acid phosphatase activity was higher inside the pitcher. Results from the feeding experiments are consistent with a primary role of microbes in the digestion of prey in pitchers of S. purpurea. However, the relationship between pitcher age and enzyme activity is not a function of microbes in the pitcher fluid and may depend on enzymes produced by the plant. Our methods would not detect microbes embedded on the inner surface of the pitcher; and if they survived the alcohol rinse and antibiotics, we cannot rule out microbes as the source of the relationship between pitcher age and acid phosphatase activity. PMID:28719666

Inhibition of Osteoclast Differentiation and Bone Resorption by N-Methylpyrrolidone*

PubMed Central

Ghayor, Chafik; Correro, Rita M.; Lange, Katrin; Karfeld-Sulzer, Lindsay S.; Grätz, Klaus W.; Weber, Franz E.

2011-01-01

Regulation of RANKL (receptor activator of nuclear factor κB ligand)-induced osteoclast differentiation is of current interest in the development of antiresorptive agents. Osteoclasts are multinucleated cells that play a crucial role in bone resorption. In this study, we investigated the effects of N-methylpyrrolidone (NMP) on the regulation of RANKL-induced osteoclastogenesis. NMP inhibited RANKL-induced tartrate-resistant acid phosphatase activity and the formation of tartrate-resistant acid phosphatase-positive multinucleated cells. The RANKL-induced expression of NFATc1 (nuclear factor of activated T cells, cytoplasmic 1) and c-Fos, which are key transcription factors for osteoclastogenesis, was also reduced by treatment with NMP. Furthermore, NMP induced disruption of the actin rings and decreased the mRNAs of cathepsin K and MMP-9 (matrix metalloproteinase-9), both involved in bone resorption. Taken together, these results suggest that NMP inhibits osteoclast differentiation and attenuates bone resorption. Therefore, NMP could prove useful for the treatment of osteoporosis or other bone diseases associated with excessive bone resorption. PMID:21613210

[Spectroscopic analysis of the interaction of ethanol and acid phosphatase from wheat germ].

PubMed

Xu, Dong-mei; Liu, Guang-shen; Wang, Li-ming; Liu, Wei-ping

2004-11-01

Conformational and activity changes of acid phosphatase from wheat germ in ethanol solutions of different concentrations were measured by fluorescence spectra and differential UV-absorption spectra. The effect of ethanol on kinetics of acid phosphatase was determined by using the double reciprocal plot. The results indicate the ethanol has a significant effect on the activity and conformation of acid phosphatase. The activity of acid phosphatase decreased linearly with increasing the concentration of ethanol. Differential UV-absorption spectra of the enzyme denatured in ethanol solutions showed two positive peaks at 213 and 234 nm, respectively. The peaks on the differential UV-absorption spectra suggested that the conformation of enzyme molecule changed from orderly structure to out-of-order crispation. The fluorescence emission peak intensity of the enzyme gradually strengthened with increasing ethanol concentration, which is in concordance with the conformational change of the microenvironments of tyrosine and tryptophan residues. The results indicate that the expression of the enzyme activity correlates with the stability and integrity of the enzyme conformation to a great degree. Ethanol is uncompetitive inhibitor of acid phosphatase.

Effects of alkaloids from Sophora flavescens on osteoblasts infected with Staphylococcus aureus and osteoclasts.

PubMed

Wang, Xuping; Zheng, Rongzong; Huang, Xiaowen; Mao, Zhujun; Wang, Nani; Li, Hongyu; Wen, Chengping; Shou, Dan

2018-03-25

Chronic osteomyelitis is primarily caused by infection with Staphylococcus aureus (S. aureus). Antibiotics are commonly administered; however, it is a challenge to promote bone healing. The aim of this study was to investigate the in vitro effects of alkaloids from the herbal remedy Sophora flavescens (ASF) on rat calvarial osteoblasts (ROBs) infected with S. aureus and healthy osteoclasts. Cell proliferation and alkaline phosphatase, interleukin-6, and tumour necrosis factor-α activity was measured in infected ROBs; tartrate-resistant acid phosphatase was evaluated in osteoclasts via enzyme-linked immunosorbent assay. The mRNA and protein expression levels of bone morphogenetic protein 2, runt-related transcription factor 2, osteoprotegerin, and receptor activator of nuclear factor kappa-B ligand were assessed in infected ROBs through reverse transcription-polymerase chain reaction and western blotting analysis, respectively. Results indicated that ASF increased the viability of uninfected ROBs and infected ROBs treated with vancomycin via regulation of bone morphogenetic protein 2, runt-related transcription factor, osteoprotegerin, and receptor activator of nuclear factor kappa-B ligand mRNA and protein expression levels. In addition, the secretion of the inflammatory factor tumour necrosis factor-α was decreased and alkaline phosphatase activity was increased, inhibiting the viability of osteoclasts and tartrate-resistant acid phosphatase activity. Therefore, the herbal remedy ASF has potential as a new treatment for chronic osteomyelitis. Copyright © 2018 John Wiley & Sons, Ltd.

Rhizobiales-like Phosphatase 2 from Arabidopsis thaliana Is a Novel Phospho-tyrosine-specific Phospho-protein Phosphatase (PPP) Family Protein Phosphatase.

PubMed

Uhrig, R Glen; Labandera, Anne-Marie; Muhammad, Jamshed; Samuel, Marcus; Moorhead, Greg B

2016-03-11

Cellular signaling through protein tyrosine phosphorylation is well established in mammalian cells. Although lacking the classic tyrosine kinases present in humans, plants have a tyrosine phospho-proteome that rivals human cells. Here we report a novel plant tyrosine phosphatase from Arabidopsis thaliana (AtRLPH2) that, surprisingly, has the sequence hallmarks of a phospho-serine/threonine phosphatase belonging to the PPP family. Rhizobiales/Rhodobacterales/Rhodospirillaceae-like phosphatases (RLPHs) are conserved in plants and several other eukaryotes, but not in animals. We demonstrate that AtRLPH2 is localized to the plant cell cytosol, is resistant to the classic serine/threonine phosphatase inhibitors okadaic acid and microcystin, but is inhibited by the tyrosine phosphatase inhibitor orthovanadate and is particularly sensitive to inhibition by the adenylates, ATP and ADP. AtRLPH2 displays remarkable selectivity toward tyrosine-phosphorylated peptides versus serine/threonine phospho-peptides and readily dephosphorylates a classic tyrosine phosphatase protein substrate, suggesting that in vivo it is a tyrosine phosphatase. To date, only one other tyrosine phosphatase is known in plants; thus AtRLPH2 represents one of the missing pieces in the plant tyrosine phosphatase repertoire and supports the concept of protein tyrosine phosphorylation as a key regulatory event in plants. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

[Chiral separation of five beta-blockers using di-n-hexyl L-tartrate-boric acid complex as mobile phase additive by reversed-phase liquid chromatography].

PubMed

Yang, Juan; Wang, Lijuan; Guo, Qiaoling; Yang, Gengliang

2012-03-01

A reversed-phase high performance liquid chromatographic (HPLC) method using the di-n-hexyl L-tartrate-boric acid complex as a chiral mobile phase additive was developed for the enantioseparation of five beta-blockers including propranolol, esmolol, metoprolol, bisoprolol and sotalol. In order to obtain a better enantioseparation, the influences of concentrations of di-n-butyl L-tartrate and boric acid, the type, concentration and pH of the buffer, methanol content as well as the molecular structure of analytes were extensively investigated. The separation of the analytes was performed on a Venusil MP-C18 column (250 mm x 4.6 mm, 5 microm). The mobile phase was 15 mmol/L ammonium acetate-methanol containing 60 mmol/L boric acid, 70 mmol/L di-n-hexyl L-tartrate (pH 6.00). The volume ratios of 15 mmol/L ammonium acetate to methanol were 20: 80 for propranolol, esmolol, metoprolol, bisoprolol and 30: 70 for sotalol. The flow rate was 0.5 mL/min and the detection wavelength was set at 214 nm. Under the optimized conditions, baseline enantioseparation was obtained separately for the five pairs of analytes.

The discovery of novel tartrate-based TNF-[alpha] converting enzyme (TACE) inhibitors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rosner, Kristin E.; Guo, Zhuyan; Orth, Peter

2010-09-17

A novel series of TNF-{alpha} convertase (TACE) inhibitors which are non-hydroxamate have been discovered. These compounds are bis-amides of L-tartaric acid (tartrate) and coordinate to the active site zinc in a tridentate manner. They are selective for TACE over other MMP's. We report the first X-ray crystal structure for a tartrate-based TACE inhibitor.

An ion-pair principle for enantioseparations of basic analytes by nonaqueous capillary electrophoresis using the di-n-butyl L-tartrate-boric acid complex as chiral selector.

PubMed

Wang, Li-Juan; Liu, Xiu-Feng; Lu, Qie-Nan; Yang, Geng-Liang; Chen, Xing-Guo

2013-04-05

A chiral recognition mechanism of ion-pair principle has been proposed in this study. It rationalized the enantioseparations of some basic analytes using the complex of di-n-butyl l-tartrate and boric acid as the chiral selector in methanolic background electrolytes (BGEs) by nonaqueous capillary electrophoresis (NACE). An approach of mass spectrometer (MS) directly confirmed that triethylamine promoted the formation of negatively charged di-n-butyl l-tartrate-boric acid complex chiral counter ion with a complex ratio of 2:1. And the negatively charged counter ion was the real chiral selector in the ion-pair principle enantioseparations. It was assumed that triethylamine should play its role by adjusting the apparent acidity (pH*) of the running buffer to a higher value. Consequently, the effects of various basic electrolytes including inorganic and organic ones on the enantioseparations in NACE were investigated. The results showed that most of the basic electrolytes tested were favorable for the enantioseparations of basic analytes using di-n-butyl l-tartrate-boric acid complex as the chiral ion-pair selector. Copyright © 2013 Elsevier B.V. All rights reserved.

Acid and alkaline phosphatase localization in the digestive tract mucosa of the Hemisorubim platyrhynchos.

PubMed

Faccioli, Claudemir Kuhn; Chedid, Renata Alari; Mori, Ricardo Hideo; Amaral, Antônio Carlos do; Franceschini-Vicentini, Irene Bastos; Vicentini, Carlos Alberto

2016-09-01

This cytochemical study investigated the acid and alkaline phosphatase of the digestive tract of Hemisorubim platyrhynchos. Acid phosphatase was detected in the lining epithelium throughout the digestive tract, whereas alkaline phosphatase was only observed in the intestine. In the esophagus, an acid phosphatase reaction occurred in the apical cytoplasm of the epithelial cells and was related to epithelial protection and freeing of superficial cells for sloughing. Similar results were also observed in epithelial cells of gastric epithelium. In the gastric glands, acid phosphatase occurred in lysosomes of the oxynticopeptic cells acting in the macromolecule degradation for use as an energy source, whereas in the vesiculotubular system, its presence could be related to secretion processes. Furthermore, acid phosphatase in the intestine occurred in microvilli and lysosomes of the enterocytes and was correlated to absorption and intracellular digestion. However, no difference was reported among the regions of the intestine. However, alkaline phosphatase reaction revealed a large number of reaction dots in the anterior intestine, with the number decreasing toward the posterior intestine. This enzyme has been related to several functions, highlighting its role in the nutrient absorption primarily in the anterior intestine but also being essential in pH regulation because this is a carnivorous species with many gastric glands with secretions that could damage the intestine. Copyright © 2016 Elsevier GmbH. All rights reserved.

Crystallization of recombinant Haemophilus influenzaee (P4) acid phosphatase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ou, Zhonghui; Felts, Richard L.; Reilly, Thomas J.

2006-05-01

Lipoprotein e (P4) is a class C acid phosphatase and a potential vaccine candidate for nontypeable H. influenzae infections. This paper reports the crystallization of recombinant e (P4) and the acquisition of a 1.7 Å resolution native X-ray diffraction data set. Haemophilus influenzae infects the upper respiratory tract of humans and can cause infections of the middle ear, sinuses and bronchi. The virulence of the pathogen is thought to involve a group of surface-localized macromolecular components that mediate interactions at the host–pathogen interface. One of these components is lipoprotein e (P4), which is a class C acid phosphatase and amore » potential vaccine candidate for nontypeable H. influenzae infections. This paper reports the crystallization of recombinant e (P4) and the acquisition of a 1.7 Å resolution native X-ray diffraction data set. The space group is P4{sub 2}2{sub 1}2, with unit-cell parameters a = 65.6, c = 101.4 Å, one protein molecule per asymmetric unit and 37% solvent content. This is the first report of the crystallization of a class C acid phosphatase.« less

Silymarin Induces Insulin Resistance through an Increase of Phosphatase and Tensin Homolog in Wistar Rats

PubMed Central

Cheng, Kai-Chun; Asakawa, Akihiro; Li, Ying-Xiao; Chung, Hsien-Hui; Amitani, Haruka; Ueki, Takatoshi; Cheng, Juei-Tang; Inui, Akio

2014-01-01

Background and aims Phosphatase and tensin homolog (PTEN) is a phosphoinositide phosphatase that regulates crucial cellular functions, including insulin signaling, lipid and glucose metabolism, as well as survival and apoptosis. Silymarin is the active ingredient in milk thistle and exerts numerous effects through the activation of PTEN. However, the effect of silymarin on the development of insulin resistance remains unknown. Methods Wistar rats fed fructose-rich chow or normal chow were administered oral silymarin to identify the development of insulin resistance using the homeostasis model assessment of insulin resistance and hyperinsulinemic- euglycemic clamping. Changes in PTEN expression in skeletal muscle and liver were compared using western blotting analysis. Further investigation was performed in L6 cells to check the expression of PTEN and insulin-related signals. PTEN deletion in L6 cells was achieved by small interfering ribonucleic acid transfection. Results Oral administration of silymarin at a dose of 200 mg/kg once daily induced insulin resistance in normal rats and enhanced insulin resistance in fructose-rich chow-fed rats. An increase of PTEN expression was observed in the skeletal muscle and liver of rats with insulin resistance. A decrease in the phosphorylation of Akt in L6 myotube cells, which was maintained in a high-glucose condition, was also observed. Treatment with silymarin aggravated high-glucose-induced insulin resistance. Deletion of PTEN in L6 cells reversed silymarin-induced impaired insulin signaling and glucose uptake. Conclusions Silymarin has the ability to disrupt insulin signaling through increased PTEN expression. Therefore, silymarin should be used carefully in type-2 diabetic patients. PMID:24404172

EVIDENCE FOR AN EXOCELLULAR SITE FOR THE ACID PHOSPHATASE OF SACCHAROMYCES MELLIS1

PubMed Central

Weimberg, Ralph; Orton, William L.

1964-01-01

Weimberg, Ralph (Northern Regional Research Laboratory, Peoria, Ill.), and William L. Orton. Evidence for an exocellular site for the acid phosphatase of Saccharomyces mellis. J. Bacteriol. 88:1743–1754. 1964.—Evidence is presented which demonstrates an exocellular location for acid phosphatase in Saccharomyces mellis. Derepressed intact cells exhibit acid phosphatase activity. The properties of the system are similar to those shown by the enzyme in cell-free extracts. There is no increase in total activity when cell-free extracts are prepared. Enzymatically active cell walls were prepared by leaching acetone-dried cells of this yeast in dilute acetate buffer (pH 6.5) plus β-mercaptoethanol. The insoluble residue, consisting mainly of cell-wall material and containing the phosphatase, was treated with a variety of hydrolytic enzymes and other chemicals. Only papain and crude snail gut extracts dissociated the enzyme from the particulate fraction in nearly quantitative amounts. The mechanism of release by these two enzymes probably differs. Of all enzymes tested, only the snail gut extract digested the cell walls. By dividing the procedure for making protoplasts of S. mellis into two steps, acid phosphatase may be dissociated from resting cells and recovered as an active soluble enzyme. The first step is to pretreat the cells with a thiol reagent. The second step is to digest the cell wall by enzymes present in crude snail gut extracts. Arsenite must be included in the second step to protect the phosphatase from inactivation. The phosphatase is quantitatively released before the cell becomes osmotically fragile. Images PMID:14240965

Biocatalysis with Sol-Gel Encapsulated Acid Phosphatase

ERIC Educational Resources Information Center

Kulkarni, Suhasini; Tran, Vu; Ho, Maggie K.-M.; Phan, Chieu; Chin, Elizabeth; Wemmer, Zeke; Sommerhalter, Monika

2010-01-01

This experiment was performed in an upper-level undergraduate biochemistry laboratory course. Students learned how to immobilize an enzyme in a sol-gel matrix and how to perform and evaluate enzyme-activity measurements. The enzyme acid phosphatase (APase) from wheat germ was encapsulated in sol-gel beads that were prepared from the precursor…

21 CFR 582.6801 - Sodium tartrate.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Sodium tartrate. 582.6801 Section 582.6801 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Sodium tartrate. (a) Product. Sodium tartrate. (b) Conditions of use. This substance is generally...

21 CFR 582.6801 - Sodium tartrate.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Sodium tartrate. 582.6801 Section 582.6801 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Sodium tartrate. (a) Product. Sodium tartrate. (b) Conditions of use. This substance is generally...

21 CFR 582.6801 - Sodium tartrate.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Sodium tartrate. 582.6801 Section 582.6801 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Sodium tartrate. (a) Product. Sodium tartrate. (b) Conditions of use. This substance is generally...

21 CFR 582.6801 - Sodium tartrate.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Sodium tartrate. 582.6801 Section 582.6801 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Sodium tartrate. (a) Product. Sodium tartrate. (b) Conditions of use. This substance is generally...

21 CFR 582.6801 - Sodium tartrate.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Sodium tartrate. 582.6801 Section 582.6801 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Sodium tartrate. (a) Product. Sodium tartrate. (b) Conditions of use. This substance is generally...

Calcium Tartrate Tetrahydrate, Case Report of a Novel Human Kidney Stone.

PubMed

Kleinguetl, Colin; Williams, James C; Ibrahim, Samar A; Daudon, Michel; Bird, Erin T; El Tayeb, Marawan M

2017-01-01

Background: Calcium tartrate tetrahydrate has been reported as the main mineral in urinary stones in rats that have significant tartrate in their diet, but in humans, there has been only one mention of calcium tartrate stones in the form of bladder stone, and that case was in Africa. Case Presentation: Patient is a 34-year-old Caucasian male who presented with typical symptoms of nephrolithiasis. CT abd/pelvis (renal stone protocol) revealed a 2 cm nonobstructing stone of the right renal pelvis. Patient underwent an uncomplicated right percutaneous nephrolithotomy and was noted to be stone free after surgery. Stone analysis was difficult with regard to determining composition, but was finally identified as calcium tartrate tetrahydrate. Conclusion: This was an unusual case, as this is the first recorded case of a calcium tartrate tetrahydrate outside of Africa. This type of stone had only been mainly described in rat models with dl- bitartrate in their diet. Our patient was an otherwise healthy, relatively muscular individual with no obvious source for this stone other than a vitamin and amino acid supplement that he takes regularly that contains l-carnitine (as tartrate) and choline (as bitartrate and citrate). The prevalence of this stone type is presently unknown, as stone analysis laboratories have not had the ability to recognize it. Although a connection between the supplement and stone formation is conjecture at this time, we believe this necessitates further investigation.

Decryptification of Acid Phosphatase in Arthrospores of Geotrichum Species Treated with Dimethyl Sulfoxide and Acetone

PubMed Central

Cotter, David A.; Martel, Anita J.; MacDonald, Paul

1975-01-01

Decryptification of acid phosphatase in Geotrichum sp. arthrospores was accomplished using acetone or dimethyl sulfoxide treatment. Both dimethyl sulfoxide and acetone irreversibly destroyed the integrity of the spore membranes without solubilizing acid phosphatase. PMID:1167386

Phosphatidate Phosphatase Activity Plays Key Role in Protection against Fatty Acid-induced Toxicity in Yeast*

PubMed Central

Fakas, Stylianos; Qiu, Yixuan; Dixon, Joseph L.; Han, Gil-Soo; Ruggles, Kelly V.; Garbarino, Jeanne; Sturley, Stephen L.; Carman, George M.

2011-01-01

The PAH1-encoded phosphatidate (PA) phosphatase in Saccharomyces cerevisiae is a pivotal enzyme that produces diacylglycerol for the synthesis of triacylglycerol (TAG) and simultaneously controls the level of PA used for phospholipid synthesis. Quantitative lipid analysis showed that the pah1Δ mutation caused a reduction in TAG mass and an elevation in the mass of phospholipids and free fatty acids, changes that were more pronounced in the stationary phase. The levels of unsaturated fatty acids in the pah1Δ mutant were unaltered, although the ratio of palmitoleic acid to oleic acid was increased with a similar change in the fatty acid composition of phospholipids. The pah1Δ mutant exhibited classic hallmarks of apoptosis in stationary phase and a marked reduction in the quantity of cytoplasmic lipid droplets. Cells lacking PA phosphatase were sensitive to exogenous fatty acids in the order of toxicity palmitoleic acid > oleic acid > palmitic acid. In contrast, the growth of wild type cells was not inhibited by fatty acid supplementation. In addition, wild type cells supplemented with palmitoleic acid exhibited an induction in PA phosphatase activity and an increase in TAG synthesis. Deletion of the DGK1-encoded diacylglycerol kinase, which counteracts PA phosphatase in controlling PA content, suppressed the defect in lipid droplet formation in the pah1Δ mutant. However, the sensitivity of the pah1Δ mutant to palmitoleic acid was not rescued by the dgk1Δ mutation. Overall, these findings indicate a key role of PA phosphatase in TAG synthesis for protection against fatty acid-induced toxicity. PMID:21708942

Powder XRD, TEM, FTIR and thermal studies of strontium tartrate nano particles

NASA Astrophysics Data System (ADS)

Lathiya, U. M.; Jethva, H. O.; Joshi, M. J.; Vyas, P. M.

2017-05-01

Strontium tartrate finds several applications, e.g., as non-linear optical and dielectric material, in tracer composition and ammunition unit, in treating structural integrity of bone. The growth of single crystals of strontium tartrate in silica gel has been widely reported. In the present study, strontium tartrate nano particles were synthesized by wet chemical method using strontium chloride, tartaric acid and sodium meta-silicate solutions in the presence of Triton X -100 surfactant. It was found that the presence of sodium meta-silicate facilitated the reaction for strontium tartrate product. The powder XRD study of strontium tartrate nano-particles suggested monoclinic crystal system and the average crystallite size was found to be 40 nm determined by applying Scherrer's formula. The TEM analysis indicated that the nano particles were spherical in nature. The FTIR spectrum confirmed the presence of various functional groups such as O-H,C-H, and C=O stretching mode. The thermal analysis was carried out by using TGA and DTA studies. The nano-particles were found to be stable up to 175°C and then decomposed through various stages. The results are compared with the bulk crystalline material available in the literature.

LEPS2, a Phosphorus Starvation-Induced Novel Acid Phosphatase from Tomato1

PubMed Central

Baldwin, James C.; Karthikeyan, Athikkattuvalasu S.; Raghothama, Kashchandra G.

2001-01-01

Phosphate (Pi) is one of the least available plant nutrients found in the soil. A significant amount of phosphate is bound in organic forms in the rhizosphere. Phosphatases produced by plants and microbes are presumed to convert organic phosphorus into available Pi, which is absorbed by plants. In this study we describe the isolation and characterization of a novel tomato (Lycopersicon esculentum) phosphate starvation-induced gene (LePS2) representing an acid phosphatase. LePS2 is a member of a small gene family in tomato. The cDNA is 942 bp long and contains an open reading frame encoding a 269-amino acid polypeptide. The amino acid sequence of LePS2 has a significant similarity with a phosphatase from chicken. Distinct regions of the peptide also share significant identity with the members of HAD and DDDD super families of phosphohydrolases. Many plant homologs of LePS2 are found in the databases. The LePS2 transcripts are induced rapidly in tomato plant and cell culture in the absence of Pi. However, the induction is repressible in the presence of Pi. Divided root studies indicate that internal Pi levels regulate the expression of LePS2. The enhanced expression of LePS2 is a specific response to Pi starvation, and it is not affected by starvation of other nutrients or abiotic stresses. The bacterially (Escherichia coli) expressed protein exhibits phosphatase activity against the synthetic substrate p-nitrophenyl phosphate. The pH optimum of the enzyme activity suggests that LePS2 is an acid phosphatase. PMID:11161030

Generation of avian cells resembling osteoclasts from mononuclear phagocytes

NASA Technical Reports Server (NTRS)

Alvarez, J. I.; Teitelbaum, S. L.; Blair, H. C.; Greenfield, E. M.; Athanasou, N. A.; Ross, F. P.

1991-01-01

Several lines of indirect evidence suggest that a monocyte family precursor gives rise to the osteoclast, although this hypothesis is controversial. Starting with a uniform population of nonspecific esterase positive, tartrate-sensitive, acid phosphatase-producing, mannose receptor-bearing mononuclear cells, prepared from dispersed marrow of calcium-deprived laying hens by cell density separation and selective cellular adherence, we generated multinucleated cells in vitro. When cultured with devitalized bone, these cells show, by electron microscopy, the characteristic osteoclast morphology in that they are mitochondria-rich, multinucleated, and, most importantly, develop characteristic ruffled membranes at the matrix attachment site. Moreover, as documented by scanning electron microscopy, these cells pit bone slices in a manner identical to freshly isolated osteoclasts. In addition, isoenzymes of acid phosphatase from generated osteoclasts, separated by 7.5% polyacrylamide gel electrophoresis at pH 4, are identical to those of mature osteoclasts in migration pattern and tartrate resistance, although the precursor cells from which the osteoclasts are generated produce an entirely different isoenzyme, which is tartrate-sensitive and migrates less rapidly at pH 4. The fused cells also exhibit a cAMP response to prostaglandin E2. Therefore, osteoclast-like cells can be derived by in vitro culture of a marrow-derived monocyte cell population.

Effect of vanadium compounds on acid phosphatase activity.

PubMed

Vescina, C M; Sálice, V C; Cortizo, A M; Etcheverry, S B

1996-01-01

The direct effect of different vanadium compounds on acid phosphatase (ACP) activity was investigated. Vanadate and vanadyl but not pervanadate inhibited the wheat germ ACP activity. These vanadium derivatives did not alter the fibroblast Swiss 3T3 soluble fraction ACP activity. Using inhibitors of tyrosine phosphatases (PTPases), the wheat germ ACP was partially characterized as a PTPase. This study suggests that the inhibitory ability of different vanadium derivatives to modulate ACP activity seems to depend on the geometry around the vanadium atom more than on the oxidation state. Our results indicate a correlation between the PTPase activity and the sensitivity to vanadate and vanadyl cation.

21 CFR 184.1801 - Sodium tartrate.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Sodium tartrate. 184.1801 Section 184.1801 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR... Specific Substances Affirmed as GRAS § 184.1801 Sodium tartrate. (a) Sodium tartrate (C4H4Na2O6·2H2O, CAS...

21 CFR 184.1801 - Sodium tartrate.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Sodium tartrate. 184.1801 Section 184.1801 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR... Specific Substances Affirmed as GRAS § 184.1801 Sodium tartrate. (a) Sodium tartrate (C4H4Na2O6·2H2O, CAS...

21 CFR 184.1801 - Sodium tartrate.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Sodium tartrate. 184.1801 Section 184.1801 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR... Specific Substances Affirmed as GRAS § 184.1801 Sodium tartrate. (a) Sodium tartrate (C4H4Na2O6·2H2O, CAS...

Preparative enantioseparation of propafenone by counter-current chromatography using di-n-butyl L-tartrate combined with boric acid as the chiral selector.

PubMed

Tong, Shengqiang; Shen, Mangmang; Zheng, Ye; Chu, Chu; Li, Xing-Nuo; Yan, Jizhong

2013-09-01

This paper extends the research of the utilization of borate coordination complexes in chiral separation by counter-current chromatography (CCC). Racemic propafenone was successfully enantioseparated by CCC with di-n-butyl l-tartrate combined with boric acid as the chiral selector. The two-phase solvent system was composed of chloroform/ 0.05 mol/L acetate buffer pH 3.4 containing 0.10 mol/L boric acid (1:1, v/v), in which 0.10 mol/L di-n-butyl l-tartrate was added in the organic phase. The influence of factors in the enantioseparation of propafenone were investigated and optimized. A total of 92 mg of racemic propafenone was completely enantioseparated using high-speed CCC in a single run, yielding 40-42 mg of (R)- and (S)-propafenone enantiomers with an HPLC purity over 90-95%. The recovery for propafenone enantiomers from fractions of CCC was in the range of 85-90%. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Phosphotyrosine as a substrate of acid and alkaline phosphatases.

PubMed

Apostoł, I; Kuciel, R; Wasylewska, E; Ostrowski, W S

1985-01-01

A new spectrophotometric method for following dephosphorylation of phosphotyrosine has been described. The absorption spectra of phosphotyrosine and tyrosine were plotted over the pH range from 3 to 9. The change in absorbance accompanying the conversion of phosphotyrosine to tyrosine was the greatest at 286 nm. The difference absorption coefficients were calculated for several pH values. Dephosphorylation of phosphotyrosine by acid phosphatases from human prostate gland, from wheat germ and potatoes obeys the Michaelis-Menten equation, whereas alkaline phosphatases calf intestine and E. coli are inhibited by excess of substrate.

21 CFR 184.1801 - Sodium tartrate.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Sodium tartrate. 184.1801 Section 184.1801 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DIRECT... GRAS § 184.1801 Sodium tartrate. (a) Sodium tartrate (C4H4Na2O6·2H2O, CAS Reg. No. 868-18-8) is the...

21 CFR 184.1801 - Sodium tartrate.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Sodium tartrate. 184.1801 Section 184.1801 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN... Substances Affirmed as GRAS § 184.1801 Sodium tartrate. (a) Sodium tartrate (C4H4Na2O6·2H2O, CAS Reg. No. 868...

Therapeutic effect of icariin combined with stem cells on postmenopausal osteoporosis in rats.

PubMed

Tang, Dao; Ju, Cuiling; Liu, Yanjie; Xu, Fei; Wang, Zhengguang; Wang, Dongbo

2018-03-01

Osteoporosis is characterized by skeletal fragility and microarchitectural deterioration. The side effects of drugs to treat osteoporosis will negatively affect the health of patients. This study aimed to investigate the therapeutic effects of icariin combined with adipose-derived stem cells on osteoporosis in a postmenopausal osteoporosis model after ovariectomy in rats. After ovariectomy the rats were treated with icariin combined with adipose-derived stem cell transplantation. The levels of alkaline phosphatase, tartrate-resistant acid phosphatase, osteoprotegerin, and bone γ-carboxyglutamate protein in serum were determined by ELISA. The bone mineral density was measured by dual-energy X-ray absorptiometry. The mechanical properties were determined by a three-point bending test. The kidney functions were evaluated by an automatic analyzer and a diagnostic kit. Icariin combined with stem cells significantly reduced body weight gain caused by ovariectomy, significantly decreased alkaline phosphatase, tartrate-resistant acid phosphatase, and bone γ-carboxyglutamate protein content in serum, significantly increased osteoprotegerin content, significantly elevated bone mineral density of the lumbar spine, left femur, and right femur, and enhanced bone biomechanical properties of the femur, including maximum bending load, bending rigidity, and fracture energy, in osteoporotic rats. In addition, icariin combined with stem cells substantially decreased the damage to the liver and kidney in osteoporotic rats. Icariin combined with stem cells can not only ameliorate reduction of bone mass and disruption of the microarchitectural structure of bone tissue caused by osteoporosis in a rat model but can also have a beneficial effect on organ functions, such as those of the liver and kidney.

21 CFR 582.1804 - Sodium potassium tartrate.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Sodium potassium tartrate. 582.1804 Section 582.1804 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... Additives § 582.1804 Sodium potassium tartrate. (a) Product. Sodium potassium tartrate. (b) Conditions of...

21 CFR 582.1804 - Sodium potassium tartrate.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Sodium potassium tartrate. 582.1804 Section 582.1804 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... Additives § 582.1804 Sodium potassium tartrate. (a) Product. Sodium potassium tartrate. (b) Conditions of...

21 CFR 582.6804 - Sodium potassium tartrate.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Sodium potassium tartrate. 582.6804 Section 582.6804 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED....6804 Sodium potassium tartrate. (a) Product. Sodium potassium tartrate. (b) Conditions of use. This...

21 CFR 582.1804 - Sodium potassium tartrate.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Sodium potassium tartrate. 582.1804 Section 582.1804 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... Additives § 582.1804 Sodium potassium tartrate. (a) Product. Sodium potassium tartrate. (b) Conditions of...

21 CFR 582.6804 - Sodium potassium tartrate.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Sodium potassium tartrate. 582.6804 Section 582.6804 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED....6804 Sodium potassium tartrate. (a) Product. Sodium potassium tartrate. (b) Conditions of use. This...

21 CFR 582.1804 - Sodium potassium tartrate.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Sodium potassium tartrate. 582.1804 Section 582.1804 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... Additives § 582.1804 Sodium potassium tartrate. (a) Product. Sodium potassium tartrate. (b) Conditions of...

21 CFR 582.6804 - Sodium potassium tartrate.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Sodium potassium tartrate. 582.6804 Section 582.6804 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED....6804 Sodium potassium tartrate. (a) Product. Sodium potassium tartrate. (b) Conditions of use. This...

21 CFR 582.1804 - Sodium potassium tartrate.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Sodium potassium tartrate. 582.1804 Section 582.1804 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... Additives § 582.1804 Sodium potassium tartrate. (a) Product. Sodium potassium tartrate. (b) Conditions of...

21 CFR 582.6804 - Sodium potassium tartrate.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Sodium potassium tartrate. 582.6804 Section 582.6804 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED....6804 Sodium potassium tartrate. (a) Product. Sodium potassium tartrate. (b) Conditions of use. This...

21 CFR 582.6804 - Sodium potassium tartrate.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Sodium potassium tartrate. 582.6804 Section 582.6804 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED....6804 Sodium potassium tartrate. (a) Product. Sodium potassium tartrate. (b) Conditions of use. This...

Zolpidem tartrate and somnambulism.

PubMed

Harazin, J; Berigan, T R

1999-09-01

A case is reported in which a patient experienced somnambulistic episodes only after taking zolpidem tartrate for insomnia. Previous to the patient's use of zolpidem tartrate he had never experienced sleepwalking, and once the medication was discontinued the sleepwalking stopped. A search of the literature revealed only two other cases of zolpidem-induced sleepwalking, both involving individuals with a previous history of somnambulism in their youth.

Isolation of Lysophosphatidic Acid Phosphatase from Developing Peanut Cotyledons1

PubMed Central

Shekar, Sunil; Tumaney, Ajay W.; Rao, T.J.V. Sreenivasa; Rajasekharan, Ram

2002-01-01

The soluble fraction of immature peanut (Arachis hypogaea) was capable of dephosphorylating [3H]lysophosphatidic acid (LPA) to generate monoacylglycerol (MAG). The enzyme responsible for the generation of MAG, LPA phosphatase, has been identified in plants and purified by successive chromatography separations on octyl-Sepharose, Blue Sepharose, Superdex-75, and heparin-agarose to apparent homogeneity from developing peanuts. This enzyme was purified 5,048-fold to a final specific activity of 858 nmol min−1 mg−1. The enzyme has a native molecular mass of approximately 39 kD determined by gel filtration and migrates as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with a subunit molecular mass of 39 ± 1.5 kD. The Km values for oleoyl-, stearoyl-, and palmitoyl-sn-glycerol-3-phosphate were determined to be 28.6, 39.3, and 47.9 μm, respectively. The LPA phosphatase was specific to LPA and did not utilize any other substrate such as glycerol-3-phosphate, phosphatidic acid, or p-nitrophenylphosphate. The enzyme activity was stimulated by the low concentrations of detergents such as Triton X-100 and octylglucoside. Cations had no effect on the enzyme activity. Fatty acids, sphingosine, and sphingomyelin at low concentrations stimulated the enzyme activity. The identification of LPA phosphatase in plants demonstrates the existence of MAG biosynthetic machinery in plants. PMID:11891254

Endocytosis of lysosomal acid phosphatase; involvement of mannose receptor and effect of lectins.

PubMed

Imai, K; Yoshimura, T

1994-08-01

Acid phosphatase and beta-glucosidase are unique among lysosomal enzymes in that they have both high mannose and complex type sugasr chains, whereas oligosaccharide chains of lysosomal enzymes in matrix are of high mannose type. We have previously shown that beta-glucosidase was endocytosed into macrophages via an unidentified receptor different from a mannose/fucose receptor (K. Imai, Cell Struct. Funct. 13, 325-332, 1988). Here, we show that uptake of acid phosphatase purified from rat liver lysosomes into rat macrophages was inhibited by ligands for a mannose/fucose receptor and was mediated via an apparently single binding site with Kuptake of 24.7 nM. These results indicate that acid phosphatase and beta-glucosidase recognize different types of receptors even if they have similar sugar chains. Polyvalent concanavalin A which binds both to the enzyme and to macrophages specifically stimulated the uptake in a dose dependent manner, whereas wheat germ agglutinin and phytohaemagglutinin did not.

Dephosphorylation of microtubule-binding sites at the neurofilament-H tail domain by alkaline, acid, and protein phosphatases.

PubMed

Hisanaga, S; Yasugawa, S; Yamakawa, T; Miyamoto, E; Ikebe, M; Uchiyama, M; Kishimoto, T

1993-06-01

The dephosphorylation-induced interaction of neurofilaments (NFs) with microtubules (MTs) was investigated by using several phosphatases. Escherichia coli alkaline and wheat germ acid phosphatases increased the electrophoretic mobility of NF-H and NF-M by dephosphorylation, and induced the binding of NF-H to MTs. The binding of NFs to MTs was observed only after the electrophoretic mobility of NF-H approached the exhaustively dephosphorylated level when alkaline phosphatase was used. The number of phosphate remaining when NF-H began to bind to MTs was estimated by measuring phosphate bound to NF-H. NF-H did not bind to MTs even when about 40 phosphates from the total of 51 had been removed by alkaline phosphatase. The removal of 6 further phosphates finally resulted in the association of NF-H with MTs. A similar finding, that the restricted phosphorylation sites in the NF-H tail domain, but not the total amount of phosphates, were important for binding to MTs, was also obtained with acid phosphatases. In contrast to alkaline and acid phosphatases, four classes of protein phosphatases (protein phosphatases 1, 2A, 2B, and 2C) were ineffective for shifting the electrophoretic mobility of NF proteins and for inducing the association of NFs to MTs.

Okadaic acid and microcystin insensitive PPP-family phosphatases may represent novel biotechnology targets.

PubMed

Uhrig, R Glen; Moorhead, Greg B

2011-12-01

Reversible protein phosphorylation is of central importance to the proper cellular functioning of all living organisms. Catalyzed by the opposing reactions of protein kinases and phosphatases, dysfunction in reversible protein phosphorylation can result in a wide variety of cellular aberrations. In eukaryotic organisms there exists four classes of protein phosphatases, of which the PPP-family protein phosphatases have documented susceptibility to a range of protein and small molecule inhibitors. These inhibitors have been of great importance to the biochemical characterization of PPP-family protein phosphatases since their discovery, but also maintain in natura biological significance with their endogenous regulatory properties (protein inhibitors) and toxicity (small molecule inhibitors). Recently, two unique PPP-family protein phosphatases, named the Shewanella-like protein phosphatases (SLP phosphatases), from Arabidopsis thaliana were characterized and found to be phylogenetically similar to the PPP-family protein phosphatases protein phosphatase 1 (PP1) and protein phosphatase 2A (PP2A), while completely lacking sensitivity to the classic PPP-family phosphatase small molecule inhibitors okadaic acid and microcystin-LR. SLP phosphatases were also found to be absent in metazoans, but present in a wide range of bacteria, fungi and protozoa responsible for human disease. The unique biochemical properties and evolutionary heritage of SLP phosphatases suggests they could not only be potential biotechnology targets for agriculture, but may also prove to be of interest for future therapeutic drug development. © 2011 Landes Bioscience

Gadolinium and didymium (praseodymium/neodymium) cations as capture agents in lightmicroscopical histochemistry of acid and alkaline phosphatase.

PubMed

Halbhuber, K J; Zimmermann, N

1987-01-01

In previous papers, cerium and lanthanum based methods for light-microscopical detection of acid and alkaline phosphatase activity were proposed. In this paper, the usefulness of other lanthanide cations such as gadolinium and praseodymium/neodymium cations as capture agents in phosphatase histochemistry is tested. It is evident that phosphate ions were sufficiently trapped by these cations. According to the lead and silver multistep procedures earlier described it is possible to visualize alkaline phosphatase activity in the brush borders of the intestine or kidney as well as acid phosphatase activity in the lysosomes. These methods can be recommended.

Alkaline and Acid Phosphatase Activity, pH and Osmotic Pressure of Boar Semen***

PubMed Central

King, G. J.; Macpherson, J. W.

1966-01-01

Alkaline phosphatase activity was recorded in forty ejaculates of the sperm rich fraction of boar semen as 9,790 ± 5,250 Klein-Babson-Read units per 100 ml. of seminal plasma. Acid phosphatase activity in the same ejaculates was 681 ± 304 Babson-Read units per 100 ml. of seminal plasma. No alkaline phosphatase activity was detected in the seminal plasma of vasectomized boars. The pH of the sperm rich fractions was 7.69 ± 0.33 and the osmotic pressure was 313.56 ± 7.98 milliosmols. PMID:4226380

PURIFICATION AND PARTIAL CHARACTERIZATION OF AN ACID PHOSPHATASE FROM SPIRODELA OLIGORRHIZA AND ITS AFFINITY FOR SELECTED ORGANOPHOSPHATE PESTICIDES

EPA Science Inventory

An acid phosphatase from the aquatic plant Spirodela oligorrhiza (duckweed) was isolated by fast protein liquid chromatography (FPLC) and partially characterized. The enzyme was purified 1871-fold with a total yield of 40%. SDS-PAGE electrophoresis of the pure acid phosphatase ...

The hppA gene of Helicobacter pylori encodes the class C acid phosphatase precursor.

PubMed

Godlewska, Renata; Bujnicki, Janusz M; Ostrowski, Jerzy; Jagusztyn-Krynicka, Elzbieta K

2002-08-14

Screening of the Helicobacter pylori genomic library with sera from infected humans and from immunized rabbits resulted in identification of the 25 kDa protein cell envelope (HppA) which exhibits acid phosphatase activity. Enzyme activity was demonstrated by specific enzymatic assays with whole-cell protein preparations of H. pylori strain N6 and from Escherichia coli carrying the hppA gene (pUWM192). HppA showed optimum activity at pH 5.6 and was resistant to inhibition by EDTA. Bioinformatics analysis and site-directed mutagenesis of two putative active site residues (D73 and D192) provide further insight into the sequence-structure-function relationships of HppA as a member of the DDDD phosphohydrolase superfamily.

The inhibitory effect of metals and other ions on acid phosphatase activity from Vigna aconitifolia seeds.

PubMed

Srivastava, Pramod Kumar; Anand, Asha

2015-01-01

Sensitivity of acid phosphatase from Vigna aconitifolia seeds to metal ions, fluoride, and phosphate was examined. All the effectors had different degree of inhibitory effect on the enzyme. Among metal ions, molybdate and ferric ion were observed to be most potent inhibitors and both exhibited mixed type of inhibition. Acid phosphatase activity was inhibited by Cu2+ in a noncompetitive manner. Zn and Mn showed mild inhibition on the enzyme activity. Inhibition kinetics analysis explored molybdate as a potent inhibitor for acid phosphatase in comparison with other effectors used in this study. Fluoride was the next most strong inhibitor for the enzyme activity, and caused a mixed type of inhibition. Phosphate inhibited the enzyme competitively, which demonstrates that inhibition due to phosphate is one of the regulatory factors for enzyme activity.

Interaction of Myosin Phosphatase Target Subunit (MYPT1) with Myosin Phosphatase-RhoA Interacting Protein (MRIP): A Role of Glutamic Acids in the Interaction.

PubMed

Lee, Eunhee; Stafford, Walter F

2015-01-01

Scaffold proteins bind to and functionally link protein members of signaling pathways. Interaction of the scaffold proteins, myosin phosphatase target subunit (MYPT1) and myosin phosphatase-RhoA interacting protein (MRIP), causes co-localization of myosin phosphatase and RhoA to actomyosin. To examine biophysical properties of interaction of MYPT1 with MRIP, we employed analytical ultracentrifugation and surface plasmon resonance. In regard to MRIP, its residues 724-837 are sufficient for the MYPT1/MRIP interaction. Moreover, MRIP binds to MYPT1 as either a monomer or a dimer. With respect to MYPT1, its leucine repeat region, LR (residues 991-1030) is sufficient to account for the MYPT1/MRIP interaction. Furthermore, point mutations that replace glutamic acids 998-1000 within LR reduced the binding affinity toward MRIP. This suggests that the glutamic acids of MYPT1 play an important role in the interaction.

Isolation and characterization of a homogeneous isoenzyme of wheat germ acid phosphatase.

PubMed

Waymack, P P; Van Etten, R L

1991-08-01

An acid phosphatase (orthophosphoric monoester phosphohydrolase, acid optimum; EC 3.1.3.2) isoenzyme from wheat germ was purified 7000-fold to homogeneity. The effect of wheat germ sources and their relationship to the isoenzyme content and purification behavior of acid phosphatases was investigated. Extensive information about the purification and stabilization of the enzyme is provided. The instability of isoenzymes in the latter stages of purification appeared to be the result of surface inactivation together with a sensitivity to dilution that could be partially offset by addition of Triton X-100 during chromatographic procedures. Added sulfhydryl protecting reagents had no effect on activity or stability, which was greatest in the pH range 4-7. The purified isoenzyme was homogeneous by polyacrylamide gel electrophoresis and exhibited the highest specific activity and turnover number reported for any acid phosphatase. The molecular weights of the pure isoenzyme and of related isoenzymes from wheat germ were found to be identical (58,000). The pure isoenzyme contained a single polypeptide chain and had a negligible carbohydrate content. The amino acid composition was determined. Of the various reasons that were considered to explain isoenzyme occurrence, a genetic basis was considered most likely. The enzyme was found to exhibit substrate inhibition with some substrates below pH 6, while above pH 8 it exhibited downwardly curving Lineweaver-Burk plots of the type that are generally described as "substrate activation". The observation of a phosphotransferase activity was consistent with the formation of a covalent phosphoenzyme intermediate, while inactivation by diethyl pyrocarbonate was consistent with the presence of an active site histidine.

Substrate specificity and pH dependence of homogeneous wheat germ acid phosphatase.

PubMed

Van Etten, R L; Waymack, P P

1991-08-01

The broad substrate specificity of a homogeneous isoenzyme of wheat germ acid phosphatase (WGAP) was extensively investigated by chromatographic, electrophoretic, NMR, and kinetic procedures. WGAP exhibited no divalent metal ion requirement and was unaffected upon incubation with EDTA or o-phenanthroline. A comparison of two catalytically homogeneous isoenzymes revealed little difference in substrate specificity. The specificity of WGAP was established by determining the Michaelis constants for a wide variety of substrates. p-Nitrophenyl phosphate, pyrophosphate, tripolyphosphate, and ATP were preferred substrates while lesser activities were seen toward sugar phosphates, trimetaphosphate, phosphoproteins, and (much less) phosphodiesters. An extensive table of Km and Vmax values is given. The pathway for the hydrolysis of trimetaphosphate was examined by colorimetric and 31P NMR methods and it was found that linear tripolyphosphate is not a free intermediate in the enzymatic reaction. In contrast to literature reports, homogeneous wheat germ acid phosphatase exhibits no measurable carboxylesterase activity, nor does it hydrolyze phenyl phosphonothioate esters or phytic acid at significant rates.

21 CFR 520.246 - Butorphanol tartrate tablets.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Butorphanol tartrate tablets. 520.246 Section 520.246 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... tartrate tablets. (a) Specifications. Each tablet contains 1, 5, or 10 milligrams of butorphanol base...

Recognition of Nucleoside Monophosphate Substrates by Haemophilus influenzae Class C Acid Phosphatase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Singh, Harkewal; Schuermann, Jonathan P.; Reilly, Thomas J.

2010-12-08

The e (P4) phosphatase from Haemophilus influenzae functions in a vestigial NAD{sup +} utilization pathway by dephosphorylating nicotinamide mononucleotide to nicotinamide riboside. P4 is also the prototype of class C acid phosphatases (CCAPs), which are nonspecific 5{prime},3{prime}-nucleotidases localized to the bacterial outer membrane. To understand substrate recognition by P4 and other class C phosphatases, we have determined the crystal structures of a substrate-trapping mutant P4 enzyme complexed with nicotinamide mononucleotide, 5{prime}-AMP, 3{prime}-AMP, and 2{prime}-AMP. The structures reveal an anchor-shaped substrate-binding cavity comprising a conserved hydrophobic box that clamps the nucleotide base, a buried phosphoryl binding site, and three solvent-filled pocketsmore » that contact the ribose and the hydrogen-bonding edge of the base. The span between the hydrophobic box and the phosphoryl site is optimal for recognizing nucleoside monophosphates, explaining the general preference for this class of substrate. The base makes no hydrogen bonds with the enzyme, consistent with an observed lack of base specificity. Two solvent-filled pockets flanking the ribose are key to the dual recognition of 5{prime}-nucleotides and 3{prime}-nucleotides. These pockets minimize the enzyme's direct interactions with the ribose and provide sufficient space to accommodate 5{prime} substrates in an anti conformation and 3{prime} substrates in a syn conformation. Finally, the structures suggest that class B acid phosphatases and CCAPs share a common strategy for nucleotide recognition.« less

Ascorbic acid induces alkaline phosphatase, type X collagen, and calcium deposition in cultured chick chondrocytes.

PubMed

Leboy, P S; Vaias, L; Uschmann, B; Golub, E; Adams, S L; Pacifici, M

1989-10-15

During the process of endochondral bone formation, proliferating chondrocytes give rise to hypertrophic chondrocytes, which then deposit a mineralized matrix to form calcified cartilage. Chondrocyte hypertrophy and matrix mineralization are associated with expression of type X collagen and the induction of high levels of the bone/liver/kidney isozyme of alkaline phosphatase. To determine what role vitamin C plays in these processes, chondrocytes derived from the cephalic portion of 14-day chick embryo sternae were grown in the absence or presence of exogenous ascorbic acid. Control untreated cells displayed low levels of type X collagen and alkaline phosphatase activity throughout the culture period. However, cells grown in the presence of ascorbic acid produced increasing levels of alkaline phosphatase activity and type X collagen mRNA and protein. Both alkaline phosphatase activity and type X collagen mRNA levels began to increase within 24 h of ascorbate treatment; by 9 days, the levels of both alkaline phosphatase activity and type X collagen mRNA were 15-20-fold higher than in non-ascorbate-treated cells. Ascorbate treatment also increased calcium deposition in the cell layer and decreased the levels of types II and IX collagen mRNAs; these effects lagged significantly behind the elevation of alkaline phosphatase and type X collagen. Addition of beta-glycerophosphate to the medium increased calcium deposition in the presence of ascorbate but had no effect on levels of collagen mRNAs or alkaline phosphatase. The results suggest that vitamin C may play an important role in endochondral bone formation by modulating gene expression in hypertrophic chondrocytes.

A Black Phosphate Conversion Coating on Steel Surface Using Antimony(III)-Tartrate as an Additive

NASA Astrophysics Data System (ADS)

Li, Feng; Wang, Guiping

2016-05-01

A novel black phosphate conversion coating was formed on steel surface through a Zn-Mn phosphating bath containing mainly ZnO, H3PO4, Mn(H2PO4)2, and Ca(NO3)2, where antimony(III)-tartrate was used as the blackening agent of phosphatization. The surface morphology and composition of the coating were characterized by scanning electron microscopy, energy dispersion spectroscopy, and x-ray photoelectron spectroscopy. Corrosion resistance of the coating was studied by potentiodynamic polarization curves and electrochemical impedance spectroscopy. The pH value of the solution had significant influence on the formation and corrosion resistance of the coating. The experimental results indicated that the Sb plays a vital role in the blackening of phosphate conversion coating. The optimal concentration of antimony(III)-tartrate in the phosphating bath used in this experiment was 1.0 g L-1, as higher values reduced the corrosion resistance of the coating. In addition, by saponification and oil seals, the corrosion duration of the black phosphate coating in a copper sulfate spot test can be as long as 20 min.

21 CFR 862.1020 - Acid phosphatase (total or prostatic) test system.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Acid phosphatase (total or prostatic) test system. 862.1020 Section 862.1020 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry...

21 CFR 862.1020 - Acid phosphatase (total or prostatic) test system.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Acid phosphatase (total or prostatic) test system. 862.1020 Section 862.1020 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry...

21 CFR 862.1020 - Acid phosphatase (total or prostatic) test system.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Acid phosphatase (total or prostatic) test system. 862.1020 Section 862.1020 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry...

21 CFR 862.1020 - Acid phosphatase (total or prostatic) test system.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Acid phosphatase (total or prostatic) test system. 862.1020 Section 862.1020 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry...

21 CFR 862.1020 - Acid phosphatase (total or prostatic) test system.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Acid phosphatase (total or prostatic) test system. 862.1020 Section 862.1020 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry...

Evidence that Speciation of Oxovanadium Complexes does not Solely Account for Inhibition of Leishmania Acid Phosphatases

NASA Astrophysics Data System (ADS)

Dorsey, Benjamin M.; McLauchlan, Craig C.; Jones, Marjorie A.

2018-04-01

Leishmaniasis is an endemic disease affecting a diverse spectra of populations, with 1.6 million new cases reported each year. Current treatment options are costly and have harsh side effects. New therapeutic options that have been previously identified, but still underappreciated as potential pharmaceutical targets, are Leishmania secreted acid phosphatases (SAP). These acid phosphatases, which are reported to play a role in the survival of the parasite in the sand fly vector, and in homing to the host macrophage, are inhibited by orthovanadate and decavanadate. Here, we use L. tarentolae to further evaluate these inhibitors. Using enzyme assays, and UV-visible spectroscopy, we investigate which oxovanadium starting material (orthovanadate or decavanadate) is a better inhibitor of L. tarentolae secreted acid phosphatase activity in vitro at the same total moles of vanadium. Considering speciation and total vanadium concentration, decavanadate is a consistently better inhibitor of SAP in our conditions, especially at low substrate:inhibitor ratios.

Evidence That Speciation of Oxovanadium Complexes Does Not Solely Account for Inhibition of Leishmania Acid Phosphatases

PubMed Central

Dorsey, Benjamin M.; McLauchlan, Craig C.; Jones, Marjorie A.

2018-01-01

Leishmaniasis is an endemic disease affecting a diverse spectra of populations, with 1.6 million new cases reported each year. Current treatment options are costly and have harsh side effects. New therapeutic options that have been previously identified, but still underappreciated as potential pharmaceutical targets, are Leishmania secreted acid phosphatases (SAP). These acid phosphatases, which are reported to play a role in the survival of the parasite in the sand fly vector, and in homing to the host macrophage, are inhibited by orthovanadate and decavanadate. Here, we use L. tarentolae to further evaluate these inhibitors. Using enzyme assays, and UV-visible spectroscopy, we investigate which oxovanadium starting material (orthovanadate or decavanadate) is a better inhibitor of L. tarentolae secreted acid phosphatase activity in vitro at the same total moles of vanadium. Considering speciation and total vanadium concentration, decavanadate is a consistently better inhibitor of SAP in our conditions, especially at low substrate:inhibitor ratios. PMID:29707535

ISOLATION AND PARTIAL CHARACTERIZATION OF AN ACID PHOSPHATASE ACTIVITY FROM SPIRODELA OLIGORHIZA

EPA Science Inventory

An acid phosphatase activity from the aquatic plant Spirodela oligorhiza (duckweed) was isolated and partially characterized. S. oligorhiza was grown in a hydroponic growth medium, harvested, and ground up in liquid nitrogen. The ground plant material was added to a biological ...

Structural basis of the inhibition of class C acid phosphatases by adenosine 5;#8242;-phosphorothioate

DOE Office of Scientific and Technical Information (OSTI.GOV)

Singh, Harkewal; Reilly, Thomas J.; Tanner, John J.

2012-01-20

The inhibition of phosphatases by adenosine 5'-phosphorothioate (AMPS) was first reported in the late 1960s; however, the structural basis for the inhibition has remained unknown. Here, it is shown that AMPS is a submicromolar inhibitor of class C acid phosphatases, a group of bacterial outer membrane enzymes belonging to the haloacid dehalogenase structural superfamily. Furthermore, the 1.35-{angstrom} resolution crystal structure of the inhibited recombinant Haemophilus influenzae class C acid phosphatase was determined; this is the first structure of a phosphatase complexed with AMPS. The conformation of AMPS is identical to that of the substrate 5'-AMP, except that steric factors forcemore » a rotation of the thiophosphoryl out of the normal phosphoryl-binding pocket. This conformation is catalytically nonproductive, because the P atom is not positioned optimally for nucleophilic attack by Asp64, and the O atom of the scissile O-P bond is too far from the Asp (Asp66) that protonates the leaving group. The structure of 5'-AMP complexed with the Asp64 {yields} Asn mutant enzyme was also determined at 1.35-{angstrom} resolution. This mutation induces the substrate to adopt the same nonproductive binding mode that is observed in the AMPS complex. In this case, electrostatic considerations, rather than steric factors, underlie the movement of the phosphoryl. The structures not only provide an explanation for the inhibition by AMPS, but also highlight the precise steric and electrostatic requirements of phosphoryl recognition by class C acid phosphatases. Moreover, the structure of the Asp64 {yields} Asn mutant illustrates how a seemingly innocuous mutation can cause an unexpected structural change.« less

Photometric and fluorometric continuous kinetic assay of acid phosphatases with new substrates possessing longwave absorption and emission maxima.

PubMed

Koller, E; Wolfbeis, O S

1984-11-15

A direct and continuous kinetic method for the photometric and fluorometric determination of various acid phosphatases is described. It is based on new coumarin-derived phosphates, which after enzymatic hydrolysis undergo dissociation to form intensely colored and strongly fluorescent phenolate anions. The latter have absorption maxima ranging from 385 to 505 nm, and fluorescence maxima between 470 and 595 nm. The new substrates were compared with respect to their rate of enzymatic hydrolysis, optimum pH, and detection limits of acid phosphatase from potato and wheat germ. Detection limits of 0.001 unit/ml were found by photometry, and as low as 0.00006 unit/ml by fluorometry. The principal advantages of the new substrates over existing ones are longwave absorptions and emissions, large Stokes shifts, and the low pKa values of the corresponding phenols, thus allowing a direct and continuous assay of acid phosphatase even in weakly acidic solutions.

Intravenous Vitamin B6 Increases Resistance to Erythropoiesis-Stimulating Agents in Hemodialysis Patients: A Randomized Controlled Trial.

PubMed

Obi, Yoshitsugu; Mikami, Satoshi; Hamano, Takayuki; Obi, Yasue; Tanaka, Hirotaka; Shimomura, Akihiro; Rakugi, Hiromi; Inoue, Toru; Isaka, Yoshitaka

2016-11-01

Vitamin B6 deficiency is common in hemodialysis patients and may contribute to anemia and abnormal bone metabolism in this population. 6-month, open-label, randomized controlled parallel-group study in hemodialysis centers. Fifty-six maintenance hemodialysis patients with relatively high resistance to erythropoiesis-stimulating agents (ESA). Intravenous vitamin B6 (60 mg of intravenous pyridoxal 5'-phosphate after each thrice-weekly hemodialysis session). The primary and secondary outcomes were changes over time in ESA resistance index and bone turnover markers, respectively. The prevalence of vitamin B6 deficiency was 40% overall. Compared with the control group, the B6 group showed an upward change in ESA resistance index over time (P interaction  = .038). At week 13 (a priori-defined time point), pyridoxal 5'-phosphate administration was associated with higher ESA resistance index by 0.97 (95% confidence interval, 0.02-1.92) ×10 -2  μg ⋅darbepoetin-α/kg per g/dL⋅hemoglobin after baseline adjustment, which was not modified by baseline vitamin B6 status. There was a trend toward increase in serum erythropoietin concentrations in the B6 group after adjustment for baseline values, hemoglobin, and weekly ESA dose (P interaction  = .06). The downward changes of bone-specific alkaline phosphatase and tartrate-resistant acid phosphatase 5b in the B6 group relative to the control group were pronounced in patients without vitamin B6 deficiency (P interaction < .001 and .017, respectively), despite nonsignificant between-group difference in 1-84 parathyroid hormone. Thrice-weekly intravenous vitamin B6 (60 mg pyridoxal 5'-phosphate hydrate) worsens the response to ESA and may blunt the response of bone to parathyroid hormone in hemodialysis patients. Copyright © 2016 National Kidney Foundation, Inc. Published by Elsevier Inc. All rights reserved.

Characterization of a unique class C acid phosphatase from Clostridium perfringens.

PubMed

Reilly, Thomas J; Chance, Deborah L; Calcutt, Michael J; Tanner, John J; Felts, Richard L; Waller, Stephen C; Henzl, Michael T; Mawhinney, Thomas P; Ganjam, Irene K; Fales, William H

2009-06-01

Clostridium perfringens is a gram-positive anaerobe and a pathogen of medical importance. The detection of acid phosphatase activity is a powerful diagnostic indicator of the presence of C. perfringens among anaerobic isolates; however, characterization of the enzyme has not previously been reported. Provided here are details of the characterization of a soluble recombinant form of this cell-associated enzyme. The denatured enzyme was approximately 31 kDa and a homodimer in solution. It catalyzed the hydrolysis of several substrates, including para-nitrophenyl phosphate, 4-methylumbelliferyl phosphate, and 3' and 5' nucleoside monophosphates at pH 6. Calculated K(m)s ranged from 0.2 to 0.6 mM with maximum velocity ranging from 0.8 to 1.6 micromol of P(i)/s/mg. Activity was enhanced in the presence of some divalent cations but diminished in the presence of others. Wild-type enzyme was detected in all clinical C. perfringens isolates tested and found to be cell associated. The described enzyme belongs to nonspecific acid phosphatase class C but is devoid of lipid modification commonly attributed to this class.

Halenaquinone inhibits RANKL-induced osteoclastogenesis.

PubMed

Tsukamoto, Sachiko; Takeuchi, Tomoharu; Kawabata, Tetsuro; Kato, Hikaru; Yamakuma, Michiko; Matsuo, Kanae; El-Desoky, Ahmed H; Losung, Fitje; Mangindaan, Remy E P; de Voogd, Nicole J; Arata, Yoichiro; Yokosawa, Hideyoshi

2014-11-15

Halenaquinone was isolated from the marine sponge Petrosia alfiani as an inhibitor of osteoclastogenic differentiation of murine RAW264 cells. It inhibited the RANKL (receptor activator of nuclear factor-κB ligand)-induced upregulation of TRAP (tartrate-resistant acid phosphatase) activity as well as the formation of multinuclear osteoclasts. In addition, halenaquinone substantially suppressed RANKL-induced IκB degradation and Akt phosphorylation. Thus, these results suggest that halenaquinone inhibits RANKL-induced osteoclastogenesis at least by suppressing the NF-κB and Akt signaling pathways. Copyright © 2014 Elsevier Ltd. All rights reserved.

Phosphatidylinositol anchor of HeLa cell alkaline phosphatase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jemmerson, R.; Low, M.G.

1987-09-08

Alkaline phosphatase from cancer cells, HeLa TCRC-1, was biosynthetically labeled with either /sup 3/H-fatty acids or (/sup 3/H)ethanolamine as analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and fluorography of immunoprecipitated material. Phosphatidylinositol-specific phospholipase C (PI-PLC) released a substantial proportion of the /sup 3/H-fatty acid label from immunoaffinity-purified alkaline phosphatase but had no effect on the radioactivity of (/sup 3/H)ethanolamine-labeled material. PI-PLC also liberated catalytically active alkaline phosphatase from viable cells, and this could be selectively blocked by monoclonal antibodies to alkaline phosphatase. However, the alkaline phosphatase released from /sup 3/H-fatty acid labeled cells by PI-PLC was not radioactive. By contrast,more » treatment with bromelain removed both the /sup 3/H-fatty acid and the (/sup 3/H)ethanolamine label from purified alkaline phosphatase. Subtilisin was also able to remove the (/sup 3/H)ethanolamine label from the purified alkaline phosphatase. The /sup 3/H radioactivity in alkaline phosphatase purified from (/sup 3/H)ethanolamine-labeled cells comigrated with authentic (/sup 3/H)ethanolamine by anion-exchange chromatography after acid hydrolysis. The data suggest that the /sup 3/H-fatty acid and (/sup 3/H)ethanolamine are covalently attached to the carboxyl-terminal segment since bromelain and subtilisin both release alkaline phosphatase from the membrane by cleavage at that end of the polypeptide chain. The data are consistent with findings for other proteins recently shown to be anchored in the membrane through a glycosylphosphatidylinositol structure and indicate that a similar structure contributes to the membrane anchoring of alkaline phosphatase.« less

[Electrophoretic forms of glucose-6-phosphate dehydrogenase, acid phosphatase and esterase in Amoeba species amoebas].

PubMed

Sopina, V A

2000-01-01

Glucose-6-phosphate dehydrogenase (G6PD), acid phosphatase and esterases in free-living amoebae of 7 Amoeba species were investigated with the use of disc-electrophoresis in polyacrylamide gel. The evidence provided is suggestive that the electrophoretic isoenzyme patterns of acid phosphatase and esterases (and G6PD in some cases), in addition to a few morphological characters, can serve as a taxonomic criterion for species identification within this genus, as well as for revealing erroneously classified species and strains. It is suggested that A. indica is an independent species whose preliminary diagnosis has been given in this paper. It is concluded that A. discoides and A. lescherae are strains of A. proteus, rather than two independent species. A and As-102 amoebian strains, kept in the collection of protozoan strains and species of the Institute of Cytology RAS and referred to as strains of A. proteus, belong in reality to another Amoeba species and even to another genus within the family Amoebidae. This conclusion has been documented by results of our analysis of electrophoretic patterns of acid phosphatase and esterases in these strains.

A Eukaryotic-Type Serine/Threonine Protein Kinase Is Required for Biofilm Formation, Genetic Competence, and Acid Resistance in Streptococcus mutans

PubMed Central

Hussain, Haitham; Branny, Pavel; Allan, Elaine

2006-01-01

We report an operon encoding a eukaryotic-type serine/threonine protein kinase (STPK) and its cognate phosphatase (STPP) in Streptococcus mutans. Mutation of the gene encoding the STPK produced defects in biofilm formation, genetic competence, and acid resistance, determinants important in caries pathogenesis. PMID:16452447

Formulation and in-vitro evaluation of floating bilayer tablet of lisinopril maleate and metoprolol tartrate.

PubMed

Ijaz, Hira; Qureshi, Junaid; Danish, Zeeshan; Zaman, Muhammad; Abdel-Daim, Mohamed; Hanif, Muhammad; Waheed, Imran; Mohammad, Imran Shair

2015-11-01

The purpose of this study was to introduce the technology for the development of rate-controlled oral drug delivery system to overcome various physiological problems. Several approaches are being used for the purpose of increasing the gastric retentive time, including floating drug delivery system. Gastric floating lisinopril maleate and metoprolol tartrate bilayer tablets were formulated by direct compression method using the sodium starch glycolate, crosscarmellose sodium for IR layer. Eudragit L100, pectin, acacia as sustained release polymers in different ratios for SR metoprolol tartrate layer and sodium bicarbonate, citric acid as gas generating agents for the floating extended release layer. The floating bilayer tablets of lisinopril maleate and metoprolol tartrate were designed to overcome the various problems associated with conventional oral dosage form. Floating tablets were evaluated for floating lag time, drug contents and in-vitro dissolution profile and different kinetic release models were applied. It was clear that the different ratios of polymers affected the drug release and floating time. L2 and M4 showed good drug release profile and floating behavior. The linear regression and model fitting showed that all formulation followed Higuchi model of drug release model except M4 that followed zero order kinetic. From the study it is evident that a promising controlled release by floating bilyer tablets of lisinopril maleate and metoprolol tartrate can be developed successfully.

Characterization of a Unique Class C Acid Phosphatase from Clostridium perfringens▿

PubMed Central

Reilly, Thomas J.; Chance, Deborah L.; Calcutt, Michael J.; Tanner, John J.; Felts, Richard L.; Waller, Stephen C.; Henzl, Michael T.; Mawhinney, Thomas P.; Ganjam, Irene K.; Fales, William H.

2009-01-01

Clostridium perfringens is a gram-positive anaerobe and a pathogen of medical importance. The detection of acid phosphatase activity is a powerful diagnostic indicator of the presence of C. perfringens among anaerobic isolates; however, characterization of the enzyme has not previously been reported. Provided here are details of the characterization of a soluble recombinant form of this cell-associated enzyme. The denatured enzyme was ∼31 kDa and a homodimer in solution. It catalyzed the hydrolysis of several substrates, including para-nitrophenyl phosphate, 4-methylumbelliferyl phosphate, and 3′ and 5′ nucleoside monophosphates at pH 6. Calculated Kms ranged from 0.2 to 0.6 mM with maximum velocity ranging from 0.8 to 1.6 μmol of Pi/s/mg. Activity was enhanced in the presence of some divalent cations but diminished in the presence of others. Wild-type enzyme was detected in all clinical C. perfringens isolates tested and found to be cell associated. The described enzyme belongs to nonspecific acid phosphatase class C but is devoid of lipid modification commonly attributed to this class. PMID:19363079

Acid phosphatase patterns in microfilariae of Onchocerca volvulus s.l. from the Upper Orinoco Basin, Venezuela.

PubMed

Yarzàbal, L; Petralanda, I; Arango, M; Lobo, L; Botto, C

1983-06-01

The patterns of acid phosphatase in strains of Onchocerca volvulus s.l. which parasitize an Amerindian population (Yanomami) in Venezuela's Upper Orinoco Basin were examined by using the naphthol AS-TR phosphate method. The study sample consisted of 40 Yanomami inhabiting a savannah area at 950 m above sea level and 21 Yanomami residents of a tropical rainforest area at an altitude of 250 m. Stained intrauterine microfilariae, still within the egg case, exhibited a diffuse distribution of the enzyme in the early stages of embryonic development and a negative reaction at a more developed stage. Four of the five enzyme staining patterns described by Omar (1978) were found in the 3157 microfilariae examined from skin snips. Their distribution was: Type I--17.2%, Type III--0.5%, Type IV--75.6% and Type V--6.6%. No examples of Type II were observed. The results indicate that acid phosphatase patterns of the Upper Orinoco Onchocerca strain most resemble those of strains from Guatemala and Yemen, and are different from the African strains found in Upper Volta and Liberia. The relative frequency of acid phosphatase patterns was modified by cryopreservation of microfilariae.

Short-term variability in biomarkers of bone metabolism in sheep.

PubMed

Sousa, Cristina P; de Azevedo, Jorge T; Reis, Rui L; Gomes, Manuela E; Dias, Isabel R

2014-01-01

Changes in bone remodeling during pathological states and during their treatment can be assessed noninvasively by measuring biomarkers of bone metabolism. Their application is limited, however, by the potential biological variability in the levels of these biomarkers over time. To determine the short-term variability in biomarkers of bone metabolism in adult sheep, the authors measured serum levels of alkaline phosphatase (ALP), bone-specific alkaline phosphatase (BALP), osteocalcin (OC), N-terminal propeptide of type-III procollagen (PIIINP), deoxypyridinoline (DPD), tartrate-resistant acid phosphatase (TRAP), calcium and phosphorus intermittently over a 12-week period. There were significant differences in mean ALP activity and in phosphorus concentrations over time, but all other biomarkers showed no significant short-term variability. The results suggest that biomarkers of bone metabolism in sheep, especially the bone resorption marker DPD and the bone formation marker BALP, can be used reliably to detect changes in bone cellular activity.

Molecular studies on di-sodium tartrate molecule

NASA Astrophysics Data System (ADS)

Divya, P.; Jayakumar, S.; George, Preethamary; Shubashree, N. S.; Ahmed. M, Anees

2015-06-01

Structural characterization is important for the development of new material. The acoustical parameters such as Free Length, Internal Pressure have been measured from ultrasonic velocity, density for di sodium tartrate an optically active molecule at different temperatures using ultrasonic interferometer of frequency (2MHZ). The ultrasonic velocity increases with increase in concentration there is an increase in solute-solvent interaction. The stability constant had been calculated. SEM with EDAX studies has been done for Di-sodium tartrate an optically active molecule.

The vacuolar channel VvALMT9 mediates malate and tartrate accumulation in berries of Vitis vinifera.

PubMed

De Angeli, Alexis; Baetz, Ulrike; Francisco, Rita; Zhang, Jingbo; Chaves, Maria Manuela; Regalado, Ana

2013-08-01

Vitis vinifera L. represents an economically important fruit species. Grape and wine flavour is made from a complex set of compounds. The acidity of berries is a major parameter in determining grape berry quality for wine making and fruit consumption. Despite the importance of malic and tartaric acid (TA) storage and transport for grape berry acidity, no vacuolar transporter for malate or tartrate has been identified so far. Some members of the aluminium-activated malate transporter (ALMT) anion channel family from Arabidopsis thaliana have been shown to be involved in mediating malate fluxes across the tonoplast. Therefore, we hypothesised that a homologue of these channels could have a similar role in V. vinifera grape berries. We identified homologues of the Arabidopsis vacuolar anion channel AtALMT9 through a TBLASTX search on the V. vinifera genome database. We cloned the closest homologue of AtALMT9 from grape berry cDNA and designated it VvALMT9. The expression profile revealed that VvALMT9 is constitutively expressed in berry mesocarp tissue and that its transcription level increases during fruit maturation. Moreover, we found that VvALMT9 is targeted to the vacuolar membrane. Using patch-clamp analysis, we could show that, besides malate, VvALMT9 mediates tartrate currents which are higher than in its Arabidopsis homologue. In summary, in the present study we provide evidence that VvALMT9 is a vacuolar malate channel expressed in grape berries. Interestingly, in V. vinifera, a tartrate-producing plant, the permeability of the channel is apparently adjusted to TA.

Single crystal growth and characterization of pure and sodium-modified copper tartrate

NASA Astrophysics Data System (ADS)

Quasim, I.; Firdous, A.; Want, B.; Khosa, S. K.; Kotru, P. N.

2008-12-01

Single crystal growth of pure and modified copper tartrate crystals bearing composition (Cu) x(Na) yC 4H 4O 6· nH 2O (where x=1, 0.77, 0.65; y=0, 0.23, 0.35) is achieved using gel technique. The optimum conditions required for the growth of these crystals are worked out. The morphological development of these crystals is studied using optical and scanning electron microscopy. The dominant habit faces of the grown copper tartrate crystals are (0 0 1) and (1 1 1). Calculation of the cell parameters using CRYSFIRE software suggests that the pure copper tartrate crystal belongs to orthorhombic system with space group P2 1/c whereas the modified copper tartrate falls under tetragonal system with the space group P4 2/nbc. The external morphological development is shown to remain unaffected in the modified copper tartrate. The stoichiometric composition of the crystals is established by EDAX analysis, CH analysis, FTIR spectroscopy and thermoanalytical techniques. Thermal analysis of the grown crystals suggests that pure copper tartrate is thermally stable up to 42.84 °C whereas the modified copper tartrate crystals are stable only up to 33.11 and 25.11 °C. Calculation of the percentage weight loss from the thermogram supplemented by EDAX/CH analysis and FTIR spectroscopy suggest that the chemical formula of pure copper tartrate crystal is CuC 4H 4O 6·3H 2O whereas the chemical formula for the modified copper tartrate crystals is (Cu) 0.77(Na) 0.23C 4H 4O 6·3H 2O and (Cu) 0.65(Na) 0.35 C 4H 4O 6·H 2O.

The Effect of Lunasin on Receptor Activator of Nuclear Factor Kappa-B Ligand-mediated Osteoclast Formation from RAW 264.7 Cells.

PubMed

Bachala, Daisy; El-Refai, Nivine; Greenfield, Edward; Aminoshariae, Anita; Mickel, Andre

2018-06-01

To date, no study has investigated the antiresorptive property of lunasin. Hence, the present study aimed to assess the ability of lunasin to inhibit the osteoclast formation using RAW 264.7 cells. We hypothesized that lunasin is able to inhibit osteoclast formation. In the present study, the murine monocytic cell line RAW 264.7 was induced to differentiate into mature osteoclasts in the presence of recombinant receptor activator of nuclear factor kappa-B ligand. Tartrate-resistant acid phosphatase, a marker of osteoclasts, was used to identify osteoclasts. Cell lines were divided into different groups and exposed to different concentrations of 50 μmol/L, 75 μmol/L, and 100 μmol/L active and inactive lunasin. The control group was RAW 264.7 cells with receptor activator of nuclear factor kappa-B ligand. Tartrate-resistant acid phosphatase-positive cells of 3 or more nuclei, indicative of mature osteoclasts, were counted by 3 observers. The mean number of the data collected was analyzed using 1-way analysis of variance and the multiple comparison post hoc Bonferroni correction. There was a significant difference in the reduction of osteoclast formation in all the active lunasin groups (P < .001) compared with the control group and the inactive lunasin group (P < .001). Considering the suppressive effect of lunasin on osteoclastogenesis, the use of lunasin as a potential antiresorptive agent can be evaluated in future studies. Copyright © 2018 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Bis-enoxacin Inhibits Bone Resorption and Orthodontic Tooth Movement

PubMed Central

Toro, E.J.; Zuo, J.; Guiterrez, A.; La Rosa, R.L.; Gawron, A.J.; Bradaschia-Correa, V.; Arana-Chavez, V.; Dolce, C.; Rivera, M.F.; Kesavalu, L.; Bhattacharyya, I.; Neubert, J.K.; Holliday, L.S.

2013-01-01

Enoxacin inhibits binding between the B-subunit of vacuolar H+-ATPase (V-ATPase) and microfilaments, and also between osteoclast formation and bone resorption in vitro. We hypothesized that a bisphosphonate derivative of enoxacin, bis-enoxacin (BE), which was previously studied as a bone-directed antibiotic, might have similar activities. BE shared a number of characteristics with enoxacin: It blocked binding between the recombinant B-subunit and microfilaments and inhibited osteoclastogenesis in cell culture with IC50s of about 10 µM in each case. BE did not alter the relative expression levels of various osteoclast-specific proteins. Even though tartrate-resistant acid phosphatase 5b was expressed, proteolytic activation of the latent pro-enzyme was inhibited. However, unlike enoxacin, BE stimulated caspase-3 activity. BE bound to bone slices and inhibited bone resorption by osteoclasts on BE-coated bone slices in cell culture. BE reduced the amount of orthodontic tooth movement achieved in rats after 28 days. Analysis of these data suggests that BE is a novel anti-resorptive molecule that is active both in vitro and in vivo and may have clinical uses. Abbreviations: BE, bis-enoxacin; V-ATPase, vacuolar H+-ATPase; TRAP, tartrate-resistant acid phosphatase; αMEM D10, minimal essential media, alpha modification with 10% fetal bovine serum; SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis; RANKL, receptor activator of nuclear factor kappa B-ligand; NFATc1, nuclear factor of activated T-cells; ADAM, a disintegrin and metalloprotease domain; OTM, orthodontic tooth movement. PMID:23958763

Molecular basis for TPR domain-mediated regulation of protein phosphatase 5.

PubMed

Yang, Jing; Roe, S Mark; Cliff, Matthew J; Williams, Mark A; Ladbury, John E; Cohen, Patricia T W; Barford, David

2005-01-12

Protein phosphatase 5 (Ppp5) is a serine/threonine protein phosphatase comprising a regulatory tetratricopeptide repeat (TPR) domain N-terminal to its phosphatase domain. Ppp5 functions in signalling pathways that control cellular responses to stress, glucocorticoids and DNA damage. Its phosphatase activity is suppressed by an autoinhibited conformation maintained by the TPR domain and a C-terminal subdomain. By interacting with the TPR domain, heat shock protein 90 (Hsp90) and fatty acids including arachidonic acid stimulate phosphatase activity. Here, we describe the structure of the autoinhibited state of Ppp5, revealing mechanisms of TPR-mediated phosphatase inhibition and Hsp90- and arachidonic acid-induced stimulation of phosphatase activity. The TPR domain engages with the catalytic channel of the phosphatase domain, restricting access to the catalytic site. This autoinhibited conformation of Ppp5 is stabilised by the C-terminal alphaJ helix that contacts a region of the Hsp90-binding groove on the TPR domain. Hsp90 activates Ppp5 by disrupting TPR-phosphatase domain interactions, permitting substrate access to the constitutively active phosphatase domain, whereas arachidonic acid prompts an alternate conformation of the TPR domain, destabilising the TPR-phosphatase domain interface.

Ultrastructural localization of acid phosphatase in some bacteria, after treatment with Lubrol W1.

PubMed

Cherepova, N; Spasova, D

1996-01-01

The ultracytochemical localization of acid phosphatase from some bacteria (Listeria monocytogenes, Salmonella typhimurium, Pseudomonas pseudomallei and Pseudomonas aeruginosa) was dependent on the changes in the lipoprotein content of the membranes as a result of the action of the Lubrol W1.

Alterations to N-linked oligosaccharides which affect intracellular transport rates and regulated secretion but not sorting of lysosomal acid phosphatase in Dictyostelium discoideum.

PubMed

Bush, J M; Ebert, D L; Cardelli, J A

1990-11-15

The importance of N-linked oligosaccharides and their associated modifications in the transport, sorting, and secretion of lysosomal acid phosphatase was investigated using three mutant Dictyostelium cell lines. These mutants synthesize altered N-linked oligosaccharides with the following properties: (i) in strain HL244 carbohydrate side chains lack mannose 6-sulfate residues, (ii) in strain M31 the side chains retain the two alpha-1,3-linked glucose residues resulting in less sulfate and methylphosphate modifications, and (iii) in strain HL243 the nonglucosylated branches are missing three of the outer mannose sugars and the oligosaccharides contain fewer sulfate and phosphate modifications. Lysosomal enzymes in both HL243 and HL244 are also missing a shared epitope termed common antigen-1 (CA-1), which consists in part of mannose 6-sulfate moieties. No increases were observed in the secretion of radiolabeled acid phosphatase or acid phosphatase activity during growth in any of the mutant cell lines, suggesting that the enzyme was correctly sorted to lysosomes. In support of this, Percoll gradient fractionations and indirect immunofluorescence microscopy indicated that acid phosphatase was transported to lysosomes in all cell lines. However, radiolabel pulse chase protocols indicated that newly synthesized acid phosphatase was transported out of the endoplasmic reticulum (ER) and into lysosomes at a two- to threefold slower rate in HL243 and at a sixfold slower rate in M31. The rate of transport of acid phosphatase from the ER to the Golgi was reduced only twofold in M31 as determined by digestion of newly synthesized enzyme with endoglycosidose H. This suggests that certain alterations in carbohydrate structure may only slightly affect transport of the enzyme from the ER to the Golgi but these alterations may greatly delay transport from the Golgi or post-Golgi compartments to lysosomes. Finally all three mutants secreted acid phosphatase at significantly lower

Acute effects of dietary fatty acids on osteclastogenesis via RANKL/RANK/OPG system.

PubMed

Naranjo, M Carmen; Garcia, Indara; Bermudez, Beatriz; Lopez, Sergio; Cardelo, Magdalena P; Abia, Rocio; Muriana, Francisco J G; Montserrat-de la Paz, Sergio

2016-11-01

Postprandial state is directly linked with chronic diseases. We hypothesized that dietary fats may have acute effects on health status by modulating osteoclast differentiation and activation in a fatty acid-dependent manner. In healthy subjects, a fat-enriched meal increased plasma levels of the RANKL (receptor activator of nuclear factor κB ligand)/OPG (osteoprotegerin) ratio (SFAs > MUFAs = PUFAs) in the postprandial state. Postprandial TRL-SFAs enhanced tartrate-resistant acid phosphatase (TRAP) activity and the expression of osteoclast marker genes (TRAP, OSCAR, RANK, and CATHK) while downregulated the expression of OPG gene in human monocyte-derived osteoclasts. These effects were not observed with monounsaturated fatty acid (MUFA)-enriched postprandial triglyceride-rich lipoproteins (TRLs). Moreover, postprandial TRL-SFAs increased the release of osteoclastogenic cytokines (TNF-α, IL-1β, and IL-6) meanwhile TRL-MUFAs and TRL-PUFAs increased the release of anti-osteoclastogenic cytokines (IL-4 and IL-10) in the medium of human monocyte-derived osteoclasts. For the first time, we show that postprandial TRLs are metabolic entities with osteoclastogenic activity and that this property is related to the type of dietary fatty acid in the meal. The osteoclastogenic potency was as follows: SFAs > MUFAs = PUFAs. These exciting findings open opportunities for developing nutritional strategies with olive oil as the principal dietary source of MUFAs, notably oleic acid, to prevent development and progression of osteoclast-related diseases. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Enzymatic Production of Ascorbic Acid-2-phosphate by Recombinant Acid Phosphatase.

PubMed

Zheng, Kai; Song, Wei; Sun, Anran; Chen, Xiulai; Liu, Jia; Luo, Qiuling; Wu, Jing

2017-05-24

In this study, an environmentally friendly and efficient enzymatic method for the synthesis of l-ascorbic acid-2-phosphate (AsA-2P) from l-ascorbic acid (AsA) was developed. The Pseudomonas aeruginosa acid phosphatase (PaAPase) was expressed in Escherichia coli BL21. The optimal temperature, optimal pH, K m , k cat , and catalytic efficiency of recombinant PaAPase were 50 °C, 5.0, 93 mM, 4.2 s -1 , and 2.7 mM -1 min -1 , respectively. The maximal dry cell weight and PaAPase phosphorylating activity reached 8.5 g/L and 1127.7 U/L, respectively. The highest AsA-2P concentration (50.0 g/L) and the maximal conversion (39.2%) were obtained by incubating 75 g/L intact cells with 88 g/L AsA and 160 g/L sodium pyrophosphate under optimal conditions (0.1 mM Ca 2+ , pH 4.0, 30 °C) for 10 h; the average AsA-2P production rate was 5.0 g/L/h, and the AsA-2P production system was successfully scaled up to a 7.5 L fermenter. Therefore, the enzymatic process showed great potential for production of AsA-2P in industry.

Inhibition of hydrolytic enzymes by gold compounds. I. beta-Glucuronidase and acid phosphatase by sodium tetrachloroaurate (III) and potassium tetrabromoaurate (III).

PubMed

Lee, M T; Ahmed, T; Friedman, M E

1989-01-01

Purified bovine liver beta-glucuronidase (beta-D-glucuronide glucuronohydrolase, EC 3.2.1.32) and wheat germ acid phosphatase (orthophosphoric monoesterphosphohydrolase, EC 3.1.3.2) were inhibited with freshly dissolved and 24 h aquated tetrahaloaurate (III) compounds. Rate and equilibrium inhibition constants were measured. From this data two acid phosphatases species were observed. Equilibrium inhibition constants ranged from 1 to 12.5 microM for the various gold compounds toward both enzymes. The first order rate constants ranged between 0.005 and 0.04 min.-1 for most reactions with the exception of the fast reacting acid phosphatase which had values as high as 2.6 and 2.8 min.-1. It is observed that the beta-glucuronidase is rapidly inhibited during the equilibrium phase before the more slower reaction covalent bond formation takes place. The acid phosphatases form the covalent bonds more rapidly, especially the faster reacting species suggesting a unique difference in the active site geometry to that of the more slowly reacting species. The tightly bonded gold (III)-enzyme complex is probably the reason for its toxicity and non-anti-inflammatory use as a drug.

Acid and Alkaline Phosphatase Levels in GCF during Orthodontic Tooth Movement

PubMed Central

Farahani, Mohammad; Safavi, Seyed Mohammadreza; Dianat, Omid; Khoramian Tusi, Somayeh; Younessian, Farnaz

2015-01-01

Statement of the Problem The present constituents of gingival crevicular fluid (GCF) can reflect the changes occurring in underlying tissues. Considering variety of biologic bone markers, alkaline phosphatase and acid phosphatase have been examined as bone turn over markers in orthodontic tooth movement. Purpose The current study designed in a longitudinal pattern to determine the changes of acid and alkaline phosphatase (ACP & ALP) in GCF during orthodontic tooth movement. Materials and Method An upper canines from twelve patients (mean age: 14±2 years) undergoing extraction orthodontic treatment for distal movement served as the test tooth (DC), and its contralateral (CC) and antagonist (AC) canines were used as controls. The CC was included in orthodontic appliance without orthodontic force; the AC was free from any orthodontic appliance. The GCF around the experimental teeth was harvested from mesial and distal tooth sites immediately before appliance placement (T0), and 14 (T2) and 28 days (T3) after it and ALP and ACP concentration were determined spectrophotometrically. Results ALP concentration was elevated significantly in DC and CC groups at days 14 and 28 compared with the AC. In DC group, the ALP was significantly greater in mesial sites than distal site, while no significant changes were found between both sites of CC. The peak level of ALP was observed in mesial sites of DC at T2. Regarding ACP, significant elevation of this enzyme was seen in DC group both in mesial and distal sites at T2 and T3. The peak level of this enzyme was seen at T2. Conclusion Monitoring simultaneous changes of ALP and ACP levels in GCF can reflect the tissue responses occur in periodontium during bone formation and bone resorption during orthodontic tooth movement, respectively. PMID:26535403

The use of acid phosphatase test papers for DNA profiling.

PubMed

Reshef, A; Barash, M; Gallili, N; Michael, A; Brauner, P

2005-01-01

The acid phosphatase (AP) test is a routine assay used to screen casework items for the possible presence of semen. This colour test is carried out on filter paper which is retained after testing. Two-year-old AP test papers were found to contain sufficient DNA for short tandem repeat (STR) profiling. Prior to polymerase chain reaction (PCR) amplification, the DNA was preferentially separated into sperm depleted and sperm enriched cell fractions. The implication of these findings for past and present cases is discussed.

Identification of an additional member of the protein-tyrosine-phosphatase family: evidence for alternative splicing in the tyrosine phosphatase domain.

PubMed Central

Matthews, R J; Cahir, E D; Thomas, M L

1990-01-01

Protein-tyrosine-phosphatases (protein-tyrosine-phosphate phosphohydrolase, EC 3.13.48) have been implicated in the regulation of cell growth; however, to date few tyrosine phosphatases have been characterized. To identify additional family members, the cDNA for the human tyrosine phosphatase leukocyte common antigen (LCA; CD45) was used to screen, under low stringency, a mouse pre-B-cell cDNA library. Two cDNA clones were isolated and sequence analysis predicts a protein sequence of 793 amino acids. We have named the molecule LRP (LCA-related phosphatase). RNA transfer analysis indicates that the cDNAs were derived from a 3.2-kilobase mRNA. The LRP mRNA is transcribed in a wide variety of tissues. The predicted protein structure can be divided into the following structural features: a short 19-amino acid leader sequence, an exterior domain of 123 amino acids that is predicted to be highly glycosylated, a 24-amino acid membrane-spanning region, and a 627-amino acid cytoplasmic region. The cytoplasmic region contains two approximately 260-amino acid domains, each with homology to the tyrosine phosphatase family. One of the cDNA clones differed in that it had a 108-base-pair insertion that, while preserving the reading frame, would disrupt the first protein-tyrosine-phosphatase domain. Analysis of genomic DNA indicates that the insertion is due to an alternatively spliced exon. LRP appears to be evolutionarily conserved as a putative homologue has been identified in the invertebrate Styela plicata. Images PMID:2162042

Improved Acetic Acid Resistance in Saccharomyces cerevisiae by Overexpression of the WHI2 Gene Identified through Inverse Metabolic Engineering

PubMed Central

Chen, Yingying; Stabryla, Lisa

2016-01-01

Development of acetic acid-resistant Saccharomyces cerevisiae is important for economically viable production of biofuels from lignocellulosic biomass, but the goal remains a critical challenge due to limited information on effective genetic perturbation targets for improving acetic acid resistance in the yeast. This study employed a genomic-library-based inverse metabolic engineering approach to successfully identify a novel gene target, WHI2 (encoding a cytoplasmatic globular scaffold protein), which elicited improved acetic acid resistance in S. cerevisiae. Overexpression of WHI2 significantly improved glucose and/or xylose fermentation under acetic acid stress in engineered yeast. The WHI2-overexpressing strain had 5-times-higher specific ethanol productivity than the control in glucose fermentation with acetic acid. Analysis of the expression of WHI2 gene products (including protein and transcript) determined that acetic acid induced endogenous expression of Whi2 in S. cerevisiae. Meanwhile, the whi2Δ mutant strain had substantially higher susceptibility to acetic acid than the wild type, suggesting the important role of Whi2 in the acetic acid response in S. cerevisiae. Additionally, overexpression of WHI2 and of a cognate phosphatase gene, PSR1, had a synergistic effect in improving acetic acid resistance, suggesting that Whi2 might function in combination with Psr1 to elicit the acetic acid resistance mechanism. These results improve our understanding of the yeast response to acetic acid stress and provide a new strategy to breed acetic acid-resistant yeast strains for renewable biofuel production. PMID:26826231

Effect of chromium on vertebrae, femur and calvaria of adult male rats.

PubMed

Sankaramanivel, S; Jeyapriya, R; Hemalatha, D; Djody, S; Arunakaran, J; Srinivasan, N

2006-06-01

Alloys of chromium have a long history of success in the surgical treatment of many orthopaedic defects. Nonetheless, prostheses loosening are commonly found around arthoplasties due to corrosion of metals. On this basis, it is hypothesized that chromium accumulation interferes with remodeling of bone. The present study aims to analyse the toxic effects of chromium on bone phosphatases in various regions of the bone in rats. Rats were treated with chromium intraperitoneally (0.5 mg/kg) in the form of potassium dichromate for 5 days. The accumulation of chromium is approximately 5.2-fold in the vertebrae, 8.9-fold in the femur and 8.7-fold in the calvaria, when compared to control. Chromium administration significantly reduced the activity of enzymes, eg, alkaline phosphatase (ALP) and tartrate-resistant acid phosphatase (TRAP). The study revealed a significant increase in the concentration of calcium, altered bone formation rate and bone morphology in the femur, vertebrae and calvaria. The interesting findings of the current study suggest altered bone turnover.

Hydrolytic and alcoholytic dephosphorylation of nucleotides by acid phosphatase in the presence of ethanol.

PubMed

Tomaszewski, M; Buchowicz, J

1971-08-01

The effect of ethanol on the activity of acid phosphatase from wheat germ was studied, by using ribonucleoside monophosphates as the enzyme substrates. The nucleotides were effectively degraded to the corresponding nucleosides in the presence of ethanol at all concentrations tested, including a 96% (v/v) solution. However, the nucleotide dephosphorylation was accompanied by the liberation of orthophosphate only when the concentration of ethanol in the assay mixture did not exceed 15%. No inorganic phosphate was liberated when ethanol was present at higher concentrations. Instead, monoethyl phosphate was formed in quantities expected for orthophosphate. The results are explained in terms of phosphatase-catalysed alcoholysis.

The IBO germination quantitative trait locus encodes a phosphatase 2C-related variant with a nonsynonymous amino acid change that interferes with abscisic acid signaling.

PubMed

Amiguet-Vercher, Amélia; Santuari, Luca; Gonzalez-Guzman, Miguel; Depuydt, Stephen; Rodriguez, Pedro L; Hardtke, Christian S

2015-02-01

Natural genetic variation is crucial for adaptability of plants to different environments. Seed dormancy prevents precocious germination in unsuitable conditions and is an adaptation to a major macro-environmental parameter, the seasonal variation in temperature and day length. Here we report the isolation of IBO, a quantitative trait locus (QTL) that governs c. 30% of germination rate variance in an Arabidopsis recombinant inbred line (RIL) population derived from the parental accessions Eilenburg-0 (Eil-0) and Loch Ness-0 (Lc-0). IBO encodes an uncharacterized phosphatase 2C-related protein, but neither the Eil-0 nor the Lc-0 variant, which differ in a single amino acid, have any appreciable phosphatase activity in in vitro assays. However, we found that the amino acid change in the Lc-0 variant of the IBO protein confers reduced germination rate. Moreover, unlike the Eil-0 variant of the protein, the Lc-0 variant can interfere with the activity of the phosphatase 2C ABSCISIC ACID INSENSITIVE 1 in vitro. This suggests that the Lc-0 variant possibly interferes with abscisic acid signaling, a notion that is supported by physiological assays. Thus, we isolated an example of a QTL allele with a nonsynonymous amino acid change that might mediate local adaptation of seed germination timing. © 2014 The Authors. New Phytologist © 2014 New Phytologist Trust.

[Phosphatase activity in Amoeba proteus at pH 9.0].

PubMed

Sopina, V A

2007-01-01

In the free-living amoeba Amoeba proteus (strain B), after PAAG disk-electrophoresis of the homogenate supernatant, at using 1-naphthyl phosphate as a substrate and pH 9.0, three forms of phosphatase activity were revealed; they were arbitrarily called "fast", "intermediate", and "slow" phosphatases. The fast phosphatase has been established to be a fraction of lysosomal acid phosphatase that preserves some low activity at alkaline pH. The question as to which particular class the intermediate phosphatase belongs to has remained unanswered: it can be both acid phosphatase and protein tyrosine phosphatase (PTP). Based on data of inhibitor analysis, large substrate specificity, results of experiments with reactivation by Zn ions after inactivation with EDTA, other than in the fast and intermediate phosphatases localization in the amoeba cell, it is concluded that only slow phosphatase can be classified as alkaline phosphatase (EC 3.1.3.1).

The erythrocyte acid phosphatase isoenzyme distribution among the negroid population of Rhodesia.

PubMed

Kobus, H J; Fowler, J C

1979-01-01

The value of the erythrocyte acid phosphatase isoenzyme system as a method for blood typing in forensic science in Rhodesia has been evaluated. Three hundred and three blood samples from negroid people were examined. The high incidence of the B phenotype (72%) results in a poor division of the population using this system. The R allele which has been found in other negroid peoples also occurs in the Rhodesian population.

Enhancement of acid phosphatase secretion and Pi acquisition in Suaeda fruticosa on calcareous soil by high saline level.

PubMed

Labidi, Nehla; Snoussi, Sana; Ammari, Manel; Metoui, Wissal; Ben Yousfi, N; Hamrouni, Lamia; Abdelly, C

2010-12-01

The aim of this study was to identify the relationship between the adaptive processes of Suaeda fruticosa for Pi acquisition and the physic-chemical and biological characteristics of two soil types under moderate and high saline conditions. Four treatments were established in pots: namely SS100, SS600, CS100 and CS600 where SS stood for sandy soil and CS for calcareous soil, and the indexes 100 and 600 were NaCl concentrations (mM) in irrigation distilled water. Assuming that Pi per g of plant biomass is an indicator of plant efficiency for P acquisition, the results showed that Pi acquisition was easiest on SS100 and was difficult on CS100. The differences in Pi acquisition between plants on SS100 and CS100 could be attributed to the low root surface area (-30%) and to the low alkaline phosphatases (Pases) activities (-50%) in calcareous rhizospheric soil. The high salinity level had no effect on the efficiency of P acquisition on SS but increased this parameter on CS (+50%). In the latter soil type, high acid phosphatase activities were observed in rhizospheric soil at high salinity level. Acid phosphatase seemed to be secreted from the roots. The higher secretion of acid phosphatase in this soil was related to the root lipid peroxidation in response to elevated salinity associated with the augmentation of unsaturated acids which might induce an oxidative damage of the root membrane. Thus we can conclude that in deficient soil such as calcareous, the efficiency of P acquisition in S. fruticosa which was difficult at moderate salinity level can be enhanced by high salinity level.

Improved Acetic Acid Resistance in Saccharomyces cerevisiae by Overexpression of the WHI2 Gene Identified through Inverse Metabolic Engineering.

PubMed

Chen, Yingying; Stabryla, Lisa; Wei, Na

2016-01-29

Development of acetic acid-resistant Saccharomyces cerevisiae is important for economically viable production of biofuels from lignocellulosic biomass, but the goal remains a critical challenge due to limited information on effective genetic perturbation targets for improving acetic acid resistance in the yeast. This study employed a genomic-library-based inverse metabolic engineering approach to successfully identify a novel gene target, WHI2 (encoding a cytoplasmatic globular scaffold protein), which elicited improved acetic acid resistance in S. cerevisiae. Overexpression of WHI2 significantly improved glucose and/or xylose fermentation under acetic acid stress in engineered yeast. The WHI2-overexpressing strain had 5-times-higher specific ethanol productivity than the control in glucose fermentation with acetic acid. Analysis of the expression of WHI2 gene products (including protein and transcript) determined that acetic acid induced endogenous expression of Whi2 in S. cerevisiae. Meanwhile, the whi2Δ mutant strain had substantially higher susceptibility to acetic acid than the wild type, suggesting the important role of Whi2 in the acetic acid response in S. cerevisiae. Additionally, overexpression of WHI2 and of a cognate phosphatase gene, PSR1, had a synergistic effect in improving acetic acid resistance, suggesting that Whi2 might function in combination with Psr1 to elicit the acetic acid resistance mechanism. These results improve our understanding of the yeast response to acetic acid stress and provide a new strategy to breed acetic acid-resistant yeast strains for renewable biofuel production. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Enantioseparation of Racemic Flurbiprofen by Aqueous Two-Phase Extraction With Binary Chiral Selectors of L-dioctyl Tartrate and L-tryptophan.

PubMed

Chen, Zhi; Zhang, Wei; Wang, Liping; Fan, Huajun; Wan, Qiang; Wu, Xuehao; Tang, Xunyou; Tang, James Z

2015-09-01

A novel method for chiral separation of flurbiprofen enantiomers was developed using aqueous two-phase extraction (ATPE) coupled with biphasic recognition chiral extraction (BRCE). An aqueous two-phase system (ATPS) was used as an extracting solvent which was composed of ethanol (35.0% w/w) and ammonium sulfate (18.0% w/w). The chiral selectors in ATPS for BRCE consideration were L-dioctyl tartrate and L-tryptophan, which were screened from amino acids, β-cyclodextrin derivatives, and L-tartrate esters. Factors such as the amounts of L-dioctyl tartrate and L-tryptophan, pH, flurbiprofen concentration, and the operation temperature were investigated in terms of chiral separation of flurbiprofen enantiomers. The optimum conditions were as follows: L-dioctyl tartrate, 80 mg; L-tryptophan, 40 mg; pH, 4.0; flurbiprofen concentration, 0.10 mmol/L; and temperature, 25 °C. The maximum separation factor α for flurbiprofen enantiomers could reach 2.34. The mechanism of chiral separation of flurbiprofen enantiomers is discussed and studied. The results showed that synergistic extraction has been established by L-dioctyl tartrate and L-tryptophan, which enantioselectively recognized R- and S-enantiomers in top and bottom phases, respectively. Compared to conventional liquid-liquid extraction, ATPE coupled with BRCE possessed higher separation efficiency and enantioselectivity without the use of any other organic solvents. The proposed method is a potential and powerful alternative to conventional extraction for separation of various enantiomers. © 2015 Wiley Periodicals, Inc.

Structure of dysprosium(111) dl-tartrate dimer in aqueous solution

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chevela, V.V.; Vulfson, S.G.; Salnikov, Y.I.

1994-10-01

The paramagnetic birefringence method was supplemented by numerical simulation to determine the molar paramagnetic-birefringence constant of the dysprosium dl-tartrate dimer Dy{sub 2}(d-L)(l-L){sup 2-} (I), where d-L{sup 4-} and l-L{sup 4-} are the deprotonated d- and l-tartaric acid molecules, respectively. The structure of the ligand and hydration surroundings of I was modeled by molecular mechanic calculations (the Dashevskii-Pylamovatyi model). It is shown that adequate results can be obtained only if one takes into account the coordination of I to the Na{sup +} ion.

Association between serum alkaline phosphatase and primary resistance to erythropoiesis stimulating agents in chronic kidney disease: a secondary analysis of the HERO trial.

PubMed

Badve, Sunil V; Zhang, Lei; Coombes, Jeff S; Pascoe, Elaine M; Cass, Alan; Clarke, Philip; Ferrari, Paolo; McDonald, Stephen P; Morrish, Alicia T; Pedagogos, Eugenie; Perkovic, Vlado; Reidlinger, Donna; Scaria, Anish; Walker, Rowan; Vergara, Liza A; Hawley, Carmel M; Johnson, David W

2015-01-01

Erythropoiesis stimulating agent (ESA)-resistant anemia is common in chronic kidney disease (CKD). To evaluate the determinants of severity of ESA resistance in patients with CKD and primary ESA-resistance. Secondary analysis of a randomized controlled trial (the Handling Erythropoietin Resistance with Oxpentifylline, HERO). 53 adult patients with CKD stage 4 or 5 and primary ESA-resistant anemia (hemoglobin ≤120 g/L, ESA resistance index [ERI] ≥1.0 IU/kg/week/gHb for erythropoietin or ≥0.005 μg/kg/week/gHb for darbepoeitin, no cause for ESA-resistance identified). Iron studies, parathyroid hormone, albumin, liver enzymes, phosphate or markers of oxidative stress and inflammation. Participants were divided into tertiles of ERI. Multinomial logistic regression was used to analyse the determinants of ERI tertiles. All patients, except one, were receiving dialysis for end-stage kidney disease. The mean ± SD ERI values in the low (n = 18), medium (n = 18) and high (n = 17) ERI tertiles were 1.4 ± 0.3, 2.3 ± 0.2 and 3.5 ± 0.8 IU/kg/week/gHb, respectively (P < 0.001). There were no significant differences observed in age, gender, ethnicity, cause of kidney disease, diabetes, iron studies, parathyroid hormone, albumin, liver enzymes, phosphate or markers of oxidative stress and inflammation between the ERI tertiles. The median [inter-quartile range] serum alkaline phosphatase concentrations in the low, medium and high ERI tertiles were 89 [64,121], 99 [76,134 and 148 [87,175] U/L, respectively (P = 0.054). There was a weak but statistically significant association between ERI and serum alkaline phosphatase (R(2) = 0.06, P = 0.03). Using multinomial logistic regression, the risk of being in the high ERI tertile relative to the low ERI tertile increased with increasing serum alkaline phosphatase levels (P = 0.02). No other variables were significantly associated with ERI. Small sample size; bone

Preparative resolution of D,L-threonine catalyzed by immobilized phosphatase.

PubMed

Scollar, M P; Sigal, G; Klibanov, A M

1985-03-01

Hydrolysis of L- and D-O-phosphothreonines catalyzed by four different phosphatases, alkaline phosphatases from calf intestine and E. coli and acid phosphatases from wheat germ and potato, has been kinetically studied. Alkaline phosphatases were found to have comparable reactivities towards the optical isomers. On the other hand, both acid phosphatases displayed a marked stereoselectivity, hydrolyzing the L-ester much faster than its D counterpart. Wheat germ acid phosphatase was the most stereoselective enzyme: V(L)/V(D) = 24 and K(m,L)/K(m,D) = 0.17. This enzyme was immobilized (in k-carrageenan gel, followed by crosslinking with glutaraldehyde) and used for the preparative resolution of D,L-threonine: the latter was first chemically O-phosphorylated and then asymmetrically hydrolyzed by the immobilized phosphatase. As a result, gram quantities of L-threonine of high optical purity and O-phospho-D-threonine were prepared. Immobilized wheat germ phosphatase has been tested for the resolution of other racemic alcohols: serine, 2-amino-1-butanol, 1-amino-2-propanol, 2-octanol, and menthol. In all those cases, the enzyme was either not sufficiently stereoselective or too slow for preparative resolutions.

Development of sustained release capsules containing "coated matrix granules of metoprolol tartrate".

PubMed

Siddique, Sabahuddin; Khanam, Jasmina; Bigoniya, Papiya

2010-09-01

The objective of this investigation was to prepare sustained release capsule containing coated matrix granules of metoprolol tartrate and to study its in vitro release and in vivo absorption. The design of dosage form was performed by choosing hydrophilic hydroxypropyl methyl cellulose (HPMC K100M) and hydrophobic ethyl cellulose (EC) polymers as matrix builders and Eudragit® RL/RS as coating polymers. Granules were prepared by composing drug with HPMC K100M, EC, dicalcium phosphate by wet granulation method with subsequent coating. Optimized formulation of metoprolol tartrate was formed by using 30% HPMC K100M, 20% EC, and ratio of Eudragit® RS/RL as 97.5:2.5 at 25% coating level. Capsules were filled with free flowing optimized granules of uniform drug content. This extended the release period upto 12 h in vitro study. Similarity factor and mean dissolution time were also reported to compare various dissolution profiles. The network formed by HPMC and EC had been coupled satisfactorily with the controlled resistance offered by Eudragit® RS. The release mechanism of capsules followed Korsemeyer-Peppas model that indicated significant contribution of erosion effect of hydrophilic polymer. Biopharmaceutical study of this optimized dosage form in rabbit model showed 10 h prolonged drug release in vivo. A close correlation (R(2) = 0.9434) was established between the in vitro release and the in vivo absorption of drug. The results suggested that wet granulation with subsequent coating by fluidized bed technique, is a suitable method to formulate sustained release capsules of metoprolol tartrate and it can perform therapeutically better than conventional immediate release dosage form.

Overexpression of Poplar Pyrabactin Resistance-Like Abscisic Acid Receptors Promotes Abscisic Acid Sensitivity and Drought Resistance in Transgenic Arabidopsis.

PubMed

Yu, Jingling; Yang, Lei; Liu, Xiaobing; Tang, Renjie; Wang, Yuan; Ge, Haiman; Wu, Mengting; Zhang, Jiang; Zhao, Fugeng; Luan, Sheng; Lan, Wenzhi

2016-01-01

Drought stress is an important environmental factor limiting productivity of plants, especially fast growing species with high water consumption like poplar. Abscisic acid (ABA) is a phytohormone that positively regulates seed dormancy and drought resistance. The PYR1 (Pyrabactin Resistance 1)/ PYRL (PYR-Like)/ RCAR (Regulatory Component of ABA Receptor) (PYR/PYL/RCAR) ABA receptor family has been identified and widely characterized in Arabidopsis thaliana. However, their functions in poplars remain unknown. Here, we report that 2 of 14 PYR/PYL/RCAR orthologues in poplar (Populus trichocarpa) (PtPYRLs) function as a positive regulator of the ABA signal transduction pathway. The Arabidopsis transient expression and yeast two-hybrid assays showed the interaction among PtPYRL1 and PtPYRL5, a clade A protein phosphatase 2C, and a SnRK2, suggesting that a core signalling complex for ABA signaling pathway exists in poplars. Phenotypic analysis of PtPYRL1 and PtPYRL5 transgenic Arabidopsis showed that these two genes positively regulated the ABA responses during the seed germination. More importantly, the overexpression of PtPYRL1 and PtPYRL5 substantially improved ABA sensitivity and drought stress tolerance in transgenic plants. In summary, we comprehensively uncovered the properties of PtPYRL1 and PtPYRL5, which might be good target genes to genetically engineer drought-Resistant plants.

Phosphatase synthesis in Klebsiella (Aerobacter) aerogenes growing in continuous culture

PubMed Central

Bolton, P. G.; Dean, A. C. R.

1972-01-01

1. Phosphatase synthesis was studied in Klebsiella aerogenes grown in a wide range of continuous-culture systems. 2. Maximum acid phosphatase synthesis was associated with nutrient-limited, particularly carbohydrate-limited, growth at a relatively low rate, glucose-limited cells exhibiting the highest activity. Compared with glucose as the carbon-limiting growth material, other sugars not only altered the activity but also changed the pH–activity profile of the enzyme(s). 3. The affinity of the acid phosphatase in glucose-limited cells towards p-nitrophenyl phosphate (Km 0.25–0.43mm) was similar to that of staphylococcal acid phosphatase but was ten times greater than that of the Escherichia coli enzyme. 4. PO43−-limitation derepressed alkaline phosphatase synthesis but the amounts of activity were largely independent of the carbon source used for growth. 5. The enzymes were further differentiated by the effect of adding inhibitors (F−, PO43−) and sugars to the reaction mixture during the assays. In particular, it was shown that adding glucose, but not other sugars, stimulated the rate of hydrolysis of p-nitrophenyl phosphate by the acid phosphatase in carbohydrate-limited cells at low pH values (<4.6) but inhibited it at high pH values (>4.6). Alkaline phosphatase activity was unaffected. 6. The function of phosphatases in general is discussed and possible mechanisms for the glucose effect are outlined. PMID:4342213

Characterization of Protein Tyrosine Phosphatase 1B Inhibition by Chlorogenic Acid and Cichoric Acid.

PubMed

Lipchock, James M; Hendrickson, Heidi P; Douglas, Bonnie B; Bird, Kelly E; Ginther, Patrick S; Rivalta, Ivan; Ten, Nicholas S; Batista, Victor S; Loria, J Patrick

2017-01-10

Protein tyrosine phosphatase 1B (PTP1B) is a known regulator of the insulin and leptin signaling pathways and is an active target for the design of inhibitors for the treatment of type II diabetes and obesity. Recently, cichoric acid (CHA) and chlorogenic acid (CGA) were predicted by docking methods to be allosteric inhibitors that bind distal to the active site. However, using a combination of steady-state inhibition kinetics, solution nuclear magnetic resonance experiments, and molecular dynamics simulations, we show that CHA is a competitive inhibitor that binds in the active site of PTP1B. CGA, while a noncompetitive inhibitor, binds in the second aryl phosphate binding site, rather than the predicted benzfuran binding pocket. The molecular dynamics simulations of the apo enzyme and cysteine-phosphoryl intermediate states with and without bound CGA suggest CGA binding inhibits PTP1B by altering hydrogen bonding patterns at the active site. This study provides a mechanistic understanding of the allosteric inhibition of PTP1B.

21 CFR 556.560 - Pyrantel tartrate.

Code of Federal Regulations, 2011 CFR

2011-04-01

... and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS TOLERANCES FOR RESIDUES OF NEW ANIMAL DRUGS IN FOOD Specific Tolerances for Residues of New Animal Drugs § 556.560 Pyrantel tartrate. Tolerances are established for...

21 CFR 556.560 - Pyrantel tartrate.

Code of Federal Regulations, 2010 CFR

2010-04-01

... and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS TOLERANCES FOR RESIDUES OF NEW ANIMAL DRUGS IN FOOD Specific Tolerances for Residues of New Animal Drugs § 556.560 Pyrantel tartrate. Tolerances are established for...

21 CFR 556.560 - Pyrantel tartrate.

Code of Federal Regulations, 2013 CFR

2013-04-01

... and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS TOLERANCES FOR RESIDUES OF NEW ANIMAL DRUGS IN FOOD Specific Tolerances for Residues of New Animal Drugs § 556.560 Pyrantel tartrate. Tolerances are established for...

21 CFR 556.560 - Pyrantel tartrate.

Code of Federal Regulations, 2012 CFR

2012-04-01

... and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS TOLERANCES FOR RESIDUES OF NEW ANIMAL DRUGS IN FOOD Specific Tolerances for Residues of New Animal Drugs § 556.560 Pyrantel tartrate. Tolerances are established for...

21 CFR 556.560 - Pyrantel tartrate.

Code of Federal Regulations, 2014 CFR

2014-04-01

... and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS TOLERANCES FOR RESIDUES OF NEW ANIMAL DRUGS IN FOOD Specific Tolerances for Residues of New Animal Drugs § 556.560 Pyrantel tartrate. Tolerances are established for...

AAS and spectrophotometric methods for the determination metoprolol tartrate in tablets

NASA Astrophysics Data System (ADS)

Alpdoğan, Güzin; Sungur, Sidika

1999-11-01

Sensitive and specific atomic adsorption spectroscopy (AAS) and spectrophotometric methods have been developed for the determination of beta adrenergic blocking drug, metoprolol tartrate.The method is based on the formation of Cu(II) dithiocarbamate complex by derivatization of the secondary amino group of metoprolol with CS 2 and CuCl 2 in the presence of ammonia.The copper-bis(dithiocarbamate) complex was extracted into chloroform and the concentration of metoprolol tartrate was determined directly by spectrophotometric and indirectly by AAS measurement of copper.The two methods developed were applied to the assay of metoprolol tartrate in commercial tablet formulations.The methods were compared statistically with each other and with the high performance liquid chromatography (HPLC) method of USPXXII using t- and F-tests.

Emerging Roles of Human Prostatic Acid Phosphatase

PubMed Central

Kong, Hoon Young; Byun, Jonghoe

2013-01-01

Prostate cancer is one of the most prevalent non-skin related cancers. It is the second leading cause of cancer deaths among males in most Western countries. If prostate cancer is diagnosed in its early stages, there is a higher probability that it will be completely cured. Prostatic acid phosphatase (PAP) is a non-specific phosphomonoesterase synthesized in prostate epithelial cells and its level proportionally increases with prostate cancer progression. PAP was the biochemical diagnostic mainstay for prostate cancer until the introduction of prostate-specific antigen (PSA) which improved the detection of early-stage prostate cancer and largely displaced PAP. Recently, however, there is a renewed interest in PAP because of its usefulness in prognosticating intermediate to high-risk prostate cancers and its success in the immunotherapy of prostate cancer. Although PAP is believed to be a key regulator of prostate cell growth, its exact role in normal prostate as well as detailed molecular mechanism of PAP regulation is still unclear. Here, many different aspects of PAP in prostate cancer are revisited and its emerging roles in other environment are discussed. PMID:24009853

Identification of soybean purple acid phosphatase genes and their expression responses to phosphorus availability and symbiosis

USDA-ARS?s Scientific Manuscript database

Background and Aims Purple acid phosphatases (PAPs) are members of the metallo-phosphoesterase family and have been known to play important roles in phosphorus (P) acquisition and recycling in plants. Low P availability is a major constraint to growth and production of soybean, Glycine max. Comparat...

Effects of gamma radiation on hard dental tissues of albino rats: investigation by light microscopy.

PubMed

El-Faramawy, Nabil; Ameen, Reham; El-Haddad, Khaled; El-Zainy, Medhat

2013-08-01

The present work aims at studying the effect of gamma radiation on the hard dental tissues. Eighty adult male albino rats with weights of about 250 g were used. The rats were irradiated at 0.2, 0.5, 1.0, 2.0, 4.0 and 6.0 Gy whole-body gamma doses. The effects on hard dental tissue samples were investigated after 48 h in histological and ground sections using light microscopy. Areas of acid phosphatase activity were detected using tartrate-resistant acid phosphatase (TRAP) stains. Observation of histological sections revealed disturbance in predentin thickness and odontoblastic layer as the irradiation dose increased. In cementum, widened cementocytes lacunae were occasionally detected even with low irradiated doses. On the other hand, relatively homogenous enamel was detected with darkened areas in enamel surface at doses over than 0.5 Gy. TRAP-positive cells were detected on the surface of the dentin of irradiated groups as well as cementum surface. Minimal detectable changes were observed in ground sections.

OPG-Fc but Not Zoledronic Acid Discontinuation Reverses Osteonecrosis of the Jaws (ONJ) in Mice

PubMed Central

de Molon, Rafael Scaf; Shimamoto, Hiroaki; Bezouglaia, Olga; Pirih, Flavia Q; Dry, Sarah M; Kostenuik, Paul; Boyce, Rogely W; Dwyer, Denise; Aghaloo, Tara L; Tetradis, Sotirios

2016-01-01

Osteonecrosis of the jaws (ONJ) is a significant complication of antiresorptive medications, such as bisphosphonates and denosumab. Antiresorptive discontinuation to promote healing of ONJ lesions remains highly controversial and understudied. Here, we investigated whether antiresorptive discontinuation alters ONJ features in mice, employing the potent bisphosphonate zoledronic acid (ZA) or the receptor activator of NF-κB ligand (RANKL) inhibitor OPG-Fc, utilizing previously published ONJ animal models. Mice were treated with vehicle (veh), ZA, or OPG-Fc for 11 weeks to induce ONJ, and antiresorptives were discontinued for 6 or 10 weeks. Maxillae and mandibles were examined by µCT imaging and histologically. ONJ features in ZA and OPG-Fc groups included periosteal bone deposition, empty osteocyte lacunae, osteonecrotic areas, and bone exposure, each of which substantially resolved 10 weeks after discontinuing OPG-Fc but not ZA. Full recovery of tartrate-resistant acid phosphatase-positive (TRAP+) osteoclast numbers occurred after discontinuing OPG-Fc but not ZA. Our data provide the first experimental evidence demonstrating that discontinuation of a RANKL inhibitor, but not a bisphosphonate, reverses features of osteonecrosis in mice. It remains unclear whether antiresorptive discontinuation increases the risk of skeletal-related events in patients with bone metastases or fracture risk in osteoporosis patients, but these preclinical data may nonetheless help to inform discussions on the rationale for a “drug holiday” in managing the ONJ patient. PMID:25727550

Structure of Thermotoga maritima Stationary Phase Survival Protein SurE: A Novel Acid Phosphatase

PubMed Central

Zhang, R.-G.; Skarina, T.; Katz, J.E.; Beasley, S.; Khachatryan, A.; Vyas, S.; Arrowsmith, C.H.; Clarke, S.; Edwards, A.; Joachimiak, A.; Savchenko, A.

2009-01-01

Summary Background The rpoS, nlpD, pcm, and surE genes are among many whose expression is induced during the stationary phase of bacterial growth. rpoS codes for the stationary-phase RNA polymerase σ subunit, and nlpD codes for a lipoprotein. The pcm gene product repairs damaged proteins by converting the atypical isoaspartyl residues back to L-aspartyls. The physiological and biochemical functions of surE are unknown, but its importance in stress is supported by the duplication of the surE gene in E. coli subjected to high-temperature growth. The pcm and surE genes are highly conserved in bacteria, archaea, and plants. Results The structure of SurE from Thermotoga maritima was determined at 2.0 Å. The SurE monomer is composed of two domains; a conserved N-terminal domain, a Rossman fold, and a C-terminal oligomerization domain, a new fold. Monomers form a dimer that assembles into a tetramer. Biochemical analysis suggests that SurE is an acid phosphatase, with an optimum pH of 5.5–6.2. The active site was identified in the N-terminal domain through analysis of conserved residues. Structure-based site-directed point mutations abolished phosphatase activity. T. maritima SurE intra- and inter-subunit salt bridges were identified that may explain the SurE thermostability. Conclusions The structure of SurE provided information about the protein’s fold, oligomeric state, and active site. The protein possessed magnesium-dependent acid phosphatase activity, but the physiologically relevant substrate(s) remains to be identified. The importance of three of the assigned active site residues in catalysis was confirmed by site-directed mutagenesis. PMID:11709173

An Acidic PATHOGENESIS-RELATED1 Gene of Oryza grandiglumis is Involved in Disease Resistance Response Against Bacterial Infection

PubMed Central

Shin, Sang Hyun; Pak, Jung-Hun; Kim, Mi Jin; Kim, Hye Jeong; Oh, Ju Sung; Choi, Hong Kyu; Jung, Ho Won; Chung, Young Soo

2014-01-01

Wild rice, Oryza grandiglumis shows hyper-resistance response to pathogen infection. In order to identify genes necessary for defense response in plants, we have carried out a subtractive hybridization coupled with a cDNA macroarray. An acidic PATHOGENESIS-RELATED1 (PR1) gene of the wild rice is highly identical to the acidic PR1 genes of different plant species. The OgPR1a cDNA has an apparent single open reading frame with a predicted molecular mass 40,621 Da and an isoelectic point of 5.14. Both in silico analysis and a transient expression assay in onion epidermal cells revealed that the OgPR1a protein could be localized in intercellular space in plants. The OgPR1a mRNA was strongly transcribed by the exogenous treatment with ethylene and jasmonic acid as well as protein phosphatase inhibitors. Additionally, ectopic expression of the OgPR1a conferred disease resistance on Arabidopsis to the bacterial and fungal infections. PMID:25289005

The stability of prostatic acid phosphatase, as measured by a capture immunoenzyme assay.

PubMed

Lin, T M; Chin-See, M W; Halbert, S P

1984-03-27

A capture immunoenzyme assay (CIEA) for prostatic acid phosphatase (PAP) was developed and used to study the stability of this isoenzyme. Immunospecifically purified goat antibodies to PAP were covalently bound to special discs and used to capture the enzyme in serum samples in a weakly acidic medium during the first incubation (2 h) at 37 degrees C. The capture enzyme was then measured by its catalytic activity with p-nitrophenyl phosphate as substrate during the second incubation (1 h) at 37 degrees C. As much as 98% of the PAP in test specimens was captured and measured by this CIEA. The test results were expressed as enzymatic activity (U/l), extrapolated from a standard curve which was linear between 0.026 and 70 U/l. In test sera stored at 4 degrees C, the PAP was variably stable for 7 to 70 days, but the enzyme was quite stable in serum when stored at -20 degrees C for at least 156 days. At room temperature, when the sera were appropriately acidified, there was no loss of enzymatic activity for periods of 15 days, and in some cases, a large proportion of activity was still intact after 70 days. At 4 degrees C, as well as -20 degrees C, acidified serum and the partially purified PAP standard showed complete stability for at least 7 months. The CIEA reactivity of positive test specimens was inhibited by L(+)-tartaric acid, but not by cupric sulfate. The acid phosphatases of blood cell extracts were non-reactive in the CIEA procedure. The CIEA results of 224 serum samples from patients with and without prostate cancer correlated very well with those obtained by two direct enzymatic and two commercial RIA procedures, with correlation coefficients between 0.960 and 0.993, and diagnostic agreement between 86% and 100%.

Arsanilic acid modified superparamagnetic iron oxide nanoparticles for Purification of alkaline phosphatase from hen's egg yolk.

PubMed

Farzi-Khajeh, Hamed; Safa, Kazem D; Dastmalchi, Siavoush

2017-09-01

Recent studies of magnetic carrier technology have focused on its applications in separation and purification technologies, due to easy separation of the target from the reaction medium by applying an external magnetic field. In the present study, Fe 3 O 4 superparamagnetic nanoparticles were prepared to utilize a chemical co-precipitation method, then the surfaces of the nanoparticles were modified with arsanilic acid derivatives which were used as the specific nanocarriers for the affinity purification of alkaline phosphatase from the hen's egg yolk. The six different types of magnetic nanocarriers with varied lengths of the linkers were obtained. All samples were characterized step by step and validated using FTIR, SEM, EDX, VSM and XRD analysis methods As the results were shown, the use of inflexible tags with long linkers on the surface of the nanocarrier could lead to better results for separation of alkaline phosphatase from the hen's egg yolk with 76.2% recovery and 1361.7-fold purification. The molecular weight of the purified alkaline phosphatase was estimated to be 68kDa by SDS-PAGE. The results of this study showed that the novel magnetic nanocarriers were capable of purifying alkaline phosphatase in a practically time and cost effective way. Copyright © 2017 Elsevier B.V. All rights reserved.

Antioxidant and Anti-Osteoporosis Activities of Chemical Constituents of the Stems of Zanthoxylum piperitum.

PubMed

Yang, Seo Young; Lee, Sang-Hyun; Tai, Bui Huu; Jang, Hae-Dong; Kim, Young Ho

2018-02-18

Two new lignans, zanthoxyloside C ( 1 ) and zanthoxyloside D ( 2 ), together with nine known compounds comprising lignans ( 3 - 5 ), flavonoids ( 6 - 8 ), and phenolics ( 9 - 11 ), were isolated from the methanol extract of the stems of Zanthoxylum piperitum. All isolates were evaluated for their antioxidant and anti-osteoporotic activities using oxygen radical absorbance capacity (ORAC), cupric reducing antioxidant capacity (CUPRAC), and tartrate-resistant acid phosphatase (TRAP) assays. Compounds 7 - 10 showed peroxyl radical-scavenging capacities and 4 , 6 - 7 , and 9 showed reducing capacities. Moreover, compounds 3 , 6 - 9 , and 11 significantly suppressed TRAP activities. These results indicated that the stems of Z. piperitum could be an excellent source for natural antioxidant and anti-osteoporosis.

21 CFR 556.425 - Morantel tartrate.

Code of Federal Regulations, 2011 CFR

2011-04-01

... DRUGS, FEEDS, AND RELATED PRODUCTS TOLERANCES FOR RESIDUES OF NEW ANIMAL DRUGS IN FOOD Specific Tolerances for Residues of New Animal Drugs § 556.425 Morantel tartrate. A tolerance of 0.7 part per million is established for N-methyl-1,3-propanediamine (MAPA, marker residue) in the liver (target tissue) of...

21 CFR 556.425 - Morantel tartrate.

Code of Federal Regulations, 2014 CFR

2014-04-01

... DRUGS, FEEDS, AND RELATED PRODUCTS TOLERANCES FOR RESIDUES OF NEW ANIMAL DRUGS IN FOOD Specific Tolerances for Residues of New Animal Drugs § 556.425 Morantel tartrate. A tolerance of 0.7 part per million is established for N-methyl-1,3-propanediamine (MAPA, marker residue) in the liver (target tissue) of...

21 CFR 556.425 - Morantel tartrate.

Code of Federal Regulations, 2012 CFR

2012-04-01

... DRUGS, FEEDS, AND RELATED PRODUCTS TOLERANCES FOR RESIDUES OF NEW ANIMAL DRUGS IN FOOD Specific Tolerances for Residues of New Animal Drugs § 556.425 Morantel tartrate. A tolerance of 0.7 part per million is established for N-methyl-1,3-propanediamine (MAPA, marker residue) in the liver (target tissue) of...

21 CFR 556.425 - Morantel tartrate.

Code of Federal Regulations, 2013 CFR

2013-04-01

... DRUGS, FEEDS, AND RELATED PRODUCTS TOLERANCES FOR RESIDUES OF NEW ANIMAL DRUGS IN FOOD Specific Tolerances for Residues of New Animal Drugs § 556.425 Morantel tartrate. A tolerance of 0.7 part per million is established for N-methyl-1,3-propanediamine (MAPA, marker residue) in the liver (target tissue) of...

21 CFR 556.425 - Morantel tartrate.

Code of Federal Regulations, 2010 CFR

2010-04-01

... DRUGS, FEEDS, AND RELATED PRODUCTS TOLERANCES FOR RESIDUES OF NEW ANIMAL DRUGS IN FOOD Specific Tolerances for Residues of New Animal Drugs § 556.425 Morantel tartrate. A tolerance of 0.7 part per million is established for N-methyl-1,3-propanediamine (MAPA, marker residue) in the liver (target tissue) of...

Effect of Soil Amendments on Microbial Resilience Capacity of Acid Soil Under Copper Stress.

PubMed

Mounissamy, Vassanda Coumar; Kundu, Samaresh; Selladurai, Rajendiran; Saha, Jayanta Kumar; Biswas, Ashish Kumar; Adhikari, Tapan; Patra, Ashok Kumar

2017-11-01

An incubation study was undertaken to study microbial resilience capacity of acid soil amended with farmyard manure (FYM), charcoal and lime under copper (Cu) perturbation. Copper stress significantly reduced enzymatic activities and microbial biomass carbon (MBC) in soil. Percent reduction in microbial activity of soil due to Cu stress was 74.7% in dehydrogenase activity, 59.9% in MBC, 48.2% in alkaline phosphatase activity and 15.1% in acid phosphatase activity. Soil treated with FYM + charcoal showed highest resistance index for enzymatic activities and MBC. Similarly, the highest resilience index for acid phosphatase activity was observed in soil amended with FYM (0.40), whereas FYM + charcoal-treated soil showed the highest resilience indices for alkaline, dehydrogenase activity and MBC: 0.50, 0.22 and 0.25, respectively. This investigation showed that FYM and charcoal application, either alone or in combination, proved to be better than lime with respect to microbial functional resistance and resilience of acid soil under Cu perturbation.

ON THE RELATIVE STABILITY OF ALUMINUM, TITANIUM, VANADIUM, IRON, AND COPPER TARTRATE COMPLEXES IN ALKALI MEDIA (in Russian)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pyatnitskii, I.V.; Kostyshina, A.P.

1959-06-01

The stability of aluminum, copper, iron, titunium, and vanadium tartrate complexes was determined using bond magnitudes as criteria (the ratio between the concentrations of complexed and free ions at a certain standard acid condition). A method is suggested for determining the ratio of the bonds combining the complexes of two metals. The partition constaats of aluminum, copper, iron(III), and vanadium hydroxyquinolinates between the aqueous solution and chloroform were 2.6 x 10/sup -33/, 7.3 x 10/sup -23/, 1.5 x 10/sup -37/, and 4.2 x 10/sup -23/, respectively. The relative stability of copper and iron turtrate complexes in alkali solution (pH 13)more » and aluminum, iron(III), titunium, and vanadium(IV) tartrate complexes in ammonium solution (pH 9.5) was determined. (R.V.J.)« less

FERONIA interacts with ABI2-type phosphatases to facilitate signaling cross-talk between abscisic acid and RALF peptide in Arabidopsis

PubMed Central

Chen, Jia; Yu, Feng; Liu, Ying; Du, Changqing; Li, Xiushan; Zhu, Sirui; Wang, Xianchun; Lan, Wenzhi; Rodriguez, Pedro L.; Liu, Xuanming; Li, Dongping; Chen, Liangbi; Luan, Sheng

2016-01-01

Receptor-like kinase FERONIA (FER) plays a crucial role in plant response to small molecule hormones [e.g., auxin and abscisic acid (ABA)] and peptide signals [e.g., rapid alkalinization factor (RALF)]. It remains unknown how FER integrates these different signaling events in the control of cell growth and stress responses. Under stress conditions, increased levels of ABA will inhibit cell elongation in the roots. In our previous work, we have shown that FER, through activation of the guanine nucleotide exchange factor 1 (GEF1)/4/10-Rho of Plant 11 (ROP11) pathway, enhances the activity of the phosphatase ABA Insensitive 2 (ABI2), a negative regulator of ABA signaling, thereby inhibiting ABA response. In this study, we found that both RALF and ABA activated FER by increasing the phosphorylation level of FER. The FER loss-of-function mutant displayed strong hypersensitivity to both ABA and abiotic stresses such as salt and cold conditions, indicating that FER plays a key role in ABA and stress responses. We further showed that ABI2 directly interacted with and dephosphorylated FER, leading to inhibition of FER activity. Several other ABI2-like phosphatases also function in this pathway, and ABA-dependent FER activation required PYRABACTIN RESISTANCE (PYR)/PYR1-LIKE (PYL)/REGULATORY COMPONENTS OF ABA RECEPTORS (RCAR)–A-type protein phosphatase type 2C (PP2CA) modules. Furthermore, suppression of RALF1 gene expression, similar to disruption of the FER gene, rendered plants hypersensitive to ABA. These results formulated a mechanism for ABA activation of FER and for cross-talk between ABA and peptide hormone RALF in the control of plant growth and responses to stress signals. PMID:27566404

Protein phosphatase 2Cm is a critical regulator of branched-chain amino acid catabolism in mice and cultured cells.

PubMed

Lu, Gang; Sun, Haipeng; She, Pengxiang; Youn, Ji-Youn; Warburton, Sarah; Ping, Peipei; Vondriska, Thomas M; Cai, Hua; Lynch, Christopher J; Wang, Yibin

2009-06-01

The branched-chain amino acids (BCAA) are essential amino acids required for protein homeostasis, energy balance, and nutrient signaling. In individuals with deficiencies in BCAA, these amino acids can be preserved through inhibition of the branched-chain-alpha-ketoacid dehydrogenase (BCKD) complex, the rate-limiting step in their metabolism. BCKD is inhibited by phosphorylation of its E1alpha subunit at Ser293, which is catalyzed by BCKD kinase. During BCAA excess, phosphorylated Ser293 (pSer293) becomes dephosphorylated through the concerted inhibition of BCKD kinase and the activity of an unknown intramitochondrial phosphatase. Using unbiased, proteomic approaches, we have found that a mitochondrial-targeted phosphatase, PP2Cm, specifically binds the BCKD complex and induces dephosphorylation of Ser293 in the presence of BCKD substrates. Loss of PP2Cm completely abolished substrate-induced E1alpha dephosphorylation both in vitro and in vivo. PP2Cm-deficient mice exhibited BCAA catabolic defects and a metabolic phenotype similar to the intermittent or intermediate types of human maple syrup urine disease (MSUD), a hereditary disorder caused by defects in BCKD activity. These results indicate that PP2Cm is the endogenous BCKD phosphatase required for nutrient-mediated regulation of BCKD activity and suggest that defects in PP2Cm may be responsible for a subset of human MSUD.

Trichoderma harzianum Produces a New Thermally Stable Acid Phosphatase, with Potential for Biotechnological Application

PubMed Central

Souza, Amanda Araújo; Leitão, Vanessa Oliveira; Ramada, Marcelo Henrique; Mehdad, Azadeh; Georg, Raphaela de Castro; Ulhôa, Cirano José; de Freitas, Sonia Maria

2016-01-01

Acid phosphatases (ACPases) are produced by a variety of fungi and have gained attention due their biotechnological potential in industrial, diagnosis and bioremediation processes. These enzymes play a specific role in scavenging, mobilization and acquisition of phosphate, enhancing soil fertility and plant growth. In this study, a new ACPase from Trichoderma harzianum, named ACPase II, was purified and characterized as a glycoprotein belonging to the acid phosphatase family. ACPase II presents an optimum pH and temperature of 3.8 and 65°C, respectively, and is stable at 55°C for 120 min, retaining 60% of its activity. The enzyme did not require metal divalent ions, but was inhibited by inorganic phosphate and tungstate. Affinity for several phosphate substrates was observed, including phytate, which is the major component of phosphorus in plant foods. The inhibition of ACPase II by tungstate and phosphate at different pH values is consistent with the inability of the substrate to occupy its active site due to electrostatic contacts that promote conformational changes, as indicated by fluorescence spectroscopy. A higher affinity for tungstate rather than phosphate at pH 4.0was observed, in accordance with its highest inhibitory effect. Results indicate considerable biotechnological potential of the ACPase II in soil environments. PMID:26938873

Trichoderma harzianum Produces a New Thermally Stable Acid Phosphatase, with Potential for Biotechnological Application.

PubMed

Souza, Amanda Araújo; Leitão, Vanessa Oliveira; Ramada, Marcelo Henrique; Mehdad, Azadeh; Georg, Raphaela de Castro; Ulhôa, Cirano José; de Freitas, Sonia Maria

2016-01-01

Acid phosphatases (ACPases) are produced by a variety of fungi and have gained attention due their biotechnological potential in industrial, diagnosis and bioremediation processes. These enzymes play a specific role in scavenging, mobilization and acquisition of phosphate, enhancing soil fertility and plant growth. In this study, a new ACPase from Trichoderma harzianum, named ACPase II, was purified and characterized as a glycoprotein belonging to the acid phosphatase family. ACPase II presents an optimum pH and temperature of 3.8 and 65 °C, respectively, and is stable at 55 °C for 120 min, retaining 60% of its activity. The enzyme did not require metal divalent ions, but was inhibited by inorganic phosphate and tungstate. Affinity for several phosphate substrates was observed, including phytate, which is the major component of phosphorus in plant foods. The inhibition of ACPase II by tungstate and phosphate at different pH values is consistent with the inability of the substrate to occupy its active site due to electrostatic contacts that promote conformational changes, as indicated by fluorescence spectroscopy. A higher affinity for tungstate rather than phosphate at pH 4.0 was observed, in accordance with its highest inhibitory effect. Results indicate considerable biotechnological potential of the ACPase II in soil environments.

Effect of Korean Red Ginseng on radiation-induced bone loss in C3H/HeN mice

PubMed Central

Lee, Jin-Hee; Lee, Hae-June; Yang, Miyoung; Moon, Changjong; Kim, Jong-Choon; Bae, Chun-Sik; Jo, Sung-Kee; Jang, Jong-Sik; Kim, Sung-Ho

2013-01-01

This study investigated the effects of Korean Red Ginseng (KRG) on radiation-induced bone loss in C3H/HeN mice. C3H/HeN mice were divided into sham and irradiation (3 Gy, gamma-ray) groups. The irradiated mice were treated for 12 wk with vehicle, KRG (per os, p.o.) or KRG (intraperitoneal). Serum alkaline phosphatase (ALP), tartrate-resistant acid phosphatase, estradiol level, and biomechanical properties were measured. Tibiae were analyzed using micro-computed tomography. Treatment of KRG (p.o., 250 mg/kg of body weight/d) significantly preserved trabecular bone volume, trabecular number, structure model index, and bone mineral density of proximal tibia metaphysic, but did not alter the uterus weight of the mice. Serum ALP level was slightly reduced by KRG treatment. However, grip strength, mechanical property, and cortical bone architecture did not differ among the experimental groups. The results indicate that KRG can prevent radiation-induced bone loss in mice. PMID:24233384

Effect of perfluorooctane sulfonate on the conformation of wheat germ acid phosphatase.

PubMed

Xu, Dongmei; Jin, Jianchang; Shen, Tong; Wang, Yanhua

2013-11-01

Fluorescence spectroscopy was used to study the quenching mechanism, the type of force and the binding sites of perfluorooctane sulfonate (PFOS) on wheat germ acid phosphatase (ACPase). The results showed that the quenching effect of PFOS on ACPase was mainly due to a static quenching mechanism that occurred via the formation of hydrogen bonds and van der Waals forces. The results from synchronous fluorescence spectroscopy demonstrated that PFOS interacts with ACPase close to the tryptophan residues. In addition, synchronous fluorescence spectroscopy also showed that PFOS increases the hydrophobicity of the microenvironment of the tyrosine residues, hence decreasing the local polarity.

Domain-to-domain coupling in voltage-sensing phosphatase.

PubMed

Sakata, Souhei; Matsuda, Makoto; Kawanabe, Akira; Okamura, Yasushi

2017-01-01

Voltage-sensing phosphatase (VSP) consists of a transmembrane voltage sensor and a cytoplasmic enzyme region. The enzyme region contains the phosphatase and C2 domains, is structurally similar to the tumor suppressor phosphatase PTEN, and catalyzes the dephosphorylation of phosphoinositides. The transmembrane voltage sensor is connected to the phosphatase through a short linker region, and phosphatase activity is induced upon membrane depolarization. Although the detailed molecular characteristics of the voltage sensor domain and the enzyme region have been revealed, little is known how these two regions are coupled. In addition, it is important to know whether mechanism for coupling between the voltage sensor domain and downstream effector function is shared among other voltage sensor domain-containing proteins. Recent studies in which specific amino acid sites were genetically labeled using a fluorescent unnatural amino acid have enabled detection of the local structural changes in the cytoplasmic region of Ciona intestinalis VSP that occur with a change in membrane potential. The results of those studies provide novel insight into how the enzyme activity of the cytoplasmic region of VSP is regulated by the voltage sensor domain.

Domain-to-domain coupling in voltage-sensing phosphatase

PubMed Central

Sakata, Souhei; Matsuda, Makoto; Kawanabe, Akira; Okamura, Yasushi

2017-01-01

Voltage-sensing phosphatase (VSP) consists of a transmembrane voltage sensor and a cytoplasmic enzyme region. The enzyme region contains the phosphatase and C2 domains, is structurally similar to the tumor suppressor phosphatase PTEN, and catalyzes the dephosphorylation of phosphoinositides. The transmembrane voltage sensor is connected to the phosphatase through a short linker region, and phosphatase activity is induced upon membrane depolarization. Although the detailed molecular characteristics of the voltage sensor domain and the enzyme region have been revealed, little is known how these two regions are coupled. In addition, it is important to know whether mechanism for coupling between the voltage sensor domain and downstream effector function is shared among other voltage sensor domain-containing proteins. Recent studies in which specific amino acid sites were genetically labeled using a fluorescent unnatural amino acid have enabled detection of the local structural changes in the cytoplasmic region of Ciona intestinalis VSP that occur with a change in membrane potential. The results of those studies provide novel insight into how the enzyme activity of the cytoplasmic region of VSP is regulated by the voltage sensor domain. PMID:28744425

Crystal structure of (S)-sec-butyl­ammonium l-tartrate monohydrate

PubMed Central

Publicover, Ernlie A.; Kolwich, Jennifer; Stack, Darcie L.; Doué, Alyssa J.; Ylijoki, Kai E. O.

2017-01-01

The title hydrated mol­ecular salt, C4H12N+·C4H5O6 −·H2O, was prepared by deprotonation of enanti­opure l-tartaric acid with racemic sec-butyl­amine in water. Only one enanti­omer was observed crystallographically, resulting from the combination of (S)-sec-butyl­amine with l-tartaric acid. The sec-butyl­ammonium moiety is disordered over two conformations related by rotation around the CH–CH2 bond; the refined occupancy ratio is 0.68 (1):0.32 (1). In the crystal, mol­ecules are linked through a network of O—H⋯O and N—H⋯O hydrogen-bonding inter­actions, between the ammonium H atoms, the tartrate hy­droxy H atoms, and the inter­stitial water, forming a three-dimensional supra­molecular structure. PMID:28529783

A Novel Phytase with Sequence Similarity to Purple Acid Phosphatases Is Expressed in Cotyledons of Germinating Soybean Seedlings 1

PubMed Central

Hegeman, Carla E.; Grabau, Elizabeth A.

2001-01-01

Phytic acid (myo-inositol hexakisphosphate) is the major storage form of phosphorus in plant seeds. During germination, stored reserves are used as a source of nutrients by the plant seedling. Phytic acid is degraded by the activity of phytases to yield inositol and free phosphate. Due to the lack of phytases in the non-ruminant digestive tract, monogastric animals cannot utilize dietary phytic acid and it is excreted into manure. High phytic acid content in manure results in elevated phosphorus levels in soil and water and accompanying environmental concerns. The use of phytases to degrade seed phytic acid has potential for reducing the negative environmental impact of livestock production. A phytase was purified to electrophoretic homogeneity from cotyledons of germinated soybeans (Glycine max L. Merr.). Peptide sequence data generated from the purified enzyme facilitated the cloning of the phytase sequence (GmPhy) employing a polymerase chain reaction strategy. The introduction of GmPhy into soybean tissue culture resulted in increased phytase activity in transformed cells, which confirmed the identity of the phytase gene. It is surprising that the soybean phytase was unrelated to previously characterized microbial or maize (Zea mays) phytases, which were classified as histidine acid phosphatases. The soybean phytase sequence exhibited a high degree of similarity to purple acid phosphatases, a class of metallophosphoesterases. PMID:11500558

Defects in the acid phosphatase ACPT cause recessive hypoplastic amelogenesis imperfecta.

PubMed

Smith, Claire El; Whitehouse, Laura LE; Poulter, James A; Brookes, Steven J; Day, Peter F; Soldani, Francesca; Kirkham, Jennifer; Inglehearn, Chris F; Mighell, Alan J

2017-08-01

We identified two homozygous missense variants (c.428C>T, p.(T143M) and c.746C>T, p.(P249L)) in ACPT, the gene encoding acid phosphatase, testicular, which segregates with hypoplastic amelogenesis imperfecta in two unrelated families. ACPT is reported to play a role in odontoblast differentiation and mineralisation by supplying phosphate during dentine formation. Analysis by computerised tomography and scanning electron microscopy of a primary molar tooth from an individual homozygous for the c.746C>T variant revealed an enamel layer that was hypoplastic, but mineralised with prismatic architecture. These findings implicate variants in ACPT as a cause of early failure of amelogenesis during the secretory phase.

Multifunctional hybrid micelles with tunable active targeting and acid/phosphatase-stimulated drug release for enhanced tumor suppression.

PubMed

Liu, Xuhan; Li, Yinghuan; Tan, Xi; Rao, Rong; Ren, Yuanyuan; Liu, Lingyan; Yang, Xiangliang; Liu, Wei

2018-03-01

Therapeutic efficacy of conventional single PEGylated polymeric micelles is significantly reduced by limited endocytosis and intracellular drug release. To improve drug delivery efficiency, poly (ethylene glycol)-block-poly (l-lactic acid)/(Arg-Gly-Asp-Phe)-poly (aminoethyl ethylene phosphate)-block-poly (l-lactic acid) (PEG-PLLA/RGDF-PAEEP-PLLA) hybrid micelles with tunable active targeting and acid/phosphatase-stimulated drug release are developed. The optimized hybrid micelles with 6 wt % of RGDF have favorable in vitro and in vivo activities. The hybrid micelles could temporarily shield the targeting efficacy of RGDF at pH 7.4 due to the steric effect exerted by concealment of RGDF peptides in the PEG corona, which strongly decreases the clearance by mononuclear phagocyte system and consequently improves the tumor accumulation. Inside the solid tumor with a lower acidic pH, the hybrid micelles restore the active tumor targeting property with exposed RGDF on the surface of the micelles because of the increased protonation and stretching degree of PAEEP blocks. RGDF-mediated endocytosis improves the tumor cell uptake. The hybrid micelles would also enhance intracellular drug release because of the hydrolysis of the acid/phosphatase-sensitivity of PAEEP blocks in endo/lysosome. Systemic administration of the hybrid micelles significantly inhibits tumor growth by 96% due to the integration of enhanced circulation time, tumor accumulation, cell uptake and intracellular drug release. Copyright © 2017 Elsevier Ltd. All rights reserved.

Ferulic Acid Attenuates the Injury-Induced Decrease of Protein Phosphatase 2A Subunit B in Ischemic Brain Injury

PubMed Central

Koh, Phil-Ok

2013-01-01

Background Ferulic acid provides a neuroprotective effect during cerebral ischemia through its anti-oxidant function. Protein phosphatase 2A (PP2A) is a serine and threonine phosphatase that contributes broadly to normal brain function. This study investigated whether ferulic acid regulates PP2A subunit B in a middle cerebral artery occlusion (MCAO) animal model and glutamate toxicity-induced neuronal cell death. Methodology/Principal Findings MCAO was surgically induced to yield permanent cerebral ischemic injury in rats. The rats were treated with either vehicle or ferulic acid (100 mg/kg, i.v.) immediately after MCAO, and cerebral cortex tissues were collected 24 h after MCAO. A proteomics approach, RT-PCR, and Western blot analyses performed to identification of PP2A subunit B expression levels. Ferulic acid significantly reduced the MCAO-induced infarct volume of the cerebral cortex. A proteomics approach elucidated the reduction of PP2A subunit B in MCAO-induced animals, and ferulic acid treatment prevented the injury-induced reduction in PP2A subunit B levels. RT-PCR and Western blot analyses also showed that ferulic acid treatment attenuates the injury-induced decrease in PP2A subunit B levels. Moreover, the number of PP2A subunit B-positive cells was reduced in MCAO-induced animals, and ferulic acid prevented these decreases. In cultured neuronal cells, ferulic acid treatment protected cells against glutamate toxicity and prevented the glutamate-induced decrease in PP2A subunit B. Conclusions/Significance These results suggest that the maintenance of PP2A subunit B by ferulic acid in ischemic brain injury plays an important role for the neuroprotective function of ferulic acid. PMID:23349830

21 CFR 558.485 - Pyrantel tartrate.

Code of Federal Regulations, 2010 CFR

2010-04-01

... containing 9.6, 19.2, 48, or 80 grams per pound pyrantel tartrate. (b) Approvals. See sponsors in § 510.600(c... grams per pound for use as in paragraph (e)(1) of this section. (2) [Reserved] (3) Nos. 010439, 011490, 011749, 012286, 016968, 017790, 043733, and 049685: 9.6 and 19.2 grams per pound for use as in paragraphs...

The Ablation of Mitochondrial Protein Phosphatase Pgam5 Confers Resistance Against Metabolic Stress.

PubMed

Sekine, Shiori; Yao, Akari; Hattori, Kazuki; Sugawara, Sho; Naguro, Isao; Koike, Masato; Uchiyama, Yasuo; Takeda, Kohsuke; Ichijo, Hidenori

2016-03-01

Phosphoglycerate mutase family member 5 (PGAM5) is a mitochondrial protein phosphatase that has been reported to be involved in various stress responses from mitochondrial quality control to cell death. However, its roles in vivo are largely unknown. Here, we show that Pgam5-deficient mice are resistant to several metabolic insults. Under cold stress combined with fasting, Pgam5-deficient mice better maintained body temperature than wild-type mice and showed an extended survival rate. Serum triglycerides and lipid content in brown adipose tissue (BAT), a center of adaptive thermogenesis, were severely reduced in Pgam5-deficient mice. Moreover, although Pgam5 deficiency failed to maintain proper mitochondrial integrity in BAT, it reciprocally resulted in the dramatic induction of fibroblast growth factor 21 (FGF21) that activates various functions of BAT including thermogenesis. Thus, the enhancement of lipid metabolism and FGF21 may contribute to the cold resistance of Pgam5-deficient mice under fasting condition. Finally, we also found that Pgam5-deficient mice are resistant to high-fat-diet-induced obesity. Our study uncovered that PGAM5 is involved in the whole-body metabolism in response to stresses that impose metabolic challenges on mitochondria.

Regulation of lipid droplet and membrane biogenesis by the acidic tail of the phosphatidate phosphatase Pah1p

PubMed Central

Karanasios, Eleftherios; Barbosa, Antonio Daniel; Sembongi, Hiroshi; Mari, Muriel; Han, Gil-Soo; Reggiori, Fulvio; Carman, George M.; Siniossoglou, Symeon

2013-01-01

Lipins are evolutionarily conserved phosphatidate phosphatases that perform key functions in phospholipid, triglyceride, and membrane biogenesis. Translocation of lipins on membranes requires their dephosphorylation by the Nem1p-Spo7p transmembrane phosphatase complex through a poorly understood mechanism. Here we identify the carboxy-terminal acidic tail of the yeast lipin Pah1p as an important regulator of this step. Deletion or mutations of the tail disrupt binding of Pah1p to the Nem1p-Spo7p complex and Pah1p membrane translocation. Overexpression of Nem1p-Spo7p drives the recruitment of Pah1p in the vicinity of lipid droplets in an acidic tail–dependent manner and induces lipid droplet biogenesis. Genetic analysis shows that the acidic tail is essential for the Nem1p-Spo7p–dependent activation of Pah1p but not for the function of Pah1p itself once it is dephosphorylated. Loss of the tail disrupts nuclear structure, INO1 gene expression, and triglyceride synthesis. Similar acidic sequences are present in the carboxy-terminal ends of all yeast lipin orthologues. We propose that acidic tail–dependent binding and dephosphorylation of Pah1p by the Nem1p-Spo7p complex is an important determinant of its function in lipid and membrane biogenesis. PMID:23657815

Disruption of Abscisic Acid Signaling Constitutively Activates Arabidopsis Resistance to the Necrotrophic Fungus Plectosphaerella cucumerina1[W

PubMed Central

Sánchez-Vallet, Andrea; López, Gemma; Ramos, Brisa; Delgado-Cerezo, Magdalena; Riviere, Marie-Pierre; Llorente, Francisco; Fernández, Paula Virginia; Miedes, Eva; Estevez, José Manuel; Grant, Murray; Molina, Antonio

2012-01-01

Plant resistance to necrotrophic fungi is regulated by a complex set of signaling pathways that includes those mediated by the hormones salicylic acid (SA), ethylene (ET), jasmonic acid (JA), and abscisic acid (ABA). The role of ABA in plant resistance remains controversial, as positive and negative regulatory functions have been described depending on the plant-pathogen interaction analyzed. Here, we show that ABA signaling negatively regulates Arabidopsis (Arabidopsis thaliana) resistance to the necrotrophic fungus Plectosphaerella cucumerina. Arabidopsis plants impaired in ABA biosynthesis, such as the aba1-6 mutant, or in ABA signaling, like the quadruple pyr/pyl mutant (pyr1pyl1pyl2pyl4), were more resistant to P. cucumerina than wild-type plants. In contrast, the hab1-1abi1-2abi2-2 mutant impaired in three phosphatases that negatively regulate ABA signaling displayed an enhanced susceptibility phenotype to this fungus. Comparative transcriptomic analyses of aba1-6 and wild-type plants revealed that the ABA pathway negatively regulates defense genes, many of which are controlled by the SA, JA, or ET pathway. In line with these data, we found that aba1-6 resistance to P. cucumerina was partially compromised when the SA, JA, or ET pathway was disrupted in this mutant. Additionally, in the aba1-6 plants, some genes encoding cell wall-related proteins were misregulated. Fourier transform infrared spectroscopy and biochemical analyses of cell walls from aba1-6 and wild-type plants revealed significant differences in their Fourier transform infrared spectratypes and uronic acid and cellulose contents. All these data suggest that ABA signaling has a complex function in Arabidopsis basal resistance, negatively regulating SA/JA/ET-mediated resistance to necrotrophic fungi. PMID:23037505

Ontogeny and distribution of alkaline and acid phosphatases in the digestive system of California halibut larvae (Paralichthys californicus).

PubMed

Zacarias-Soto, Magali; Barón-Sevilla, Benjamín; Lazo, Juan P

2013-10-01

Studies aimed to assess the digestive physiology of marine fish larvae under culture conditions are important to further understand the functional characteristics and digestive capacities of the developing larvae. Most studies to date concentrate on intestinal lumen digestion and little attention to the absorption process. Thus, the objectives of this study were to histochemically detect and quantify some of the enzymes responsible for absorption and intracellular digestion of nutrients in the anterior and posterior intestine of California halibut larvae. Alkaline and acid phosphatases were detected from the first days post-hatch (dph). Alkaline phosphatase maintained a high level of activity during the first 20 dph in both intestinal regions. Thereafter, a clear intestinal regionalization of the activity was observed with the highest levels occurring in the anterior intestine. Acid phosphatase activity gradually increased in both intestinal regions during development, and a regionalization of the activity was not observed until late in development, once the ocular migration began. Highest levels were observed in the anterior intestine at the end of metamorphosis concomitant with the stomach development. The results from this study show some morphological and physiological changes are occurring during larval development and a clear regionalization of the absorption process as the larvae develops. These ontological changes must be considered in the elaboration of diets according to the digestive capacity of the larvae.

21 CFR 184.1804 - Sodium potassium tartrate.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Sodium potassium tartrate. 184.1804 Section 184.1804 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... has a cooling saline taste. It is obtained as a byproduct of wine manufacture. (b) The ingredient...

Final product analysis in the e-beam and gamma radiolysis of aqueous solutions of metoprolol tartrate

NASA Astrophysics Data System (ADS)

Slegers, Catherine; Tilquin, Bernard

2006-09-01

The radiostability of metoprolol tartrate aqueous solutions and the influence of the absorbed dose (0-50 kGy), dose rate (e-beam (EB) vs. gamma ( γ)) and radioprotectors (pharmaceutical excipients) are investigated by HPLC-UV analyses and through computer simulations. The use of radioprotecting excipients is more promising than an increase in the dose rate to lower the degradation of metoprolol tartrate aqueous solutions for applications such as radiosterilization. The decontamination of metoprolol tartrate from waste waters by EB processing appears highly feasible.

Simplified preparation of a phosphatase inhibitor and further studies of its action.

PubMed

Coburn, S P; Schaltenbrand, W E

1978-05-01

1-Pyrrolidinecarbothioic acid (2-pyridylmethylene) hydrazide chelates Zn2+ but not Mg2+. This compound is about twice as effective as EDTA for inhibiting alkaline phosphatase from calf mucosa, and approx. 1000-fold more effective than EDTA for inhibiting acid phosphatase from wheat germ. The compound did not inhibit pyridoxine kinase activity in human leucocytes at the highest concentration tested (33 micron). Therefore it may be a useful tool for either examining or eliminating the effects of phosphatases in complex enzyme systems.

Delayed bone regeneration and low bone mass in a rat model of insulin-resistant type 2 diabetes mellitus is due to impaired osteoblast function.

PubMed

Hamann, Christine; Goettsch, Claudia; Mettelsiefen, Jan; Henkenjohann, Veit; Rauner, Martina; Hempel, Ute; Bernhardt, Ricardo; Fratzl-Zelman, Nadja; Roschger, Paul; Rammelt, Stefan; Günther, Klaus-Peter; Hofbauer, Lorenz C

2011-12-01

Patients with diabetes mellitus have an impaired bone metabolism; however, the underlying mechanisms are poorly understood. Here, we analyzed the impact of type 2 diabetes mellitus on bone physiology and regeneration using Zucker diabetic fatty (ZDF) rats, an established rat model of insulin-resistant type 2 diabetes mellitus. ZDF rats develop diabetes with vascular complications when fed a Western diet. In 21-wk-old diabetic rats, bone mineral density (BMD) was 22.5% (total) and 54.6% (trabecular) lower at the distal femur and 17.2% (total) and 20.4% (trabecular) lower at the lumbar spine, respectively, compared with nondiabetic animals. BMD distribution measured by backscattered electron imaging postmortem was not different between diabetic and nondiabetic rats, but evaluation of histomorphometric indexes revealed lower mineralized bone volume/tissue volume, trabecular thickness, and trabecular number. Osteoblast differentiation of diabetic rats was impaired based on lower alkaline phosphatase activity (-20%) and mineralized matrix formation (-55%). In addition, the expression of the osteoblast-specific genes bone morphogenetic protein-2, RUNX2, osteocalcin, and osteopontin was reduced by 40-80%. Osteoclast biology was not affected based on tartrate-resistant acidic phosphatase staining, pit formation assay, and gene profiling. To validate the implications of these molecular and cellular findings in a clinically relevant model, a subcritical bone defect of 3 mm was created at the left femur after stabilization with a four-hole plate, and bone regeneration was monitored by X-ray and microcomputed tomography analyses over 12 wk. While nondiabetic rats filled the defects by 57%, diabetic rats showed delayed bone regeneration with only 21% defect filling. In conclusion, we identified suppressed osteoblastogenesis as a cause and mechanism for low bone mass and impaired bone regeneration in a rat model of type 2 diabetes mellitus.

Identification of amino acids in the tetratricopeptide repeat and C-terminal domains of protein phosphatase 5 involved in autoinhibition and lipid activation.

PubMed

Kang, H; Sayner, S L; Gross, K L; Russell, L C; Chinkers, M

2001-09-04

Protein phosphatase 5 (PP5) exhibits low basal activity due to the autoinhibitory properties of its N-terminal and C-terminal domains but can be activated approximately 40-fold in vitro by polyunsaturated fatty acids. To identify residues involved in regulating PP5 activity, we performed scanning mutagenesis of its N-terminal tetratricopeptide repeat (TPR) domain and deletion mutagenesis of its C-terminal domain. Mutating residues in a groove of the TPR domain that binds to heat shock protein 90 had no effect on basal phosphatase activity. Mutation of Glu-76, however, whose side chain projects away from this groove, resulted in a 10-fold elevation of basal activity without affecting arachidonic acid-stimulated activity. Thus, the interface of the TPR domain involved in PP5 autoinhibition appears to be different from that involved in heat shock protein 90 binding. We also observed a 10-fold elevation of basal phosphatase activity upon removing the C-terminal 13 amino acids of PP5, with a concomitant 50% decrease in arachidonic acid-stimulated activity. These two effects were accounted for by two distinct amino acid deletions: deleting the four C-terminal residues (496-499) of PP5 had no effect on its activity, but removing Gln-495 elevated basal activity 10-fold. Removal of a further three amino acids had no additional effect, but deleting Asn-491 resulted in a 50% reduction in arachidonic acid-stimulated activity. Thus, Glu-76 in the TPR domain and Gln-495 at the C-terminus were implicated in maintaining the low basal activity of PP5. While the TPR domain alone has been thought to mediate fatty acid activation of PP5, our data suggest that Asn-491, near its C-terminus, may also be involved in this process.

The Role of DmCatD, a Cathepsin D-Like Peptidase, and Acid Phosphatase in the Process of Follicular Atresia in Dipetalogaster maxima (Hemiptera: Reduviidae), a Vector of Chagas' Disease

PubMed Central

Leyria, Jimena; Fruttero, Leonardo L.; Nazar, Magalí; Canavoso, Lilián E.

2015-01-01

In this work, we have investigated the involvement of DmCatD, a cathepsin D-like peptidase, and acid phosphatase in the process of follicular atresia of Dipetalogaster maxima, a hematophagous insect vector of Chagas’ disease. For the studies, fat bodies, ovaries and hemolymph were sampled from anautogenous females at representative days of the reproductive cycle: pre-vitellogenesis, vitellogenesis as well as early and late atresia. Real time PCR (qPCR) and western blot assays showed that DmCatD was expressed in fat bodies and ovaries at all reproductive stages, being the expression of its active form significantly higher at the atretic stages. In hemolymph samples, only the immunoreactive band compatible with pro-DmCatD was observed by western blot. Acid phosphatase activity in ovarian tissues significantly increased during follicular atresia in comparison to pre-vitellogenesis and vitellogenesis. A further enzyme characterization with inhibitors showed that the high levels of acid phosphatase activity in atretic ovaries corresponded mainly to a tyrosine phosphatase. Immunofluorescence assays demonstrated that DmCatD and tyrosine phosphatase were associated with yolk bodies in vitellogenic follicles, while in atretic stages they displayed a different cellular distribution. DmCatD and tyrosine phosphatase partially co-localized with vitellin. Moreover, their interaction was supported by FRET analysis. In vitro assays using homogenates of atretic ovaries as the enzyme source and enzyme inhibitors demonstrated that DmCatD, together with a tyrosine phosphatase, were necessary to promote the degradation of vitellin. Taken together, the results strongly suggested that both acid hydrolases play a central role in early vitellin proteolysis during the process of follicular atresia. PMID:26091289

Plasma fluctuation in estradiol-17β and bone resorption markers around parturition in dairy cows.

PubMed

Devkota, Bhuminad; Takahashi, Masahiro; Sato, Saori; Sasaki, Kouya; Ueki, Atsushi; Osawa, Takeshi; Takahashi, Masahiro; Yamagishi, Norio

2015-07-01

Blood samples were obtained sequentially from 10 dairy cows around the time of parturition to assess plasma fluctuations in estradiol-17β (E2) levels in association with those of several bone resorption markers. Plasma E2 concentration increased sharply a few days prepartum and decreased quickly after parturition. In terms of bone resorption markers, the plasma level of tartrate-resistant acid phosphatase isoform 5b (TRAP5b) rose significantly, commencing 1 week prepartum, and was maintained at this level to a few days postpartum. The plasma concentration of carboxyterminal collagen cross-links of type-I collagen (CTx) increased significantly after parturition. These observations suggest that osteoclast-mediated bone resorption was activated after parturition when plasma E2 concentrations decreased.

Plasma fluctuation in estradiol-17β and bone resorption markers around parturition in dairy cows

PubMed Central

DEVKOTA, Bhuminad; TAKAHASHI, Masahiro; SATO, Saori; SASAKI, Kouya; UEKI, Atsushi; OSAWA, Takeshi; TAKAHASHI, Masahiro; YAMAGISHI, Norio

2015-01-01

Blood samples were obtained sequentially from 10 dairy cows around the time of parturition to assess plasma fluctuations in estradiol-17β (E2) levels in association with those of several bone resorption markers. Plasma E2 concentration increased sharply a few days prepartum and decreased quickly after parturition. In terms of bone resorption markers, the plasma level of tartrate-resistant acid phosphatase isoform 5b (TRAP5b) rose significantly, commencing 1 week prepartum, and was maintained at this level to a few days postpartum. The plasma concentration of carboxyterminal collagen cross-links of type-I collagen (CTx) increased significantly after parturition. These observations suggest that osteoclast-mediated bone resorption was activated after parturition when plasma E2 concentrations decreased. PMID:25755022

Osthole Promotes Endochondral Ossification and Accelerates Fracture Healing in Mice.

PubMed

Zhang, Zhongrong; Leung, Wing Nang; Li, Gang; Lai, Yau Ming; Chan, Chun Wai

2016-12-01

Osthole has been found to restore bone mass in preclinical osteoporotic models. In the present study, we investigated the effects of osthole on bone fracture repair in mice. Adult C57BL/6 mice were subjected to transverse femoral fractures and administrated orally with 20 mg/kg osthole and vehicle solvent daily from week 1 post-operation. Fracture callus were analyzed by plain radiography, micro-computed tomography, histology, molecular imaging and immunohistochemistry and tartrate-resistant acid phosphatase staining. Results demonstrated that osthole treatment enhanced removal of cartilage and bony union during reparative stage without significant interfering on remodeling process. In vivo molecular imaging showed bone formation rate of the treatment group was almost twofold of control group at week 2 post-operation. Osthole augmented the expression of alkaline phosphatase and collagen type X in hypertrophic chondrocytes as well as expression of bone morphogenetic protein-2, osteocalcin and alkaline phosphatase in osteoblastic cells, indicating it promoted mineralization of hypertrophic cartilage and woven bone growth simultaneously during endochondral healing. In summary, osthole promotes endochondral ossification via upregulation of maturation osteogenic marker genes in chondrocytes and subsequently accelerates fracture repair and bony fusion.

Evolution of Enzymatic Activities in the Enolase Superfamily: D-Tartrate Dehydratase from Bradyrhizobium japonicum

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yew,W.; Fedorov, A.; Fedorov, E.

2006-01-01

We focus on the assignment of function to and elucidation of structure-function relationships for a member of the mechanistically diverse enolase superfamily encoded by the Bradyrhizobium japonicum genome (bll6730; GI:27381841). As suggested by sequence alignments, the active site contains the same functional groups found in the active site of mandelate racemase (MR) that catalyzes a 1,1-proton transfer reaction: two acid/base catalysts, Lys 184 at the end of the second {beta}-strand, and a His 322-Asp 292 dyad at the ends of the seventh and sixth -strands, respectively, as well as ligands for an essential Mg{sup 2+}, Asp 213, Glu 239, andmore » Glu 265 at the ends of the third, fourth, and fifth {beta}-strands, respectively. We screened a library of 46 acid sugars and discovered that only D-tartrate is dehydrated, yielding oxaloacetate as product. The kinetic constants (k{sub cat} = 7.3 s{sup -1}; k{sub cat}/K{sub M} = 8.5 x 10{sup 4} M{sup -1} s{sup -1}) are consistent with assignment of the D-tartrate dehydratase (TarD) function. The kinetic phenotypes of mutants as well as the structures of liganded complexes are consistent with a mechanism in which Lys 184 initiates the reaction by abstraction of the {alpha}-proton to generate a Mg{sup 2+}-stabilized enediolate intermediate, and the vinylogous -elimination of the 3-OH group is general acid-catalyzed by the His 322, accomplishing the anti-elimination of water. The replacement of the leaving group by solvent-derived hydrogen is stereorandom, suggesting that the enol tautomer of oxaloacetate is the product; this expectation was confirmed by its observation by {sup 1}H NMR spectroscopy. Thus, the TarD-catalyzed reaction is a 'simple' extension of the two-step reaction catalyzed by MR: base-catalyzed proton abstraction to generate a Mg{sup 2+}-stabilized enediolate intermediate followed by acid-catalyzed decomposition of that intermediate to yield the product.« less

Effect of salts on the kinetic parameters and thermal stability of bovine brain acid phosphatase.

PubMed

Bittencourt, H M; Chaimovich, H

1976-08-01

Bovine brain acid phosphatase is inhibited, at any pH, by an increase in ionic strength. The rate decrease is associated at pH 5, with a marked decrease in Km and, at pH 8, with a noticeable decrease in Vm. The rate of thermal inactivation of the enzyme is unaffected by increasing ionic strength up to 300 mM. These results are discussed in terms of interactions at the active site of the enzyme.

Diabetes reversal by inhibition of the low molecular weight tyrosine phosphatase

PubMed Central

Stanford, Stephanie M; Aleshin, Alexander E; Zhang, Vida; Ardecky, Robert J; Hedrick, Michael P; Zou, Jiwen; Ganji, Santhi R.; Bliss, Matthew R; Yamamoto, Fusayo; Bobkov, Andrey A.; Kiselar, Janna; Liu, Yingge; Cadwell, Gregory W; Khare, Shilpi; Yu, Jinghua; Barquilla, Antonio; Chung, Thomas DY; Mustelin, Tomas; Schenk, Simon; Bankston, Laurie A; Liddington, Robert C; Pinkerton, Anthony B; Bottini, Nunzio

2017-01-01

Obesity-associated insulin resistance plays a central role in type 2 diabetes. As such, tyrosine phosphatases that dephosphorylate the insulin receptor (IR) are potential therapeutic targets. The low molecular weight protein tyrosine phosphatase (LMPTP) is a proposed IR phosphatase, yet its role in insulin signaling in vivo has not been defined. Here we show that global and liver-specific LMPTP deletion protects mice from high-fat diet-induced diabetes without affecting body weight. To examine the role of the catalytic activity of LMPTP, we developed a small-molecule inhibitor with a novel uncompetitive mechanism, a unique binding site at the opening of the catalytic pocket, and exquisite selectivity over other phosphatases. This inhibitor is orally bioavailable, increases liver IR phosphorylation in vivo, and reverses high-fat diet induced diabetes. Our findings suggest that LMPTP is a key promoter of insulin resistance and that LMPTP inhibitors would be beneficial for treating type 2 diabetes. PMID:28346406

Diabetes reversal by inhibition of the low-molecular-weight tyrosine phosphatase.

PubMed

Stanford, Stephanie M; Aleshin, Alexander E; Zhang, Vida; Ardecky, Robert J; Hedrick, Michael P; Zou, Jiwen; Ganji, Santhi R; Bliss, Matthew R; Yamamoto, Fusayo; Bobkov, Andrey A; Kiselar, Janna; Liu, Yingge; Cadwell, Gregory W; Khare, Shilpi; Yu, Jinghua; Barquilla, Antonio; Chung, Thomas D Y; Mustelin, Tomas; Schenk, Simon; Bankston, Laurie A; Liddington, Robert C; Pinkerton, Anthony B; Bottini, Nunzio

2017-06-01

Obesity-associated insulin resistance plays a central role in type 2 diabetes. As such, tyrosine phosphatases that dephosphorylate the insulin receptor (IR) are potential therapeutic targets. The low-molecular-weight protein tyrosine phosphatase (LMPTP) is a proposed IR phosphatase, yet its role in insulin signaling in vivo has not been defined. Here we show that global and liver-specific LMPTP deletion protects mice from high-fat diet-induced diabetes without affecting body weight. To examine the role of the catalytic activity of LMPTP, we developed a small-molecule inhibitor with a novel uncompetitive mechanism, a unique binding site at the opening of the catalytic pocket, and an exquisite selectivity over other phosphatases. This inhibitor is orally bioavailable, and it increases liver IR phosphorylation in vivo and reverses high-fat diet-induced diabetes. Our findings suggest that LMPTP is a key promoter of insulin resistance and that LMPTP inhibitors would be beneficial for treating type 2 diabetes.

Acute response of plasma markers of bone turnover to a single bout of resistance training or plyometrics.

PubMed

Rogers, Robert S; Dawson, Andrew W; Wang, Ze; Thyfault, John P; Hinton, Pamela S

2011-11-01

The time course of changes in plasma bone turnover markers following an acute bout of resistance training (RT) or plyometrics (PLY) has not been well characterized. This study is the first to compare the acute response of bone formation and resorption markers to a single bout of RT or PLY. Using a partially randomized, cross-over study design, 12 recreationally active men, aged 43 ± 5 yr, each completed four exercise trials: RT (Fed/Fasted) and PLY (Fed/Fasted). In addition to the RT and PLY trials, 5 of the original 12 participants also completed a fasted, no-exercise control trial to examine time-of-day variation. For each trial, blood was drawn immediately before exercise (PRE), immediately following exercise, and 15 min, 30 min, 1 h, 2 h, and 24 h following PRE for determination of plasma bone-specific alkaline phosphatase (BAP), osteocalcin (OC), tartrate-resistant acid phosphatase 5b (TRAP5b), COOH-terminal telopeptide of type I collagen (CTX), testosterone, parathyroid hormone, and cortisol. A one-factor repeated-measures ANOVA was performed for each trial to detect changes in bone markers during the 2 h following RT or PLY. TRAP5b transiently decreased during the 2 h following all exercise trials (main effect for time, P < 0.05), but returned to PRE concentrations 2 h postexercise. BAP, CTX, and OC remained unchanged, except for reductions in BAP and CTX following PLY-Fasted and PLY-Fed, respectively. During the control trial, BAP decreased, while TRAP5b, CTX, and OC remained unchanged. In general, plasma hormone concentrations decreased during the 2 h following PLY or RT, and cumulative decreases in TRAP5b during the 2 h following exercise were positively correlated with cumulative decreases in parathyroid hormone. The results of the present study suggest that the timing of the measurement of bone turnover markers relative to the last exercise bout is important for detection of exercise-associated changes in bone turnover markers, as the markers returned

Colorimetric detection of trivalent chromium in aqueous solution using tartrate-capped silver nanoparticles as probe.

PubMed

Xu, Yunbo; Dong, Yangjun; Jiang, Xue; Zhu, Ningning

2013-10-01

This study describes a simple and highly selective method for the colorimetric detection of trivalent chromium (Cr3+) using tartrate-capped silver nanoparticles (AgNPs) as probe. The addition of tartrate to the initially prepared AgNPs gives tartrate-stabilized AgNPs ascribing to the electrostatic repulsion of the highly negatively charged tartrate ions covered on the surface of AgNPs. It is found that, in the presence of Cr3+ in aqueous solution, the aggregation of tartrate-stabilized AgNPs occurs. The color of AgNPs suspension changes from yellow to pink and the surface plasmon absorption band broadens and red shifts, which could be applied for the colorimetric detection of Cr3+ in aqueous solution. The utilization of tartrate-stabilized AgNPs as probe substantially increases the selectivity and sensitivity for colorimetric detection of Cr3+. Control experiments with the addition of over 14 other metal ions, such as Pb2+, Zn2+, Cr2O7(2-), Cd2+, Co2+, Cu2+, Al3+, Ni2+, Mn2+, Ba2+, Fe3+, Ca2+, Mg2+, Sr+ do not result in a distinct change in the color or in the spectrum of the suspension, indicating that these metal ions do not interfere with the colorimetric detection of Cr3+. Under the conditions employed here, A502/A393 (ratio of absorption value at 502 nm to 393 nm) is linear with the concentration of Cr3+ within a concentration range from 0.1 to 1.17 microM with a detection limit of 0.06 microM. This study may offer a simple, rapid and sensitive approach to colorimetric detection of Cr3+ in aqueous solution.

Soil properties influence kinetics of soil acid phosphatase in response to arsenic toxicity.

PubMed

Wang, Ziquan; Tan, Xiangping; Lu, Guannan; Liu, Yanju; Naidu, Ravi; He, Wenxiang

2018-01-01

Soil phosphatase, which plays an important role in phosphorus cycling, is strongly inhibited by Arsenic (As). However, the inhibition mechanism in kinetics is not adequately investigated. In this study, we investigated the kinetic characteristics of soil acid phosphatase (ACP) in 14 soils with varied properties, and also explored how kinetic properties of soil ACP changed with different spiked As concentrations. The results showed that the Michaelis constant (K m ) and maximum reaction velocity (V max ) values of soil ACP ranged from 1.18 to 3.77mM and 0.025-0.133mMh -1 in uncontaminated soils. The kinetic parameters of soil ACP in different soils changed differently with As contamination. The K m remained unchanged and V max decreased with increase of As concentration in most acid and neutral soils, indicating a noncompetitive inhibition mechanism. However, in alkaline soils, the K m increased linearly and V max decreased with increase of As concentration, indicating a mixed inhibition mechanism that include competitive and noncompetitive. The competitive inhibition constant (K ic ) and noncompetitive inhibition constant (K iu ) varied among soils and ranged from 0.38 to 3.65mM and 0.84-7.43mM respectively. The inhibitory effect of As on soil ACP was mostly affected by soil organic matter and cation exchange capacity. Those factors influenced the combination of As with enzyme, which resulted in a difference of As toxicity to soil ACP. Catalytic efficiency (V max /K m ) of soil ACP was a sensitive kinetic parameter to assess the ecological risks of soil As contamination. Copyright © 2017 Elsevier Inc. All rights reserved.

Cytochemical localization of phosphatases in the germ- and Sertoli cells of Odontophrynus cultripes (Amphibia, Anura, Leptodactylidae).

PubMed

Fernandes, A P; Báo, S N

1998-08-01

Ultrastructural cytochemical techniques were used for the localization of phosphatases in spermatid and spermatozoon, as well as in Sertoli cells of Odontophrynus cultripes (Amphibia, Anura, Leptodactylidae). Acid phosphatase was found in the acrosome. Thiamine pyrophosphatase was observed in the Golgi cisternae and in the tail spermatozoon surface. Glucose-6-phosphatase was located in the membrane complex of the acrosomal region. Already, in the Sertoli cells acid phosphatase was located in the lysosomes and glucose-6-phosphatase was observed in association with the endoplasmic reticulum and Golgi complex. These observations support the idea that various phosphatases may play some role in spermatid differentiation and in the interactions germ cells--Sertoli cells during spermiogenesis process.

Incidence of Quinolone Resistance Over the Period 1986 to 1998 in Veterinary Salmonella Isolates from Germany

PubMed Central

Malorny, Burkhard; Schroeter, Andreas; Helmuth, Reiner

1999-01-01

A total of 24,591 nonhuman salmonella strains isolated in Germany between 1986 and 1998 were examined for their resistance to nalidixic acid by an agar diffusion method. The rate of resistance (inhibition zone, ≤13 mm) ranged from 0.2% in 1986 to a peak of 14.8% in 1990. Between 1991 and 1998 the MICs for nalidixic acid-resistant strains ranged from more than 256 μg/ml for nalidixic acid to between 0.25 and 128 μg/ml for enrofloxacin. In the early 1990s a particularly high incidence of fluoroquinolone resistance (49.5%) was seen among isolates of Salmonella enterica serotype Typhimurium (Salmonella Typhimurium) definitive phage type 204c that mainly originated from cattle. Among isolates from poultry an increase in the incidence of nalidixic acid resistance to a peak of 14.4% was observed in 1994. This peak was due to the presence of specific resistant serotypes, mainly serotypes Hadar, Saintpaul, Paratyphi B (d-tartrate positive; formerly serotype Java) and Newport. Such strains exhibited a decreased susceptibility to enrofloxacin (MIC, 1 μg/ml). Among isolates from pigs the peak incidence of resistance was reached in 1993, with 7.5% of isolates resistant to nalidixic acid and enrofloxacin. The study demonstrates an increase in the incidence of strains that are resistant to nalidixic acid and that have decreased susceptibility to enrofloxacin after the licensing of enrofloxacin. In addition, the number of other serotypes that exhibited nalidixic acid resistance or reduced enrofloxacin susceptibility increased among the total number of isolates investigated between 1992 and 1998. PMID:10471579

Growth, structural, spectroscopic and optical characterization of barium doped calcium tartrate

NASA Astrophysics Data System (ADS)

Verma, Seema; Raina, Bindu; Gupta, Vandana; Bamzai, K. K.

2018-05-01

Barium doped calcium tartrates synthesized by controlled diffusion using silica gel technique at ambient temperature was characterized by single crystal X-ray diffraction which establishes monoclinic crystal system with volume of the unit cell 923.97(10) Ǻ3 and the space group being P21. UV - Vis characterization gives various linear optical constants like absorption, transmittance, reflectance, band gap, extinction coefficient, urbach energy, complex dielectric constant, optical and electrical conductivity. These constants are considered to be essential in characterizing materials that are used in various applications like fabrication of optoelectronic devices. FTIR spectrum establishes the presence of various bands of functional groups expected from metal tartrate with water of crystallization.

Kidney function and population-based outcomes of initiating oral atenolol versus metoprolol tartrate in older adults.

PubMed

Fleet, Jamie L; Weir, Matthew A; McArthur, Eric; Ozair, Sundus; Devereaux, Philip J; Roberts, Matthew A; Jain, Arsh K; Garg, Amit X

2014-12-01

Atenolol and metoprolol tartrate are commonly prescribed β-blockers. Atenolol elimination depends on kidney function, whereas metoprolol tartrate does not. We hypothesized that compared to metoprolol tartrate, initiating oral atenolol treatment would be associated with more adverse events in older adults, with the association most pronounced in patients with lower baseline estimated glomerular filtration rates (eGFRs). Population-based matched retrospective cohort study. Older adults (mean age, 75 years) in Ontario, Canada, prescribed oral atenolol versus metoprolol tartrate from April 2002 through December 2011. The 2 groups were well matched (n=75,257 in each group), with no difference in 31 measured baseline characteristics. Patients with end-stage renal disease were ineligible, and 4.6% of patients had chronic kidney disease (median eGFR, 38mL/min/1.73m(2) assessed through a database algorithm). β-Blocker type and eGFR. A composite outcome of hospitalization with bradycardia or hypotension and all-cause mortality were assessed in 90-day follow-up. Compared to metoprolol tartrate, initiating atenolol treatment was not associated with higher risk of hospitalization with bradycardia or hypotension (incidence, 0.71% vs 0.79%; relative risk, 0.90; 95%CI, 0.80-1.01). Atenolol treatment initiation was associated with lower 90-day risk of mortality than metoprolol tartrate (incidence, 0.97% vs 1.44%; relative risk, 0.68; 95%CI, 0.61-0.74). Lower eGFR did not modify either association (P for interaction=0.5 and 0.6, respectively). Heart rate and blood pressure were not available in our data sources, and effects ascertained from observational studies are subject to residual confounding. Contrary to our expectation, we found that atenolol versus metoprolol tartrate was associated with lower 90-day risk of mortality in patients regardless of eGFR, with no difference in risk of hospitalization with bradycardia or hypotension. Copyright © 2014 National Kidney Foundation

Identification of a novel phosphatase with high affinity for nucleotides monophosphate from common bean (Phaseolus vulgaris).

PubMed

Cabello-Díaz, Juan Miguel; Quiles, Francisco Antonio; Lambert, Rocío; Pineda, Manuel; Piedras, Pedro

2012-04-01

Common bean (Phaseolus vulgaris) seedlings accumulate ureides derived from purines after germination. The first step in the conversion of purines to ureides is the removal of the 5'-phosphate group by a phosphatase that has not been established yet. Two main phosphatase activities were detected in the embryonic axes of common bean using inosine monophosphate as substrate in an in-gel assay. Both activities differed in their sensitive to the common phosphatase inhibitor molybdate, with the molybdate-resistant as the first enzyme induced after radicle protrusion. The molybdate-resistant phosphatase has been purified to electrophoretic homogeneity and this is the first enzyme which shows this resistance purified and characterized from plant tissues. The native enzyme was a monomer of 55 kDa and it showed highest activity with nucleotides as substrates, with the K(m) values in the micromolar range. Among nucleotides, the highest specific constant (V(max)/K(m)) was observed for adenosine monophosphate. Furthermore, the enzyme was inhibited by nucleosides, the products of the enzymatic reaction, with maximum effect for adenosine. Common bean seedlings imbibed in the presence of adenosine monophosphate in vivo showed the highest molybdate-resistant phosphatase activity in the axes in addition to increased ureide content. The data presented suggests that purified phosphatase is involved in nucleotide metabolism in embryonic axes from common bean. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

Identification of Open Stomata1-Interacting Proteins Reveals Interactions with Sucrose Non-fermenting1-Related Protein Kinases2 and with Type 2A Protein Phosphatases That Function in Abscisic Acid Responses

DOE PAGES

Waadt, Rainer; Manalansan, Bianca; Rauniyar, Navin; ...

2015-09-04

The plant hormone abscisic acid (ABA) controls growth and development and regulates plant water status through an established signaling pathway. In the presence of ABA, pyrabactin resistance/regulatory component of ABA receptor proteins inhibit type 2C protein phosphatases (PP2Cs). This, in turn, enables the activation of Sucrose Nonfermenting1-Related Protein Kinases2 (SnRK2). Open Stomata1 (OST1)/SnRK2.6/SRK2E is a major SnRK2-type protein kinase responsible for mediating ABA responses. Arabidopsis (Arabidopsis thaliana) expressing an epitope-tagged OST1 in the recessive ost1-3 mutant background was used for the copurification and identification of OST1-interacting proteins after osmotic stress and ABA treatments. Furthemore, these analyses, which were confirmed usingmore » bimolecular fluorescence complementation and coimmunoprecipitation, unexpectedly revealed homo- and heteromerization of OST1 with SnRK2.2, SnRK2.3, OST1, and SnRK2.8. Furthermore, several OST1-complexed proteins were identified as type 2A protein phosphatase (PP2A) subunits and as proteins involved in lipid and galactolipid metabolism. More detailed analyses suggested an interaction network between ABA-activated SnRK2-type protein kinases and several PP2A-type protein phosphatase regulatory subunits. pp2a double mutants exhibited a reduced sensitivity to ABA during seed germination and stomatal closure and an enhanced ABA sensitivity in root growth regulation. Our analyses add PP2A-type protein phosphatases as another class of protein phosphatases to the interaction network of SnRK2-type protein kinases.« less

Identification of Open Stomata1-Interacting Proteins Reveals Interactions with Sucrose Non-fermenting1-Related Protein Kinases2 and with Type 2A Protein Phosphatases That Function in Abscisic Acid Responses1[OPEN

PubMed Central

Waadt, Rainer; Manalansan, Bianca; Rauniyar, Navin; Munemasa, Shintaro; Booker, Matthew A.; Brandt, Benjamin; Waadt, Christian; Nusinow, Dmitri A.; Kay, Steve A.; Kunz, Hans-Henning; Schumacher, Karin; DeLong, Alison; Yates, John R.; Schroeder, Julian I.

2015-01-01

The plant hormone abscisic acid (ABA) controls growth and development and regulates plant water status through an established signaling pathway. In the presence of ABA, pyrabactin resistance/regulatory component of ABA receptor proteins inhibit type 2C protein phosphatases (PP2Cs). This, in turn, enables the activation of Sucrose Nonfermenting1-Related Protein Kinases2 (SnRK2). Open Stomata1 (OST1)/SnRK2.6/SRK2E is a major SnRK2-type protein kinase responsible for mediating ABA responses. Arabidopsis (Arabidopsis thaliana) expressing an epitope-tagged OST1 in the recessive ost1-3 mutant background was used for the copurification and identification of OST1-interacting proteins after osmotic stress and ABA treatments. These analyses, which were confirmed using bimolecular fluorescence complementation and coimmunoprecipitation, unexpectedly revealed homo- and heteromerization of OST1 with SnRK2.2, SnRK2.3, OST1, and SnRK2.8. Furthermore, several OST1-complexed proteins were identified as type 2A protein phosphatase (PP2A) subunits and as proteins involved in lipid and galactolipid metabolism. More detailed analyses suggested an interaction network between ABA-activated SnRK2-type protein kinases and several PP2A-type protein phosphatase regulatory subunits. pp2a double mutants exhibited a reduced sensitivity to ABA during seed germination and stomatal closure and an enhanced ABA sensitivity in root growth regulation. These analyses add PP2A-type protein phosphatases as another class of protein phosphatases to the interaction network of SnRK2-type protein kinases. PMID:26175513

A pleîotropic acid phosphatase-deficient mutant of Escherichia coli shows premature termination in the dsbA gene. Use of dsbA::phoA fusions to localize a structurally important domain in DsbA.

PubMed

Belin, P; Quéméneur, E; Boquet, P L

1994-01-01

A one-step mutant of Escherichia coli K-12 lacking both glucose-1-phosphatase (Agp) and pH 2.5 acid phosphatase (AppA) activities in the periplasmic space was isolated. The mutation which mapped close to chlB, at 87 min on the E. coli linkage map, also caused the loss of alkaline phosphatase (PhoA) activity, even when this activity was expressed from TnphoA fusions to genes encoding periplasmic or membrane proteins. A DNA fragment that complements the mutation was cloned and shown to carry the dsbA gene, which encodes a periplasmic disulphide bond-forming factor. The mutant had an ochre triplet in dsbA, truncating the protein at amino acid 70. Introduction of TnphoA fusions into a plasmid-borne dsbA gene resulted in DsbA-PhoA hybrid proteins that were all exported to the periplasmic space in both dsbA+ and dsbA strains. They belong to three different classes, depending on the length of the DsbA fragment fused to PhoA. When PhoA was fused to an amino-terminal DsbA heptapeptide, the protein was only seen in the periplasm of a dsbA+ strain, as in the case of wild-type PhoA. Hybrid proteins missing up to 29 amino acids at the carboxy-terminus of DsbA were stable and retained both the DsbA and PhoA activities. Those with shorter DsbA fragments that still carried the -Cys-Pro-His-Cys- motif were rapidly degraded (no DsbA activity). The presence is discussed of a structural domain lying around amino acid 170 of DsbA and which is probably essential for its folding into a proteolytic-resistant and enzymatically active form.

Ergotism related to a single dose of ergotamine tartrate in an AIDS patient treated with ritonavir

PubMed Central

Blanche, P; Rigolet, A; Gombert, B; Ginsburg, C; Salmon, D; Sicard, D

1999-01-01

We report a rare case of ergotism related to a single dose of ergotamine tartrate in a man with AIDS being treated with ritonavir. He was treated with a prostacyclin analogue and made a complete recovery.


Keywords: ergotism; ergotamine tartrate; AIDS; ritonavir; adverse drug reaction; HIV infection PMID:10616689

An extract of Urtica dioica L. mitigates obesity induced insulin resistance in mice skeletal muscle via protein phosphatase 2A (PP2A).

PubMed

Obanda, Diana N; Ribnicky, David; Yu, Yongmei; Stephens, Jacqueline; Cefalu, William T

2016-02-26

The leaf extract of Urtica dioica L. (UT) has been reported to improve glucose homeostasis in vivo, but definitive studies on efficacy and mechanism of action are lacking. We investigated the effects of UT on obesity- induced insulin resistance in skeletal muscle. Male C57BL/6J mice were divided into three groups: low-fat diet (LFD), high-fat diet (HFD) and HFD supplemented with UT. Body weight, body composition, plasma glucose and plasma insulin were monitored. Skeletal muscle (gastrocnemius) was analyzed for insulin sensitivity, ceramide accumulation and the post translational modification and activity of protein phosphatase 2A (PP2A). PP2A is activated by ceramides and dephosphorylates Akt. C2C12 myotubes exposed to excess free fatty acids with or without UT were also evaluated for insulin signaling and modulation of PP2A. The HFD induced insulin resistance, increased fasting plasma glucose, enhanced ceramide accumulation and PP2A activity in skeletal muscle. Supplementation with UT improved plasma glucose homeostasis and enhanced skeletal muscle insulin sensitivity without affecting body weight and body composition. In myotubes, UT attenuated the ability of FFAs to induce insulin resistance and PP2A hyperactivity without affecting ceramide accumulation and PP2A expression. UT decreased PP2A activity through posttranslational modification that was accompanied by a reduction in Akt dephosphorylation.

An extract of Urtica dioica L. mitigates obesity induced insulin resistance in mice skeletal muscle via protein phosphatase 2A (PP2A)

PubMed Central

Obanda, Diana N.; Ribnicky, David; Yu, Yongmei; Stephens, Jacqueline; Cefalu, William T.

2016-01-01

The leaf extract of Urtica dioica L. (UT) has been reported to improve glucose homeostasis in vivo, but definitive studies on efficacy and mechanism of action are lacking. We investigated the effects of UT on obesity- induced insulin resistance in skeletal muscle. Male C57BL/6J mice were divided into three groups: low-fat diet (LFD), high-fat diet (HFD) and HFD supplemented with UT. Body weight, body composition, plasma glucose and plasma insulin were monitored. Skeletal muscle (gastrocnemius) was analyzed for insulin sensitivity, ceramide accumulation and the post translational modification and activity of protein phosphatase 2A (PP2A). PP2A is activated by ceramides and dephosphorylates Akt. C2C12 myotubes exposed to excess free fatty acids with or without UT were also evaluated for insulin signaling and modulation of PP2A. The HFD induced insulin resistance, increased fasting plasma glucose, enhanced ceramide accumulation and PP2A activity in skeletal muscle. Supplementation with UT improved plasma glucose homeostasis and enhanced skeletal muscle insulin sensitivity without affecting body weight and body composition. In myotubes, UT attenuated the ability of FFAs to induce insulin resistance and PP2A hyperactivity without affecting ceramide accumulation and PP2A expression. UT decreased PP2A activity through posttranslational modification that was accompanied by a reduction in Akt dephosphorylation. PMID:26916435

Cellular prostatic acid phosphatase, a PTEN-functional homologue in prostate epithelia, functions as a prostate-specific tumor suppressor

PubMed Central

Muniyan, Sakthivel; Ingersoll, Matthew A.; Batra, Surinder K.; Lin, Ming-Fong

2014-01-01

The inactivation of tumor suppressor genes (TSGs) plays a vital role in the progression of human cancers. Nevertheless, those ubiquitous TSGs have been shown with limited roles in various stages of diverse carcinogenesis. Investigation on identifying unique TSG, especially for early stage of carcinogenesis, is imperative. As such, the search for organ-specific TSGs has emerged as a major strategy in cancer research. Prostate cancer (PCa) has the highest incidence in solid tumors in US males. Cellular prostatic acid phosphatase (cPAcP) is a prostate-specific differentiation antigen. Despite intensive studies over the past several decades on PAcP as a PCa biomarker, the role of cPAcP as a PCa-specific tumor suppressor has only recently been emerged and validated. The mechanism underlying the pivotal role of cPAcP as a prostate-specific TSG is, in part, due to its function as a protein tyrosine phosphatase (PTP) as well as a phosphoinositide phosphatase (PIP), an apparent functional homologue to Phosphatase and tensin homolog (PTEN) in PCa cells. This review is focused on discussing the function of this authentic prostate-specific tumor suppressor and the mechanism behind the loss of cPAcP expression leading to prostate carcinogenesis. We review other phosphatases’ roles as TSGs which regulate oncogenic PI3K signaling in PCa and discuss the functional similarity between cPAcP and PTEN in prostate carcinogenesis. PMID:24747769

Diminished Progression of Periapical Lesions with Zoledronic Acid in Ovariectomized Rats.

PubMed

Wayama, Marcelo Tadahiro; Yoshimura, Hitoshi; Ohba, Seigo; Yoshida, Hisato; Matsuda, Shinpei; Kobayashi, Junichi; Kobayashi, Motohiro; Gomes Filho, João Eduardo; Sano, Kazuo

2015-12-01

The aim of this study was to investigate the effects of systemically administered zoledronic acid (ZOL) on the progression of periapical lesions in estrogen-deficient rats. Female Wistar rats were divided into the following groups: SHAM-veh, sham surgery treated with vehicle (physiological saline); OVX-veh, ovariectomy treated with vehicle; SHAM-ZOL, sham surgery treated with ZOL; and OVX-ZOL, ovariectomy treated with ZOL. Vehicle or ZOL was administered intravenously once a week for 4 weeks. The pulp of the mandibular first molar of all rats was exposed to the oral environment to induce a periapical lesion, and the lesions were analyzed after 7 and 30 days. The mandibles were examined by micro-computed tomographic imaging and histopathologic, histometric, and immunohistochemical analyses. Histopathologically, the OVX-veh group had more severe inflammation and bone loss and a larger number of cells that were positive for tartrate-resistant acid phosphatase compared with the SHAM-veh and OVX-ZOL groups; the SHAM-veh and OVX-ZOL groups were similar to each other. The SHAM-ZOL group had the lowest magnitude of these conditions. Tomographically, the OVX-veh group had greater bone loss than the other groups at both time points. The SHAM-veh, SHAM-ZOL, and OVX-ZOL groups had similar bone loss at both time points. In the sagittal section on day 30, the SHAM-ZOL group had lower bone loss compared with the SHAM-veh and OVX-ZOL groups. The hypoestrogenic condition aggravates the progression of periapical lesions. ZOL therapy may help contain bone destruction of periapical lesions. Copyright © 2015 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Phosphatase activities as biosignatures of extant life

NASA Astrophysics Data System (ADS)

Kobayashi, K.; Itoh, Y.; Edazawa, Y.; Moroi, A.; Takano, Y.

It has been recognized that terrestrial biosphere expands to such extreme environments as deep subsurface lithosphere high temperature hot springs and stratosphere Possible extraterrestrial biospheres in Mars Europa and Titan are being discussed Many biosignatures or biomarkers have been proposed to detect microbial activities in such extreme environments Phosphate esters are essential for the terrestrial life since they are constituents of nucleic acids and cell mebranes Thus all the terrestrial organisms have phosphatases that are enzymes catalyzing hydrolysis of phosphate esters We analyzed phosphatase activities in the samples obtained in extreme environments such as submarine hydrothermal systems and discussed whether they can be used as biosignatures for extant life Core samples and chimney samples were collected at the Suiyo Seamount Izu-Bonin Arc the Pacific Ocean in 2001 and 2002 and in South Mariana hydrothermal systems the Pacific Oceanas in 2003 both in a part of the Archaean Park Project Phosphatase activity in solid rock samples was measured spectrometrically by using 25 mM p-nitrophenyl phosphate pH 8 0 or pH 6 5 as a substrate as follows Pulverized samples were incuvated with substrate solution for an hour and then production rate of p-nitrophenol was calculated with absorbance at 410 nm Phosphatase activity in extracts was measured fluorometrically by using 4-methylumberyferryl phosphate as a substrate Concentration of amino acids and their enantiomeric ratio were determined by HPLC after HF digestion of the

Vanillic acid derivatives from the green algae Cladophora socialis as potent protein tyrosine phosphatase 1B inhibitors.

PubMed

Feng, Yunjiang; Carroll, Anthony R; Addepalli, Rama; Fechner, Gregory A; Avery, Vicky M; Quinn, Ronald J

2007-11-01

A novel vanillic acid derivative (1) and its sulfate adduct (2) were isolated from a green algae, Cladophora socialis. The structures of 1 and 2 were elucidated from NMR and HRESIMS experiments. Both compounds showed potent inhibitory activity against protein tyrosine phosphatase 1B (PTP1B), an enzyme involved in the regulation of insulin cell signaling. Compounds 1 and 2 had IC50 values of 3.7 and 1.7 microM, respectively.

Development of modified-release tablets of zolpidem tartrate by biphasic quick/slow delivery system.

PubMed

Mahapatra, Anjan Kumar; Sameeraja, N H; Murthy, P N

2015-06-01

Zolpidem tartrate is a non-benzodiazepine analogue of imidazopyridine of sedative and hypnotic category. It has a short half-life with usual dosage regimen being 5 mg, two times a day, or 10 mg, once daily. The duration of action is considered too short in certain circumstances. Thus, it is desirable to lengthen the duration of action. The formulation design was implemented by preparing extended-release tablets of zolpidem tartrate using the biphasic delivery system technology, where sodium starch glycolate acts as a superdisintegrant in immediate-release part and hydroxypropyl methyl cellulose as a release retarding agent in extended-release core. Tablets were prepared by direct compression. Both the core and the coat contained the drug. The pre-compression blends were evaluated for angle of repose, bulk density, and compressibility index. The tablets were evaluated for thickness, hardness, weight variation test, friability, and in vitro release studies. No interaction was observed between zolpidem tartrate and excipients from the Fourier transform infrared spectroscopy and differential scanning calorimetry analysis. The results of all the formulations prepared were compared with reference product Stilnoct®. Optimized formulations showed release patterns that match the United States Pharmacopeia (USP) guidelines for zolpidem tartrate extended-release tablets. The mechanism of drug release was studied using different mathematical models, and the optimized formulation has shown Fickian diffusion. Accelerated stability studies were performed on the optimized formulation.

Co-administration of α-lipoic acid and glutathione is associated with no significant changes in serum bilirubin, alkaline phosphatase or γ-glutamyltranspeptidase levels during the treatment of neuroborreliosis with intravenous ceftriaxone.

PubMed

Puri, Basant K; Hakkarainen-Smith, Jaana S; Derham, Anne; Monro, Jean A

2015-09-01

While pharmacotherapy with intravenous ceftriaxone, a third-generation cephalosporin, is a potential treatment of Lyme neuroborreliosis, there is concern that it can cause the formation of biliary sludge, leading to hepatobiliary complications such as biliary colic, jaundice and cholelithiasis, which are reflected in changes in serum levels of bilirubin and markers of cholestatic liver injury (alkaline phosphatase and γ-glutamyltranspeptidase). It has been suggested that the naturally occurring substances α-lipoic acid and glutathione may be helpful in preventing hepatic disease. α-Lipoic acid exhibits antioxidant, anti-inflammatory and anti-apoptotic activities in the liver, while glutathione serves as a sulfhydryl buffer. The aim of this study was to determine whether co-administration of α-lipoic acid and glutathione is associated with significant changes in serum levels of bilirubin, alkaline phosphatase and γ-glutamyltranspeptidase during the treatment of Lyme neuroborreliosis with long-term intravenous ceftriaxone. Serum levels of bilirubin, alkaline phosphatase and γ-glutamyltranspeptidase were measured in 42 serologically positive Lyme neuroborreliosis patients before and after long-term treatment with intravenous ceftriaxone (2-4 g daily) with co-administration of oral/intravenous α-lipoic acid (600 mg daily) and glutathione (100 mg orally or 0.6-2.4 g intravenously daily). None of the patients developed biliary colic and there were no significant changes in serum bilirubin, alkaline phosphatase or γ-glutamyltranspeptidase levels over the course of the intravenous ceftriaxone treatment (mean length 75.0 days). Co-administration of α-lipoic acid and glutathione is associated with no significant changes in serum bilirubin, alkaline phosphatase or γ-glutamyltranspeptidase levels during the treatment of neuroborreliosis with intravenous ceftriaxone.

Molecular Characterization of Multiresistant d-Tartrate-Positive Salmonella enterica Serovar Paratyphi B Isolates

PubMed Central

Miko, Angelika; Guerra, Beatriz; Schroeter, Andreas; Dorn, Christina; Helmuth, Reiner

2002-01-01

Since 1996, the National Salmonella Reference Laboratory of Germany has received an increasing number of Salmonella enterica subsp. enterica serovar Paratyphi B isolates. Nearly all of these belonged to the dextrorotatory tartrate-positive variant (S. enterica subsp. enterica serovar Paratyphi B dT+), formerly called S. enterica subsp. enterica serovar Java. A total of 55 selected contemporary and older S. enterica subsp. enterica serovar Paratyphi B dT+ isolates were analyzed by plasmid profiling, antimicrobial resistance testing, pulsed-field gel electrophoresis, IS200 profiling, and PCR-based detection of integrons. The results showed a high genetic heterogeneity among 10 old strains obtained from 1960 to 1993. In the following years, however, new distinct multiresistant S. enterica subsp. enterica serovar Paratyphi B dT+ clones emerged, and one clonal lineage successfully displaced the older ones. Since 1994, 88% of the isolates investigated were multiple drug resistant. Today, a particular clone predominates in some German poultry production lines, poultry products, and various other sources. It was also detected in contemporary isolates from two neighboring countries as well. PMID:12202551

Older adults with heart failure treated with carvedilol, bisoprolol, or metoprolol tartrate: risk of mortality.

PubMed

Perreault, Sylvie; de Denus, Simon; White, Michel; White-Guay, Brian; Bouvier, Michel; Dorais, Marc; Dubé, Marie-Pierre; Rouleau, Jean-Lucien; Tardif, Jean-Claude; Jenna, Sarah; Haibe-Kains, Benjamin; Leduc, Richard; Deblois, Denis

2017-01-01

The long-term use of β-blockers has been shown to improve clinical outcomes among patients with heart failure (HF). However, a lack of data persists in assessing whether carvedilol or bisoprolol are superior to metoprolol tartrate in clinical practice. We endeavored to compare the effectiveness of β-blockers among older adults following a primary hospital admission for HF. We conducted a cohort study using Quebec administrative databases to identify patients who were using β-blockers, carvedilol, bisoprolol, or metoprolol tartrate after the diagnosis of HF. We characterized the patients by the type of β-blocker prescribed at discharge of their first HF hospitalization. An adjusted multivariate Cox proportional hazards model was used to compare the primary outcome of all-cause mortality. We also conducted analyses by matching for a propensity score for initiation of β-blocker therapy and assessed the effect on primary outcome. Among 3197 patients with HF with a median follow-up of 2.8 years, the crude annual mortality rates (per 100 person-years) were at 16, 14.9, and 17.7 for metoprolol tartrate, carvedilol, and bisoprolol, respectively. Adjusted hazard ratios of carvedilol (hazard ratio 0.92; 0.78-1.09) and bisoprolol (hazard ratio 1.04; 0.93-1.16) were not significantly different from that of metoprolol tartrate in improving survival. After matching for propensity score, carvedilol and bisoprolol showed no additional benefit with respect to all-cause mortality compared with metoprolol tartrate. Our evidence suggests no differential effect of β-blockers on all-cause mortality among older adults with HF. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

The structure of a purple acid phosphatase involved in plant growth and pathogen defence exhibits a novel immunoglobulin-like fold.

PubMed

Antonyuk, Svetlana Vladimirovna; Olczak, Mariusz; Olczak, Teresa; Ciuraszkiewicz, Justyna; Strange, Richard William

2014-03-01

Phosphatases function in the production, transport and recycling of inorganic phosphorus, which is crucial for cellular metabolism and bioenergetics, as well as in bacterial killing, since they are able to generate reactive oxygen species via Fenton chemistry. Diphosphonucleotide phosphatase/phosphodiesterase (PPD1), a glycoprotein plant purple acid phosphatase (PAP) from yellow lupin seeds, contains a bimetallic Fe-Mn catalytic site which is most active at acidic pH. Unlike other plant PAPs, PPD1 cleaves the pyrophosphate bond in diphosphonucleotides and the phosphodiester bond in various phosphodiesters. The homohexameric organization of PPD1, as revealed by a 1.65 Å resolution crystal structure and confirmed by solution X-ray scattering, is unique among plant PAPs, for which only homodimers have previously been reported. A phosphate anion is bound in a bidentate fashion at the active site, bridging the Fe and Mn atoms in a binding mode similar to that previously reported for sweet potato PAP, which suggests that common features occur in their catalytic mechanisms. The N-terminal domain of PPD1 has an unexpected and unique fibronectin type III-like fold that is absent in other plant PAPs. Here, the in vitro DNA-cleavage activity of PPD1 is demonstrated and it is proposed that the fibronectin III-like domain, which 'overhangs' the active site, is involved in DNA selectivity, binding and activation. The degradation of DNA by PPD1 implies a role for PPD1 in plant growth and repair and in pathogen defence.

Identification of caffeoylquinic acid derivatives as natural protein tyrosine phosphatase 1B inhibitors from Artemisia princeps.

PubMed

Zhang, Jie; Sasaki, Tatsunori; Li, Wei; Nagata, Kazuya; Higai, Koji; Feng, Feng; Wang, Jian; Cheng, Maosheng; Koike, Kazuo

2018-04-15

Considerable attention has been paid to protein tyrosine phosphatase 1B (PTP1B) inhibitors as a potential therapy for diabetes, obesity, and cancer. Ten caffeoylquinic acid derivatives (1-10) from leaves of Artemisia princeps Pamp. (Asteraceae) were identified as natural PTP1B inhibitors. Among them, chlorogenic acid (3) showed the most potent inhibitory activity (IC 50 11.1 μM). Compound 3 was demonstrated to be a noncompetitive inhibitor by a kinetic analysis. Molecular docking simulation suggested that compound 3 bound to the allosteric site of PTP1B. Furthermore, compound 3 showed remarkable selectivity against four homologous PTPs. According to these findings, compound 3 might be potentially valuable for further drug development. Copyright © 2018 Elsevier Ltd. All rights reserved.

Assessment and kinetics of soil phosphatase in Brazilian Savanna systems.

PubMed

Ferreira, Adão S; Espíndola, Suéllen P; Campos, Maria Rita C

2016-05-31

The activity and kinetics of soil phosphatases are important indicators to evaluate soil quality in specific sites such as the Cerrado (Brazilian Savanna). This study aimed to determine the activity and kinetic parameters of soil phosphatase in Cerrado systems. Soil phosphatase activity was assessed in samples of native Cerrado (NC), no-tillage (NT), conventional tillage (CT) and pasture with Brachiaria brizantha (PBb) and evaluated with acetate buffer (AB), tris-HCl buffer (TB), modified universal buffer (MUB) and low MUB. The Michaelis-Menten equation and Eadie-Hofstee model were applied to obtain the kinetic parameters of soil phosphatase using different concentrations of p-nitrophenol phosphate (p-NPP). MUB showed the lowest soil phosphatase activity in all soils whereas AB in NC and NT presented the highest. Low MUB decreased interferences in the assessment of soil phosphatase activity when compared to MUB, suggesting that organic acids interfere on the soil phosphatase activity. In NC and NT, soil phosphatase activity performed with TB was similar to AB and low MUB. Km values from the Michaels-Menten equation were higher in NC than in NT, which indicate a lower affinity of phosphatase activity for the substrate in NC. Vmax values were also higher in NC than in NT. The Eadie-Hofstee model suggests that NC had more phosphatase isoforms than NT. The study showed that buffer type is of fundamental importance when assessing soil phosphatase activity in Cerrado soils.

Phelligridin D-loaded oral nanotube titanium implant enhances osseointegration and prevents osteolysis in rat mandible.

PubMed

Kim, Ji-Eun; Takanche, Jyoti Shrestha; Kim, Jeong-Seok; Lee, Min-Ho; Jeon, Jae-Gyu; Park, Il-Song; Yi, Ho-Keun

2018-04-12

Poor bone quality and osteolysis are the major causes of implant failure in dentistry. Here, this study tested the effect of phelligridin D-loaded nanotubes titanium (Ti) for bone formation around the dental implants. The purpose of this study was to enhance osseointegration of phelligridin D-loaded implant into the bone for bone formation and prevention of osteolysis. Cell viability, crystal violet staining, Western blot, alizarin red S staining, alkaline phosphatase activity, tartrate-resistant acid phosphatase staining, micro-computed tromography (μ-CT), hematoxylin and eosin (H&E) and immunohistochemical staining were used in vitro and in vivo to test the biocompatibility of phelligridin D. Phelligridin D enhanced osteoblast differentiation and mineralization by increasing bone morphogenic protein-2/7 (BMP-2/7), Osterix, Runx-2, osteoprotegerin (OPG), alkaline phosphatase and inhibited osteoclast differentiation by decreasing receptor activator of nuclear factor kappa-B ligand (RANKL) in MC-3T3 E1 cells. Further, phelligridin D promoted bone regeneration around nanotube Ti implant surface by increasing the levels of BMP-2/7 and OPG in a rat model. Phelligridin D also inhibited osteolysis by suppressing the expression of RANKL. These findings strongly suggest that phelligridin D is a new compound representing a potential therapeutic candidate for implant failure caused by osteolysis and poor bone quality of teeth.

A New Rapid and Sensitive Stability-Indicating UPLC Assay Method for Tolterodine Tartrate: Application in Pharmaceuticals, Human Plasma and Urine Samples.

PubMed

Yanamandra, Ramesh; Vadla, Chandra Sekhar; Puppala, Umamaheshwar; Patro, Balaram; Murthy, Yellajyosula L N; Ramaiah, Parimi Atchuta

2012-01-01

A new rapid, simple, sensitive, selective and accurate reversed-phase stability-indicating Ultra Performance Liquid Chromatography (RP-UPLC) technique was developed for the assay of Tolterodine Tartrate in pharmaceutical dosage form, human plasma and urine samples. The developed UPLC method is superior in technology to conventional HPLC with respect to speed, solvent consumption, resolution and cost of analysis. Chromatographic run time was 6 min in reversed-phase mode and ultraviolet detection was carried out at 220 nm for quantification. Efficient separation was achieved for all the degradants of Tolterodine Tartrate on BEH C18 sub-2-μm Acquity UPLC column using Trifluoroacetic acid and acetonitrile as organic solvent in a linear gradient program. The active pharmaceutical ingredient was extracted from tablet dosage form using a mixture of acetonitrile and water as diluent. The calibration graphs were linear and the method showed excellent recoveries for bulk and tablet dosage form. The test solution was found to be stable for 40 days when stored in the refrigerator between 2 and 8 °C. The developed UPLC method was validated and meets the requirements delineated by the International Conference on Harmonization (ICH) guidelines with respect to linearity, accuracy, precision, specificity and robustness. The intra-day and inter-day variation was found be less than 1%. The method was reproducible and selective for the estimation of Tolterodine Tartrate. Because the method could effectively separate the drug from its degradation products, it can be employed as a stability-indicating one.

The maize (Zea mays ssp. mays var. B73) genome encodes 33 members of the purple acid phosphatase family

PubMed Central

González-Muñoz, Eliécer; Avendaño-Vázquez, Aida-Odette; Montes, Ricardo A. Chávez; de Folter, Stefan; Andrés-Hernández, Liliana; Abreu-Goodger, Cei; Sawers, Ruairidh J. H.

2015-01-01

Purple acid phosphatases (PAPs) play an important role in plant phosphorus nutrition, both by liberating phosphorus from organic sources in the soil and by modulating distribution within the plant throughout growth and development. Furthermore, members of the PAP protein family have been implicated in a broader role in plant mineral homeostasis, stress responses and development. We have identified 33 candidate PAP encoding gene models in the maize (Zea mays ssp. mays var. B73) reference genome. The maize Pap family includes a clear single-copy ortholog of the Arabidopsis gene AtPAP26, shown previously to encode both major intracellular and secreted acid phosphatase activities. Certain groups of PAPs present in Arabidopsis, however, are absent in maize, while the maize family contains a number of expansions, including a distinct radiation not present in Arabidopsis. Analysis of RNA-sequencing based transcriptome data revealed accumulation of maize Pap transcripts in multiple plant tissues at multiple stages of development, and increased accumulation of specific transcripts under low phosphorus availability. These data suggest the maize PAP family as a whole to have broad significance throughout the plant life cycle, while highlighting potential functional specialization of individual family members. PMID:26042133

Short term effects of Glomus claroideum and Azospirillum brasilense on growth and root acid phosphatase activity of Carica papaya L. under phosphorus stress.

PubMed

Alarcón, Alejandro; Davies, Frederick T; Egilla, Johnatan N; Fox, Theodore C; Estrada-Luna, Arturo A; Ferrera-Cerrato, Ronald

2002-01-01

Arbuscular mycorrhizal fungi (AMF) are able to increase root enzymatic activity of acid and alkaline phosphatases. However, the role of AMF on phosphatase activity has not been reported in papaya (Carica papaya L.), which is frequently established at places with soil phosphorus (P) deficiencies. The goals of this research were to determine the effect of Glomus claroideum (Gc), and plant growth promoting rhizobacterium Azospirillum brasilense strain VS7 [Ab]) on root phosphatase activity and seedling growth of Carica papaya L. cv. Red Maradol under low P conditions. There were four treatments-colonization with: 1) Gc, 2) Ab, 3) Gc+Ab, and 4) non-inoculated seedlings. Plants were established in a coarse sand:sandy loam substrate under P-limitation (11 microg P ml(-1)), supplied with a modified Long Ashton Nutrient Solution. Seedling growth was severely reduced by low P. Gc+Ab inoculated plants had greater total dry matter and leaf area than non-colonized plants. Gc-inoculated plants had greater leaf area than non-colonized plants. Treatments did not differ in leaf area ratio, specific leaf area and, total chlorophyll content. There was a non-significant effect on stem relative growth rate with Gc and Gc+Ab plants. Mycorrhizal colonization enhanced the bacterial population 3.4-fold in the Gc+Ab treatment compared with the population quantified in Ab treatment. Soluble and extractable root acid phosphatase activity (RAPA) was higher in Gc inoculated plants. We discussed on the possible relation among both inoculated microorganisms and also with the P-limitation which plants were established.

Characterization of a soluble phosphatidic acid phosphatase in bitter melon (Momordica charantia).

PubMed

Cao, Heping; Sethumadhavan, Kandan; Grimm, Casey C; Ullah, Abul H J

2014-01-01

Momordica charantia is often called bitter melon, bitter gourd or bitter squash because its fruit has a bitter taste. The fruit has been widely used as vegetable and herbal medicine. Alpha-eleostearic acid is the major fatty acid in the seeds, but little is known about its biosynthesis. As an initial step towards understanding the biochemical mechanism of fatty acid accumulation in bitter melon seeds, this study focused on a soluble phosphatidic acid phosphatase (PAP, 3-sn-phosphatidate phosphohydrolase, EC 3.1.3.4) that hydrolyzes the phosphomonoester bond in phosphatidate yielding diacylglycerol and P(i). PAPs are typically categorized into two subfamilies: Mg(2+)-dependent soluble PAP and Mg(2+)-independent membrane-associated PAP. We report here the partial purification and characterization of an Mg(2+)-independent PAP activity from developing cotyledons of bitter melon. PAP protein was partially purified by successive centrifugation and UNOsphere Q and S columns from the soluble extract. PAP activity was optimized at pH 6.5 and 53-60 °C and unaffected by up to 0.3 mM MgCl2. The K(m) and Vmax values for dioleoyl-phosphatidic acid were 595.4 µM and 104.9 ηkat/mg of protein, respectively. PAP activity was inhibited by NaF, Na(3)VO(4), Triton X-100, FeSO4 and CuSO4, but stimulated by MnSO4, ZnSO4 and Co(NO3)2. In-gel activity assay and mass spectrometry showed that PAP activity was copurified with a number of other proteins. This study suggests that PAP protein is probably associated with other proteins in bitter melon seeds and that a new class of PAP exists as a soluble and Mg(2+)-independent enzyme in plants.

Phosphate forms an unusual tripodal complex with the Fe–Mn center of sweet potato purple acid phosphatase

PubMed Central

Schenk, Gerhard; Gahan, Lawrence R.; Carrington, Lyle E.; Mitić, Nataša; Valizadeh, Mohsen; Hamilton, Susan E.; de Jersey, John; Guddat, Luke W.

2005-01-01

Purple acid phosphatases (PAPs) are a family of binuclear metalloenzymes that catalyze the hydrolysis of phosphoric acid esters and anhydrides. A PAP in sweet potato has a unique, strongly antiferromagnetically coupled Fe(III)–Mn(II) center and is distinguished from other PAPs by its increased catalytic efficiency for a range of activated and unactivated phosphate esters, its strict requirement for Mn(II), and the presence of a μ-oxo bridge at pH 4.90. This enzyme displays maximum catalytic efficiency (kcat/Km) at pH 4.5, whereas its catalytic rate constant (kcat) is maximal at near-neutral pH, and, in contrast to other PAPs, its catalytic parameters are not dependent on the pKa of the leaving group. The crystal structure of the phosphate-bound Fe(III)–Mn(II) PAP has been determined to 2.5-Å resolution (final Rfree value of 0.256). Structural comparisons of the active site of sweet potato, red kidney bean, and mammalian PAPs show several amino acid substitutions in the sweet potato enzyme that can account for its increased catalytic efficiency. The phosphate molecule binds in an unusual tripodal mode to the two metal ions, with two of the phosphate oxygen atoms binding to Fe(III) and Mn(II), a third oxygen atom bridging the two metal ions, and the fourth oxygen pointing toward the substrate binding pocket. This binding mode is unique among the known structures in this family but is reminiscent of phosphate binding to urease and of sulfate binding to λ protein phosphatase. The structure and kinetics support the hypothesis that the bridging oxygen atom initiates hydrolysis. PMID:15625111

Reduction of the degradation activity of umami-enhancing purinic ribonucleotide supplement in miso by the targeted suppression of acid phosphatases in the Aspergillus oryzae starter culture.

PubMed

Marui, Junichiro; Tada, Sawaki; Fukuoka, Mari; Wagu, Yutaka; Shiraishi, Yohei; Kitamoto, Noriyuki; Sugimoto, Tatsuya; Hattori, Ryota; Suzuki, Satoshi; Kusumoto, Ken-Ichi

2013-09-02

Miso (fermented soybean paste) is a traditional Japanese fermented food, and is now used worldwide. The solid-state culture of filamentous fungus, Aspergillus oryzae, grown on rice is known as rice-koji, and is important as a starter for miso fermentation because of its prominent hydrolytic enzyme activities. Recently, commercial miso products have been supplemented with purinic ribonucleotides, such as inosine monophosphate (IMP) and guanine monophosphate, to enhance the characteristic umami taste of glutamate in miso. Because the purinic ribonucleotides are degraded by enzymes such as acid phosphatases in miso, heat inactivation is required prior to the addition of these flavorings. However, heat treatment is a costly process and reduces the quality of miso. Therefore, an approach to lower acid phosphatase activities in koji culture is necessary. Transcriptional analysis using an A. oryzae KBN8048 rice-koji culture showed that eight of the 13 acid phosphatase (aph) genes were significantly down-regulated by the addition of phosphoric acid in the preparation of the culture in a concentration-dependent manner, while aphC expression was markedly up-regulated under the same conditions. The eight down-regulated genes might be under the control of the functional counterpart of the Saccharomyces cerevisiae transcriptional activator Pho4, which specifically regulates phosphatase genes in response to the ambient phosphate availability. However, the regulatory mechanism of aphC was not clear. The IMP dephosphorylation activities in rice-koji cultures of KBN8048 and the aphC deletion mutant (ΔaphC) were reduced by up to 30% and 70%, respectively, in cultures with phosphoric acid, while protease and amylase activity, which is important for miso fermentation, was minimally affected. The miso products fermented using the rice-koji cultures of KBN8048 and ΔaphC prepared with phosphoric acid had reductions in IMP dephosphorylation activity of 80% and 90%, respectively, without

Overexpression of Human Bone Alkaline Phosphatase in Pichia Pastoris

NASA Technical Reports Server (NTRS)

Karr, Laurel; Malone, Christine, C.; Rose, M. Franklin (Technical Monitor)

2000-01-01

The Pichiapastoris expression system was utilized to produce functionally active human bone alkaline phosphatase in gram quantities. Bone alkaline phosphatase is a key enzyme in bone formation and biomineralization, yet important questions about its structural chemistry and interactions with other cellular enzymes in mineralizing tissues remain unanswered. A soluble form of human bone alkaline phosphatase was constructed by deletion of the 25 amino acid hydrophobic C-terminal region of the encoding cDNA and inserted into the X-33 Pichiapastoris strain. An overexpression system was developed in shake flasks and converted to large-scale fermentation. Alkaline phosphatase was secreted into the medium to a level of 32mgAL when cultured in shake flasks. Enzyme activity was 12U/mg measured by a spectrophotometric assay. Fermentation yielded 880mgAL with enzymatic activity of 968U/mg. Gel electrophoresis analysis indicates that greater than 50% of the total protein in the fermentation is alkaline phosphatase. A purification scheme has been developed using ammonium sulfate precipitation followed by hydrophobic interaction chromatography. We are currently screening crystallization conditions of the purified recombinant protein for subsequent X-ray diffraction analyses. Structural data should provide additional information on the role of alkaline phosphatase in normal bone mineralization and in certain bone mineralization anomalies.

Relative changes in phosphatase activities as influenced by source and application rate of organic composts in field crops.

PubMed

Saha, Supradip; Mina, B L; Gopinath, K A; Kundu, S; Gupta, H S

2008-04-01

Potential impact of different levels and sources of organic composts on activities of phosphatases (acid and alkaline phosphatase, phosphodiesterase, and inorganic pyrophosphatase) was studied after three years of continuous application. Enzyme activities were compared with microbial biomass P and available P. Experimental plots were divided based on the organic source into three groups: those receiving farmyard manure (FYM), vermicompost (VC) and Lantana compost (LC). Microbial biomass P (11.7 g kg(-1) soil), available P (24.0 g kg(-1) soil) and acid phosphatase (1.3 mg g(-1) p-NP g(-1) soil h(-1)) was highest in highest dose of VC. Acid phosphatase activity was high in all plots, including those where microbial biomass P levels were low. Most of the phosphatase activities were significantly correlated with available P in FYM and VC. These relationships were negative for LC treatments. Results showed that application of earthworm casts is helpful in faster transformation of organic P by facilitating better environment to microbes and plant roots.

Nucleic Acid-Induced Resistance to Viral Infection

PubMed Central

Takano, Kouichi; Warren, Joel; Jensen, Keith E.; Neal, Alan L.

1965-01-01

Takano, Kouichi (Chas. Pfizer & Co., Inc., Terre Haute, Ind.), Joel Warren, Keith E. Jensen, and Alan L. Neal. Nucleic acid resistance to viral infection. J. Bacteriol. 90:1542–1547. 1965.—Administration of nonviral nucleic acids to mice increased their resistance to a subsequent infection with influenza or encephalomyocarditis viruses. Injection of ribonucleic acid or deoxyribonucleic acid by peripheral routes did not modify susceptibility to intranasal infection. Lung tissue extracts from animals previously treated with yeast nucleic acid inhibited the growth of vaccinia and influenza viruses. The protective effect of exogenous nucleic acids persisted in mice for several days, but gradually diminished to undetectable levels. PMID:4285332

Unfolding and inactivation during thermal denaturation of an enzyme that exhibits phytase and acid phosphatase activities.

PubMed

Wang, Xiao-Yun; Meng, Fan-Guo; Zhou, Hai-Meng

2004-03-01

The thermostability of an enzyme that exhibits phytase and acid phosphatase activities was studied. Kinetics of inactivation and unfolding during thermal denaturation of the enzyme were compared. The loss of phytase activity on thermal denaturation is most suggestive of a reversible process. As for acid phosphatase activities, an interesting phenomenon was observed; there are two phases in thermal inactivation: when the temperature was between 45 and 50 degrees C, the thermal inactivation could be characterized as an irreversible inactivation which had some residual activity and when the temperature was above 55 degrees C, the thermal inactivation could be characterized as an irreversible process which had no residual activity. The microscopic rate constants for the free enzyme and substrate-enzyme complex were determined by Tsou's method [Adv. Enzymol. Relat. Areas Mol. Biol. 61 (1988) 381]. Fluorescence analyses indicate that when the enzyme was treated at temperatures below 60 degrees C for 60 min, the conformation of the enzyme had no detectable change; when the temperatures were above 60 degrees C, some fluorescence red-shift could be observed with a decrease in emission intensity. The inactivation rates (k(+0)) of free enzymes were faster than those of conformational changes during thermal denaturation at the same temperature. The rapid inactivation and slow conformational changes of phytase during thermal denaturation suggest that inactivation occurs before significant conformational changes of the enzyme, and the active site of this enzyme is situated in a relatively fragile region which makes the active site more flexible than the molecule as a whole.

Recessive Mutations in ACPT, Encoding Testicular Acid Phosphatase, Cause Hypoplastic Amelogenesis Imperfecta.

PubMed

Seymen, Figen; Kim, Youn Jung; Lee, Ye Ji; Kang, Jenny; Kim, Tak-Heun; Choi, Hwajung; Koruyucu, Mine; Kasimoglu, Yelda; Tuna, Elif Bahar; Gencay, Koray; Shin, Teo Jeon; Hyun, Hong-Keun; Kim, Young-Jae; Lee, Sang-Hoon; Lee, Zang Hee; Zhang, Hong; Hu, Jan C-C; Simmer, James P; Cho, Eui-Sic; Kim, Jung-Wook

2016-11-03

Amelogenesis imperfecta (AI) is a heterogeneous group of genetic disorders affecting tooth enamel. The affected enamel can be hypoplastic and/or hypomineralized. In this study, we identified ACPT (testicular acid phosphatase) biallelic mutations causing non-syndromic, generalized hypoplastic autosomal-recessive amelogenesis imperfecta (AI) in individuals from six apparently unrelated Turkish families. Families 1, 4, and 5 were affected by the homozygous ACPT mutation c.713C>T (p.Ser238Leu), family 2 by the homozygous ACPT mutation c.331C>T (p.Arg111Cys), family 3 by the homozygous ACPT mutation c.226C>T (p.Arg76Cys), and family 6 by the compound heterozygous ACPT mutations c.382G>C (p.Ala128Pro) and 397G>A (p.Glu133Lys). Analysis of the ACPT crystal structure suggests that these mutations damaged the activity of ACPT by altering the sizes and charges of key amino acid side chains, limiting accessibility of the catalytic core, and interfering with homodimerization. Immunohistochemical analysis confirmed localization of ACPT in secretory-stage ameloblasts. The study results provide evidence for the crucial function of ACPT during amelogenesis. Copyright © 2016 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

Immunocytochemical detection of the microsomal glucose-6-phosphatase in human brain astrocytes.

PubMed

Bell, J E; Hume, R; Busuttil, A; Burchell, A

1993-10-01

Using an antibody raised against the catalytic subunit of glucose-6-phosphatase, this enzyme was immunolocalized in many astrocytes in 20 normal human brains. Double immunofluorescence studies showed co-localization of glial fibrillary acidic protein (GFAP) with glucose-6-phosphatase in astrocytes. However, not all GFAP-positive cells were also glucose-6-phosphatase positive, indicating that some astrocytes do not contain demonstrable expression of this enzyme. Reactive astrocytes in a variety of abnormal brains were strongly glucose-6-phosphatase positive, but neoplastic astrocytes were often only weakly positive. Expression of the enzyme could not be demonstrated in radial glia, neurons or oligodendroglia. Astrocytes normally contain glycogen and the demonstration that some astrocytes also contain glucose-6-phosphatase indicates that they are competent for both glycogenolysis and gluconeogenesis, which may be critical for neuronal welfare.

Flame and acid resistant polymide fibers

NASA Technical Reports Server (NTRS)

Stringham, R. S.; Toy, M. S.

1977-01-01

Economical process improves flame resistance and resistance to acids of polyamide fibers, without modifying colors of mechanical properties. Process improves general safety of garments and other items made from polyamide fibers and makes them suitable for applications requiring exposure to oxygen-rich atmosphere or corrosive acids. Halo-olefins are added to surface of fibers by photoadditon in sealed chamber. Process could be used with films and other forms of polyamide.

The discrimination of d-tartrate positive and d-tartrate negative S. enterica subsp. enterica serovar Paratyphi B isolated in Malaysia by phenotypic and genotypic methods.

PubMed

Ahmad, Norazah; Hoon, Shirley Tang Gee; Ghani, Mohamed Kamel Abd; Tee, Koh Yin

2012-06-01

Serotyping is not sufficient to differentiate between Salmonella species that cause paratyphoid fever from the strains that cause milder gastroenteritis as these organisms share the same serotype Salmonella Paratyphi B (S. Paratyphi B). Strains causing paratyphoid fever do not ferment d-tartrate and this key feature was used in this study to determine the prevalence of these strains among the collection of S. Paratyphi B strains isolated from patients in Malaysia. A total of 105 isolates of S. Paratyphi B were discriminated into d-tartrate positive (dT+) and d-tartrate negative (dT) variants by two lead acetate test protocols and multiplex PCR. The lead acetate test protocol 1 differed from protocol 2 by a lower inoculum size and different incubation conditions while the multiplex PCR utilized 2 sets of primers targeting the ATG start codon of the gene STM3356. Lead acetate protocol 1 discriminated 97.1% of the isolates as S. Paratyphi B dT+ and 2.9% as dT while test protocol 2 discriminated all the isolates as S. Paratyphi B dT+. The multiplex PCR test identified all 105 isolates as S. Paratyphi B dT+ strains. The concordance of the lead acetate test relative to that of multiplex PCR was 97.7% and 100% for protocol 1 and 2 respectively. This study showed that S. Paratyphi B dT+ is a common causative agent of gastroenteritis in Malaysia while paratyphoid fever appears to be relatively uncommon. Multiplex PCR was shown to be a simpler, more rapid and reliable method to discriminate S. Paratyphi B than the phenotypic lead acetate test.

Protein phosphatase 2A in stretch-induced endothelial cell proliferation

NASA Technical Reports Server (NTRS)

Murata, K.; Mills, I.; Sumpio, B. E.

1996-01-01

We previously proposed that activation of protein kinase C is a key mechanism for control of cell growth enhanced by cyclic strain [Rosales and Sumpio (1992): Surgery 112:459-466]. Here we examined protein phosphatase 1 and 2A activity in bovine aortic endothelial cells exposed to cyclic stain. Protein phosphatase 2A activity in the cytosol was decreased by 36.1% in response to cyclic strain for 60 min, whereas the activity in the membrane did not change. Treatment with low concentration (0.1 nM) of okadaic acid enhanced proliferation of both static and stretched endothelial cells in 10% fetal bovine serum. These data suggest that protein phosphatase 2A acts as a growth suppressor and cyclic strain may enhance cellular proliferation by inhibiting protein phosphatase 2A as well as stimulating protein kinase C.

Reduced bone breakage and increased bone strength in free range laying hens fed omega-3 polyunsaturated fatty acid supplemented diets.

PubMed

Tarlton, John F; Wilkins, Lindsay J; Toscano, Michael J; Avery, Nick C; Knott, Lynda

2013-02-01

The omega-3 and omega-6 polyunsaturated fatty acids (PUFAs) are the immediate precursors to a number of important mediators of immunity, inflammation and bone function, with products of omega-6 generally thought to promote inflammation and favour bone resorption. Western diets generally provide a 10 to 20-fold deficit in omega-3 PUFAs compared with omega-6, and this is thought to have contributed to the marked rise in incidence of disorders of modern human societies, such as heart disease, colitis and perhaps osteoporosis. Many of our food production animals, fed on grains rich in omega-6, are also exposed to a dietary deficit in omega-3, with perhaps similar health consequences. Bone fragility due to osteoporotic changes in laying hens is a major economic and welfare problem, with our recent estimates of breakage rates indicating up to 95% of free range hens suffer breaks during lay. Free range hens housed in full scale commercial systems were provided diets supplemented with omega-3 alpha linolenic acid, and the skeletal benefits were investigated by comparison to standard diets rich in omega-6. There was a significant 40-60% reduction in keel bone breakage rate, and a corresponding reduction in breakage severity in the omega-3 supplemented hens. There was significantly greater bone density and bone mineral content, alongside increases in total bone and trabecular volumes. The mechanical properties of the omega-3 supplemented hens were improved, with strength, energy to break and stiffness demonstrating significant increases. Alkaline phosphatase (an osteoblast marker) and tartrate-resistant acid phosphatase (an osteoclast marker) both showed significant increases with the omega-3 diets, indicating enhanced bone turnover. This was corroborated by the significantly lower levels of the mature collagen crosslinks, hydroxylysyl pyridinoline, lysyl pyridinoline and histidinohydroxy-lysinonorleucine, with a corresponding significant shift in the mature

A New Rapid and Sensitive Stability-Indicating UPLC Assay Method for Tolterodine Tartrate: Application in Pharmaceuticals, Human Plasma and Urine Samples

PubMed Central

Yanamandra, Ramesh; Vadla, Chandra Sekhar; Puppala, Umamaheshwar; Patro, Balaram; Murthy, Yellajyosula. L. N.; Ramaiah, Parimi Atchuta

2012-01-01

A new rapid, simple, sensitive, selective and accurate reversed-phase stability-indicating Ultra Performance Liquid Chromatography (RP-UPLC) technique was developed for the assay of Tolterodine Tartrate in pharmaceutical dosage form, human plasma and urine samples. The developed UPLC method is superior in technology to conventional HPLC with respect to speed, solvent consumption, resolution and cost of analysis. Chromatographic run time was 6 min in reversed-phase mode and ultraviolet detection was carried out at 220 nm for quantification. Efficient separation was achieved for all the degradants of Tolterodine Tartrate on BEH C18 sub-2-μm Acquity UPLC column using Trifluoroacetic acid and acetonitrile as organic solvent in a linear gradient program. The active pharmaceutical ingredient was extracted from tablet dosage form using a mixture of acetonitrile and water as diluent. The calibration graphs were linear and the method showed excellent recoveries for bulk and tablet dosage form. The test solution was found to be stable for 40 days when stored in the refrigerator between 2 and 8 °C. The developed UPLC method was validated and meets the requirements delineated by the International Conference on Harmonization (ICH) guidelines with respect to linearity, accuracy, precision, specificity and robustness. The intra-day and inter-day variation was found be less than 1%. The method was reproducible and selective for the estimation of Tolterodine Tartrate. Because the method could effectively separate the drug from its degradation products, it can be employed as a stability-indicating one. PMID:22396907

Comparison of methods for acid quantification: impact of resist components on acid-generating efficiency

NASA Astrophysics Data System (ADS)

Cameron, James F.; Fradkin, Leslie; Moore, Kathryn; Pohlers, Gerd

2000-06-01

Chemically amplified deep UV (CA-DUV) positive resists are the enabling materials for manufacture of devices at and below 0.18 micrometer design rules in the semiconductor industry. CA-DUV resists are typically based on a combination of an acid labile polymer and a photoacid generator (PAG). Upon UV exposure, a catalytic amount of a strong Bronsted acid is released and is subsequently used in a post-exposure bake step to deprotect the acid labile polymer. Deprotection transforms the acid labile polymer into a base soluble polymer and ultimately enables positive tone image development in dilute aqueous base. As CA-DUV resist systems continue to mature and are used in increasingly demanding situations, it is critical to develop a fundamental understanding of how robust these materials are. One of the most important factors to quantify is how much acid is photogenerated in these systems at key exposure doses. For the purpose of quantifying photoacid generation several methods have been devised. These include spectrophotometric methods, ion conductivity methods and most recently an acid-base type titration similar to the standard addition method. This paper compares many of these techniques. First, comparisons between the most commonly used acid sensitive dye, tetrabromophenol blue sodium salt (TBPB) and a less common acid sensitive dye, Rhodamine B base (RB) are made in several resist systems. Second, the novel acid-base type titration based on the standard addition method is compared to the spectrophotometric titration method. During these studies, the make up of the resist system is probed as follows: the photoacid generator and resist additives are varied to understand the impact of each of these resist components on the acid generation process.

The possible involvement of protein phosphatase 1 in thrombin-induced Ca2+ influx of human platelets.

PubMed

Murata, K; Sakon, M; Kambayashi, J; Yukawa, M; Yano, Y; Fujitani, K; Kawasaki, T; Shiba, E; Mori, T

1993-04-01

Protein phosphatase 1 is considered to be involved in thrombin-induced platelet activation (Murata et al., Biochem Int 26:327-334, 1992). To clarify the mechanism, we examined the effects of protein phosphatase 1 and 2A inhibitors (calyculin A, tautomycin, okadaic acid) on Ca2+ influx. In the presence of 1 mM Ca2+, thrombin- (0.1 U/ml) induced platelet aggregation and ATP release were inhibited by calyculin A, while this inhibitory effect was abolished in the absence of Ca2+ (EGTA 1 mM). Furthermore, thrombin-induced Mn2+ influx but not intracellular Ca2+ mobilization was inhibited by calyculin A in a dose-related manner. Calyculin A also blocked the ongoing Ca2+ influx when added 3 min after thrombin stimulation. Similar inhibitory effects were observed with okadaic acid and tautomycin in the same potency sequence as the reported one for protein phosphatase 1 (calyculin A > tautomycin > okadaic acid). These results suggest that the anti-platelet effects of phosphatase inhibitors are due to the inhibition of Ca2+ influx and that protein phosphatase 1 plays a key role in the regulation of receptor operated Ca2+ channel of human platelets.

Synthesis of carbohydrates in a continuous flow reactor by immobilized phosphatase and aldolase.

PubMed

Babich, Lara; Hartog, Aloysius F; van Hemert, Lieke J C; Rutjes, Floris P J T; Wever, Ron

2012-12-01

Herein, we report a new flow process with immobilized enzymes to synthesize complex chiral carbohydrate analogues from achiral inexpensive building blocks in a three-step cascade reaction. The first reactor contained immobilized acid phosphatase, which phosphorylated dihydroxyacetone to dihydroxyacetone phosphate using pyrophosphate as the phosphate donor. The second flow reactor contained fructose-1,6-diphosphate aldolase (RAMA, rabbit muscle aldolase) or rhamnulose-1-phosphate aldolase (RhuA from Thermotoga maritima) and acid phosphatase. The immobilized aldolases coupled the formed dihydroxyacetone phosphate to aldehydes, resulting in phosphorylated carbohydrates. A final reactor containing acid phosphatase that dephosphorylated the phosphorylated product yielded the final product. Different aldehydes were used to synthesize carbohydrates on a gram scale. To demonstrate the feasibility of the flow systems, we synthesized 0.6 g of the D-fagomine precursor. By using immobilized aldolase RhuA we were also able to obtain other stereoisomers of the D-fagomine precursor. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Diversity and Gene Expression of Phosphatase Genes Provide Insight into Soil Phosphorus Dynamics in a New Zealand Managed Grassland

NASA Astrophysics Data System (ADS)

Dunfield, K. E.; Gaiero, J. R.; Condron, L.

2017-12-01

Healthy and diverse communities of soil organisms influence key soil ecosystem services such as carbon sequestration, water quality protection, climate regulation and nutrient cycling. Microbially driven mineralization of organic phosphorus is an important contributor to plant available inorganic orthophosphates. In acidic soils, microbes produce non-specific acid phosphatases (NSAPs) which act on common forms of organic phosphorus (P). Our current understanding of P turnover in soils has been limited by lack of research tools capable of targeting these genes. Thus, we developed a set of oligonucleotide PCR primers that targeted bacteria with the genetic potential for acid phosphatase production. A long term randomized-block pasture trial was sampled following 22 years of continued aerial biomass removal and retention. Primers were used to target genes encoding alkaline phosphatase (phoD) and the three classes (CAAP, CBAP, CCAP) of non-specific acid phosphatases. PCR amplicons targeting total genes and gene transcripts were sequenced using Illumina MiSeq to understand the diversity of the bacterial phosphatase producing communities. In general, the majority of operational taxonomic units (OTUs) were shared across both treatments and across metagenomes and transcriptomes. However, analysis of DNA OTUs revealed significantly different communities driven by treatment differences (P < 0.05). Transcript expression was highest in the removed biomass treatment which corresponded the reduced Olsen P levels (15 vs. 36 mg kg-1 in retained treatment). Acid phosphatase activity was measured in all samples, and found to be highest in the biomass retained treatment (16.8 vs. 11.4 µmol g-1 dry soil h-1), likely elevated due to plant-derived enzymes; however, was still correlated to bacterial gene abundances. Overall, the phosphatase producing microbial communities responded to the effect of consistent P limitation as expected, through alteration in the composition of the

The effects of a 6-month resistance training and dried plum consumption intervention on strength, body composition, blood markers of bone turnover, and inflammation in breast cancer survivors.

PubMed

Simonavice, Emily; Liu, Pei-Yang; Ilich, Jasminka Z; Kim, Jeong-Su; Arjmandi, Bahram; Panton, Lynn B

2014-06-01

The purpose of this study was to examine the effects of resistance training (RT) and dried plum (DP) consumption on strength, body composition, blood markers of bone, and inflammation in breast cancer survivors (BCS). Twenty-three BCS (RT, n = 12; RT+DP, n = 11), aged 64 ± 7 years, were evaluated at baseline and after 6 months of intervention on the following: muscular strength (chest press and leg extension) via 1-repetition maximums (1RMs); body composition, specifically bone mineral density (BMD) by dual energy X-ray absorptiometry; biochemical markers of bone turnover (bone-specific alkaline phosphatase (BAP), tartrate resistant acid phosphatase (TRAP-5b)); and inflammation (C-reactive protein (CRP)). Target RT prescription was 2 days/week of 10 exercises, including 2 sets of 8-12 repetitions at ∼60%-80% of 1RM. RT+DP also consumed 90 g of DP daily. There were no baseline differences between groups or any group-by-time interactions for any of the variables. BCS increased upper (p < 0.05) (RT: 64 ± 14 to 80 ± 17 kg; RT+DP: 72 ± 23 to 91 ± 20 kg) and lower (p < 0.05) (RT: 69 ± 20 to 87 ± 28 kg; RT+DP: 78 ± 19 to 100 ± 21 kg) body strength. Body composition and BMD improvements were not observed. TRAP-5b decreased in the RT group (p < 0.05) (4.55 ± 1.57 to 4.04 ± 1.63 U/L) and the RT+DP group (p = 0.07) (5.10 ± 2.75 to 4.27 ± 2.03 U/L). Changes in BAP and CRP were not observed. RT was effective for improving biochemical markers of bone turnover and muscular strength in BCS. A longer and higher intensity intervention may be needed to reveal the true effects of RT and DP on body composition and biochemical markers of inflammation.

The cell wall-targeted purple acid phosphatase AtPAP25 is critical for acclimation of Arabidopsis thaliana to nutritional phosphorus deprivation.

PubMed

Del Vecchio, Hernan A; Ying, Sheng; Park, Joonho; Knowles, Vicki L; Kanno, Satomi; Tanoi, Keitaro; She, Yi-Min; Plaxton, William C

2014-11-01

Plant purple acid phosphatases (PAPs) belong to a relatively large gene family whose individual functions are poorly understood. Three PAP isozymes that are up-regulated in the cell walls of phosphate (Pi)-starved (-Pi) Arabidopsis thaliana suspension cells were purified and identified by MS as AtPAP12 (At2g27190), AtPAP25 (At4g36350) and AtPAP26 (At5g34850). AtPAP12 and AtPAP26 were previously isolated from the culture medium of -Pi cell cultures, and shown to be secreted by roots of Arabidopsis seedlings to facilitate Pi scavenging from soil-localized organophosphates. AtPAP25 exists as a 55 kDa monomer containing complex NX(S/T) glycosylation motifs at Asn172, Asn367 and Asn424. Transcript profiling and immunoblotting with anti-AtPAP25 immune serum indicated that AtPAP25 is exclusively synthesized under -Pi conditions. Coupled with potent mixed-type inhibition of AtPAP25 by Pi (I50 = 50 μm), this indicates a tight feedback control by Pi that prevents AtPAP25 from being synthesized or functioning as a phosphatase except when Pi levels are quite low. Promoter-GUS reporter assays revealed AtPAP25 expression in shoot vascular tissue of -Pi plants. Development of an atpap25 T-DNA insertion mutant was arrested during cultivation on soil lacking soluble Pi, but rescued upon Pi fertilization or complementation with AtPAP25. Transcript profiling by quantitative RT-PCR indicated that Pi starvation signaling was attenuated in the atpap25 mutant. AtPAP25 exhibited near-optimal phosphatase activity with several phosphoproteins and phosphoamino acids as substrates. We hypothesize that AtPAP25 plays a key signaling role during Pi deprivation by functioning as a phosphoprotein phosphatase rather than as a non-specific scavenger of Pi from extracellular P-monoesters. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

Fates of Acid-Resistant and Non-Acid-Resistant Shiga Toxin-Producing Escherichia coli Strains in Ruminant Digestive Contents in the Absence and Presence of Probiotics ▿

PubMed Central

Chaucheyras-Durand, Frédérique; Faqir, Fahima; Ameilbonne, Aurélie; Rozand, Christine; Martin, Christine

2010-01-01

Healthy ruminants are the main reservoir of Shiga toxin-producing Escherichia coli (STEC). During their transit through the ruminant gastrointestinal tract, STEC encounters a number of acidic environments. As all STEC strains are not equally resistant to acidic conditions, the purpose of this study was to investigate whether acid resistance confers an ecological advantage to STEC strains in ruminant digestive contents and whether acid resistance mechanisms are induced in the rumen compartment. We found that acid-resistant STEC survived at higher rates during prolonged incubation in rumen fluid than acid-sensitive STEC and that they resisted the highly acidic conditions of the abomasum fluid, whereas acid-sensitive strains were killed. However, transit through the rumen contents allowed acid-sensitive strains to survive in the abomasum fluid at levels similar to those of acid-resistant STEC. The acid resistance status of the strains had little influence on STEC growth in jejunal and cecal contents. Supplementation with the probiotic Saccharomyces cerevisiae CNCM I-1077 or Lactobacillus acidophilus BT-1386 led to killing of all of the strains tested during prolonged incubation in the rumen contents, but it did not have any influence in the other digestive compartments. In addition, S. cerevisiae did not limit the induction of acid resistance in the rumen fluid. Our results indicate that the rumen compartment could be a relevant target for intervention strategies that could both limit STEC survival and eliminate induction of acid resistance mechanisms in order to decrease the number of viable STEC cells reaching the hindgut and thus STEC shedding and food contamination. PMID:19948865

Relationship between bone turnover markers and the heel stiffness index measured by quantitative ultrasound in middle-aged and elderly Japanese men

PubMed Central

Nishimura, Takayuki; Arima, Kazuhiko; Abe, Yasuyo; Kanagae, Mitsuo; Mizukami, Satoshi; Okabe, Takuhiro; Tomita, Yoshihito; Goto, Hisashi; Horiguchi, Itsuko; Aoyagi, Kiyoshi

2018-01-01

Abstract The aim of the present study was to investigate the age-related patterns and the relationships between serum levels of tartrate-resistant acid phosphatase-5b (TRACP-5b) or bone-specific alkaline phosphatase (BAP), and the heel stiffness index measured by quantitative ultrasound (QUS) in 429 Japanese men, with special emphasis on 2 age groups (40–59 years and 60 years or over). The heel stiffness index (bone mass) was measured by QUS. Serum samples were collected, and TRACP-5b and BAP levels were measured. The stiffness index was significantly decreased with age. Log (TRACP-5b) was significantly increased with age, but Log (BAP) was stable. Generalized linear models showed that higher levels of Log (TRACP-5b) and Log (BAP) were correlated with a lower stiffness index after adjusting for covariates in men aged 60 years or over, but not in men aged 40 to 59 years. In conclusion, higher rates of bone turnover markers were associated with a lower stiffness index only in elderly men. These results may indicate a different mechanism of low bone mass among different age groups of men. PMID:29465590

The acid-base resistant zone in three dentin bonding systems.

PubMed

Inoue, Go; Nikaido, Toru; Foxton, Richard M; Tagami, Junji

2009-11-01

An acid-base resistant zone has been found to exist after acid-base challenge adjacent to the hybrid layer using SEM. The aim of this study was to examine the acid-base resistant zone using three different bonding systems. Dentin disks were applied with three different bonding systems, and then a resin composite was light-cured to make dentin disk sandwiches. After acid-base challenge, the polished surfaces were observed using SEM. For both one- and two-step self-etching primer systems, an acid-base resistant zone was clearly observed adjacent to the hybrid layer - but with differing appearances. For the wet bonding system, the presence of an acid-base resistant zone was unclear. This was because the self-etching primer systems etched the dentin surface mildly, such that the remaining mineral phase of dentin and the bonding agent yielded clear acid-base resistant zones. In conclusion, the acid-base resistant zone was clearly observed when self-etching primer systems were used, but not so for the wet bonding system.

Prophylactic treatment with alkaline phosphatase in cardiac surgery induces endogenous alkaline phosphatase release.

PubMed

Kats, Suzanne; Brands, Ruud; Hamad, Mohamed A Soliman; Seinen, Willem; Scharnhorst, Volkher; Wulkan, Raymond W; Schönberger, Jacques P; Oeveren, Wim van

2012-02-01

Laboratory and clinical data have implicated endotoxin as an important factor in the inflammatory response to cardiopulmonary bypass. We assessed the effects of the administration of bovine intestinal alkaline phosphatase (bIAP), an endotoxin detoxifier, on alkaline phosphatase levels in patients undergoing coronary artery bypass grafting. A total of 63 patients undergoing coronary artery bypass grafting were enrolled and prospectively randomized. Bovine intestinal alkaline phosphatase (n=32) or placebo (n=31) was administered as an intravenous bolus followed by continuous infusion for 36 hours. The primary endpoint was to evaluate alkaline phosphatase levels in both groups and to find out if administration of bIAP to patients undergoing CABG would lead to endogenous alkaline phosphatase release. No significant adverse effects were identified in either group. In all the 32 patients of the bIAP-treated group, we found an initial rise of plasma alkaline phosphatase levels due to bolus administration (464.27±176.17 IU/L). A significant increase of plasma alkaline phosphatase at 4-6 hours postoperatively was observed (354.97±95.00 IU/L) as well. Using LHA inhibition, it was shown that this second peak was caused by the generation of tissue non specific alkaline phosphatase (TNSALP-type alkaline phosphatase). Intravenous bolus administration plus 8 hours continuous infusion of alkaline phosphatase in patients undergoing coronary artery bypass grafting with cardiopulmonary bypass results in endogenous alkaline phosphatase release. This endogenous alkaline phosphatase may play a role in the immune defense system.

Cancerous inhibitor of protein phosphatase 2A determines bortezomib-induced apoptosis in leukemia cells

PubMed Central

Liu, Chun-Yu; Shiau, Chung-Wai; Kuo, Hsin-Yu; Huang, Hsiang-Po; Chen, Ming-Huang; Tzeng, Cheng-Hwai; Chen, Kuen-Feng

2013-01-01

The multiple cellular targets affected by proteasome inhibition implicate a potential role for bortezomib, a first-in-class proteasome inhibitor, in enhancing antitumor activities in hematologic malignancies. Here, we examined the antitumor activity and drug targets of bortezomib in leukemia cells. Human leukemia cell lines were used for in vitro studies. Drug efficacy was evaluated by apoptosis assays and associated molecular events assessed by Western Blot. Gene silencing was performed by small interference RNA. Drug was tested in vivo in xenograft models of human leukemia cell lines and in primary leukemia cells. Clinical samples were assessed by immunohistochemical staining. Bortezomib differentially induced apoptosis in leukemia cells that was independent of its proteasome inhibition. Cancerous inhibitor of protein phosphatase 2A, a cellular inhibitor of protein phosphatase 2A, mediated the apoptotic effect of bortezomib. Bortezomib increased protein phosphatase 2A activity in sensitive leukemia cells (HL-60 and KG-1), but not in resistant cells (MOLT-3 and K562). Bortezomib’s downregulation of cancerous inhibitor of protein phosphatase 2A and phospho-Akt correlated with its drug sensitivity. Furthermore, cancerous inhibitor of protein phosphatase 2A negatively regulated protein phosphatase 2A activity. Ectopic expression of CIP2A up-regulated phospho-Akt and protected HL-60 cells from bortezomib-induced apoptosis, whereas silencing CIP2A overcame the resistance to bortezomib-induced apoptosis in MOLT3 and K562 cells. Importantly, bortezomib exerted in vivo antitumor activity in HL-60 xenografted tumors and induced cell death in some primary leukemic cells. Cancerous inhibitor of protein phosphatase 2A was expressed in leukemic blasts from bone marrow samples. Cancerous inhibitor of protein phosphatase 2A plays a major role in mediating bortezomib-induced apoptosis in leukemia cells. PMID:22983581

Alkaline phosphatase binds tenaciously to titanium; implications for biological surface evaluation following bone implant retrieval.

PubMed

Mansell, J P; Shiel, A I; Harwood, C; Stephens, D

2017-07-01

Enhancing the performance and longevity of titanium (Ti) implants continues to be a significant developmental theme in contemporary biomaterials design. Our specific focus pertains to the surface functionalisation of Ti using the bioactive lipid, lysophosphatidic acid (LPA) and certain phosphatase-resistant analogues of LPA. Coating survivorship to a plethora of testing regimens is required to align with due regulatory process before novel biomaterials can enter clinical trials. One of the key acceptance criteria is coating retention to the physical stresses experienced during implantation. In assessing coating stability to insertion into porcine bone we found that a subsequent in vitro assessment to confirm coating persistence was masked by abundant alkaline phosphatase (ALP) contamination adsorbed to the metal surface. Herein we report that ALP can bind to Ti in a matter of minutes by simply immersing Ti samples in aqueous solutions of the enzyme. We strongly discourage the in vitro monitoring of osteoblast and stromal cell ALP expression when assessing bioactive coating survivorship following Ti implant retrieval form native bone tissue. Copyright © 2017 Elsevier B.V. All rights reserved.

A c-fms tyrosine kinase inhibitor, Ki20227, suppresses osteoclast differentiation and osteolytic bone destruction in a bone metastasis model.

PubMed

Ohno, Hiroaki; Kubo, Kazuo; Murooka, Hideko; Kobayashi, Yoshiko; Nishitoba, Tsuyoshi; Shibuya, Masabumi; Yoneda, Toshiyuki; Isoe, Toshiyuki

2006-11-01

In bone metastatic lesions, osteoclasts play a key role in the development of osteolysis. Previous studies have shown that macrophage colony-stimulating factor (M-CSF) is important for the differentiation of osteoclasts. In this study, we investigated whether an inhibitor of M-CSF receptor (c-Fms) suppresses osteoclast-dependent osteolysis in bone metastatic lesions. We developed small molecule inhibitors against ligand-dependent phosphorylation of c-Fms and examined the effects of these compounds on osteolytic bone destruction in a bone metastasis model. We discovered a novel quinoline-urea derivative, Ki20227 (N-{4-[(6,7-dimethoxy-4-quinolyl)oxy]-2-methoxyphenyl}-N'-[1-(1,3-thiazole-2-yl)ethyl]urea), which is a c-Fms tyrosine kinase inhibitor. The IC(50)s of Ki20227 to inhibit c-Fms, vascular endothelial growth factor receptor-2 (KDR), stem cell factor receptor (c-Kit), and platelet-derived growth factor receptor beta were found to be 2, 12, 451, and 217 nmol/L, respectively. Ki20227 did not inhibit other kinases tested, such as fms-like tyrosine kinase-3, epidermal growth factor receptor, or c-Src (c-src proto-oncogene product). Ki20227 was also found to inhibit the M-CSF-dependent growth of M-NFS-60 cells but not the M-CSF-independent growth of A375 human melanoma cells in vitro. Furthermore, in an osteoclast-like cell formation assay using mouse bone marrow cells, Ki20227 inhibited the development of tartrate-resistant acid phosphatase-positive osteoclast-like cells in a dose-dependent manner. In in vivo studies, oral administration of Ki20227 suppressed osteoclast-like cell accumulation and bone resorption induced by metastatic tumor cells in nude rats following intracardiac injection of A375 cells. Moreover, Ki20227 decreased the number of tartrate-resistant acid phosphatase-positive osteoclast-like cells on bone surfaces in ovariectomized (ovx) rats. These findings suggest that Ki20227 inhibits osteolytic bone destruction through the suppression of M

Ectopic mineralization of cartilage and collagen-rich tendons and ligaments in Enpp1asj-2J mice.

PubMed

Zhang, Jieyu; Dyment, Nathaniel A; Rowe, David W; Siu, Sarah Y; Sundberg, John P; Uitto, Jouni; Li, Qiaoli

2016-03-15

Generalized arterial calcification of infancy (GACI), an autosomal recessive disorder caused by mutations in the ENPP1 gene, manifests with extensive mineralization of the cardiovascular system. A spontaneous asj-2J mutant mouse has been characterized as a model for GACI. Previous studies focused on phenotypic characterization of skin and vascular tissues. This study further examined the ectopic mineralization phenotype of cartilage, collagen-rich tendons and ligaments in this mouse model. The mice were placed on either control diet or the "acceleration diet" for up to 12 weeks of age. Soft connective tissues, such as ear (elastic cartilage) and trachea (hyaline cartilage), were processed for standard histology. Assessment of ectopic mineralization in articular cartilage and fibrocartilage as well as tendons and ligaments which are attached to long bones were performed using a novel cryo-histological method without decalcification. These analyses demonstrated ectopic mineralization in cartilages as well as tendons and ligaments in the homozygous asj-2J mice at 12 weeks of age, with the presence of immature osteophytes displaying alkaline phosphatase and tartrate-resistant acid phosphatase activities as early as at 6 weeks of age. Alkaline phosphatase activity was significantly increased in asj-2J mouse serum as compared to wild type mice, indicating increased bone formation rate in these mice. Together, these data highlight the key role of ENPP1 in regulating calcification of both soft and skeletal tissues.

Surface phosphatase in Rhinocladiella aquaspersa: biochemical properties and its involvement with adhesion.

PubMed

Kneipp, Lucimar F; Magalhães, Andressa S; Abi-Chacra, Erika A; Souza, Lucieri O P; Alviano, Celuta S; Santos, André L S; Meyer-Fernandes, José R

2012-08-01

Rhinocladiella aquaspersa is an etiologic agent of chromoblastomycosis, a subcutaneous chronic infectious disease. In the present work, we found that the three morphological forms of this fungus (conidia, mycelia and sclerotic bodies) expressed different levels of ecto-phosphatase activity. Our results demonstrated that surface conidial enzyme is an acid phosphatase, inhibited by sodium salts of molybdate, orthovanadate and fluoride and that the inhibition caused by orthovanadate and molybdate was irreversible. The conidial ecto-phosphatase efficiently released phosphate groups from different phosphorylated substrates, causing a higher rate of phosphate removal when p-nitrophenylphosphate was used as substrate. This ecto-enzyme of R. aquaspersa is modulated by Co(2 +) ions and inorganic phosphate (Pi). Accordingly, removal of Pi from the culture medium resulted in a marked (121-fold) increase of ecto-phosphatase activity. Surface phosphatase activity is apparently involved in fungal adhesive properties, since the attachment of R. aquaspersa to epithelial cells was reversed by the pre-treatment of the conidia with orthovanadate, molybdate and anti-phosphatase antibody. Corroborating this finding, conidia with greater ecto-phosphatase activity (grown in Pi-depleted medium) showed higher adherence to epithelial cells than fungi cultivated in the presence of Pi.

Increasing efficacy and reducing systemic absorption of brimonidine tartrate ophthalmic gels in rabbits.

PubMed

Pang, Xiaochen; Li, Jiawei; Pi, Jiaxin; Qi, Dongli; Guo, Pan; Li, Nan; Wu, Yumei; Liu, Zhidong

2018-03-01

Systemic absorption of ocularly administered Brimonidine Tartrate has been reported to give rise to several side-effects. Hence, it has become crucial to develop a delivery system that could increase efficacy and reduce systemic absorption. Therefore, the present work aims to develop Brimonidine Tartrate gels with different concentrations (0.05%, 0.1%, and 0.2% w/v, respectively) using Carbopol 974 P and HPMC E4M, and compare the therapeutic efficacy and systemic absorption with that of eye drop (0.2%, w/v) by UPLC-MS/MS. The result of histological analysis did not show any morphological or structural changes after the administration of formulations. In vitro residence time studies demonstrated that the gels exhibited a better precorneal residence time as compared with the eye drop. The gels with lower concentrations of the drug (0.05% and 0.1%, w/v) could significantly decrease intraocular pressure (IOP) in both normal and water-loaded rabbits as compared to the eye drop. Finally, the values of the ratio of AUC (0→∞) in comparison to eye drop showed the gels with lower concentrations of Brimonidine Tartrate could decrease the systemic absorption. From the result, it can be concluded the 0.1% ophthalmic gel has a potential to improve therapeutic efficacy and reduce the potential toxicity caused by systemic absorption.

Cdk1 and okadaic acid-sensitive phosphatases control assembly of nuclear pore complexes in Drosophila embryos.

PubMed

Onischenko, Evgeny A; Gubanova, Natalia V; Kiseleva, Elena V; Hallberg, Einar

2005-11-01

Disassembly and reassembly of the nuclear pore complexes (NPCs) is one of the major events during open mitosis in higher eukaryotes. However, how this process is controlled by the mitotic machinery is not clear. To investigate this we developed a novel in vivo model system based on syncytial Drosophila embryos. We microinjected different mitotic effectors into the embryonic cytoplasm and monitored the dynamics of disassembly/reassembly of NPCs in live embryos using fluorescently labeled wheat germ agglutinin (WGA) or in fixed embryos using electron microscopy and immunostaining techniques. We found that in live embryos Cdk1 activity was necessary and sufficient to induce disassembly of NPCs as well as their cytoplasmic mimics: annulate lamellae pore complexes (ALPCs). Cdk1 activity was also required for keeping NPCs and ALPCs disassembled during mitosis. In agreement recombinant Cdk1/cyclin B was able to induce phosphorylation and dissociation of nucleoporins from the NPCs in vitro. Conversely, reassembly of NPCs and ALPCs was dependent on the activity of protein phosphatases, sensitive to okadaic acid (OA). Our findings suggest a model where mitotic disassembly/reassembly of the NPCs is regulated by a dynamic equilibrium of Cdk1 and OA-sensitive phosphatase activities and provide evidence that mitotic phosphorylation mediates disassembly of the NPC.

Phosphatase activity and culture conditions of the yeast Candida mycoderma sp. and analysis of organic phosphorus hydrolysis ability.

PubMed

Yan, Mang; Yu, Liufang; Zhang, Liang; Guo, Yuexia; Dai, Kewei; Chen, Yuru

2014-11-01

Orthophosphate is an essential but limiting macronutrient for plant growth. About 67% cropland in China lacks sufficient phosphorus, especially that with red soil. Extensive soil phosphorus reserves exist in the form of organic phosphorus, which is unavailable for root uptake unless hydrolyzed by secretory acid phosphatases. Thus, many microorganisms with the ability to produce phosphatase have been exploited. In this work, the activity of an extracellular acid phosphatase and yeast biomass from Candida mycoderma was measured under different culture conditions, such as pH, temperature, and carbon source. A maximal phosphatase activity of 8.47×10(5)±0.11×10(5)U/g was achieved by C. Mycoderma in 36 hr under the optimal conditions. The extracellular acid phosphatase has high activity over a wide pH tolerance range from 2.5 to 5.0 (optimum pH3.5). The effects of different phosphorus compounds on the acid phosphatase production were also studied. The presence of phytin, lecithin or calcium phosphate reduced the phosphatase activity and biomass yield significantly. In addition, the pH of the culture medium was reduced significantly by lecithin. The efficiency of the strain in releasing orthophosphate from organic phosphorus was studied in red soil (used in planting trees) and rice soil (originating as red soil). The available phosphorus content was increased by 230% after inoculating 20 days in rice soil and decreased by 50% after inoculating 10 days in red soil. This work indicates that the yeast strain C. mycoderma has potential application for enhancing phosphorus utilization in plants that grow in rice soil. Copyright © 2014. Published by Elsevier B.V.

[Inhibition of osthole for resorption of rats femur tissue in vitro].

PubMed

Zhou, Jian; Ren, Xue-mei; Ma, Xiao-ni; Gao, Yu-hai; Yan, Li-juan; Shi, Wen-gui; Chen, Ke-ming

2015-09-01

To investigate osthole effect on femoral tissue resorption activity of rat in vitro. Six SD rats weighted (80 ± 5) g were used to isolate and culture femoral tissue (diaphyses and metaphysis) in vitro. The cultured tissue were devided into control group, estradiol group and osthole group. The femoral tissue was treated with final concentration of 1 x 10(-5) mol/L osthole and 1 x 10(-8) mol/L estradiol culture in vitro at 48 hours after cultured. Tartrate-resistant acid phosphatase (StrACP) activity, glucose and Lactic acid content, StrACP, MCSF (Macrophage colony stimulating factor) and CTSK (Cathepsin K) mRNA was detected by Real-Time RT-PCR were detected. Concetration of Alkaline phosphatase activity were 2226 and 2498 in 1 x 10(-5) mol/L osthole and 1 x 10(-8) mol/L estradiol respectively. As compared with control group, the activity of StrACP of 1 x 10(-5) mol/L osthole and 1 x 10(-8) mol/L estradiol were inhibited at 6, 9, 12 days (P < 0.05); under treatment of in l x 10(-5) mol/L osthole, the content of Lactic acid were increased and the content of glucose were decreased at 3, 6, 9 days (P < 0.05); StrACP, MCSF and CTSK mRNA expression level were inhibited at 6, 9 days (P < 0.05). Osthole can inhibit bone resorption and raise the level of nutrition metabolism of femurs tissue.

Changes in Sugars, Enzymic Activities and Acid Phosphatase Isoenzyme Profiles of Bananas Ripened in Air or Stored in 2.5% O2 with and without Ethylene 1

PubMed Central

Kanellis, Angelos K.; Solomos, Theophanes; Mattoo, Autar K.

1989-01-01

This study investigates the effect of 2.5% O2, both alone and in combination with ethylene, on respiration, sugar accumulation and activities of pectin methylesterase and acid phosphatase during ripening of bananas (Musa paradisiaca sapientum). In addition, the changes in the phosphatase isoenzyme profiles are also analyzed. Low oxygen diminished respiration and slowed down the accumulation of sugars and development of the yellow color. Furthermore, low O2 prevented the rise in acid phosphatase activities and this suppression was not reversed by the inclusion of 100 microliters per liter ethylene in 2.5% O2 atmosphere. Gel electrophoresis of both the soluble and particulate cell-free fractions under nondenaturing conditions revealed the presence of 8 and 9 isoenzymes in the soluble and particulate fractions, respectively. Low O2 suppressed the appearance of all isoenzymes, and the addition of 500 microliters per liter ethylene to the low oxygen atmosphere did not reverse this effect. Similarly, the decline in pectin methylesterase that was observed in air-ripened fruits was prevented by 2.5% O2 alone and in combination with 500 microliters per liter ethylene. Images Figure 5 Figure 6 Figure 7 PMID:16666745

Loss of muscle mass: Current developments in cachexia and sarcopenia focused on biomarkers and treatment.

PubMed

Drescher, Cathleen; Konishi, Masaaki; Ebner, Nicole; Springer, Jochen

2016-01-01

Loss of muscle mass arises from an imbalance of protein synthesis and protein degradation. Potential triggers of muscle wasting and function are immobilization, loss of appetite, dystrophies and chronic diseases as well as aging. All these conditions lead to increased morbidity and mortality in patients, which makes it a timely matter to find new biomarkers to get a fast clinical diagnosis and to develop new therapies. This mini-review covers current developments in the field of biomarkers and drugs on cachexia and sarcopenia. Here, we reported about promising markers, e.g. tartrate-resistant acid phosphatase 5a (TRACP5a), and novel substances like Epigallocatechin-3-gallate (EGCg). In summary, the progress to combat muscle wasting is in full swing and perhaps diagnosis of muscle atrophy and of course patient treatments could be soon supported by improved and more helpful strategies. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

A New Piezoelectric Actuator Induces Bone Formation In Vivo: A Preliminary Study

PubMed Central

Reis, Joana; Frias, Clara; Canto e Castro, Carlos; Botelho, Maria Luísa; Marques, António Torres; Simões, José António Oliveira; Capela e Silva, Fernando; Potes, José

2012-01-01

This in vivo study presents the preliminary results of the use of a novel piezoelectric actuator for orthopedic application. The innovative use of the converse piezoelectric effect to mechanically stimulate bone was achieved with polyvinylidene fluoride actuators implanted in osteotomy cuts in sheep femur and tibia. The biological response around the osteotomies was assessed through histology and histomorphometry in nondecalcified sections and histochemistry and immunohistochemistry in decalcified sections, namely, through Masson's trichrome, and labeling of osteopontin, proliferating cell nuclear antigen, and tartrate-resistant acid phosphatase. After one-month implantation, total bone area and new bone area were significantly higher around actuators when compared to static controls. Bone deposition rate was also significantly higher in the mechanically stimulated areas. In these areas, osteopontin increased expression was observed. The present in vivo study suggests that piezoelectric materials and the converse piezoelectric effect may be used to effectively stimulate bone growth. PMID:22701304

Bioactivity of calcium phosphate bioceramic coating fabricated by laser cladding

NASA Astrophysics Data System (ADS)

Zhu, Yizhi; Liu, Qibin; Xu, Peng; Li, Long; Jiang, Haibing; Bai, Yang

2016-05-01

There were always strong expectations for suitable biomaterials used for bone regeneration. In this study, to improve the biocompatiblity of titanium alloy, calcium phosphate bioceramic coating was obtained by laser cladding technology. The microstructure, phases, bioactivity, cell differentiation, morphology and resorption lacunae were investigated by optical microscope (OM), x-ray diffraction (XRD), methyl thiazolyl tetrazolium (MTT) assay, tartrate-resistant acid phosphatase (TRAP) staining and scanning electronic microscope (SEM), respectively. The results show that bioceramic coating consists of three layers, which are a substrate, an alloyed layer and a ceramic layer. Bioactive phases of β-tricalcium phosphate (β-TCP) and hydroxyapatite (HA) were found in ceramic coating. Osteoclast precursors have excellent proliferation on the bioceramic surface. The bioceramics coating could be digested by osteoclasts, which led to the resorption lacunae formed on its surface. It revealed that the gradient bioceramic coating has an excellent bioactivity.

Purification and characterization of a phosphotyrosyl-protein phosphatase from wheat seedlings.

PubMed

Cheng, H F; Tao, M

1989-10-19

A neutral phosphatase which catalyzes the hydrolysis of p-nitrophenylphosphate has been purified to homogeneity from wheat seedlings. The enzyme is a monomeric glycoprotein exhibiting a molecular weight of 35,000, frictional ratio of 1.22, Stokes' radius of 260 nm, and sedimentation coefficient of 3.2 S. That the enzyme is a glycoprotein is surmised from its chromatographic property on Concanavalin A-Sepharose column. An examination of the substrate specificity indicates that the enzyme exhibits a preference for phosphotyrosine over a number of phosphocompounds, including p-nitrophenylphosphate and several glycolytic intermediates. Both phosphoserine and phosphothreonine are not hydrolyzed by the enzyme. The phosphatase activity is not affected by high concentrations of chelating agents and does not require metal ions. Molybdate, orthovanadate, Zn2+, and Hg2+ are all potent inhibitors of the phosphatase activity. The ability of the phosphatase to dephosphorylate protein phosphotyrosine has been investigated. [32P-Tyr]poly(Glu,Tyr)n, [32P-Tyr]alkylated bovine serum albumin, [32P-Tyr]angiotensin-I, and [32P-Tyr]band 3 (from human erythrocyte) are all substrates of the phosphatase. On the other hand, the enzyme has no activity toward protein phosphoserine and phosphothreonine. Our result further indicates that the neutral phosphatase is distinct from the wheat germ acid phosphatase. The latter enzyme is found to dephosphorylate phosphotyrosyl as well as phosphoseryl and phosphothreonyl groups in proteins. In light of the many similarities in properties to phosphotyrosyl protein phosphatases isolated from several sources, it is suggested that the wheat seedling phosphatase may participate in cellular regulation involving protein tyrosine phosphorylation.

Identification of a novel transposon-associated phosphoethanolamine transferase gene, mcr-5, conferring colistin resistance in d-tartrate fermenting Salmonella enterica subsp. enterica serovar Paratyphi B.

PubMed

Borowiak, Maria; Fischer, Jennie; Hammerl, Jens A; Hendriksen, Rene S; Szabo, Istvan; Malorny, Burkhard

2017-12-01

Plasmid-mediated mobilized colistin resistance is currently known to be caused by phosphoethanolamine transferases termed MCR-1, MCR-2, MCR-3 and MCR-4. However, this study focuses on the dissection of a novel resistance mechanism in mcr-1-, mcr-2- and mcr-3-negative d-tartrate fermenting Salmonella enterica subsp. enterica serovar Paratyphi B (Salmonella Paratyphi B dTa+) isolates with colistin MIC values >2 mg/L. A selected isolate from the strain collection of the German National Reference Laboratory for Salmonella was investigated by WGS and bioinformatical analysis to identify novel phosphoethanolamine transferase genes involved in colistin resistance. Subsequently PCR screening, S1-PFGE and DNA-DNA hybridization were performed to analyse the prevalence and location of the identified mcr-5 gene. Cloning and transformation experiments in Escherichia coli DH5α and Salmonella Paratyphi B dTa+ control strains were carried out and the activity of MCR-5 was determined in vitro by MIC testing. In this study, we identified a novel phosphoethanolamine transferase in 14 mcr-1-, mcr-2- and mcr-3-negative Salmonella Paratyphi B dTa+ isolates with colistin MIC values >2 mg/L that were received during 2011-13. The respective gene, further termed as mcr-5 (1644 bp), is part of a 7337 bp transposon of the Tn3 family and usually located on related multi-copy ColE-type plasmids. Interestingly, in one isolate an additional subclone with a chromosomal location of the mcr-5 transposon was observed. Our findings suggest that the transfer of colistin-resistance-mediating phosphoethanolamine transferase genes from bacterial chromosomes to mobile genetic elements has occurred in multiple independent events raising concern regarding their variety, prevalence and impact on public health. © The Author 2017. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Behavior of soluble and immobilized acid phosphatase in hydro-organic media.

PubMed

Wan, H; Horvath, C

1975-11-20

The hydrolysis of p-nitrophenyl phosphate by wheat germ acid phosphatase (orthophosphoric monoester phosphohydrolase, EC 3.1.3.2) has been investigated in mixtures of aqueous buffers with acetone, dioxane and acetonitrile. The enzyme was either in free solution or immobilized on a pellicular support which consisted of a porous carbonaceous layer on solid glass beads. The highest enzyme activity was obtained in acetone and acetonitrile mixed with citrate buffer over a wide range of organic solvent concentration. In 50% (v/v) acetone both V and Km of the immobilized enzyme were about half of the values in the neat aqueous buffer, but the Ki for inorganic phosphate was unchanged. In 50% (v/v) mixtures of various solvents and citrate buffers of different pH, the enzymic activity was found to depend on the pH of the aqueous buffer component rather than the pH of the hydro-organic mixture as measured with the glass-calomel electrode. The relatively high rates of p-nitrophenol liberation in the presence of glucose even at high organic solvent concentrations suggest that transphosphorylation is facilitated at low water activity.

Crystal structure of rivastigmine hydrogen tartrate Form I (Exelon®), C 14H 23N 2O 2(C 4H 5O 6)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kaduk, James A.; Zhong, Kai; Gindhart, Amy M.

2016-03-08

The crystal structure of rivastigmine hydrogen tartrate has been solved and refined using synchrotron X-ray powder diffraction data, and optimized using density functional techniques. Rivastigmine hydrogen tartrate crystallizes in space groupP2 1(#4) witha= 17.538 34(5),b= 8.326 89(2),c= 7.261 11(2) Å,β= 98.7999(2)°,V= 1047.929(4) Å 3, andZ= 2. The un-ionized end of the hydrogen tartrate anions forms a very strong hydrogen bond with the ionized end of another anion to form a chain. The ammonium group of the rivastigmine cation forms a strong discrete hydrogen bond with the carbonyl oxygen atom of the un-ionized end of the tartrate anion. These hydrogen bondsmore » form a corrugated network in thebc-plane. Both hydroxyl groups of the tartrate anion form intramolecular O–H···O hydrogen bonds. Several C–H···O hydrogen bonds appear to contribute to the crystal energy. The powder pattern is included in the Powder Diffraction File ™as entry 00-064-1501.« less

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. Copyright © 2015 Elsevier Inc. All rights reserved.

Teaching resources. Protein phosphatases.

PubMed

Salton, Stephen R

2005-03-01

This Teaching Resource provides lecture notes and slides for a class covering the structure and function of protein phosphatases and is part of the course "Cell Signaling Systems: A Course for Graduate Students." The lecture begins with a discussion of the importance of phosphatases in physiology, recognized by the award of a Nobel Prize in 1992, and then proceeds to describe the two types of protein phosphatases: serine/threonine and tyrosine phosphatases. The information covered includes the structure, regulation, and substrate specificity of protein phosphatases, with an emphasis on their importance in disease and clinical settings.

Biomineralization of Uranium by PhoY Phosphatase Activity Aids Cell Survival in Caulobacter crescentus

PubMed Central

Yung, Mimi C.

2014-01-01

Caulobacter crescentus is known to tolerate high levels of uranium [U(VI)], but its detoxification mechanism is poorly understood. Here we show that C. crescentus is able to facilitate U(VI) biomineralization through the formation of U-Pi precipitates via its native alkaline phosphatase activity. The U-Pi precipitates, deposited on the cell surface in the form of meta-autunite structures, have a lower U/Pi ratio than do chemically produced precipitates. The enzyme that is responsible for the phosphatase activity and thus the biomineralization process is identified as PhoY, a periplasmic alkaline phosphatase with broad substrate specificity. Furthermore, PhoY is shown to confer a survival advantage on C. crescentus toward U(VI) under both growth and nongrowth conditions. Results obtained in this study thus highlight U(VI) biomineralization as a resistance mechanism in microbes, which not only improves our understanding of bacterium-mineral interactions but also aids in defining potential ecological niches for metal-resistant bacteria. PMID:24878600

Biomineralization of Uranium by PhoY Phosphatase Activity Aids Cell Survival in Caulobacter crescentus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yung, M C; Jiao, Y

2014-07-22

Caulobacter crescentus is known to tolerate high levels of uranium [U(VI)], but its detoxification mechanism is poorly understood. Here we show that C. crescentus is able to facilitate U(VI) biomineralization through the formation of U-P i precipitates via its native alkaline phosphatase activity. The U-P i precipitates, deposited on the cell surface in the form of meta-autunite structures, have a lower U/P i ratio than do chemically produced precipitates. The enzyme that is responsible for the phosphatase activity and thus the biomineralization process is identified as PhoY, a periplasmic alkaline phosphatase with broad substrate specificity. Furthermore, PhoY is shown tomore » confer a survival advantage on C. crescentus toward U(VI) under both growth and nongrowth conditions. Results obtained in this study thus highlight U(VI) biomineralization as a resistance mechanism in microbes, which not only improves our understanding of bacterium-mineral interactions but also aids in defining potential ecological niches for metal-resistant bacteria.« less

Protein tyrosine phosphatase 1B inhibitory activity of lavandulyl flavonoids from roots of Sophora flavescens.

PubMed

Sasaki, Tatsunori; Li, Wei; Higai, Koji; Quang, Tran Hong; Kim, Young Ho; Koike, Kazuo

2014-05-01

Protein tyrosine phosphatase 1B is a non-transmembrane protein tyrosine phosphatase and major negative regulator in insulin signaling cascades, and much attention has been paid to protein tyrosine phosphatase 1B inhibitors as potential therapies for diabetes. The screening of a natural compound library led to the discovery of five lavandulyl flavonoids, which were isolated from the roots of Sophora flavescens, as novel PTP1B inhibitors: kuraridin (1), norkurarinone (2), kurarinone (3), 2'-methoxykurarinone (4), and kushenol T (5). The three most potent compounds, 1, 2, and 4 (IC50 < 30 µM), were demonstrated to be noncompetitive inhibitors of protein tyrosine phosphatase 1B based on a kinetic analysis, and they exhibited different inhibitory selectivities against four homologous protein tyrosine phosphatases (T cell protein tyrosine phosphatase, vaccinia H1-related phosphatase, and Src homology domain 2-containing protein tyrosine phosphatases 1 and 2). Compounds 1, 2, and 4 also exhibited cellular activity in the insulin signaling pathway by increasing the insulin-stimulated Akt phosphorylation level in human hepatocellular liver carcinoma HepG2 cells, suggesting their potential for new anti-insulin-resistant drug developments. Georg Thieme Verlag KG Stuttgart · New York.

Fungal extracellular phosphatases: their role in P cycling under different pH and P sources availability.

PubMed

Della Mónica, I F; Godoy, M S; Godeas, A M; Scervino, J M

2018-01-01

The aim of this work is to analyse the effect of pH, fungal identity and P chemical nature on microbial development and phosphatase release, discussing solubilization and mineralization processes in P cycling. P solubilizing fungi (Talaromyces flavus, T. helicus L, T. helicus N, T. diversus and Penicillium purpurogenum) were grown under three pH conditions (6, 6·5 and 8·5) and with different inorganic (calcium, iron, aluminium and rock) and organic (lecithin and phytate) P sources. P solubilization, mineralization, growth and phosphatase production were recorded. Acid and neutral environments maximized fungal development and P recycling. P chemical nature changed the phosphatases release pattern depending on the fungal identity. Acid phosphatase activity was higher than alkaline phosphatases, regardless of pH or sample times. Alkaline phosphatases were affected by a combination of those factors. P chemical nature and pH modify fungal growth, P mineralization and solubilization processes. The underlying fungal identity-dependent metabolism governs the capacity and efficiency of P solubilization and mineralization. P solubilization and mineralization processes are interrelated and simultaneously present in soil fungi. This study constitutes a reference work to improve the selection of fungal bioinoculants in different environmental conditions, highlighting their role in P cycling. © 2017 The Society for Applied Microbiology.

MECHANISMS IN ENDOCRINOLOGY: Diabetes mellitus, a state of low bone turnover - a systematic review and meta-analysis.

PubMed

Hygum, Katrine; Starup-Linde, Jakob; Harsløf, Torben; Vestergaard, Peter; Langdahl, Bente L

2017-03-01

To investigate the differences in bone turnover between diabetic patients and controls. A systematic review and meta-analysis. A literature search was conducted using the databases Medline at PubMed and EMBASE. The free text search terms 'diabetes mellitus' and 'bone turnover', 'sclerostin', 'RANKL', 'osteoprotegerin', 'tartrate-resistant acid' and 'TRAP' were used. Studies were eligible if they investigated bone turnover markers in patients with diabetes compared with controls. Data were extracted by two reviewers. A total of 2881 papers were identified of which 66 studies were included. Serum levels of the bone resorption marker C-terminal cross-linked telopeptide (-0.10 ng/mL (-0.12, -0.08)) and the bone formation markers osteocalcin (-2.51 ng/mL (-3.01, -2.01)) and procollagen type 1 amino terminal propeptide (-10.80 ng/mL (-12.83, -8.77)) were all lower in patients with diabetes compared with controls. Furthermore, s-tartrate-resistant acid phosphatase was decreased in patients with type 2 diabetes (-0.31 U/L (-0.56, -0.05)) compared with controls. S-sclerostin was significantly higher in patients with type 2 diabetes (14.92 pmol/L (3.12, 26.72)) and patients with type 1 diabetes (3.24 pmol/L (1.52, 4.96)) compared with controls. Also, s-osteoprotegerin was increased among patients with diabetes compared with controls (2.67 pmol/L (0.21, 5.14)). Markers of both bone formation and bone resorption are decreased in patients with diabetes. This suggests that diabetes mellitus is a state of low bone turnover, which in turn may lead to more fragile bone. Altered levels of sclerostin and osteoprotegerin may be responsible for this. © 2017 European Society of Endocrinology.

Overview on mechanisms of acetic acid resistance in acetic acid bacteria.

PubMed

Wang, Bin; Shao, Yanchun; Chen, Fusheng

2015-02-01

Acetic acid bacteria (AAB) are a group of gram-negative or gram-variable bacteria which possess an obligate aerobic property with oxygen as the terminal electron acceptor, meanwhile transform ethanol and sugar to corresponding aldehydes, ketones and organic acids. Since the first genus Acetobacter of AAB was established in 1898, 16 AAB genera have been recorded so far. As the main producer of a world-wide condiment, vinegar, AAB have evolved an elegant adaptive system that enables them to survive and produce a high concentration of acetic acid. Some researches and reviews focused on mechanisms of acid resistance in enteric bacteria and made the mechanisms thoroughly understood, while a few investigations did in AAB. As the related technologies with proteome, transcriptome and genome were rapidly developed and applied to AAB research, some plausible mechanisms conferring acetic acid resistance in some AAB strains have been published. In this review, the related mechanisms of AAB against acetic acid with acetic acid assimilation, transportation systems, cell morphology and membrane compositions, adaptation response, and fermentation conditions will be described. Finally, a framework for future research for anti-acid AAB will be provided.

Modulation of vinblastine cytotoxicity by dilantin (phenytoin) or the protein phosphatase inhibitor okadaic acid involves the potentiation of anti-mitotic effects and induction of apoptosis in human tumour cells.

PubMed Central

Kawamura, K. I.; Grabowski, D.; Weizer, K.; Bukowski, R.; Ganapathi, R.

1996-01-01

Cellular insensitivity to vinca alkaloids is suggested to be primarily due to drug efflux by P-glycoprotein (P-gp). The anti-epileptic phenytoin (DPH), which does not bind to P-gp, can selectively enhance vincristine (VCR) cytotoxicity in wild-type (WT) or multidrug-resistant (MDR) cells. We now demonstrate that the protein phosphatase inhibitor okadaic acid (OKA) can mimic the effect of DPH by selectively enhancing cytotoxicity of vinblastine (VBL), but not taxol and doxorubicin, in human leukaemia HL-60 cells. Both DPH and OKA potentiate the anti-mitotic effects of VBL by enhanced damage to the mitotic spindle, resulting in prolonged growth arrest. Also, unlike VBL alone, in human leukaemia or non-small-cell lung carcinoma cells treated with VBL plus DPH, recovery from damage to the mitotic spindle is compromised in drug-free medium and cell death by apoptosis in interphase ensues. Since protein phosphatases are involved with the regulation of metaphase to anaphase transit of cells during the mitotic cycle, enhanced VBL cytotoxicity in the presence of DPH or OKA may involve effects during metaphase on the mitotic spindle tubulin leading to growth arrest and apoptosis in interphase. These novel results suggest that DPH or OKA could be powerful tools to study cellular effects of vinca alkaloids and possibly for the development of novel therapeutic strategies. Images Figure 6 PMID:8546904

Phosphatase inhibitors remove the run-down of γ-aminobutyric acid type A receptors in the human epileptic brain

PubMed Central

Palma, E.; Ragozzino, D. A.; Di Angelantonio, S.; Spinelli, G.; Trettel, F.; Martinez-Torres, A.; Torchia, G.; Arcella, A.; Di Gennaro, G.; Quarato, P. P.; Esposito, V.; Cantore, G.; Miledi, R.; Eusebi, F.

2004-01-01

The properties of γ-aminobutyric acid (GABA) type A receptors (GABAA receptors) microtransplanted from the human epileptic brain to the plasma membrane of Xenopus oocytes were compared with those recorded directly from neurons, or glial cells, in human brains slices. Cell membranes isolated from brain specimens, surgically obtained from six patients afflicted with drug-resistant temporal lobe epilepsy (TLE) were injected into frog oocytes. Within a few hours, these oocytes acquired GABAA receptors that generated GABA currents with an unusual run-down, which was inhibited by orthovanadate and okadaic acid. In contrast, receptors derived from membranes of a nonepileptic hippocampal uncus, membranes from mouse brain, or recombinant rat α1β2γ2-GABA receptors exhibited a much less pronounced GABA-current run-down. Moreover, the GABAA receptors of pyramidal neurons in temporal neocortex slices from the same six epileptic patients exhibited a stronger run-down than the receptors of rat pyramidal neurons. Interestingly, the GABAA receptors of neighboring glial cells remained substantially stable after repetitive activation. Therefore, the excessive GABA-current run-down observed in the membrane-injected oocytes recapitulates essentially what occurs in neurons, rather than in glial cells. Quantitative RT-PCR analyses from the same TLE neocortex specimens revealed that GABAA-receptor β1, β2, β3, and γ2 subunit mRNAs were significantly overexpressed (8- to 33-fold) compared with control autopsy tissues. Our results suggest that an abnormal GABA-receptor subunit transcription in the TLE brain leads to the expression of run-down-enhanced GABAA receptors. Blockage of phosphatases stabilizes the TLE GABAA receptors and strengthens GABAergic inhibition. It may be that this process can be targeted to develop new treatments for intractable epilepsy. PMID:15218107

A Novel Phosphatidic Acid-Protein-tyrosine Phosphatase D2 Axis Is Essential for ERBB2 Signaling in Mammary Epithelial Cells*

PubMed Central

Ramesh, Mathangi; Krishnan, Navasona; Muthuswamy, Senthil K.; Tonks, Nicholas K.

2015-01-01

We used a loss-of-function screen to investigate the role of classical protein-tyrosine phosphatases (PTPs) in three-dimensional mammary epithelial cell morphogenesis and ERBB2 signaling. The study revealed a novel role for PTPD2 as a positive regulator of ERBB2 signaling. Suppression of PTPD2 attenuated the ERBB2-induced multiacinar phenotype in three-dimensional cultures specifically by inhibiting ERBB2-mediated loss of polarity and lumen filling. In contrast, overexpression of PTPD2 enhanced the ERBB2 phenotype. We also found that a lipid second messenger, phosphatidic acid, bound PTPD2 in vitro and enhanced its catalytic activity. Small molecule inhibitors of phospholipase D (PLD), an enzyme that produces phosphatidic acid in cells, also attenuated the ERBB2 phenotype. Exogenously added phosphatidic acid rescued the PLD-inhibition phenotype, but only when PTPD2 was present. These findings illustrate a novel pathway involving PTPD2 and the lipid second messenger phosphatidic acid that promotes ERBB2 function. PMID:25681440

Identification, synthesis, isolation and characterization of new impurity in metoprolol tartrate tablets.

PubMed

Reddy, R Buchi; More, Kishor R; Gupta, Leena; Jha, Mukesh S; Magar, Laki

2016-01-05

A new unknown impurity was observed in accelerated stability studies of Metoprolol tartrate tablets. This impurity has been identified, synthesized and characterized through different spectral studies and confirmed as an adduct of lactose and Metoprolol formed by Maillard reaction. Copyright © 2015 Elsevier B.V. All rights reserved.

Toxic impact of aldrin on acid and alkaline phosphatase activity of penaeid prawn, Metapenaeus monoceros: In vitro study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Reddy, M.S.; Jayaprada, P.; Rao, K.V.R.

1991-03-01

The increasing contamination of the aquatic environment by the indiscriminate and widespread use of different kinds of pesticides is a serious problem for environmental biologists. Organochlorine insecticides are more hazardous since they are not only more toxic but also leave residues in nature. The deleterious effects of aldrin on several crustaceans have been studied. But studies concerning the impact of aldrin on biochemical aspects of crustaceans are very much limited. The present study is aimed at probing the in vitro effects of aldrin on the acid and alkaline phosphatase activity levels in selected tissues of penaeid prawn, Metapenaeus monoceros (Fabricius).

Relation of fatty acid composition in lead-exposed mallards to fat mobilization, lipid peroxidation and alkaline phosphatase activity

USGS Publications Warehouse

Mateo, R.; Beyer, W.N.; Spann, J.W.; Hoffman, D.J.

2003-01-01

The increase of n-6 polyunsaturated fatty acids (PUFA) in animal tissues has been proposed as a mechanism of lead (Pb) poisoning through lipid peroxidation or altered eicosanoids metabolism. We have studied fatty acid (FA) composition in liver and brain of mallards (Anas platyrhynchos) feeding for 3 weeks on diets containing combinations of low or high levels of vitamin E (20 or 200 UI/kg) and Pb (0 or 2 g/kg). Saturated FA, n-6 PUFA and total concentrations of FA were higher in livers of Pb-exposed mallards, but not in their brains. The percentage of n-6 PUFA in liver and brain was slightly higher in Pb-exposed mallards. The increase of n-6 PUFA in liver was associated with decreased triglycerides and increased cholesterol in plasma, thus could be in part attributed to feed refusal and fat mobilization. The hepatic ratios between adrenic acid (22:4 n-6) and arachidonic acid (20:4 n-6) or between adrenic acid and linoleic acid (18:2 n-6) were higher in Pb exposed birds, supporting the existing hypothesis of increased fatty acid elongation by Pb. Among the possible consequences of increased n-6 PUFA concentration in tissues, we found increased lipid peroxidation in liver without important histopathological changes, and decreased plasma alkaline phosphatase activity that may reflect altered bone metabolism in birds.

Dual actions of lysergic acid diethylamide tartrate (LSD), 2-bromo-D-lysergic acid diethylamide bitartrate (BOL) and methysergide on dorsal root potentials evoked by stimulation of raphe nuclei.

PubMed

Larson, A A; Chinn, C; Proudfit, H K; Anderson, E G

1981-04-01

A variety of drugs reported to antagonize serotonin were found to affect spinal cord potentials evoked by electrical stimulation of the caudal raphe nuclei of the cat. These brain stem-evoked dorsal root potentials (DRPs) consisted of a short latency depolarization (DRP-1), which was evoked by stimulation of a wide variety of sites in the medial brain stem and a long latency potential (DRP-2), which was elicited only when stimuli were applied near the raphe. The ability of serotonergic antagonists to increase or decrease these DRPs was dependent on the dose of the drug administered. High doses of lysergic acid diethylamide tartrate (LSD), 2-bromo-D-lysergic acid diethylamide bitartrate (BOL), methysergide and cinanserin each produced an immediate inhibition of DRP-2 and a simultaneous enhancement of DRP-1, both of which recovered by approximately 30 min. Each of the drugs produced a dose-related inhibition of DRP-2 at high doses, with LSD being the most potent and cinanserin the least potent. In contrast, low doses of LSD, BOL and methysergide elicited little or no immediate change in either DRP-2 or DRP-1, but produced an enhancement of DRP-2 which developed slowly over a period of 60 to 90 min. This increase in DRP-2 was most dramatic after administration of LSD and was not accompanied by changes in DRP-1. The inhibition of DRP-2 by high doses of LSD, BOL, methysergide and cinanserin may result primarily from inhibition of postsynaptic serotonergic receptors located on the primary afferent terminals. The increase in DRP-2 produced by low doses of LSD, BOL and methysergide is postulated to result from an interaction with receptors distinct from those which produced the inhibition of DRP-2 at higher doses.

Characterization of dolichol and dolichyl phosphate phosphatase from soya beans (Glycine max).

PubMed Central

Ravi, K; Rip, J W; Carroll, K K

1983-01-01

A series of polyprenols, ranging in length from 15 to 22 isoprene units, has been isolated from soya beans (Glycine max) and purified by high-pressure liquid chromatography. N.m.r., i.r. and mass spectra of the compounds indicated that they are alpha-saturated polyprenols of the dolichol type. The amount present in dry seeds was about 9 mg/100 g, whereas dolichyl phosphate (Dol-P) was present only in trace amounts. Dol-P phosphatase activity was detected in the microsomal fraction of 5-day-old germinating soya-bean cotyledons. The Dol-P phosphatase activity was linear with respect to time and protein concentration and exhibited a broad pH optimum (pH 7-9). Triton X-100 was necessary for significant enzyme activity. Enzyme activity was slightly enhanced by EDTA, whereas dithiothreitol was without effect. An apparent Km of 5 microM was determined for Dol-P. Bivalent metal ions were not required for enzyme activity. A number of phosphorylated compounds tested as enzyme substrates (including a number of nucleoside phosphates, glucose 6-phosphate, sodium beta-glycerophosphate and Na4P2O7) did not compete with [1-3H]Dol-P as substrate. A number of phospholipids were also tested for their ability to act as Dol-P phosphatase substrates. At 1 mM concentration, phosphatidylcholine, phosphatidylethanolamine, phosphatidic acid and lysophosphatidic acid each inhibited enzymic activity. However, at 0.1 mM concentration, phosphatidylcholine and phosphatidylethanolamine were slightly stimulatory, whereas phosphatidic acid and lysophosphatidic acid were still inhibitory. Phosphatidic acid showed competitive inhibition. PMID:6311165

Identification of protein tyrosine phosphatase SHP-2 as a new target of perfluoroalkyl acids in HepG2 cells.

PubMed

Yang, Yu; Lv, Qi-Yan; Guo, Liang-Hong; Wan, Bin; Ren, Xiao-Min; Shi, Ya-Li; Cai, Ya-Qi

2017-04-01

Perfluoroalkyl acids (PFAAs) are widespread environmental contaminants which have been detected in humans and linked to adverse health effects. Previous toxicological studies mostly focused on nuclear receptor-mediated pathways and did not support the observed toxic effects. In this study, we aimed to investigate the molecular mechanisms of PFAA toxicities by identifying their biological targets in cells. Using a novel electrochemical biosensor, 16 PFAAs were evaluated for inhibition of protein tyrosine phosphatase SHP-2 activity. Their potency increased with PFAA chain length, with perfluorooctadecanoic acid (PFODA) showing the strongest inhibition. Three selected PFAAs, 25 μM perfluorooctanoic acid (PFOA), perfluorooctane sulfonic acid, and PFODA, also inhibited SHP-2 activity in HepG2 cells and increased paxillin phosphorylation level. PFOA was detected in the immunoprecipitated SHP-2 from the cells exposed to 250 μM PFOA, providing unequivocal evidence for the direct binding of PFOA with SHP-2 in the cell. Molecular docking rationalized the formation of PFAA/SHP-2 complex and chain length-dependent inhibition potency. Our results have established SHP-2 as a new cellular target of PFAAs.

Influence of cyclopropane fatty acids on heat, high pressure, acid and oxidative resistance in Escherichia coli.

PubMed

Chen, Yuan Yao; Gänzle, Michael G

2016-04-02

Heat and high pressure resistant strains of Escherichia coli are a challenge to food safety. This study investigated effects of cyclopropane fatty acids (CFAs) on stress tolerance in the heat- and pressure-resistant strain E. coli AW1.7 and the sensitive strain E. coli MG1655. The role of CFAs was explored by disruption of cfa coding for CFA synthase with an in-frame, unmarked deletion method. Both wild-type strains consumed all the unsaturated fatty acids (C16:1 and C18:1) that were mostly converted to CFAs and a low proportion to saturated fatty acid (C16:0). Moreover, E. coli AW1.7 contained a higher proportion of membrane C19:0 cyclopropane fatty acid than E. coli MG1655 (P<0.05). The Δcfa mutant strains did not produce CFAs, and the corresponding substrates C16:1 and C18:1 accumulated in membrane lipids. The deletion of cfa did not alter resistance to H2O2 but increased the lethality of heat, high pressure and acid treatments in E. coli AW1.7, and E. coli MG1655. E. coli AW1.7 and its Δcfa mutant were more resistant to pressure and heat but less resistant to acid stress than E. coli MG1655. Heat resistance of wild-type strains and their Δcfa mutant was also assessed in beef patties grilled to an internal temperature of 71 °C. After treatment, cell counts of wild type strains were higher than those of the Δcfa mutant strains. In conclusion, CFA synthesis in E. coli increases heat, high pressure and acid resistance, and increases heat resistance in food. This knowledge on mechanisms of stress resistance will facilitate the design of intervention methods for improved pathogen control in food production. Copyright © 2016 Elsevier B.V. All rights reserved.

Voltage-dependent motion of the catalytic region of voltage-sensing phosphatase monitored by a fluorescent amino acid

PubMed Central

Sakata, Souhei; Jinno, Yuka; Kawanabe, Akira; Okamura, Yasushi

2016-01-01

The cytoplasmic region of voltage-sensing phosphatase (VSP) derives the voltage dependence of its catalytic activity from coupling to a voltage sensor homologous to that of voltage-gated ion channels. To assess the conformational changes in the cytoplasmic region upon activation of the voltage sensor, we genetically incorporated a fluorescent unnatural amino acid, 3-(6-acetylnaphthalen-2-ylamino)-2-aminopropanoic acid (Anap), into the catalytic region of Ciona intestinalis VSP (Ci-VSP). Measurements of Anap fluorescence under voltage clamp in Xenopus oocytes revealed that the catalytic region assumes distinct conformations dependent on the degree of voltage-sensor activation. FRET analysis showed that the catalytic region remains situated beneath the plasma membrane, irrespective of the voltage level. Moreover, Anap fluorescence from a membrane-facing loop in the C2 domain showed a pattern reflecting substrate turnover. These results indicate that the voltage sensor regulates Ci-VSP catalytic activity by causing conformational changes in the entire catalytic region, without changing their distance from the plasma membrane. PMID:27330112

Voltage-dependent motion of the catalytic region of voltage-sensing phosphatase monitored by a fluorescent amino acid.

PubMed

Sakata, Souhei; Jinno, Yuka; Kawanabe, Akira; Okamura, Yasushi

2016-07-05

The cytoplasmic region of voltage-sensing phosphatase (VSP) derives the voltage dependence of its catalytic activity from coupling to a voltage sensor homologous to that of voltage-gated ion channels. To assess the conformational changes in the cytoplasmic region upon activation of the voltage sensor, we genetically incorporated a fluorescent unnatural amino acid, 3-(6-acetylnaphthalen-2-ylamino)-2-aminopropanoic acid (Anap), into the catalytic region of Ciona intestinalis VSP (Ci-VSP). Measurements of Anap fluorescence under voltage clamp in Xenopus oocytes revealed that the catalytic region assumes distinct conformations dependent on the degree of voltage-sensor activation. FRET analysis showed that the catalytic region remains situated beneath the plasma membrane, irrespective of the voltage level. Moreover, Anap fluorescence from a membrane-facing loop in the C2 domain showed a pattern reflecting substrate turnover. These results indicate that the voltage sensor regulates Ci-VSP catalytic activity by causing conformational changes in the entire catalytic region, without changing their distance from the plasma membrane.

Discovery and structure-activity relationship of oxalylarylaminobenzoic acids as inhibitors of protein tyrosine phosphatase 1B.

PubMed

Liu, Gang; Szczepankiewicz, Bruce G; Pei, Zhonghua; Janowick, David A; Xin, Zhili; Hajduk, Philip J; Abad-Zapatero, Cele; Liang, Heng; Hutchins, Charles W; Fesik, Stephen W; Ballaron, Steve J; Stashko, Mike A; Lubben, Tom; Mika, Amanda K; Zinker, Bradley A; Trevillyan, James M; Jirousek, Michael R

2003-05-22

Protein Tyrosine phosphatase 1B (PTP1B) has been implicated as a key negative regulator of both insulin and leptin signaling pathways. Using an NMR-based screening approach with 15N- and 13C-labeled PTP1B, we have identified 2,3-dimethylphenyloxalylaminobenzoic acid (1) as a general, reversible, and competitive PTPase inhibitor. Structure-based approach guided by X-ray crystallography facilitated the development of 1 into a novel series of potent and selective PTP1B inhibitors occupying both the catalytic site and a portion of the noncatalytic, second phosphotyrosine binding site. Interestingly, oral biovailability has been observed in rats for some compounds. Furthermore, we demonstrated in vivo plasma glucose lowering effects with compound 12d in ob/ob mice.

Differential Processing of Propeptide Inhibitors of Rap Phosphatases in Bacillus subtilis†

PubMed Central

Jiang, Min; Grau, Roberto; Perego, Marta

2000-01-01

In the phosphorelay signal transduction system for sporulation initiation in Bacillus subtilis, the opposing activities of histidine kinases and aspartyl phosphate phosphatases determine the cell's decision whether to continue with vegetative growth or to initiate the differentiation process. Regulated dephosphorylation of the Spo0A and Spo0F response regulators allows a variety of negative signals from physiological processes that are antithetical to sporulation to impact on the activation level of the phosphorelay. Spo0F∼P is the known target of two related phosphatases, RapA and RapB. In addition to RapA and RapB, a third member of the Rap family of phosphatases, RapE, specifically dephosphorylated the Spo0F∼P intermediate in response to competence development. RapE phosphatase activity was found to be controlled by a pentapeptide (SRNVT) generated from within the carboxy-terminal domain of the phrE gene product. A synthetic PhrE pentapeptide could (i) complement the sporulation deficiency caused by deregulated RapE activity of a phrE mutant and (ii) inhibit RapE-dependent dephosphorylation of Spo0F∼P in in vitro experiments. The PhrE pentapeptide did not inhibit the phosphatase activity of RapA and RapB. These results confirm previous conclusions that the specificity for recognition of the target phosphatase is contained within the amino acid sequence of the pentapeptide inhibitor. PMID:10629174

Conservation of the PTEN catalytic motif in the bacterial undecaprenyl pyrophosphate phosphatase, BacA/UppP.

PubMed

Bickford, Justin S; Nick, Harry S

2013-12-01

Isoprenoid lipid carriers are essential in protein glycosylation and bacterial cell envelope biosynthesis. The enzymes involved in their metabolism (synthases, kinases and phosphatases) are therefore critical to cell viability. In this review, we focus on two broad groups of isoprenoid pyrophosphate phosphatases. One group, containing phosphatidic acid phosphatase motifs, includes the eukaryotic dolichyl pyrophosphate phosphatases and proposed recycling bacterial undecaprenol pyrophosphate phosphatases, PgpB, YbjB and YeiU/LpxT. The second group comprises the bacterial undecaprenol pyrophosphate phosphatase, BacA/UppP, responsible for initial formation of undecaprenyl phosphate, which we predict contains a tyrosine phosphate phosphatase motif resembling that of the tumour suppressor, phosphatase and tensin homologue (PTEN). Based on protein sequence alignments across species and 2D structure predictions, we propose catalytic and lipid recognition motifs unique to BacA/UppP enzymes. The verification of our proposed active-site residues would provide new strategies for the development of substrate-specific inhibitors which mimic both the lipid and pyrophosphate moieties, leading to the development of novel antimicrobial agents.

A paralogue of the phosphomutase-like gene family in Candida glabrata, CgPmu2, gained broad-range phosphatase activity due to a small number of clustered substitutions.

PubMed

Orlando, Kelly A; Iosue, Christine L; Leone, Sarah G; Davies, Danielle L; Wykoff, Dennis D

2015-10-15

Inorganic phosphate is required for a range of cellular processes, such as DNA/RNA synthesis and intracellular signalling. The phosphate starvation-inducible phosphatase activity of Candida glabrata is encoded by the gene CgPMU2 (C. glabrata phosphomutase-like protein). CgPMU2 is part of a three-gene family (∼75% identical) created through gene duplication in the C. glabrata clade; only CgPmu2 is a PHO-regulated broad range acid phosphatase. We identified amino acids that confer broad range phosphatase activity on CgPmu2 by creating fusions of sections of CgPMU2 with CgPMU1, a paralogue with little broad range phosphatase activity. We used site-directed mutagenesis on various fusions to sequentially convert CgPmu1 to CgPmu2. Based on molecular modelling of the Pmu proteins on to a histidine phosphatase crystal structure, clusters of amino acids were found in two distinct regions that were able to confer phosphatase activity. Substitutions in these two regions together conferred broad phosphatase activity on CgPmu1. Interestingly, one change is a histidine adjacent to the active site histidine of CgPmu2 and it exhibits a novel ability to partially replace the conserved active site histidine in CgPmu2. Additionally, a second amino acid change was able to confer nt phosphatase activity to CgPmu1, suggesting single amino acid changes neofunctionalize CgPmu2. © 2015 Authors; published by Portland Press Limited.

RANKL-induced DC-STAMP Is Essential for Osteoclastogenesis

PubMed Central

Kukita, Toshio; Wada, Naohisa; Kukita, Akiko; Kakimoto, Takashi; Sandra, Ferry; Toh, Kazuko; Nagata, Kengo; Iijima, Tadahiko; Horiuchi, Madoka; Matsusaki, Hiromi; Hieshima, Kunio; Yoshie, Osamu; Nomiyama, Hisayuki

2004-01-01

Osteoclasts are bone-resorbing, multinucleated giant cells that are essential for bone remodeling and are formed through cell fusion of mononuclear precursor cells. Although receptor activator of nuclear factor–κB ligand (RANKL) has been demonstrated to be an important osteoclastogenic cytokine, the cell surface molecules involved in osteoclastogenesis are mostly unknown. Here, we report that the seven-transmembrane receptor-like molecule, dendritic cell–specific transmembrane protein (DC-STAMP) is involved in osteoclastogenesis. Expression of DC-STAMP is rapidly induced in osteoclast precursor cells by RANKL and other osteoclastogenic stimulations. Targeted inhibition of DC-STAMP by small interfering RNAs and specific antibody markedly suppressed the formation of multinucleated osteoclast-like cells. Overexpression of DC-STAMP enhanced osteoclastogenesis in the presence of RANKL. Furthermore, DC-STAMP directly induced the expression of the osteoclast marker tartrate-resistant acid phosphatase. These data demonstrate for the first time that DC-STAMP has an essential role in osteoclastogenesis. PMID:15452179

Effect of amorphous silica nanoparticles on in vitro RANKL-induced osteoclast differentiation in murine macrophages

NASA Astrophysics Data System (ADS)

Nabeshi, Hiromi; Yoshikawa, Tomoaki; Akase, Takanori; Yoshida, Tokuyuki; Tochigi, Saeko; Hirai, Toshiro; Uji, Miyuki; Ichihashi, Ko-Ichi; Yamashita, Takuya; Higashisaka, Kazuma; Morishita, Yuki; Nagano, Kazuya; Abe, Yasuhiro; Kamada, Haruhiko; Tsunoda, Shin-Ichi; Itoh, Norio; Yoshioka, Yasuo; Tsutsumi, Yasuo

2011-07-01

Amorphous silica nanoparticles (nSP) have been used as a polishing agent and/or as a remineralization promoter for teeth in the oral care field. The present study investigates the effects of nSP on osteoclast differentiation and the relationship between particle size and these effects. Our results revealed that nSP exerted higher cytotoxicity in macrophage cells compared with submicron-sized silica particles. However, tartrate-resistant acid phosphatase (TRAP) activity and the number of osteoclast cells (TRAP-positive multinucleated cells) were not changed by nSP treatment in the presence of receptor activator of nuclear factor κB ligand (RANKL) at doses that did not induce cytotoxicity by silica particles. These results indicated that nSP did not cause differentiation of osteoclasts. Collectively, the results suggested that nanosilica exerts no effect on RANKL-induced osteoclast differentiation of RAW264.7 cells, although a detailed mechanistic examination of the nSP70-mediated cytotoxic effect is needed.

Increased hypothalamic protein tyrosine phosphatase 1B contributes to leptin resistance with age.

PubMed

Morrison, Christopher D; White, Christy L; Wang, Zhong; Lee, Seung-Yub; Lawrence, David S; Cefalu, William T; Zhang, Zhong-Yin; Gettys, Thomas W

2007-01-01

Animals at advanced ages exhibit a reduction in central leptin sensitivity. However, changes in growth, metabolism, and obesity risk occur much earlier in life, particularly during the transition from youth to middle age. To determine when initial decreases in central leptin sensitivity occur, leptin-dependent suppression of food intake was tested in 8-, 12-, and 20-wk-old male, chow-fed Sprague Dawley rats. Intracerebroventricular leptin injection (3 microg) suppressed 24-h food intake in 8- and 12-wk-old rats (P < 0.05) but not 20-wk-old rats. To identify potential cellular mediators of this resistance, we focused on protein tyrosine phosphatase 1B (PTP1B), a recently described inhibitor of leptin signaling. PTP1B protein levels, as determined by Western blot, were significantly higher in mediobasal hypothalamic punches collected from 20-wk-old rats, compared with 8-wk-old rats (P < 0.05). When 20-wk-old rats were fasted for 24 h, levels of hypothalamic PTP1B decreased (P < 0.05), coincident with a restoration of leptin sensitivity. To directly test whether inhibition of PTP1B restores leptin sensitivity, 20-wk-old chow-fed rats were pretreated with a pharmacological PTP1B inhibitor 1 h before leptin, and 24-h food intake was recorded. As expected, leptin alone produced a small but nonsignificant reduction in food intake. However, pretreatment with the PTP1B inhibitor resulted in a marked improvement in leptin-dependent suppression of food intake (P < 0.05). These data are consistent with the hypothesis that increases in PTP1B contribute to hypothalamic leptin resistance as rats transition into middle age.

Acetic acid fermentation of acetobacter pasteurianus: relationship between acetic acid resistance and pellicle polysaccharide formation.

PubMed

Kanchanarach, Watchara; Theeragool, Gunjana; Inoue, Taketo; Yakushi, Toshiharu; Adachi, Osao; Matsushita, Kazunobu

2010-01-01

Acetobacter pasteurianus strains IFO3283, SKU1108, and MSU10 were grown under acetic acid fermentation conditions, and their growth behavior was examined together with their capacity for acetic acid resistance and pellicle formation. In the fermentation process, the cells became aggregated and covered by amorphous materials in the late-log and stationary phases, but dispersed again in the second growth phase (due to overoxidation). The morphological change in the cells was accompanied by changes in sugar contents, which might be related to pellicle polysaccharide formation. To determine the relationship between pellicle formation and acetic acid resistance, a pellicle-forming R strain and a non-forming S strain were isolated, and their fermentation ability and acetic acid diffusion activity were compared. The results suggest that pellicle formation is directly related to acetic acid resistance ability, and thus is important to acetic acid fermentation in these A. pasteurianus strains.

Legionella pneumophila effector WipA, a bacterial PPP protein phosphatase with PTP activity.

PubMed

Jia, Qian; Lin, Yun; Gou, Xuejing; He, Lei; Shen, Dong; Chen, Dongni; Xie, Wei; Lu, Yongjun

2018-04-26

The gram-negative bacterium Legionella pneumophila invades human's lung and causes Legionnaires' disease. To benefit its survival and replication in cellular milieu, L. pneumophila secrets at least 330 effector proteins into host cells. We found that the effector WipA has the protein tyrosine phosphatase (PTP) activity but does not depend on the classical CX5R motif for activity, suggesting that WipA is an unconventional PTP. Meanwhile, the presence of three other highly conserved motifs typically seen in protein serine/threonine phosphatases and the poor inhibition of WipA activity by okadaic acid led us to propose that WipA is a bacterial protein phosphatase. In addition, the determination of the 2.55-Å crystal structure of WipA revealed that WipA resembles cold-active protein tyrosine phosphatase (CAPTPase), and therefore very likely shares the same catalytic mechanism.

Effects of synthetic detergents on in vivo activity of tissue phosphatases and succinic dehydrogenase from Mystus vittatus.

PubMed

Mohan, D; Verma, S R

1981-05-01

African catfish (Mystus vittatus) were exposed to three sub-lethal concentrations of Swascofix E45 (13.8, 9.2 and 4.6 mg/l) and Swascol 3L (69.3, 46.2 and 23.1 mg/l) for 15 and 30 days, and their effects on alkaline and acid phosphatase, and succinic dehydrogenase in liver, kidney and intestine were measured. The enzymes were found to be inhibited in all the tissues. Maximum inhibition (38.44%) was observed in liver alkaline phosphatase activity after 30 days with the highest concentration of Swascofix E45 and the lowest inhibition (0.118%) was found in kidney acid phosphatase activity with the lowest concentration of Swascol 3L after 15 days. Insignificant enzyme stimulation in some cases was also observed.

Histochemical assessment for osteoblastic activity coupled with dysfunctional osteoclasts in c-src deficient mice.

PubMed

Toray, Hisashi; Hasegawa, Tomoka; Sakagami, Naoko; Tsuchiya, Erika; Kudo, Ai; Zhao, Shen; Moritani, Yasuhito; Abe, Miki; Yoshida, Taiji; Yamamoto, Tomomaya; Yamamoto, Tsuneyuki; Oda, Kimimitsu; Udagawa, Nobuyuki; Luiz de Freitas, Paulo Henrique; Li, Minqi

2017-01-01

Since osteoblastic activities are believed to be coupled with osteoclasts, we have attempted to histologically verify which of the distinct cellular circumstances, the presence of osteoclasts themselves or bone resorption by osteoclasts, is essential for coupled osteoblastic activity, by examining c-fos -/- or c-src -/- mice. Osteopetrotic c-fos deficient (c-fos -/- ) mice have no osteoclasts, while c-src deficient (c-src -/- ) mice, another osteopetrotic model, develop dysfunctional osteoclasts due to a lack of ruffled borders. c-fos -/- mice possessed no tartrate-resistant acid phosphatase (TRAPase)-reactive osteoclasts, and showed very weak tissue nonspecific alkaline phosphatase (TNALPase)-reactive mature osteoblasts. In contrast, c-src -/- mice had many TNALPase-positive osteoblasts and TRAPase-reactive osteoclasts. Interestingly, the parallel layers of TRAPase-reactive/osteopontin-positive cement lines were observed in the superficial region of c-src -/- bone matrix. This indicates the possibility that in c-src -/- mice, osteoblasts were activated to deposit new bone matrices on the surfaces that osteoclasts previously passed along, even without bone resorption. Transmission electron microscopy demonstrated cell-to-cell contacts between mature osteoblasts and neighboring ruffled border-less osteoclasts, and osteoid including many mineralized nodules in c-src -/- mice. Thus, it seems likely that osteoblastic activities would be maintained in the presence of osteoclasts, even if they are dysfunctional.

Long-Term Administration of High-Fat Diet Corrects Abnormal Bone Remodeling in the Tibiae of Interleukin-6-Deficient Mice

PubMed Central

Feng, Wei; Liu, Bo; Liu, Di; Hasegawa, Tomoka; Wang, Wei; Han, Xiuchun; Cui, Jian; Yimin; Oda, Kimimitsu; Amizuka, Norio; Li, Minqi

2015-01-01

In this study, we aimed to evaluate the influence of diet-induced obesity on IL-6 deficiency-induced bone remodeling abnormality. Seven-week-old IL-6-/- mice and their wild type (WT) littermates were fed a standard diet (SD) or high-fat diet (HFD) for 25 weeks. Lipid formation and bone metabolism in mice tibiae were investigated by histochemical analysis. Both IL-6-/- and WT mice fed the HFD showed notable body weight gain, thickened cortical bones, and adipose accumulation in the bone marrow. Notably, the HFD normalized the bone phenotype of IL-6-/- mice to that of their WT counterpart, as characterized by a decrease in bone mass and the presence of an obliquely arranged, plate-like morphology in the trabecular bone. Alkaline phosphatase and osteocalcin expressions were attenuated in both genotypes after HFD feeding, especially for the IL-6-/- mice. Meanwhile, tartrate-resistant acid phosphatase staining was inhibited, osteoclast apoptosis rate down-regulated (revealed by TUNEL assay), and the proportion of cathepsin K (CK)-positive osteoclasts significantly increased in IL-6-/- mice on a HFD as compared with IL-6-/- mice on standard chow. Our results demonstrate that HFD-induced obesity reverses IL-6 deficiency-associated bone metabolic disorders by suppressing osteoblast activity, upregulating osteoclastic activity, and inhibiting osteoclast apoptosis. PMID:26416243

ACP5: its structure, distribution, regulation and novel functions.

PubMed

Ren, Xin; Shan, Wen-Hua; Wei, Lu-Lu; Gong, Chan-Chan; Pei, Dong-Sheng

2018-04-11

Tartrate-resistant acid phosphatase 5 (ACP5) is an evolutionarily conserved and multifunctional protein that is involved in generations of reactive oxygen species, normal bone development, osteoblast regulation and macrophage function, affecting a series of pathways, as well as reflecting bone resorption and osteoclast activity. Literature searches, systematic reviews and assessments about the structure, distribution, regulation and novel functions of ACP5 were performed in this review from PubMed and Medline databases. Studies demonstrate that RANKL can increase the expression of ACP5 through NFATc1 and c-Fos to accelerate osteoclastogenesis, which also can be regulated by many regulators. Based on aforementioned information, it is shown that ACP5, together with the phosphatase activity, can medicate the progression and development of human genetic diseases and cancer. As a novel target, ACP5 plays a critical role in preventing, monitoring and treating various kinds of tumors, as well as accelerating the development of a promising therapeutic strategy for human genetic diseases. However, the explicit mechanism between ACP5 and cancer is not so clear. It is necessary and significant for us to pay more in-depth attention. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Accelerated craniofacial bone regeneration through dense collagen gel scaffolds seeded with dental pulp stem cells

NASA Astrophysics Data System (ADS)

Chamieh, Frédéric; Collignon, Anne-Margaux; Coyac, Benjamin R.; Lesieur, Julie; Ribes, Sandy; Sadoine, Jérémy; Llorens, Annie; Nicoletti, Antonino; Letourneur, Didier; Colombier, Marie-Laure; Nazhat, Showan N.; Bouchard, Philippe; Chaussain, Catherine; Rochefort, Gael Y.

2016-12-01

Therapies using mesenchymal stem cell (MSC) seeded scaffolds may be applicable to various fields of regenerative medicine, including craniomaxillofacial surgery. Plastic compression of collagen scaffolds seeded with MSC has been shown to enhance the osteogenic differentiation of MSC as it increases the collagen fibrillary density. The aim of the present study was to evaluate the osteogenic effects of dense collagen gel scaffolds seeded with mesenchymal dental pulp stem cells (DPSC) on bone regeneration in a rat critical-size calvarial defect model. Two symmetrical full-thickness defects were created (5 mm diameter) and filled with either a rat DPSC-containing dense collagen gel scaffold (n = 15), or an acellular scaffold (n = 15). Animals were imaged in vivo by microcomputer tomography (Micro-CT) once a week during 5 weeks, whereas some animals were sacrificed each week for histology and histomorphometry analysis. Bone mineral density and bone micro-architectural parameters were significantly increased when DPSC-seeded scaffolds were used. Histological and histomorphometrical data also revealed significant increases in fibrous connective and mineralized tissue volume when DPSC-seeded scaffolds were used, associated with expression of type I collagen, osteoblast-associated alkaline phosphatase and osteoclastic-related tartrate-resistant acid phosphatase. Results demonstrate the potential of DPSC-loaded-dense collagen gel scaffolds to benefit of bone healing process.

Antiosteoclastic activity of milk thistle extract after ovariectomy to suppress estrogen deficiency-induced osteoporosis.

PubMed

Kim, Jung-Lye; Kim, Yun-Ho; Kang, Min-Kyung; Gong, Ju-Hyun; Han, Seoung-Jun; Kang, Young-Hee

2013-01-01

Bone integrity abnormality and imbalance between bone formation by osteoblasts and bone resorption by osteoclasts are known to result in metabolic bone diseases such as osteoporosis. Silymarin-rich milk thistle extract (MTE) and its component silibinin enhanced alkaline phosphatase activity of osteoblasts but reduced tartrate-resistant acid phosphatase (TRAP) activity of osteoclasts. The osteoprotective effects of MTE were comparable to those of estrogenic isoflavone. Low-dose combination of MTE and isoflavone had a pharmacological synergy that may be useful for osteogenic activity. This study attempted to reveal the suppressive effects of MTE on bone loss. C57BL/6 female mice were ovariectomized (OVX) as a model for postmenopausal osteopenia and orally administered 10 mg/kg MTE or silibinin for 8 weeks. The sham-operated mice served as estrogen controls. The treatment of ovariectomized mice with nontoxic MTE and silibinin improved femoral bone mineral density and serum receptor activator of nuclear factor- κB ligand/osteoprotegerin ratio, an index of osteoclastogenic stimulus. In addition, the administration of MTE or silibinin inhibited femoral bone loss induced by ovariectomy and suppressed femoral TRAP activity and cathepsin K induction responsible for osteoclastogenesis and bone resorption. Collectively, oral dosage of MTE containing silibinin in the preclinical setting is effective in preventing estrogen deficiency-induced bone loss.

Antiosteoclastic Activity of Milk Thistle Extract after Ovariectomy to Suppress Estrogen Deficiency-Induced Osteoporosis

PubMed Central

Kim, Jung-Lye; Kim, Yun-Ho; Kang, Min-Kyung; Gong, Ju-Hyun; Han, Seoung-Jun; Kang, Young-Hee

2013-01-01

Bone integrity abnormality and imbalance between bone formation by osteoblasts and bone resorption by osteoclasts are known to result in metabolic bone diseases such as osteoporosis. Silymarin-rich milk thistle extract (MTE) and its component silibinin enhanced alkaline phosphatase activity of osteoblasts but reduced tartrate-resistant acid phosphatase (TRAP) activity of osteoclasts. The osteoprotective effects of MTE were comparable to those of estrogenic isoflavone. Low-dose combination of MTE and isoflavone had a pharmacological synergy that may be useful for osteogenic activity. This study attempted to reveal the suppressive effects of MTE on bone loss. C57BL/6 female mice were ovariectomized (OVX) as a model for postmenopausal osteopenia and orally administered 10 mg/kg MTE or silibinin for 8 weeks. The sham-operated mice served as estrogen controls. The treatment of ovariectomized mice with nontoxic MTE and silibinin improved femoral bone mineral density and serum receptor activator of nuclear factor-κB ligand/osteoprotegerin ratio, an index of osteoclastogenic stimulus. In addition, the administration of MTE or silibinin inhibited femoral bone loss induced by ovariectomy and suppressed femoral TRAP activity and cathepsin K induction responsible for osteoclastogenesis and bone resorption. Collectively, oral dosage of MTE containing silibinin in the preclinical setting is effective in preventing estrogen deficiency-induced bone loss. PMID:23781510

Levels of eicosapentaenoic acid in obese schoolchildren with and without insulin resistance.

PubMed

Sánchez Meza, Karmina; Tene Pérez, Carlos Enrique; Sánchez Ramírez, Carmen Alicia; Muñiz Valencia, Roberto; Del Toro Equihua, Mario

2014-09-12

Obesity in children is now an increasing health risk worldwide in which the insulin-resistance can be present. Studies have linked a diet rich in n-3 fatty acids with a lower prevalence of insulin-resistance. To compare the levels of eicosapentaenoic acid among obese children with and without insulin-resistance. In 56 randomly school-age children with obesity, insulin-resistance was determined by the homeostasis model assessment for insulin-resistance index and the serum levels of eicosapentaenoic acid were determined by gas chromatography. Insulin-resistance was established when the index was >6.0, non- insulin- resistance when that index was within the range of 1.4-5.9. The serum levels of eicosapentaenoic acid were compared with the Kruskal-Wallis and Mann-Whitney U tests, as needed. No differences in age or sex were identified among the groups studied. The anthropometric parameters were significantly higher in the group of children with insulin-resistance than in the other two groups. The children with insulin- resistance had significantly lower levels of eicosapentaenoic acid than the non- insulin-resistance group [12.4% area under the curve vs. 37.4%, p = 0.031], respectively. Obese primary school-aged children with insulin-resistance had lower plasma levels of eicosapentaenoic acid. Copyright AULA MEDICA EDICIONES 2014. Published by AULA MEDICA. All rights reserved.

Phage & phosphatase: a novel phage-based probe for rapid, multi-platform detection of bacteria.

PubMed

Alcaine, S D; Pacitto, D; Sela, D A; Nugen, S R

2015-11-21

Genetic engineering of bacteriophages allows for the development of rapid, highly specific, and easily manufactured probes for the detection of bacterial pathogens. A challenge for novel probes is the ease of their adoption in real world laboratories. We have engineered the bacteriophage T7, which targets Escherichia coli, to carry the alkaline phosphatase gene, phoA. This inclusion results in phoA overexpression following phage infection of E. coli. Alkaline phosphatase is commonly used in a wide range of diagnostics, and thus a signal produced by our phage-based probe could be detected using common laboratory equipment. Our work demonstrates the successful: (i) modification of T7 phage to carry phoA; (ii) overexpression of alkaline phosphatase in E. coli; and (iii) detection of this T7-induced alkaline phosphatase activity using commercially available colorimetric and chemilumiscent methods. Furthermore, we demonstrate the application of our phage-based probe to rapidly detect low levels of bacteria and discern the antibiotic resistance of E. coli isolates. Using our bioengineered phage-based probe we were able to detect 10(3) CFU per mL of E. coli in 6 hours using a chemiluminescent substrate and 10(4) CFU per mL within 7.5 hours using a colorimetric substrate. We also show the application of this phage-based probe for antibiotic resistance testing. We were able to determine whether an E. coli isolate was resistant to ampicillin within 4.5 hours using chemiluminescent substrate and within 6 hours using a colorimetric substrate. This phage-based scheme could be readily adopted in labs without significant capital investments and can be translated to other phage-bacteria pairs for further detection.

ACP5 (Uteroferrin): Phylogeny of an Ancient and Conserved Gene Expressed in the Endometrium of Mammals1

PubMed Central

Padua, Maria B.; Lynch, Vincent J.; Alvarez, Natalia V.; Garthwaite, Mark A.; Golos, Thaddeus G.; Bazer, Fuller W.; Kalkunte, Satyan; Sharma, Surendra; Wagner, Gunter P.; Hansen, Peter J.

2012-01-01

ABSTRACT Type 5 acid phosphatase (ACP5; also known as tartrate-resistant acid phosphatase or uteroferrin) is a metalloprotein secreted by the endometrial glandular epithelium of pigs, mares, sheep, and water buffalo. In this paper, we describe the phylogenetic distribution of endometrial expression of ACP5 and demonstrate that endometrial expression arose early in evolution (i.e., before divergence of prototherian and therian mammals ∼166 million years ago). To determine expression of ACP5 in the pregnant endometrium, RNA was isolated from rhesus, mouse, rat, dog, sheep, cow, horse, armadillo, opossum, and duck-billed platypus. Results from RT-PCR and RNA-Seq experiments confirmed that ACP5 is expressed in all species examined. ACP5 was also demonstrated immunochemically in endometrium of rhesus, marmoset, sheep, cow, goat, and opossum. Alignment of inferred amino acid sequences shows a high conservation of ACP5 throughout speciation, with species-specific differences most extensive in the N-terminal and C-terminal regions of the protein. Analysis by Selecton indicated that most of the sites in ACP5 are undergoing purifying selection, and no sites undergoing positive selection were found. In conclusion, endometrial expression of ACP5 is a common feature in all orders of mammals and has been subjected to purifying selection. Expression of ACP5 in the uterus predates the divergence of therians and prototherians. ACP5 is an evolutionary conserved gene that likely exerts a common function important for pregnancy in mammals using a wide range of reproductive strategies. PMID:22278982

Altered cultivar resistance of kimchi cabbage seedlings mediated by salicylic Acid, jasmonic Acid and ethylene.

PubMed

Lee, Young Hee; Kim, Sang Hee; Yun, Byung-Wook; Hong, Jeum Kyu

2014-09-01

Two cultivars Buram-3-ho (susceptible) and CR-Hagwang (moderate resistant) of kimchi cabbage seedlings showed differential defense responses to anthracnose (Colletotrichum higginsianum), black spot (Alternaria brassicicola) and black rot (Xanthomonas campestris pv. campestris, Xcc) diseases in our previous study. Defense-related hormones salicylic acid (SA), jasmonic acid (JA) and ethylene led to different transcriptional regulation of pathogenesis-related (PR) gene expression in both cultivars. In this study, exogenous application of SA suppressed basal defenses to C. higginsianum in the 1st leaves of the susceptible cultivar and cultivar resistance of the 2nd leaves of the resistant cultivar. SA also enhanced susceptibility of the susceptible cultivar to A. brassicicola. By contrast, SA elevated disease resistance to Xcc in the resistant cultivar, but not in the susceptible cultivar. Methyl jasmonate (MJ) treatment did not affect the disease resistance to C. higginsianum and Xcc in either cultivar, but it compromised the disease resistance to A. brassicicola in the resistant cultivar. Treatment with 1-aminocyclopropane-1-carboxylic acid (ACC) ethylene precursor did not change resistance of the either cultivar to C. higginsianum and Xcc. Effect of ACC pretreatment on the resistance to A. brassicicola was not distinguished between susceptible and resistant cultivars, because cultivar resistance of the resistant cultivar was lost by prolonged moist dark conditions. Taken together, exogenously applied SA, JA and ethylene altered defense signaling crosstalk to three diseases of anthracnose, black spot and black rot in a cultivar-dependent manner.

Effects of the phosphatase inhibitors, okadaic acid, ATPgammaS, and calyculin A on the dividing sand dollar egg.

PubMed

Hamaguchi, Yukihisa; Kuriyama, Ryoko

2002-06-01

The effects of the phosphatase inhibitors, okadaic acid (OA), adenosine 5'-O-(3-thiotriphosphate) (ATPgammaS), and calyculin A (CL-A) on anaphase chromosome movement, cytokinesis, and cytoskeletal structures at cell division were examined by being microinjected into mitotic sand dollar eggs. When OA was injected, chromosome movement was inhibited and, moreover, chromosomes were ejected from the polar regions of the mitotic apparatus. By immunofluorescence, microtubules were observed to be severed in the OA-injected eggs, causing the smooth cell surface to be changed to an irregular surface. When ATPgammaS and CL-A were injected, the effect on cell shape was remarkable: In dividing eggs, furrowing stopped within several seconds after injection, small blebs appeared on the cell surface and became large, spherical or dumbbell cell shapes then changed to irregular forms, and subsequently cytoplasmic flow occurred. Microfilament detection revealed that actin accumulation in the cortex, which was not limited to the furrow cortex, occurred shortly after injection. Cortical accumulation of actin is thought to induce force generation and random cortical contraction, and accordingly to result in bleb extrusion from the cortex. Consequently, the phosphatase inhibitors inhibited the transition from mitosis to interphase by mediating cortical accumulation of actin filaments and/or fragmentation of microtubules.

A novel phosphatidic acid-protein-tyrosine phosphatase D2 axis is essential for ERBB2 signaling in mammary epithelial cells.

PubMed

Ramesh, Mathangi; Krishnan, Navasona; Muthuswamy, Senthil K; Tonks, Nicholas K

2015-04-10

We used a loss-of-function screen to investigate the role of classical protein-tyrosine phosphatases (PTPs) in three-dimensional mammary epithelial cell morphogenesis and ERBB2 signaling. The study revealed a novel role for PTPD2 as a positive regulator of ERBB2 signaling. Suppression of PTPD2 attenuated the ERBB2-induced multiacinar phenotype in three-dimensional cultures specifically by inhibiting ERBB2-mediated loss of polarity and lumen filling. In contrast, overexpression of PTPD2 enhanced the ERBB2 phenotype. We also found that a lipid second messenger, phosphatidic acid, bound PTPD2 in vitro and enhanced its catalytic activity. Small molecule inhibitors of phospholipase D (PLD), an enzyme that produces phosphatidic acid in cells, also attenuated the ERBB2 phenotype. Exogenously added phosphatidic acid rescued the PLD-inhibition phenotype, but only when PTPD2 was present. These findings illustrate a novel pathway involving PTPD2 and the lipid second messenger phosphatidic acid that promotes ERBB2 function. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Low-Molecular-Weight Protein Tyrosine Phosphatase Predicts Prostate Cancer Outcome by Increasing the Metastatic Potential.

PubMed

Ruela-de-Sousa, Roberta R; Hoekstra, Elmer; Hoogland, A Marije; Queiroz, Karla C Souza; Peppelenbosch, Maikel P; Stubbs, Andrew P; Pelizzaro-Rocha, Karin; van Leenders, Geert J L H; Jenster, Guido; Aoyama, Hiroshi; Ferreira, Carmen V; Fuhler, Gwenny M

2016-04-01

Low-risk patients suffering from prostate cancer (PCa) are currently placed under active surveillance rather than undergoing radical prostatectomy. However, clear parameters for selecting the right patient for each strategy are not available, and new biomarkers and treatment modalities are needed. Low-molecular-weight protein tyrosine phosphatase (LMWPTP) could present such a target. To correlate expression levels of LMWPTP in primary PCa to clinical outcome, and determine the role of LMWPTP in prostate tumor cell biology. Acid phosphatase 1, soluble (ACP1) expression was analyzed on microarray data sets, which were subsequently used in Ingenuity Pathway Analysis. Immunohistochemistry was performed on a tissue microarray containing material of 481 PCa patients whose clinicopathologic data were recorded. PCa cell line models were used to investigate the role of LMWPTP in cell proliferation, migration, adhesion, and anoikis resistance. The association between LMWPTP expression and clinical and pathologic outcomes was calculated using chi-square correlations and multivariable Cox regression analysis. Functional consequences of LMWPTP overexpression or downregulation were determined using migration and adhesion assays, confocal microscopy, Western blotting, and proliferation assays. LMWPTP expression was significantly increased in human PCa and correlated with earlier recurrence of disease (hazard ratio [HR]:1.99; p<0.001) and reduced patient survival (HR: 1.53; p=0.04). Unbiased Ingenuity analysis comparing cancer and normal prostate suggests migratory propensities in PCa. Indeed, overexpression of LMWPTP increases PCa cell migration, anoikis resistance, and reduces activation of focal adhesion kinase/paxillin, corresponding to decreased adherence. Overexpression of LMWPTP in PCa confers a malignant phenotype with worse clinical outcome. Prospective follow-up should determine the clinical potential of LMWPTP overexpression. These findings implicate low

Crystal growth and physical characterization of picolinic acid cocrystallized with dicarboxylic acids

NASA Astrophysics Data System (ADS)

Somphon, Weenawan; Haller, Kenneth J.

2013-01-01

Pharmaceutical cocrystals are multicomponent materials containing an active pharmaceutical ingredient with another component in well-defined stoichiometry within the same unit cell. Such cocrystals are important in drug design, particularly for improving physicochemical properties such as solubility, bioavailability, or chemical stability. Picolinic acid is an endogenous metabolite of tryptophan and is widely used for neuroprotective, immunological, and anti-proliferative effects within the body. In this paper we present cocrystallization experiments of a series of dicarboxylic acids, oxalic acid, succinic acid, DL-tartaric acid, pimelic acid, and phthalic acid, with picolinic acid. Characterization by FT-IR and Raman spectroscopy, DSC and TG/DTG analysis, and X-ray powder diffraction show that new compounds are formed, including a 1:1 picolinium tartrate monohydrate, a 2:1 monohydrate adduct of picolinic acid and oxalic acid, and a 2:1 picolinic acid-succinic acid monohydrate cocrystal.

Purification and characterization of the glycogen-bound protein phosphatase from rat liver.

PubMed

Wera, S; Bollen, M; Stalmans, W

1991-01-05

Glycogen-bound protein phosphatase G from rat liver was transferred from glycogen to beta-cyclodextrin (cycloheptaamylose) linked to Sepharose 6B. After removal of the catalytic subunit and of contaminating proteins with 2 M NaCl, elution with beta-cyclodextrin yielded a single protein on native polyacrylamide gel electrophoresis and two polypeptides (161 and 54 kDa) on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Several lines of evidence indicate that the latter polypeptides are subunits of the protein phosphatase G holoenzyme. First, these polypeptides were also present, together with the catalytic subunit, in the extensively purified holoenzyme. Also, polyclonal antibodies against these polypeptides were able to bind the holoenzyme. Further, while bound to cyclodextrin-Sepharose, the polypeptides were able to recombine with separately purified type-1 (AMD) catalytic subunit, but not with type-2A (PCS) catalytic subunit. The characteristics of the reconstituted enzyme resembled those of the nonpurified protein phosphatase G. At low dilutions, the spontaneous phosphorylase phosphatase activity of the reconstituted enzyme was about 10 times lower than that of the catalytic subunit, but it was about 1000-fold more resistant to inhibition by the modulator protein (inhibitor-2). In contrast with the free catalytic subunit, the reconstituted enzyme co-sedimented with glycogen, and it was able to activate purified liver glycogen synthase b. Also, the synthase phosphatase activity was synergistically increased by a cytosolic phosphatase and inhibited by physiological concentrations of phosphorylase alpha and of Ca2+.

Identification and characterization of a pyridoxal 5'-phosphate phosphatase in the silkworm (Bombyx mori).

PubMed

Huang, ShuoHao; Han, CaiYun; Ma, ZhenQiao; Zhou, Jie; Zhang, JianYun; Huang, LongQuan

2017-03-01

Vitamin B 6 comprises six interconvertible pyridine compounds, among which pyridoxal 5'-phosphate (PLP) is a coenzyme for over 140 enzymes. PLP is also a very reactive aldehyde. The most well established mechanism for maintaining low levels of free PLP is its dephosphorylation by phosphatases. A human PLP-specific phosphatase has been identified and characterized. However, very little is known about the phosphatase in other living organisms. In this study, a cDNA clone of putative PLP phosphatase was identified from B. mori and characterized. The cDNA encodes a polypeptide of 343 amino acid residues, and the recombinant enzyme purified from E. coli exhibited properties similar to that of human PLP phosphatase. B. mori has a single copy of the PLPP gene, which is located on 11th chromosome, spans a 5.7kb region and contains five exons and four introns. PLP phosphatase transcript was detected in every larva tissue except hemolymph, and was most highly represented in Malpighian tube. We further down-regulated the gene expression of the PLP phosphatase in 5th instar larvae with the RNA interference. However, no significant changes in the gene expression of PLP biosynthetic enzymes and composition of B 6 vitamers were detected as compared with the control. Copyright © 2017 Elsevier Inc. All rights reserved.

Differential regulation of the androgen receptor by protein phosphatase regulatory subunits

PubMed Central

Grey, James; Jones, Dominic; Wilson, Laura; Nakjang, Sirintra; Clayton, Jake; Temperley, Richard; Clark, Emma; Gaughan, Luke; Robson, Craig

2018-01-01

The Androgen Receptor (AR) is a key molecule in the development, maintenance and progression of prostate cancer (PC). However, the relationship between the AR and co-regulatory proteins that facilitate AR activity in castrate resistant settings remain understudied. Here we show that protein phosphatase 1 regulatory subunits, identified from a phosphatase RNAi screen, direct PP1 catalytic subunits to a varied yet significant response in AR function. As such, we have characterised the PP1β holoenzyme, myosin phosphatase (MLCP), as a novel ligand independent regulator of the AR. Sustained MLCP activity through down-regulation of the MLCP inhibitory subunit, PPP1R14C, results in impaired AR nuclear translocation, protein stability and transcriptional activity in distinct models of PC progression, culminating in restoration of a non-malignant prostate genotype. Phenotypically, a marked reduction in cell proliferation and migration, characterised by G1 cell cycle arrest is observed, confirming PP1 holoenzyme disruption as a novel treatment approach in PC. PMID:29423094

Dairy products and the French paradox: Could alkaline phosphatases play a role?

PubMed

Lallès, Jean-Paul

2016-07-01

The French paradox - high saturated fat consumption but low incidence of cardiovascular disease (CVD) and mortality - is still unresolved and continues to be a matter of debate and controversy. Recently, it was hypothesised that the high consumption of dairy products, and especially cheese by the French population might contribute to the explanation of the French paradox, in addition to the "(red) wine" hypothesis. Most notably this would involve milk bioactive peptides and biomolecules from cheese moulds. Here, we support the "dairy products" hypothesis further by proposing the "alkaline phosphatase" hypothesis. First, intestinal alkaline phosphatase (IAP), a potent endogenous anti-inflammatory enzyme, is directly stimulated by various components of milk (e.g. casein, calcium, lactose and even fat). This enzyme dephosphorylates and thus detoxifies pro-inflammatory microbial components like lipopolysaccharide, making them unable to trigger inflammatory responses and generate chronic low-grade inflammation leading to insulin resistance, glucose intolerance, type-2 diabetes, metabolic syndrome and obesity, known risk factors for CVD. Various vitamins present in high amounts in dairy products (e.g. vitamins A and D; methyl-donors: folate and vitamin B12), and also fermentation products such as butyrate and propionate found e.g. in cheese, all stimulate intestinal alkaline phosphatase. Second, moulded cheeses like Roquefort contain fungi producing an alkaline phosphatase. Third, milk itself contains a tissue nonspecific isoform of alkaline phosphatase that may function as IAP. Milk alkaline phosphatase is present in raw milk and dairy products increasingly consumed in France. It is deactivated by pasteurization but it can partially reactivate after thermal treatment. Experimental consolidation of the "alkaline phosphatase" hypothesis will require further work including: systematic alkaline phosphatase activity measurements in dairy products, live dairy ferments and

Effect of aspartic acid and glutamate on metabolism and acid stress resistance of Acetobacter pasteurianus.

PubMed

Yin, Haisong; Zhang, Renkuan; Xia, Menglei; Bai, Xiaolei; Mou, Jun; Zheng, Yu; Wang, Min

2017-06-15

Acetic acid bacteria (AAB) are widely applied in food, bioengineering and medicine fields. However, the acid stress at low pH conditions limits acetic acid fermentation efficiency and high concentration of vinegar production with AAB. Therefore, how to enhance resistance ability of the AAB remains as the major challenge. Amino acids play an important role in cell growth and cell survival under severe environment. However, until now the effects of amino acids on acetic fermentation and acid stress resistance of AAB have not been fully studied. In the present work the effects of amino acids on metabolism and acid stress resistance of Acetobacter pasteurianus were investigated. Cell growth, culturable cell counts, acetic acid production, acetic acid production rate and specific production rate of acetic acid of A. pasteurianus revealed an increase of 1.04, 5.43, 1.45, 3.30 and 0.79-folds by adding aspartic acid (Asp), and cell growth, culturable cell counts, acetic acid production and acetic acid production rate revealed an increase of 0.51, 0.72, 0.60 and 0.94-folds by adding glutamate (Glu), respectively. For a fully understanding of the biological mechanism, proteomic technology was carried out. The results showed that the strengthening mechanism mainly came from the following four aspects: (1) Enhancing the generation of pentose phosphates and NADPH for the synthesis of nucleic acid, fatty acids and glutathione (GSH) throughout pentose phosphate pathway. And GSH could protect bacteria from low pH, halide, oxidative stress and osmotic stress by maintaining the viability of cells through intracellular redox equilibrium; (2) Reinforcing deamination of amino acids to increase intracellular ammonia concentration to maintain stability of intracellular pH; (3) Enhancing nucleic acid synthesis and reparation of impaired DNA caused by acid stress damage; (4) Promoting unsaturated fatty acids synthesis and lipid transport, which resulted in the improvement of cytomembrane

Tomato PYR/PYL/RCAR abscisic acid receptors show high expression in root, differential sensitivity to the abscisic acid agonist quinabactin, and the capability to enhance plant drought resistance.

PubMed

González-Guzmán, Miguel; Rodríguez, Lesia; Lorenzo-Orts, Laura; Pons, Clara; Sarrión-Perdigones, Alejandro; Fernández, Maria A; Peirats-Llobet, Marta; Forment, Javier; Moreno-Alvero, Maria; Cutler, Sean R; Albert, Armando; Granell, Antonio; Rodríguez, Pedro L

2014-08-01

Abscisic acid (ABA) plays a crucial role in the plant's response to both biotic and abiotic stress. Sustainable production of food faces several key challenges, particularly the generation of new varieties with improved water use efficiency and drought tolerance. Different studies have shown the potential applications of Arabidopsis PYR/PYL/RCAR ABA receptors to enhance plant drought resistance. Consequently the functional characterization of orthologous genes in crops holds promise for agriculture. The full set of tomato (Solanum lycopersicum) PYR/PYL/RCAR ABA receptors have been identified here. From the 15 putative tomato ABA receptors, 14 of them could be grouped in three subfamilies that correlated well with corresponding Arabidopsis subfamilies. High levels of expression of PYR/PYL/RCAR genes was found in tomato root, and some genes showed predominant expression in leaf and fruit tissues. Functional characterization of tomato receptors was performed through interaction assays with Arabidopsis and tomato clade A protein phosphatase type 2Cs (PP2Cs) as well as phosphatase inhibition studies. Tomato receptors were able to inhibit the activity of clade A PP2Cs differentially in an ABA-dependent manner, and at least three receptors were sensitive to the ABA agonist quinabactin, which inhibited tomato seed germination. Indeed, the chemical activation of ABA signalling induced by quinabactin was able to activate stress-responsive genes. Both dimeric and monomeric tomato receptors were functional in Arabidopsis plant cells, but only overexpression of monomeric-type receptors conferred enhanced drought resistance. In summary, gene expression analyses, and chemical and transgenic approaches revealed distinct properties of tomato PYR/PYL/RCAR ABA receptors that might have biotechnological implications. © The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Genetic variation of an acid phosphatase (Acp-2) in the laboratory rat: possible homology with mouse AP-1 and human ACP2.

PubMed

Bender, K; Bissbort, S; Kuhn, A; Nagel, M; Günther, E

1986-02-01

A genetic locus controlling the electrophoretic mobility of an acid phosphatase in the rat (Rattus norvegicus) is described. The locus, designed Acp-2, is not expressed in erythrocytes but is expressed in all other tissues studied. The product of Acp-2 hydrolyzes a wide variety of phosphate monoesters and is inhibited by L(+)-tartaric acid. Inbred rat strains have fixed either allele Acp-2a or allele Acp-2b. Codominant expression is observed in the respective F1 hybrids. Backcross progenies revealed the expected 1:1 segregation ratio. Possible loose linkage was found between the Acp-2 and the Pep-3 gene loci at a recombination frequency of 0.36 +/- 0.06.

Abscisic Acid-Cytokinin Antagonism Modulates Resistance Against Pseudomonas syringae in Tobacco.

PubMed

Großkinsky, Dominik K; van der Graaff, Eric; Roitsch, Thomas

2014-12-01

Phytohormones are known as essential regulators of plant defenses, with ethylene, jasmonic acid, and salicylic acid as the central immunity backbone, while other phytohormones have been demonstrated to interact with this. Only recently, a function of the classic phytohormone cytokinin in plant immunity has been described in Arabidopsis, rice, and tobacco. Although interactions of cytokinins with salicylic acid and auxin have been indicated, the complete network of cytokinin interactions with other immunity-relevant phytohormones is not yet understood. Therefore, we studied the interaction of kinetin and abscisic acid as a negative regulator of plant immunity to modulate resistance in tobacco against Pseudomonas syringae. By analyzing infection symptoms, pathogen proliferation, and accumulation of the phytoalexin scopoletin as a key mediator of kinetin-induced resistance in tobacco, antagonistic interaction of these phytohormones in plant immunity was identified. Kinetin reduced abscisic acid levels in tobacco, while increased abscisic acid levels by exogenous application or inhibition of abscisic acid catabolism by diniconazole neutralized kinetin-induced resistance. Based on these results, we conclude that reduction of abscisic acid levels by enhanced abscisic acid catabolism strongly contributes to cytokinin-mediated resistance effects. Thus, the identified cytokinin-abscisic acid antagonism is a novel regulatory mechanism in plant immunity.

Dietary phosphate restriction normalizes biochemical and skeletal abnormalities in a murine model of tumoral calcinosis.

PubMed

Ichikawa, Shoji; Austin, Anthony M; Gray, Amie K; Allen, Matthew R; Econs, Michael J

2011-12-01

Mutations in the GALNT3 gene cause tumoral calcinosis characterized by ectopic calcifications due to persistent hyperphosphatemia. We recently developed Galnt3 knockout mice in a mixed background, which had hyperphosphatemia with increased bone mineral density (BMD) and infertility in males. To test the effect of dietary phosphate intake on their phenotype, Galnt3 knockout mice were generated in the C57BL/6J strain and fed various phosphate diets: 0.1% (low), 0.3% (low normal), 0.6% (normal), and 1.65% (high). Sera were analyzed for calcium, phosphorus, alkaline phosphatase, creatinine, blood urine nitrogen, 1,25-dihydroxyvitamin D, osteocalcin, tartrate-resistant acid phosphatase 5b, and fibroblast growth factor 23 (Fgf23). Femurs were evaluated by dual-energy x-ray absorptiometry, dynamic histomorphometry, and/or microcomputed tomography. Galnt3 knockout mice in C57BL/6J had the same biochemical phenotype observed in our previous study: hyperphosphatemia, inappropriately normal 1,25-dihydroxyvitamin D level, decreased alkaline phosphatase activity, and low intact Fgf23 concentration but high Fgf23 fragments. Skeletal analyses of their femurs revealed significantly high BMD with increased cortical bone area and trabecular bone volume. On all four phosphate diets, Galnt3 knockout mice had consistently higher phosphorus levels and lower alkaline phosphatase and intact Fgf23 concentrations than littermate controls. The low-phosphate diet normalized serum phosphorus, alkaline phosphatase, and areal BMD but failed to correct male infertility in Galnt3 knockout mice. The high-phosphate diet did not increase serum phosphorus concentration in either mutant or control mice due to a compensatory increase in circulating intact Fgf23 levels. In conclusion, dietary phosphate restriction normalizes biochemical and skeletal phenotypes of Galnt3 knockout mice and, thus, can be an effective therapy for tumoral calcinosis.

Dietary Phosphate Restriction Normalizes Biochemical and Skeletal Abnormalities in a Murine Model of Tumoral Calcinosis

PubMed Central

Austin, Anthony M.; Gray, Amie K.; Allen, Matthew R.; Econs, Michael J.

2011-01-01

Mutations in the GALNT3 gene cause tumoral calcinosis characterized by ectopic calcifications due to persistent hyperphosphatemia. We recently developed Galnt3 knockout mice in a mixed background, which had hyperphosphatemia with increased bone mineral density (BMD) and infertility in males. To test the effect of dietary phosphate intake on their phenotype, Galnt3 knockout mice were generated in the C57BL/6J strain and fed various phosphate diets: 0.1% (low), 0.3% (low normal), 0.6% (normal), and 1.65% (high). Sera were analyzed for calcium, phosphorus, alkaline phosphatase, creatinine, blood urine nitrogen, 1,25-dihydroxyvitamin D, osteocalcin, tartrate-resistant acid phosphatase 5b, and fibroblast growth factor 23 (Fgf23). Femurs were evaluated by dual-energy x-ray absorptiometry, dynamic histomorphometry, and/or microcomputed tomography. Galnt3 knockout mice in C57BL/6J had the same biochemical phenotype observed in our previous study: hyperphosphatemia, inappropriately normal 1,25-dihydroxyvitamin D level, decreased alkaline phosphatase activity, and low intact Fgf23 concentration but high Fgf23 fragments. Skeletal analyses of their femurs revealed significantly high BMD with increased cortical bone area and trabecular bone volume. On all four phosphate diets, Galnt3 knockout mice had consistently higher phosphorus levels and lower alkaline phosphatase and intact Fgf23 concentrations than littermate controls. The low-phosphate diet normalized serum phosphorus, alkaline phosphatase, and areal BMD but failed to correct male infertility in Galnt3 knockout mice. The high-phosphate diet did not increase serum phosphorus concentration in either mutant or control mice due to a compensatory increase in circulating intact Fgf23 levels. In conclusion, dietary phosphate restriction normalizes biochemical and skeletal phenotypes of Galnt3 knockout mice and, thus, can be an effective therapy for tumoral calcinosis. PMID:22009723

Nd:YAG laser ablation and acid resistance of enamel.

PubMed

Kwon, Yong Hoon; Kwon, Oh-Won; Kim, Hyung-Il; Kim, Kyo-Han

2003-09-01

The acid resistance of Nd:YAG laser-ablated enamel surfaces was studied by evaluating crystal structure, mineral distribution, and fluorescence radiance and image in the present study. For comparison, 37% phosphoric acid etching was performed. The formation of beta-tricalcium phosphate (beta-TCP) was confirmed in the laser-ablated surface. The Ca/P ratio increased after ablation due to mineral re-distribution. In contrast, the Ca/P ratio decreased after acid etching due to mineral loss. The laser-ablated enamels showed a smaller increase of fluorescence radiances and less clear laser confocal scanning microscope images than those observed in the acid-etched enamels. The former suggests a minimized mineral loss. The Nd:YAG laser irradiation will enhance the acid resistance and retard the carious progression in enamel.

Hydrofluoric acid-resistant composite window and method for its fabrication

DOEpatents

Ostenak, C.A.; Mackay, H.A.

1985-07-18

A hydrofluoric acid-resistant composite window and method for its fabrication are disclosed. The composite window comprises a window having first and second sides. The first side is oriented towards an environment containing hydrofluoric acid. An adhesive is applied to the first side. A layer of transparent hydrofluoric acid-resistant material, such as Mylar, is applied to the adhesive and completely covers the first side. The adhesive is then cured.

Hydrofluoric acid-resistant composite window and method for its fabrication

DOEpatents

Ostenak, Carl A.; Mackay, Harold A.

1987-01-01

A hydrofluoric acid-resistant composite window and method for its fabrication are disclosed. The composite window comprises a window having first and second sides. The first side is oriented towards an environment containing hydrofluoric acid. An adhesive is applied to the first side. A layer of transparent hydrofluoric acid-resistant material, such as Mylar, is applied to the adhesive and completely covers the first side. The adhesive is then cured.

Phylogenetic and genetic linkage between novel atypical dual-specificity phosphatases from non-metazoan organisms.

PubMed

Romá-Mateo, Carlos; Sacristán-Reviriego, Almudena; Beresford, Nicola J; Caparrós-Martín, José Antonio; Culiáñez-Macià, Francisco A; Martín, Humberto; Molina, María; Tabernero, Lydia; Pulido, Rafael

2011-04-01

Dual-specificity phosphatases (DSPs) constitute a large protein tyrosine phosphatase (PTP) family, with examples in distant evolutive phyla. PFA-DSPs (Plant and Fungi Atypical DSPs) are a group of atypical DSPs present in plants, fungi, kinetoplastids, and slime molds, the members of which share structural similarity with atypical- and lipid phosphatase DSPs from mammals. The analysis of the PFA-DSPs from the plant Arabidopsis thaliana (AtPFA-DSPs) showed differential tissue mRNA expression, substrate specificity, and catalytic activity for these proteins, suggesting different functional roles among plant PFA-DSPs. Bioinformatic analysis revealed the existence of novel PFA-DSP-related proteins in fungi (Oca1, Oca2, Oca4 and Oca6 in Saccharomyces cerevisiae) and protozoa, which were segregated from plant PFA-DSPs. The closest yeast homolog for these proteins was the PFA-DSP from S. cerevisiae ScPFA-DSP1/Siw14/Oca3. Oca1, Oca2, Siw14/Oca3, Oca4, and Oca6 were involved in the yeast response to caffeine and rapamycin stresses. Siw14/Oca3 was an active phosphatase in vitro, whereas no phosphatase activity could be detected for Oca1. Remarkably, overexpression of Siw14/Oca3 suppressed the caffeine sensitivity of oca1, oca2, oca4, and oca6 deleted strains, indicating a genetic linkage and suggesting a functional relationship for these proteins. Functional studies on mutations targeting putative catalytic residues from the A. thaliana AtPFA-DSP1/At1g05000 protein indicated the absence of canonical amino acids acting as the general acid/base in the phosphor-ester hydrolysis, which suggests a specific mechanism of reaction for PFA-DSPs and related enzymes. Our studies demonstrate the existence of novel phosphatase protein families in fungi and protozoa, with active and inactive enzymes linked in common signaling pathways. This illustrates the catalytic and functional complexity of the expanding family of atypical dual-specificity phosphatases in non-metazoans, including

Inhibition of AcpA phosphatase activity with ascorbate attenuates Francisella tularensis intramacrophage survival.

PubMed

McRae, Steven; Pagliai, Fernando A; Mohapatra, Nrusingh P; Gener, Alejandro; Mahmou, Asma Sayed Abdelgeliel; Gunn, John S; Lorca, Graciela L; Gonzalez, Claudio F

2010-02-19

Acid phosphatase activity in the highly infectious intracellular pathogen Francisella tularensis is directly related with the ability of these bacteria to survive inside host cells. Pharmacological inactivation of acid phosphatases could potentially help in the treatment of tularemia or even be utilized to neutralize the infection. In the present work, we report inhibitory compounds for three of the four major acid phosphatases produced by F. tularensis SCHU4: AcpA, AcpB, and AcpC. The inhibitors were identified using a catalytic screen from a library of chemicals approved for use in humans. The best results were obtained against AcpA. The two compounds identified, ascorbate (K(i) = 380 +/- 160 microM) and 2-phosphoascorbate (K(i) = 3.2 +/- 0.85 microM) inhibit AcpA in a noncompetitive, nonreversible fashion. A potential ascorbylation site in the proximity of the catalytic pocket of AcpA was identified using site-directed mutagenesis. The effects of the inhibitors identified in vitro were evaluated using bioassays determining the ability of F. tularensis to survive inside infected cells. The presence of ascorbate or 2-phosphoascorbate impaired the intramacrophage survival of F. tularensis in an AcpA-dependent manner as it was probed using knockout strains. The evidence presented herein indicated that ascorbate could be a good alternative to be used clinically to improve treatments against tularemia.

Inhibition of AcpA Phosphatase Activity with Ascorbate Attenuates Francisella tularensis Intramacrophage Survival

PubMed Central

McRae, Steven; Pagliai, Fernando A.; Mohapatra, Nrusingh P.; Gener, Alejandro; Abdelgeliel Mahmou, Asma Sayed; Gunn, John S.; Lorca, Graciela L.; Gonzalez, Claudio F.

2010-01-01

Acid phosphatase activity in the highly infectious intracellular pathogen Francisella tularensis is directly related with the ability of these bacteria to survive inside host cells. Pharmacological inactivation of acid phosphatases could potentially help in the treatment of tularemia or even be utilized to neutralize the infection. In the present work, we report inhibitory compounds for three of the four major acid phosphatases produced by F. tularensis SCHU4: AcpA, AcpB, and AcpC. The inhibitors were identified using a catalytic screen from a library of chemicals approved for use in humans. The best results were obtained against AcpA. The two compounds identified, ascorbate (Ki = 380 ± 160 μm) and 2-phosphoascorbate (Ki = 3.2 ± 0.85 μm) inhibit AcpA in a noncompetitive, nonreversible fashion. A potential ascorbylation site in the proximity of the catalytic pocket of AcpA was identified using site-directed mutagenesis. The effects of the inhibitors identified in vitro were evaluated using bioassays determining the ability of F. tularensis to survive inside infected cells. The presence of ascorbate or 2-phosphoascorbate impaired the intramacrophage survival of F. tularensis in an AcpA-dependent manner as it was probed using knockout strains. The evidence presented herein indicated that ascorbate could be a good alternative to be used clinically to improve treatments against tularemia. PMID:20028980

Low bioavailability of ergotamine tartrate after oral and rectal administration in migraine sufferers.

PubMed Central

Ibraheem, J J; Paalzow, L; Tfelt-Hansen, P

1983-01-01

Fifteen migraine patients were administered 2 mg ergotamine tartrate in a partial cross-over design as a single, oral tablet, rectal suppository and rectal solution. Eight of these patients were in a previous investigation given 0.5 mg ergotamine tartrate intravenously. The blood samples were taken up to 54 h after oral and suppository while it was followed for only 3 h after rectal solution. The chemical analysis was performed by applying h.p.l.c. method with a limit of sensitivity of 0.1 ng/ml ergotamine base in plasma. No ergotamine was detected in the blood samples after the oral route, whereas small and very variable quantities was found in blood after the rectal route. Regular calculation of bioavailability could therefore not be performed. An estimate of the maximal possible bioavailability was found to yield a mean value of 2% (tablets); 5% (suppositories) and 6% (rectal solution). Rectal solution elicited faster absorption and the extent of absorption was significantly higher (P less than 0.05) than for the suppository. PMID:6419759

Spinal cord projections of the rat main forelimb nerves, studied by transganglionic transport of WGA-HRP and by the disappearance of acid phosphatase.

PubMed

Castro-Lopes, J M; Coimbra, A

1991-03-01

The spinal cord projections of the 3 main forelimb nerves-median, radial and ulnar, were studied in the rat dorsal horn with transganglionic transport of wheat germ agglutinin-horseradish peroxidase (WGA-HRP), or using the disappearance of fluoride resistant acid phosphatase (FRAP) after nerve section. The projection patterns in lamina II were similar following the two procedures. The median and the radial nerve fibers projected to the medial and the intermediate thirds, respectively, of the dorsal horn lamina II in spinal cord segments C4-C8. The ulnar nerve projected to segments C6-C8 between the areas occupied by the other two nerves. The FRAP method also showed that the lateral part of lamina II, which was not filled by radial nerve fibers, received the projections from the dorsal cutaneous branches of cervical spinal nerves. In addition, FRAP disappeared from the medial end of segment T1 after skin incisions extending from the medial brachium to the axilla, which seemed due to severance of the cutaneous branchlets of the lateral anterior thoracic nerve. The FRAP procedure is thus sensitive enough to detect fibers in lamina II arising from small peripheral nerves, and may be used as an alternative to the anterograde tracing methods whenever there are no overlapping projections.

Ferulic acid reverses ABCB1-mediated paclitaxel resistance in MDR cell lines.

PubMed

Muthusamy, Ganesan; Balupillai, Agilan; Ramasamy, Karthikeyan; Shanmugam, Mohana; Gunaseelan, Srithar; Mary, Beaulah; Prasad, N Rajendra

2016-09-05

Multidrug resistance (MDR) remains a major obstacle in cancer chemotherapy. The use of the dietary phytochemicals as chemosensitizing agents to enhance the efficacy of conventional cytostatic drugs has recently gained the attention as a plausible approach for overcoming the drug resistance. The aim of this study was to investigate whether a naturally occurring diet-based phenolic acid, ferulic acid, could sensitize paclitaxel efficacy in ABCB1 overexpressing (P-glycoprotein) colchicine selected KB Ch(R)8-5 cell line. In vitro drug efflux assays demonstrated that ferulic acid inhibits P-glycoprotein transport function in drug resistant KB Ch(R)8-5 cell lines. However, ferulic acid significantly downregulates ABCB1 expression in a concentration dependent manner. Cytotoxicity assay reveals that ferulic acid decreased paclitaxel resistance in KBCh(R)8-5 and HEK293/ABCB1 cells, which indicates its chemosensitizing potential. Clonogenic cell survival assay and apoptotic morphological staining further confirm the chemosensitizing potential of ferulic acid in drug resistant KB Ch(R)8-5 cell lines. Ferulic acid treatment enhances paclitaxel mediated cell cycle arrest and upregulates paclitaxel-induced apoptotic signaling in KB resistant cells. Hence, it has been concluded that downregulation of ABCB1 and subsequent induction of paclitaxel-mediated cell cycle arrest and apoptotic signaling may be the cause for the chemosensitizing potential of ferulic acid in P-gp overexpressing cell lines. Copyright © 2016 Elsevier B.V. All rights reserved.

DELAY OF GERMINATION1 requires PP2C phosphatases of the ABA signalling pathway to control seed dormancy.

PubMed

Née, Guillaume; Kramer, Katharina; Nakabayashi, Kazumi; Yuan, Bingjian; Xiang, Yong; Miatton, Emma; Finkemeier, Iris; Soppe, Wim J J

2017-07-13

The time of seed germination is a major decision point in the life of plants determining future growth and development. This timing is controlled by seed dormancy, which prevents germination under favourable conditions. The plant hormone abscisic acid (ABA) and the protein DELAY OF GERMINATION 1 (DOG1) are essential regulators of dormancy. The function of ABA in dormancy is rather well understood, but the role of DOG1 is still unknown. Here, we describe four phosphatases that interact with DOG1 in seeds. Two of them belong to clade A of type 2C protein phosphatases: ABA-HYPERSENSITIVE GERMINATION 1 (AHG1) and AHG3. These phosphatases have redundant but essential roles in the release of seed dormancy epistatic to DOG1. We propose that the ABA and DOG1 dormancy pathways converge at clade A of type 2C protein phosphatases.The DOG1 protein is a major regulator of seed dormancy in Arabidopsis. Here, Née et al. provide evidence that DOG1 can interact with the type 2C protein phosphatases AHG1 and AHG3 and that this represents the convergence point of the DOG1-regulated dormancy pathway and signalling by the plant hormone abscisic acid.

A New Protein Phosphatase 2C (FsPP2C1) Induced by Abscisic Acid Is Specifically Expressed in Dormant Beechnut Seeds1

PubMed Central

Lorenzo, Oscar; Rodríguez, Dolores; Nicolás, Gregorio; Rodríguez, Pedro L.; Nicolás, Carlos

2001-01-01

An abscisic acid (ABA)-induced cDNA fragment encoding a putative protein phosphatase 2C (PP2C) was obtained by means of differential reverse transcriptase-polymerase chain reaction approach. The full-length clone was isolated from a cDNA library constructed using mRNA from ABA-treated beechnut (Fagus sylvatica) seeds. This clone presents all the features of plant type PP2C and exhibits homology to members of this family such as AthPP2CA (61%), ABI1 (48%), or ABI2 (47%), therefore it was named FsPP2C1. The expression of FsPP2C1 is detected in dormant seeds and increases after ABA treatment, when seeds are maintained dormant, but it decreases and tends to disappear when dormancy is being released by stratification or under gibberellic acid treatment. Moreover, drought stress seems to have no effect on FsPP2C1 transcript accumulation. The FsPP2C1 transcript expression is tissue specific and was found to accumulate in ABA-treated seeds rather than in other ABA-treated vegetative tissues examined. These results suggest that the corresponding protein could be related to ABA-induced seed dormancy. By expressing FsPP2C1 in Escherichia coli as a histidine tag fusion protein, we have obtained direct biochemical evidence supporting Mg2+-dependent phosphatase activity of this protein. PMID:11299374

Moraxella catarrhalis Synthesizes an Autotransporter That Is an Acid Phosphatase▿

PubMed Central

Hoopman, Todd C.; Wang, Wei; Brautigam, Chad A.; Sedillo, Jennifer L.; Reilly, Thomas J.; Hansen, Eric J.

2008-01-01

Moraxella catarrhalis O35E was shown to synthesize a 105-kDa protein that has similarity to both acid phosphatases and autotransporters. The N-terminal portion of the M. catarrhalis acid phosphatase A (MapA) was most similar (the BLAST probability score was 10−10) to bacterial class A nonspecific acid phosphatases. The central region of the MapA protein had similarity to passenger domains of other autotransporter proteins, whereas the C-terminal portion of MapA resembled the translocation domain of conventional autotransporters. Cloning and expression of the M. catarrhalis mapA gene in Escherichia coli confirmed the presence of acid phosphatase activity in the MapA protein. The MapA protein was shown to be localized to the outer membrane of M. catarrhalis and was not detected either in the soluble cytoplasmic fraction from disrupted M. catarrhalis cells or in the spent culture supernatant fluid from M. catarrhalis. Use of the predicted MapA translocation domain in a fusion construct with the passenger domain from another predicted M. catarrhalis autotransporter confirmed the translocation ability of this MapA domain. Inactivation of the mapA gene in M. catarrhalis strain O35E reduced the acid phosphatase activity expressed by this organism, and this mutation could be complemented in trans with the wild-type mapA gene. Nucleotide sequence analysis of the mapA gene from six M. catarrhalis strains showed that this protein was highly conserved among strains of this pathogen. Site-directed mutagenesis of a critical histidine residue (H233A) in the predicted active site of the acid phosphatase domain in MapA eliminated acid phosphatase activity in the recombinant MapA protein. This is the first description of an autotransporter protein that expresses acid phosphatase activity. PMID:18065547

Enantioselective toxicities of chiral ionic liquids 1-alkyl-3-methyl imidazolium tartrate on Scenedesmus obliquus.

PubMed

Liu, Huijun; Zhang, Xiaoqiang; Dong, Ying; Chen, Caidong; Zhu, Shimin; Ma, Xiangjuan

2015-12-01

Ionic liquids (ILs) are being used in various industries during the last few decades, while the good solubility and high stability of ILs may pose a potential threat to the aquatic environment. Effect of chiral ionic liquids (CILs) 1-alkyl-3-methyl imidazolium tartrate (RMIM T) on Scenedesmus obliquus (S.obliquus) was studied. The growth rate inhibition and cell membrane permeability increased with increasing RMIM T concentration and increasing alkyl chain lengths. The IC50 values of D-(-)-tartrate 1-hexyl-3-methyl imidazolium (D-(-)-HMIM T) were 28.30, 12.23,10.15 and 14.41 mg/L, respectively, at 24, 48, 72 and 96h. While that of L-(+)-tartrate 1-hexyl-3-methyl imidazolium (L-(+)-HMIM T) were 15.97, 7.91, 9.43 and 12.04 mg/L respectively. The concentration of chl a, chl b and chl (a+b) decreased with increasing RMIM T concentration. The chlorophyll fluorescence parameters (F0, Fv/Fm, Fv/F0, Y(II), ETR and NPQ) were affected by RMIM T, indicating that the RMIM T will damage the PSII, inhibit the transmission of excitation energy, decrease the efficiency of photosynthesis. The results showed that there were enantioselective toxicity of RMIM T to algae, and the toxicity of L-(+)-RMIM T was greater than that of D-(-)-RMIM T, but the enantioselective difference becomes smaller with increasing exposure time, and with the increasing carbon chain length of cation, indicating that cation properties may have a larger effect on toxicity than anion properties. Copyright © 2015 Elsevier B.V. All rights reserved.

Hypolipidemic and hypoglycemic activities of a oleanolic acid derivative from Malva parviflora on streptozotocin-induced diabetic mice.

PubMed

Gutiérrez, Rosa Martha Pérez

2017-05-01

One new oleanolic acid derivative, 2α,3β,23α,29α tetrahydroxyolean-12(13)-en-28-oic acid (1) was isolated from the aerial parts of Malva parviflora. Their structure was characterized by spectroscopic methods. The hypolipidemic and hypoglycemic activities of 1 was analyzed in in streptozotocin (STZ)-nicotinamide-induced type 2 diabetes in mice (MD) and type 1 diabetes in streptozotocin-induced diabetic mice (SD). Triterpene was administered orally at doses of 20 mg/kg for 4 weeks. Organ weight, body weight, glucose, fasting insulin, cholesterol-related lipid profile parameters, glutamate oxaloacetate transaminase (SGOT), glutamate pyruvate transaminase (SGPT), serum alkaline phosphatase (SALP), glucokinase, hexokinase, glucose-6-phosphatase activities and glycogen in liver were measured after 4 weeks of treatment. The results indicated that 1 regulate glucose metabolism, lipid profile, lipid peroxidation, increased body weight, glucokinase and hexokinase activities inhibited triglycerides, total cholesterol, low density lipoproteins level, SGOT, SGPT, SALP, glycogen in liver and glucose-6-phosphatase. In addition, improvement of insulin resistance and protective effect for pancreatic β-cells, also 1 may changes the expression of pro-inflammatory cytokine (IL-6 and TNF-α levels) and enzymes (PAL2, COX-2, and LOX). The results suggest that 1 has hypolipidemic and hypoglycemic, anti-inflammatory, activities, improve insulin resistance and hepatic enzymes in streptozotocin-induced diabetic mice.

The Role of PTP1B O-GlcNAcylation in Hepatic Insulin Resistance.

PubMed

Zhao, Yun; Tang, Zhuqi; Shen, Aiguo; Tao, Tao; Wan, Chunhua; Zhu, Xiaohui; Huang, Jieru; Zhang, Wanlu; Xia, Nana; Wang, Suxin; Cui, Shiwei; Zhang, Dongmei

2015-09-22

Protein tyrosine phosphatase 1B (PTP1B), which can directly dephosphorylate both the insulin receptor and insulin receptor substrate 1 (IRS-1), thereby terminating insulin signaling, reportedly plays an important role in insulin resistance. Accumulating evidence has demonstrated that O-GlcNAc modification regulates functions of several important components of insulin signal pathway. In this study, we identified that PTP1B is modified by O-GlcNAcylation at three O-GlcNAc sites (Ser104, Ser201, and Ser386). Palmitate acid (PA) impaired the insulin signaling, indicated by decreased phosphorylation of both serine/threonine-protein kinase B (Akt) and glycogen synthase kinase 3 beta (GSK3β) following insulin administration, and upregulated PTP1B O-GlcNAcylation in HepG2 cells. Compared with the wild-type, intervention PTP1B O-GlcNAcylation by site-directed gene mutation inhibited PTP1B phosphatase activity, resulted in a higher level of phosphorylated Akt and GSK3β, recovered insulin sensitivity, and improved lipid deposition in HepG2 cells. Taken together, our research showed that O-GlcNAcylation of PTP1B can influence insulin signal transduction by modulating its own phosphatase activity, which participates in the process of hepatic insulin resistance.

Structural simulation of adenosine phosphate via plumbagin and zoledronic acid competitively targets JNK/Erk to synergistically attenuate osteoclastogenesis in a breast cancer model

PubMed Central

Qiao, H; Wang, T-y; Yu, Z-f; Han, X-g; Liu, X-q; Wang, Y-g; Fan, Q-m; Qin, A; Tang, T-t

2016-01-01

The treatment of breast cancer-induced osteolysis remains a challenge in clinical settings. Here, we explored the effect and mechanism of combined treatment with zoledronic acid (ZA) and plumbagin (PL), a widely investigated component derived from Plumbago zeylanica, against breast cancer-induced osteoclastogenesis. We found that the combined treatment with PL and ZA suppressed cell viability of precursor osteoclasts and synergistically inhibited MDA-MB-231-induced osteoclast formation (combination index=0.28) with the abrogation of recombinant mouse receptor activator of nuclear factor-κB ligand (RANKL)-induced activation of NF-κB/MAPK (nuclear factor-κB/mitogen-activated protein kinase) pathways. Molecular docking suggested a putative binding area within c-Jun N-terminal kinase/extracellular signal-regulated kinase (JNK/Erk) protease active sites through the structural mimicking of adenosine phosphate (ANP) by the spatial combination of PL with ZA. A homogeneous time-resolved fluorescence assay further illustrated the direct competitiveness of the dual drugs against ANP docking to phosphorylated JNK/Erk, contributing to the inhibited downstream expression of c-Jun/c-Fos/NFATc-1 (nuclear factor of activated T cells, cytoplasmic, calcineurin-dependent 1). Then, in vivo testing demonstrated that the combined administration of PL and ZA attenuated breast cancer growth in the bone microenvironment. Additionally, these molecules prevented the destruction of proximal tibia, with significant reduction of tartrate-resistant acid phosphatase (TRAcP)-positive osteoclast cells and potentiation of apoptotic cancer cells, to a greater extent when combined than when the drugs were applied independently. Altogether, the combination treatment with PL and ZA could significantly and synergistically suppress osteoclastogenesis and inhibit tumorigenesis both in vitro and in vivo by simulating the spatial structure of ANP to inhibit competitively phosphorylation of c-Jun N

Overexpression of ESBP6 improves lactic acid resistance and production in Saccharomyces cerevisiae.

PubMed

Sugiyama, Minetaka; Akase, Shin-Pei; Nakanishi, Ryota; Kaneko, Yoshinobu; Harashima, Satoshi

2016-10-01

Polylactic acid plastics are receiving increasing attention for the control of atmospheric CO2 emissions. Lactic acid, the building block for polylactic acid, is produced by fermentation technology from renewable carbon sources. The yeast Saccharomyces cerevisiae, harboring the lactate dehydrogenases gene (LDH), produces lactic acid at a large scale due to its strong acid resistance, to its simple nutritional requirements and to its ease of genetic engineering. Since improvement of lactic acid resistance is correlated with an increase of lactic acid production under non-neutralizing condition, we isolated a novel gene that enhances lactic acid resistance using a multi-copy yeast genomic DNA library. In this study, we identified the ESBP6 gene, which increases lactic acid resistance when overexpressed and which encodes a protein with similarity to monocarboxylate permeases. Although ESBP6 was not induced in response to lactic acid stress, it caused weak but reproducible sensitivity to lactic acid when disrupted. Furthermore, intracellular pH in the ESBP6 overexpressing strain was higher than that in the wild-type strain under lactic acid stressed condition, suggesting that Esbp6 plays some roles in lactic acid adaptation response. The ESBP6 overexpressing strain carrying the LDH gene induced 20% increase in lactic acid production compared with the wild-type strain carrying the LDH gene under non-neutralizing conditions. These results indicate that overexpression of ESBP6 provides a novel and useful tool to improve lactic acid resistance and lactic acid production in yeast. Copyright © 2016 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

A branched-chain amino acid metabolite drives vascular fatty acid transport and causes insulin resistance.

PubMed

Jang, Cholsoon; Oh, Sungwhan F; Wada, Shogo; Rowe, Glenn C; Liu, Laura; Chan, Mun Chun; Rhee, James; Hoshino, Atsushi; Kim, Boa; Ibrahim, Ayon; Baca, Luisa G; Kim, Esl; Ghosh, Chandra C; Parikh, Samir M; Jiang, Aihua; Chu, Qingwei; Forman, Daniel E; Lecker, Stewart H; Krishnaiah, Saikumari; Rabinowitz, Joshua D; Weljie, Aalim M; Baur, Joseph A; Kasper, Dennis L; Arany, Zoltan

2016-04-01

Epidemiological and experimental data implicate branched-chain amino acids (BCAAs) in the development of insulin resistance, but the mechanisms that underlie this link remain unclear. Insulin resistance in skeletal muscle stems from the excess accumulation of lipid species, a process that requires blood-borne lipids to initially traverse the blood vessel wall. How this trans-endothelial transport occurs and how it is regulated are not well understood. Here we leveraged PPARGC1a (also known as PGC-1α; encoded by Ppargc1a), a transcriptional coactivator that regulates broad programs of fatty acid consumption, to identify 3-hydroxyisobutyrate (3-HIB), a catabolic intermediate of the BCAA valine, as a new paracrine regulator of trans-endothelial fatty acid transport. We found that 3-HIB is secreted from muscle cells, activates endothelial fatty acid transport, stimulates muscle fatty acid uptake in vivo and promotes lipid accumulation in muscle, leading to insulin resistance in mice. Conversely, inhibiting the synthesis of 3-HIB in muscle cells blocks the ability of PGC-1α to promote endothelial fatty acid uptake. 3-HIB levels are elevated in muscle from db/db mice with diabetes and from human subjects with diabetes, as compared to those without diabetes. These data unveil a mechanism in which the metabolite 3-HIB, by regulating the trans-endothelial flux of fatty acids, links the regulation of fatty acid flux to BCAA catabolism, providing a mechanistic explanation for how increased BCAA catabolic flux can cause diabetes.

The relationship between glucose metabolism, metabolic syndrome, and bone-specific alkaline phosphatase: a structural equation modeling approach.

PubMed

Cheung, Ching-Lung; Tan, Kathryn C B; Lam, Karen S L; Cheung, Bernard M Y

2013-09-01

Serum alkaline phosphatase plays a role in vascular calcification. It is found in various tissues, whereas bone-specific alkaline phosphatase (BAP) more specifically reflects mineral metabolism. The relationship of serum alkaline phosphatase (total and bone-specific) with diabetes and metabolic syndrome (MetS), 2 major risk factors of vascular calcification, is largely unknown. We aimed to investigate the relationships between glucose metabolism, components of the MetS, and alkaline phosphatase. This was a cross-sectional study of a nationally representative sample of the U.S. population in 1999 through 2004. Participants were 3773 nondiabetic participants of the National Health and Nutrition Examination Survey 1999-2004. We measured serum BAP and total alkaline phosphatase. In multivariable linear regression, updated homeostasis model assessment (HOMA2) for insulin resistance (β = 0.068), HOMA2 for β-cell function (β = 0.081), insulin (β = 0.065), mean arterial pressure (β = 0.15), and high-density lipoprotein (HDL)-cholesterol (β = 0.209) were positively associated with BAP, whereas HOMA2 for insulin sensitivity (β = -0.065) was negatively associated with BAP. On the other hand, only mean arterial pressure and HDL-cholesterol were significantly associated with total alkaline phosphatase. Moreover, a structural equation model revealed that hypertension, low HDL, and insulin resistance had significant direct effects on serum BAP levels, whereas obesity and inflammation might have indirect effects on serum BAP levels. The overall model showed very good fit to the data (comparative fit index = 0.995, root mean square error of approximation = 0.037, and standardized root mean square residual = 0.006). Glucose metabolism and MetS are significantly related to serum BAP levels. How BAP mediates vascular calcification in diabetes and MetS warrants further studies.

The RCN1-encoded A subunit of protein phosphatase 2A increases phosphatase activity in vivo

NASA Technical Reports Server (NTRS)

Deruere, J.; Jackson, K.; Garbers, C.; Soll, D.; Delong, A.; Evans, M. L. (Principal Investigator)

1999-01-01

Protein phosphatase 2A (PP2A), a heterotrimeric serine/threonine-specific protein phosphatase, comprises a catalytic C subunit and two distinct regulatory subunits, A and B. The RCN1 gene encodes one of three A regulatory subunits in Arabidopsis thaliana. A T-DNA insertion mutation at this locus impairs root curling, seedling organ elongation and apical hypocotyl hook formation. We have used in vivo and in vitro assays to gauge the impact of the rcn1 mutation on PP2A activity in seedlings. PP2A activity is decreased in extracts from rcn1 mutant seedlings, and this decrease is not due to a reduction in catalytic subunit expression. Roots of mutant seedlings exhibit increased sensitivity to the phosphatase inhibitors okadaic acid and cantharidin in organ elongation assays. Shoots of dark-grown, but not light-grown seedlings also show increased inhibitor sensitivity. Furthermore, cantharidin treatment of wild-type seedlings mimics the rcn1 defect in root curling, root waving and hypocotyl hook formation assays. In roots of wild-type seedlings, RCN1 mRNA is expressed at high levels in root tips, and accumulates to lower levels in the pericycle and lateral root primordia. In shoots, RCN1 is expressed in the apical hook and the basal, rapidly elongating cells in etiolated hypocotyls, and in the shoot meristem and leaf primordia of light-grown seedlings. Our results show that the wild-type RCN1-encoded A subunit functions as a positive regulator of the PP2A holoenzyme, increasing activity towards substrates involved in organ elongation and differential cell elongation responses such as root curling.

Effect of soil acidity, soil strength and macropores on root growth and morphology of perennial grass species differing in acid-soil resistance.

PubMed

Haling, Rebecca E; Simpson, Richard J; Culvenor, Richard A; Lambers, Hans; Richardson, Alan E

2011-03-01

It is unclear whether roots of acid-soil resistant plants have significant advantages, compared with acid-soil sensitive genotypes, when growing in high-strength, acid soils or in acid soils where macropores may allow the effects of soil acidity and strength to be avoided. The responses of root growth and morphology to soil acidity, soil strength and macropores by seedlings of five perennial grass genotypes differing in acid-soil resistance were determined, and the interaction of soil acidity and strength for growth and morphology of roots was investigated. Soil acidity and strength altered root length and architecture, root hair development, and deformed the root tip, especially in acid-soil sensitive genotypes. Root length was restricted to some extent by soil acidity in all genotypes, but the adverse impact of soil acidity on root growth by acid-soil resistant genotypes was greater at high levels of soil strength. Roots reacted to soil acidity when growing in macropores, but elongation through high-strength soil was improved. Soil strength can confound the effect of acidity on root growth, with the sensitivity of acid-resistant genotypes being greater in high-strength soils. This highlights the need to select for genotypes that resist both acidity and high soil strength. © 2010 Blackwell Publishing Ltd.

Structural characterization of tartrate dehydrogenase: a versatile enzyme catalyzing multiple reactions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Malik, Radhika; Viola, Ronald E.

2010-10-28

The first structure of an NAD-dependent tartrate dehydrogenase (TDH) has been solved to 2 {angstrom} resolution by single anomalous diffraction (SAD) phasing as a complex with the intermediate analog oxalate, Mg{sup 2+} and NADH. This TDH structure from Pseudomonas putida has a similar overall fold and domain organization to other structurally characterized members of the hydroxy-acid dehydrogenase family. However, there are considerable differences between TDH and these functionally related enzymes in the regions connecting the core secondary structure and in the relative positioning of important loops and helices. The active site in these complexes is highly ordered, allowing the identificationmore » of the substrate-binding and cofactor-binding groups and the ligands to the metal ions. Residues from the adjacent subunit are involved in both the substrate and divalent metal ion binding sites, establishing a dimer as the functional unit and providing structural support for an alternating-site reaction mechanism. The divalent metal ion plays a prominent role in substrate binding and orientation, together with several active-site arginines. Functional groups from both subunits form the cofactor-binding site and the ammonium ion aids in the orientation of the nicotinamide ring of the cofactor. A lysyl amino group (Lys192) is the base responsible for the water-mediated proton abstraction from the C2 hydroxyl group of the substrate that begins the catalytic reaction, followed by hydride transfer to NAD. A tyrosyl hydroxyl group (Tyr141) functions as a general acid to protonate the enolate intermediate. Each substrate undergoes the initial hydride transfer, but differences in substrate orientation are proposed to account for the different reactions catalyzed by TDH.« less

Insulin resistance and the metabolism of branched-chain amino acids.

PubMed

Lu, Jingyi; Xie, Guoxiang; Jia, Weiping; Jia, Wei

2013-03-01

Insulin resistance (IR) is a key pathological feature of metabolic syndrome and subsequently causes serious health problems with an increased risk of several common metabolic disorders. IR related metabolic disturbance is not restricted to carbohydrates but impacts global metabolic network. Branched-chain amino acids (BCAAs), namely valine, leucine and isoleucine, are among the nine essential amino acids, accounting for 35% of the essential amino acids in muscle proteins and 40% of the preformed amino acids required by mammals. The BCAAs are particularly responsive to the inhibitory insulin action on amino acid release by skeletal muscle and their metabolism is profoundly altered in insulin resistant conditions and/or insulin deficiency. Although increased circulating BCAA concentration in insulin resistant conditions has been noted for many years and BCAAs have been reported to be involved in the regulation of glucose homeostasis and body weight, it is only recently that BCAAs are found to be closely associated with IR. This review will focus on the recent findings on BCAAs from both epidemic and mechanistic studies.

Single-label kinase and phosphatase assays for tyrosine phosphorylation using nanosecond time-resolved fluorescence detection.

PubMed

Sahoo, Harekrushna; Hennig, Andreas; Florea, Mara; Roth, Doris; Enderle, Thilo; Nau, Werner M

2007-12-26

The collision-induced fluorescence quenching of a 2,3-diazabicyclo[2.2.2]oct-2-ene-labeled asparagine (Dbo) by hydrogen atom abstraction from the tyrosine residue in peptide substrates was introduced as a single-labeling strategy to assay the activity of tyrosine kinases and phosphatases. The assays were tested for 12 different combinations of Dbo-labeled substrates and with the enzymes p60c-Src Src kinase, EGFR kinase, YOP protein tyrosine phosphatase, as well as acid and alkaline phosphatases, thereby demonstrating a broad application potential. The steady-state fluorescence changed by a factor of up to 7 in the course of the enzymatic reaction, which allowed for a sufficient sensitivity of continuous monitoring in steady-state experiments. The fluorescence lifetimes (and intensities) were found to be rather constant for the phosphotyrosine peptides (ca. 300 ns in aerated water), while those of the unphosphorylated peptides were as short as 40 ns (at pH 7) and 7 ns (at pH 13) as a result of intramolecular quenching. Owing to the exceptionally long fluorescence lifetime of Dbo, the assays were alternatively performed by using nanosecond time-resolved fluorescence (Nano-TRF) detection, which leads to an improved discrimination of background fluorescence and an increased sensitivity. The potential for inhibitor screening was demonstrated through the inhibition of acid and alkaline phosphatases by molybdate.

The Saccharomyces cerevisiae Lipin Homolog is a Mg2+-dependent Phosphatidate Phosphatase Enzyme*

PubMed Central

Han, Gil-Soo; Wu, Wen-I; Carman, George M.

2006-01-01

Mg2+-dependent phosphatidate (PA) phosphatase (3-sn-phosphatidate phosphohydrolase, EC 3.1.3.4) catalyzes the dephosphorylation of PA to yield diacylglycerol and Pi. In this work, we identified the Saccharomyces cerevisiae PAH1 (previously known as SMP2) gene that encodes Mg2+-dependent PA phosphatase using amino acid sequence information derived from a purified preparation of the enzyme (Lin, Y.-P., and Carman, G.M. (1989) J. Biol. Chem. 264, 8641–8645). Overexpression of PAH1 in S. cerevisiae directed elevated levels of Mg2+-dependent PA phosphatase activity, whereas the pah1Δ mutation caused reduced levels of enzyme activity. Heterologous expression of PAH1 in Escherichia coli confirmed that Pah1p is a Mg2+-dependent PA phosphatase enzyme, and showed that its enzymological properties were very similar to those of the enzyme purified from S. cerevisiae. The PAH1-encoded enzyme activity was associated with both the membrane and cytosolic fractions of the cell, and the membrane-bound form of the enzyme was salt-extractable. Lipid analysis showed that mutants lacking PAH1 accumulated PA, and had reduced amounts of diacylglycerol and its derivative triacylglycerol. The PAH1-encoded Mg2+-dependent PA phosphatase shows homology to mammalian lipin, a fat-regulating protein whose molecular function is unknown. Heterologous expression of human LPIN1 in E. coli showed that lipin 1 is also a Mg2+-dependent PA phosphatase enzyme. PMID:16467296

Two ancient bacterial-like PPP family phosphatases from Arabidopsis are highly conserved plant proteins that possess unique properties.

PubMed

Uhrig, R Glen; Moorhead, Greg B

2011-12-01

Protein phosphorylation, catalyzed by the opposing actions of protein kinases and phosphatases, is a cornerstone of cellular signaling and regulation. Since their discovery, protein phosphatases have emerged as highly regulated enzymes with specificity that rivals their counteracting kinase partners. However, despite years of focused characterization in mammalian and yeast systems, many protein phosphatases in plants remain poorly or incompletely characterized. Here, we describe a bioinformatic, biochemical, and cellular examination of an ancient, Bacterial-like subclass of the phosphoprotein phosphatase (PPP) family designated the Shewanella-like protein phosphatases (SLP phosphatases). The SLP phosphatase subcluster is highly conserved in all plants, mosses, and green algae, with members also found in select fungi, protists, and bacteria. As in other plant species, the nucleus-encoded Arabidopsis (Arabidopsis thaliana) SLP phosphatases (AtSLP1 and AtSLP2) lack genetic redundancy and phylogenetically cluster into two distinct groups that maintain different subcellular localizations, with SLP1 being chloroplastic and SLP2 being cytosolic. Using heterologously expressed and purified protein, the enzymatic properties of both AtSLP1 and AtSLP2 were examined, revealing unique metal cation preferences in addition to a complete insensitivity to the classic serine/threonine PPP protein phosphatase inhibitors okadaic acid and microcystin. The unique properties and high conservation of the plant SLP phosphatases, coupled to their exclusion from animals, red algae, cyanobacteria, archaea, and most bacteria, render understanding the function(s) of this new subclass of PPP family protein phosphatases of particular interest.

Phosphate or phosphite addition promotes the proteolytic turnover of phosphate-starvation inducible tomato purple acid phosphatase isozymes.

PubMed

Bozzo, Gale G; Singh, Vinay K; Plaxton, William C

2004-08-27

Within 48 h of the addition of 2.5 mM phosphate (HPO42-, Pi) or phosphite (H2PO3-, Phi) to 8-day-old Pi-starved (-Pi) tomato suspension cells: (i) secreted and intracellular purple acid phosphatase (PAP) activities decreased by about 12- and 6-fold, respectively and (ii) immunoreactive PAP polypeptides either disappeared (secreted PAPs) or were substantially reduced (intracellular PAP). The degradation of both secreted PAP isozymes was correlated with the de novo synthesis of two extracellular serine proteases having M(r)s of 137 and 121 kDa. In vitro proteolysis of purified secreted tomato PAP isozymes occurred following their 24 h incubation with culture filtrate from Pi-resupplied cells. The results indicate that Pi or Phi addition to -Pi tomato cells induces serine proteases that degrade Pi-starvation inducible extracellular proteins.

[Effects of several low-molecular-weight organic acids on the release kinetic of endosulfan from red soil].

PubMed

Zhao, Zhen-hua; Wu, Yu; Jiang, Xin; Xia, Li-ling; Ni, Li-xiao

2009-10-15

The kinetic release behaviors of a-endosulfan from red soil with three kinds of low-molecular-weight organic acids (LMWOA: oxalate, tartrate and citrate) solution and water leaching were investigated by kinetic device designed by ourselves and batch method. The results show that: the release percentage of endosulfan from red soil by tartrate and citrate solution (10 mmol/L) can increase by 7%-18% more than that by distilled water and oxalate solution, especially for tartrate solution. There is no significant difference between distilled water and oxalate solution for the release percentage of endosulfan (p > 0.05). There are two stages of quick and slow for the release of endosulfan from red soil, and the leaching speed is quicker especially for the initial 200 mL leaching solution. When using distilled water or oxalate solution as leaching solution, the best equations that described the kinetic release behavior of endosulfan from red soil were parabola diffuse equation and double constant equation, and weren't the apparent first dynamics equation that represented the simple surface diffusion mechanism. The kinetic release behavior of endosulfan in tartrate or citrate leaching system can be described by Elovich equation (R2 > 0.99, p < 0.0001), it implied that the simple surface diffusion mechanism is not the primary factor that effected the release of endosulfan, which three-dimensional molecule structure is complex, from red soil in aqueous phase leaching systems, and it maybe related to the outward diffuse mechanism from soil particle, activation and deactivation function of soil particles surface, the dissolution of soil mineral surface and structure change of inherent organic matter that coating onto the soil mineral surface induced by LMW organic acid. It suggested that the tartrate and citrate induced the complication of the release mechanisms of the pesticides from red soil.

Corrosion resistance of NiTi in fluoride and acid environments.

PubMed

Benyahia, Hicham; Ebntouhami, Mohamed; Forsal, Issam; Zaoui, Fatima; Aalloula, Elhoussine

2009-12-01

The aim of our study was to assess in the laboratory the electrochemical behavior of nickel-titanium alloy (NiTi) by simulating the aggressive conditions found in the mouth (notably fluoride and acidity) in order to determine its biocompatibility. The impact of fluoride and pH acid on the corrosion resistance of orthodontic NiTi was studied using classic electrochemical measurement techniques including follow-up over time of the corrosion potential, polarization measurements and impedance spectroscopy. In addition, scanning electron microscopy was used to evaluate the status of the alloy surface before and after immersion in the different media. The results demonstrated the particularly low corrosion resistance of NiTi alloy in the presence of fluorides. In an acidic environment, the alloy showed greater resistance thanks to the passivation phenomenon. The synergistic action of fluoride and ph Acid on NiTi corrosion was not clearly demonstrated. Copyright 2009 Collège Européen d'Orthodontie. Published by Elsevier Masson SAS.. All rights reserved.

Crystal structure of the cytoplasmic phosphatase and tensin homolog (PTEN)-like region of Ciona intestinalis voltage-sensing phosphatase provides insight into substrate specificity and redox regulation of the phosphoinositide phosphatase activity.

PubMed

Matsuda, Makoto; Takeshita, Kohei; Kurokawa, Tatsuki; Sakata, Souhei; Suzuki, Mamoru; Yamashita, Eiki; Okamura, Yasushi; Nakagawa, Atsushi

2011-07-01

Ciona intestinalis voltage-sensing phosphatase (Ci-VSP) has a transmembrane voltage sensor domain and a cytoplasmic region sharing similarity to the phosphatase and tensin homolog (PTEN). It dephosphorylates phosphatidylinositol 4,5-bisphosphate and phosphatidylinositol 3,4,5-trisphosphate upon membrane depolarization. The cytoplasmic region is composed of a phosphatase domain and a putative membrane interaction domain, C2. Here we determined the crystal structures of the Ci-VSP cytoplasmic region in three distinct constructs, wild-type (248-576), wild-type (236-576), and G365A mutant (248-576). The crystal structure of WT-236 and G365A-248 had the disulfide bond between the catalytic residue Cys-363 and the adjacent residue Cys-310. On the other hand, the disulfide bond was not present in the crystal structure of WT-248. These suggest the possibility that Ci-VSP is regulated by reactive oxygen species as found in PTEN. These structures also revealed that the conformation of the TI loop in the active site of the Ci-VSP cytoplasmic region was distinct from the corresponding region of PTEN; Ci-VSP has glutamic acid (Glu-411) in the TI loop, orienting toward the center of active site pocket. Mutation of Glu-411 led to acquirement of increased activity toward phosphatidylinositol 3,5-bisphosphate, suggesting that this site is required for determining substrate specificity. Our results provide the basic information of the enzymatic mechanism of Ci-VSP.

Effects of 25-hydroxycholecalciferol supplementation in maternal diets on milk quality and serum bone status markers of sows and bone quality of piglets.

PubMed

Zhou, Hui; Chen, Yuling; Zhuo, Yong; Lv, Gang; Lin, Yan; Feng, Bin; Fang, Zhengfeng; Che, Lianqiang; Li, Jian; Xu, Shengyu; Wu, De

2017-03-01

Twenty primiparous sows were allocated to two treatments to evaluate the effects of maternal 25-hydroxycholecalciferol (25OHD 3 ) supplementation during gestation and lactation on milk quality and serum bone status markers of sows and bone quality of piglets. Immediately after mating, sows were randomly allotted to one of two diets supplemented with 50 µg/kg 25OHD 3 or basal diets without 25OHD 3 . Blood and milk samples were obtained. At birth and weaning, 10 piglets from each treatment were killed for bone quality analysis. 25OHD 3 -fed sows provided one more piglet at farrowing and 1.17 more piglets at weaning than sows fed basal diets. The contents of solids not-fat, protein, fat or lactose were increased in milk from days 7 and 14 of lactation in 25OHD 3 -supplemented sows and 25OHD 3 concentrations in milk were increased by dietary 25OHD 3 supplementation. Dietary 25OHD 3 supplementation increased serum alkaline phosphatase activity but had no effect on serum tartrate-resistant acid phosphatase activity of sows. Maternal 25OHD 3 supplementation improved bone strength, density and ash content of newborn piglets rather than those of weaning piglets. In conclusion, 25OHD 3 supplementation in maternal diets improved reproductive performance, milk quality and bone status of sows as well as bone quality of newborn piglets. © 2016 Japanese Society of Animal Science.

Appliance-induced osteopenia of dentoalveolar bone in the rat: effect of reduced bone strains on serum bone markers and the multifunctional hormone leptin.

PubMed

Vinoth, Jayaseelan K; Patel, Kaval J; Lih, Wei-Song; Seow, Yian-San; Cao, Tong; Meikle, Murray C

2013-12-01

To understand, in greater detail, the molecular mechanisms regulating the complex relationship between mechanical strain and alveolar bone metabolism during orthodontic treatment, passive cross-arch palatal springs were bonded to the maxillary molars of 6-wk-old rats, which were killed after 4 and 8 d. Outcome measures included serum assays for markers of bone formation and resorption and for the multifunctional hormone leptin, and histomorphometry of the inter-radicular bone. The concentration of the bone-formation marker alkaline phosphatase (ALP) was significantly reduced at both time points in the appliance group, accompanied by a 50% reduction in inter-radicular bone volume; however, osteocalcin (bone Gla protein) levels remained unaffected. Bone collagen deoxypyridinoline (DPD) crosslinks increased 2.3-fold at 4 d only, indicating a transient increase in bone resorption; in contrast, the level of the osteoclast-specific marker, tartrate-resistant acid phosphatase 5b (TRACP 5b), was unchanged. Leptin levels closely paralleled ALP reductions at both time points, suggesting an important role in the mechanostat negative-feedback loop required to normalize bone mass. These data suggest that an orthodontic appliance, in addition to remodeling the periodontal ligament (PDL)-bone interface, may exert unexpected side-effects on the tooth-supporting alveolar bone, and highlights the importance of recognizing that bone strains can have negative, as well as positive, effects on bone mass. © 2013 Eur J Oral Sci.

Accelerated craniofacial bone regeneration through dense collagen gel scaffolds seeded with dental pulp stem cells

PubMed Central

Chamieh, Frédéric; Collignon, Anne-Margaux; Coyac, Benjamin R.; Lesieur, Julie; Ribes, Sandy; Sadoine, Jérémy; Llorens, Annie; Nicoletti, Antonino; Letourneur, Didier; Colombier, Marie-Laure; Nazhat, Showan N.; Bouchard, Philippe; Chaussain, Catherine; Rochefort, Gael Y.

2016-01-01

Therapies using mesenchymal stem cell (MSC) seeded scaffolds may be applicable to various fields of regenerative medicine, including craniomaxillofacial surgery. Plastic compression of collagen scaffolds seeded with MSC has been shown to enhance the osteogenic differentiation of MSC as it increases the collagen fibrillary density. The aim of the present study was to evaluate the osteogenic effects of dense collagen gel scaffolds seeded with mesenchymal dental pulp stem cells (DPSC) on bone regeneration in a rat critical-size calvarial defect model. Two symmetrical full-thickness defects were created (5 mm diameter) and filled with either a rat DPSC-containing dense collagen gel scaffold (n = 15), or an acellular scaffold (n = 15). Animals were imaged in vivo by microcomputer tomography (Micro-CT) once a week during 5 weeks, whereas some animals were sacrificed each week for histology and histomorphometry analysis. Bone mineral density and bone micro-architectural parameters were significantly increased when DPSC-seeded scaffolds were used. Histological and histomorphometrical data also revealed significant increases in fibrous connective and mineralized tissue volume when DPSC-seeded scaffolds were used, associated with expression of type I collagen, osteoblast-associated alkaline phosphatase and osteoclastic-related tartrate-resistant acid phosphatase. Results demonstrate the potential of DPSC-loaded-dense collagen gel scaffolds to benefit of bone healing process. PMID:27934940

Zoledronate promotes bone formation by blocking osteocyte-osteoblast communication during bone defect healing.

PubMed

Cui, Pingping; Liu, Hongrui; Sun, Jing; Amizuka, Norio; Sun, Qinfeng; Li, Minqi

2018-01-01

Nitrogen-containing bisphosphonates (N-BPs) are potent antiresorptive drugs and their actions on osteoclasts have been studied extensively. Recent studies have suggested that N-BPs also target bone-forming cells. However, the precise mechanism of N-BPs in osteoblasts is paradoxical, and the specific role of osteocytes is worthy of in-depth study. Here, we investigated the cellular mechanisms of N-BPs regulating bone defect healing by zoledronate (ZA). Bone histomorphometry confirmed an increase in new bone formation by systemic ZA administration. ZA induced more alkaline phosphatase-positive osteoblasts and tartrate-resistant acid phosphatase-positive osteoclasts residing on the bone surface. Inexplicably, ZA increased SOST expression in osteocytes embedded in the bone matrix, which was not compatible with the intense osteoblast activity on the bone surface. ZA induced heterogeneous osteocytes and disturbed the distribution of the osteocytic-canalicular system (OLCS). Furthermore, according to the degree of OLCS regularity, dentin matrix protein 1 reactivity had accumulated around osteocytes in the ZA group, but it was distributed evenly in the OLCS of the control group. The control group showed a dense array of the gap junction protein connexin 43. However, connexin 43 was extremely sparse after ZA administration. In summary, ZA treatment reduces gap junction connections and blocks cellular communication between osteocytes and osteoblasts. Retaining SOST expression in osteocytes leads to activation of the Wnt signaling pathway and subsequent bone formation.

Mangiferin positively regulates osteoblast differentiation and suppresses osteoclast differentiation

PubMed Central

Sekiguchi, Yuusuke; Mano, Hiroshi; Nakatani, Sachie; Shimizu, Jun; Kataoka, Aya; Ogura, Kana; Kimira, Yoshifumi; Ebata, Midori; Wada, Masahiro

2017-01-01

Mangiferin is a polyphenolic compound present in Salacia reticulata. It has been reported to reduce bone destruction and inhibit osteoclastic differentiation. This study aimed to determine whether mangiferin directly affects osteoblast and osteoclast proliferation and differentiation, and gene expression in MC3T3-E1 osteoblastic cells and osteoclast-like cells derived from primary mouse bone marrow macrophage cells. Mangiferin induced significantly greater WST-1 activity, indicating increased cell proliferation. Mangiferin induced significantly increased alkaline phosphatase staining, indicating greater cell differentiation. Reverse transcription-polymerase chain reaction (RT-PCR) demonstrated that mangiferin significantly increased the mRNA level of runt-related transcription factor 2 (RunX2), but did not affect RunX1 mRNA expression. Mangiferin significantly reduced the formation of tartrate-resistant acid phosphatase-positive multinuclear cells. RT-PCR demonstrated that mangiferin significantly increased the mRNA level of estrogen receptor β (ERβ), but did not affect the expression of other osteoclast-associated genes. Mangiferin may inhibit osteoclastic bone resorption by suppressing differentiation of osteoclasts and promoting expression of ERβ mRNA in mouse bone marrow macrophage cells. It also has potential to promote osteoblastic bone formation by promoting cell proliferation and inducing cell differentiation in preosteoblast MC3T3-E1 cells via RunX2. Mangiferin may therefore be useful in improving bone disease outcomes. PMID:28627701

Phosphatase activity in Antarctica soil samples as a biosignature of extant life

NASA Astrophysics Data System (ADS)

Sato, Shuji; Itoh, Yuki; Takano, Yoshinori; Fukui, Manabu; Kaneko, Takeo; Kobayashi, Kensei

Microbial activities have been detected in such extreme terrestrial environments as deep lithosphere, a submarine hydrothermal systems, stratosphere, and Antarctica. Microorganisms have adapted to such harsh environments by evolving their biomolecules. Some of these biomolecules such as enzymes might have different characteristics from those of organisms in ordinary environments. Many biosignatures (or biomarkers) have been proposed to detect microbial activities in such extreme environments. A number of techniques are proposed to evaluate biological activities in extreme environments including cultivation methods, assay of metabolism, and analysis of bioorganic compounds like amino acids and DNA. Enzyme activities are useful signature of extant life in extreme environments. Among many enzymes, phosphatase could be a good indicator of biological activities, since phosphate esters are essential for all the living terrestrial organisms. In addition, alkaline phosphatase is known as a typical zinc-containing metalloenzyme and quite stable in environments. We analyzed phosphatase activities in Antarctica soil samples to see whether they can be used as biosignatures for extant life. In addition, we characterized phosphatases extracted from the Antarctica soil samples, and compared with those obtained from other types of environments. Antarctica surface environments are quite severe environments for life since it is extremely cold and dry and exposed to strong UV and cosmic rays. We tried to evaluate biological activities in Antarctica by measuring phosphatase activities. Surface soil samples are obtained at the Sites 1-8 near Showa Base in Antarctica during the 47th Japan Antarctic exploration mission in 2005-6. Activities of acid phosphatase (ACP) and alkaline phosphatase (ALP) are measured spectrophotometrically after mixing the powdered sample and p-nitrophenyl phosphate solution (pH 6.5 for ACP, pH 8.0 for ALP). ALP was characterized after extraction from soils with

Cultured hypothalamic neurons are resistant to inflammation and insulin resistance induced by saturated fatty acids.

PubMed

Choi, Sun Ju; Kim, Francis; Schwartz, Michael W; Wisse, Brent E

2010-06-01

Hypothalamic inflammation induced by high-fat feeding causes insulin and leptin resistance and contributes to the pathogenesis of obesity. Since in vitro exposure to saturated fatty acids causes inflammation and insulin resistance in many cultured cell types, we determined how cultured hypothalamic neurons respond to this stimulus. Two murine hypothalamic neuronal cell cultures, N43/5 and GT1-7, were exposed to escalating concentrations of saturated fatty acids for up to 24 h. Harvested cells were evaluated for activation of inflammation by gene expression and protein content. Insulin-treated cells were evaluated for induction of markers of insulin receptor signaling (p-IRS, p-Akt). In both hypothalamic cell lines, inflammation was induced by prototypical inflammatory mediators LPS and TNFalpha, as judged by induction of IkappaBalpha (3- to 5-fold) and IL-6 (3- to 7-fold) mRNA and p-IkappaBalpha protein, and TNFalpha pretreatment reduced insulin-mediated p-Akt activation by 30% (P < 0.05). By comparison, neither mixed saturated fatty acid (100, 250, or 500 microM for resistance in cultured hypothalamic neurons, whereas they did in control muscle and endothelial cell lines. Despite the lack of evidence of inflammatory signaling, saturated fatty acid exposure in cultured hypothalamic neurons causes endoplasmic reticulum stress, induces mitogen-activated protein kinase, and causes apoptotic cell death with prolonged exposure. We conclude that saturated fatty acid exposure does not induce inflammatory signaling or insulin resistance in cultured hypothalamic neurons. Therefore, hypothalamic neuronal inflammation in the setting of DIO may involve an indirect mechanism mediated by saturated fatty acids on nonneuronal cells.

Pediatric reference intervals for alkaline phosphatase.

PubMed

Zierk, Jakob; Arzideh, Farhad; Haeckel, Rainer; Cario, Holger; Frühwald, Michael C; Groß, Hans-Jürgen; Gscheidmeier, Thomas; Hoffmann, Reinhard; Krebs, Alexander; Lichtinghagen, Ralf; Neumann, Michael; Ruf, Hans-Georg; Steigerwald, Udo; Streichert, Thomas; Rascher, Wolfgang; Metzler, Markus; Rauh, Manfred

2017-01-01

Interpretation of alkaline phosphatase activity in children is challenging due to extensive changes with growth and puberty leading to distinct sex- and age-specific dynamics. Continuous percentile charts from birth to adulthood allow accurate consideration of these dynamics and seem reasonable for an analyte as closely linked to growth as alkaline phosphatase. However, the ethical and practical challenges unique to pediatric reference intervals have restricted the creation of such percentile charts, resulting in limitations when clinical decisions are based on alkaline phosphatase activity. We applied an indirect method to generate percentile charts for alkaline phosphatase activity using clinical laboratory data collected during the clinical care of patients. A total of 361,405 samples from 124,440 patients from six German tertiary care centers and one German laboratory service provider measured between January 2004 and June 2015 were analyzed. Measurement of alkaline phosphatase activity was performed on Roche Cobas analyzers using the IFCC's photometric method. We created percentile charts for alkaline phosphatase activity in girls and boys from birth to 18 years which can be used as reference intervals. Additionally, data tables of age- and sex-specific percentile values allow the incorporation of these results into laboratory information systems. The percentile charts provided enable the appropriate differential diagnosis of changes in alkaline phosphatase activity due to disease and changes due to physiological development. After local validation, integration of the provided percentile charts into result reporting facilitates precise assessment of alkaline phosphatase dynamics in pediatrics.

Purification and characterization of a polyisoprenyl phosphate phosphatase from pig brain. Possible dual specificity.

PubMed

Frank, D W; Waechter, C J

1998-05-08

Microsomal fractions from pig and calf brain catalyze the enzymatic dephosphorylation of endogenous and exogenous dolichyl monophosphate (Dol-P) (Sumbilla, C. A., and Waechter, C. J. (1985) Methods Enzymol. 111, 471-482). The Dol-P phosphatase (EC 3.1.3.51) has been solubilized by extracting pig brain microsomes with the nonionic detergent Nonidet P-40 and purified approximately 1,107-fold by a combination of anion exchange chromatography, polyethylene glycol fractionation, dye-ligand chromatography, and wheat germ agglutinin affinity chromatography. Treatment of the enzyme with neuraminidase prevented binding to wheat germ agglutinin-Sepharose, indicating the presence of one or more N-acetylneuraminyl residues per molecule of enzyme. When the highly purified polyisoprenyl phosphate phosphatase was analyzed by SDS-polyacrylamide gel electrophoresis, a major 33-kDa polypeptide was observed. Enzymatic dephosphorylation of Dol-P by the purified phosphatase was 1) optimal at pH 7; 2) potently inhibited by F-, orthovanadate, and Zn2+ > Co2+ > Mn2+ but unaffected by Mg2+; 3) exhibited an approximate Km for C95-Dol-P of 45 microM; and 4) was sensitive to N-ethylmaleimide, phenylglyoxal, and diethylpyrocarbonate. The pig brain phosphatase did not dephosphorylate glucose 6-phosphate, mannose 6-phosphate, 5'-AMP, or p-nitrophenylphosphate, but it dephosphorylated dioleoyl-phosphatidic acid at initial rates similar to those determined for Dol-P. Based on the virtually identical sensitivity of Dol-P and phosphatidic acid dephosphorylation by the highly purified enzyme to N-ethylmaleimide, F-, phenylglyoxal, and diethylpyrocarbonate, both substrates appear to be hydrolyzed by a single enzyme with an apparent dual specificity. This is the first report of the purification of a neutral Dol-P phosphatase from mammalian tissues. Although the enzyme is Mg2+-independent and capable of dephosphorylating Dol-P and PA, several enzymological properties distinguish this lipid

Growth and Survival of Acid-Resistant and Non-Acid-Resistant Shiga-Toxin-Producing Escherichia coli Strains during the Manufacture and Ripening of Camembert Cheese.

PubMed

Montet, M P; Jamet, E; Ganet, S; Dizin, M; Miszczycha, S; Dunière, L; Thevenot, D; Vernozy-Rozand, C

2009-01-01

Growth and survival of acid-resistant (AR) and non-acid-resistant (NAR) Shiga-toxin-producing Escherichia coli (STEC) strains were investigated during the manufacture and ripening of microfiltered milk Camembert cheeses. The induction of acid resistance of the STEC strains in cheeses was also studied. Six different mixtures of AR and/or NAR STEC strains were inoculated separately into microfiltered milk at a level of 10(3) CFU mL(-1). The STEC counts (AR and NAR) initially increased by 1 to 2 log(10) CFU g(-1) during cheese-making. Thereafter, the populations stabilized during salting/drying and then decreased during the early stages of ripening. Exposing the STEC strains in artificially inoculated cheeses to simulated gastric fluid (SGF - pH: 2.0) reduced the number of NAR strains to undetectable levels within 40 minutes, versus 120 minutes for the AR STEC strains. AR and NAR STEC were able to survive during the manufacture and ripening of Camembert cheese prepared from microfiltered milk with no evidence of induced acid tolerance in NAR STEC strains.

Growth and Survival of Acid-Resistant and Non-Acid-Resistant Shiga-Toxin-Producing Escherichia coli Strains during the Manufacture and Ripening of Camembert Cheese

PubMed Central

Montet, M. P.; Jamet, E.; Ganet, S.; Dizin, M.; Miszczycha, S.; Dunière, L.; Thevenot, D.; Vernozy-Rozand, C.

2009-01-01

Growth and survival of acid-resistant (AR) and non-acid-resistant (NAR) Shiga-toxin-producing Escherichia coli (STEC) strains were investigated during the manufacture and ripening of microfiltered milk Camembert cheeses. The induction of acid resistance of the STEC strains in cheeses was also studied. Six different mixtures of AR and/or NAR STEC strains were inoculated separately into microfiltered milk at a level of 103 CFU mL−1. The STEC counts (AR and NAR) initially increased by 1 to 2 log10 CFU g−1 during cheese-making. Thereafter, the populations stabilized during salting/drying and then decreased during the early stages of ripening. Exposing the STEC strains in artificially inoculated cheeses to simulated gastric fluid (SGF - pH: 2.0) reduced the number of NAR strains to undetectable levels within 40 minutes, versus 120 minutes for the AR STEC strains. AR and NAR STEC were able to survive during the manufacture and ripening of Camembert cheese prepared from microfiltered milk with no evidence of induced acid tolerance in NAR STEC strains. PMID:20016668

Immunohistochemical characterization of nanocrystalline hydroxyapatite silica gel (NanoBone(r)) osteogenesis: a study on biopsies from human jaws.

PubMed

Götz, Werner; Gerber, Thomas; Michel, Barbara; Lossdörfer, Stefan; Henkel, Kai-Olaf; Heinemann, Friedhelm

2008-10-01

Bone substitute biomaterials may be osteogenic, osteoconductive or osteoinductive. To test for these probable characteristics in a new nanoporous grafting material consisting of nanocrystalline hydroxyapatite embedded in a porous silica gel matrix (NanoBone(s)), applied in humans, we studied biopsies from 12 patients before dental implantation following various orofacial augmentation techniques with healing times of between 3.5 and 12 months. Sections from decalcified specimens were investigated using histology, histochemistry [periodic acid Schiff, alcian blue staining and tartrate-resistant acid phosphatase (TRAP)] and immunohistochemistry, with markers for osteogenesis, bone remodelling, resorption and vessel walls (alkaline phosphatase, bone morphogenetic protein-2, collagen type I, ED1, osteocalcin, osteopontin, runx2 and Von-Willebrand factor). Histologically, four specific stages of graft transformation into lamellar bone could be characterized. During early stages of healing, bone matrix proteins were absorbed by NanoBone(s) granules, forming a proteinaceous matrix, which was invaded by small vessels and cells. We assume that the deposition of these molecules promotes early osteogenesis in and around NanoBone(s) and supports the concomitant degradation probably by osteoclast-like cells. TRAP-positive osteoclast-like cells were localized directly on the granular surfaces. Runx2-immunoreactive pre-osteoblasts, which are probably involved in direct osteogenesis forming woven bone that is later transformed into lamellar bone, were attracted. Graft resorption and bone apposition around the graft granules appear concomitantly. We postulate that NanoBone(s) has osteoconductive and biomimetic properties and is integrated into the host's physiological bone turnover at a very early stage.

Plant Adaptation to Acid Soils: The Molecular Basis for Crop Aluminum Resistance.

PubMed

Kochian, Leon V; Piñeros, Miguel A; Liu, Jiping; Magalhaes, Jurandir V

2015-01-01

Aluminum (Al) toxicity in acid soils is a significant limitation to crop production worldwide, as approximately 50% of the world's potentially arable soil is acidic. Because acid soils are such an important constraint to agriculture, understanding the mechanisms and genes conferring resistance to Al toxicity has been a focus of intense research interest in the decade since the last article on crop acid soil tolerance was published in this journal. An impressive amount of progress has been made during that time that has greatly increased our understanding of the diversity of Al resistance genes and mechanisms, how resistance gene expression is regulated and triggered by Al and Al-induced signals, and how the proteins encoded by these genes function and are regulated. This review examines the state of our understanding of the physiological, genetic, and molecular bases for crop Al tolerance, looking at the novel Al resistance genes and mechanisms that have been identified over the past ten years. Additionally, it examines how the integration of molecular and genetic analyses of crop Al resistance is starting to be exploited for the improvement of crop plants grown on acid soils via both molecular-assisted breeding and biotechnology approaches.

Spectrophotometric and cytochemical analyses of phosphatase activity in Beta vulgaris L.

PubMed

Pesacreta, T C; Bennett, A B; Lucas, W J

1986-03-01

Spectrophotometric and cytochemical methods were used to investigate the localization and/or the sensitivity of phosphatase activities in aldehyde-fixed beet leaves and membrane fractions. The nonspecific acid phosphatase substrates, p-nitrophenyl phosphate and beta-glycerol phosphate, each exhibited unique spectrophotometric patterns of hydrolysis as a function of pH. Additionally, beta-glycerol phosphatase activity was primarily present on the tonoplast, whereas p-nitrophenyl phosphatase was present on the plasma membrane. Because of the unique pH response of each enzyme and their different localization, we conclude that they cannot be entirely "nonspecific." The spectrophotometric pattern of ATP hydrolysis differed from that of p-nitrophenol phosphate in that it decreased at pH 5.0-5.5 and was greatly inhibited by 10 mM sodium fluoride; however, both activities were on the plasma membrane. Therefore, we conclude that these activities represent either two enzymes or only one enzyme that differs in its ability to hydrolyze these two substrates. Generally, enzymatically produced lead deposits on the plasma membrane of non-vascular cells were as frequent and large as those on phloem cells; frequently, deposits on sieve element plasma membranes were relatively small. We therefore conclude that there is no evidence for the presence of relatively intense ATPase activity on the plasma membrane of phloem cells in beet leaf, in contrast to other species. Studies with membrane fractions indicated that formaldehyde could completely inhibit the inhibitor-sensitive phosphatase activities in mitochondrial and vacuolar fractions while preserving significant activity in the plasma membrane fraction.

THE REGENERATIVE CYCLE OF MOTONEURONS, WITH SPECIAL REFERENCE TO PHOSPHATASE ACTIVITY.

PubMed

Bodian, D; Mellors, R C

1945-05-01

1. The regenerative cycle of motoneurons after axon amputation is described, and an attempt made to correlate morphological and chemical events in cell bodies with the growth requirements of regenerating axons. 2. The "normal" pattern of Nissl material in the cell is considered to be the resultant of a steady state in cytoplasmic nucleoprotein. Chromatol is then interpreted as a shift of the balance of nucleoprotein turnover in fa of degradation. The rapid early depletion of Nissl substance in chromatolysis is ascribed to the increased growth requirements created by the active early sprouting of the regenerating axon. Acid phosphatase activity begins to increase above normal levels during this period in the region of nucleopro degradation. 3. The recovery period of chromatolysis due to axon section coincide in time with the phase of gradual lengthening of the regenerating axon, and is thought to represent a gradual restoration of the balance of nucleoprotein degradation and synthesis. During this period acid phosphatase activity is at its height in the region of transformation of Nissl substance, later declines to normal levels when the original pattern of Nissl bodie is restored. 4. The transformation of cytoplasmic nucleoprotein which occurs in chromatolysis after axon section, with the probable liberation (46), and depletion (44), of nucleotides, associated with acid phosphatase activity, suggests the hypothesis that liberated nucleotides or nucleotide compounds may pass down the axon in which they take part in enzymatic activity associated with growth and organization of the newly formed axon. This type of activity would not be incompatible with the ideas previously expressed (30, 81) of a continual function of Nissl substance in maintaining the integrity of the large volume of cytoplasm represented by the axon, as well perhaps as the associated myelin sheath.

Phosphatase activity and its relationship with physical and chemical parameters during vermicomposting of filter cake and cattle manure.

PubMed

Busato, Jader Galba; Papa, Gabriella; Canellas, Luciano Pasqualoto; Adani, Fabrizio; de Oliveira, Aline Lima; Leão, Tairone Paiva

2016-03-15

Recycling of phosphorus (P) from organic residues (ORs) is important to develop environmentally friendly agriculture. The use of this P source depends on phosphatase enzymes, which can be affected by a chain of parameters during maturation of ORs. In this study the phosphatase activity levels throughout vermicomposting of filter cake (FC) and cattle manure (CM) were correlated with different physical and chemical parameters in an effort to increase the knowledge about recycling of P from ORs. FC presented higher total nitrogen content (TNC), total organic carbon (TOC), humic acid (HA) content, water-soluble P (WSP), phosphatase activities and nanopore volume than CM during vermicomposting. Decreases in TOC of CM resulted from carbohydrate mineralization, which was not observed for FC. CM showed increased hydrophobic index during vermicomposting while FC showed a slight decrease. Phosphatase activities correlated positively with TOC, pH and WSP and negatively with HA content for both vermicomposts. Nanopore volume was negatively correlated with phosphatase activities for FC but not for CM. No correlations between hydrophobicity and phosphatase activities were found for FC. Increased hydrophobicity throughout vermicomposting of CM could be partially associated with decreases in phosphatase levels. © 2015 Society of Chemical Industry.

Sulfur amino acid metabolism in doxorubicin-resistant breast cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ryu, Chang Seon; Kwak, Hui Chan; Lee, Kye Sook

2011-08-15

Although methionine dependency is a phenotypic characteristic of tumor cells, it remains to be determined whether changes in sulfur amino acid metabolism occur in cancer cells resistant to chemotherapeutic medications. We compared expression/activity of sulfur amino acid metabolizing enzymes and cellular levels of sulfur amino acids and their metabolites between normal MCF-7 cells and doxorubicin-resistant MCF-7 (MCF-7/Adr) cells. The S-adenosylmethionine/S-adenosylhomocysteine ratio, an index of transmethylation potential, in MCF-7/Adr cells decreased to {approx} 10% relative to that in MCF-7 cells, which may have resulted from down-regulation of S-adenosylhomocysteine hydrolase. Expression of homocysteine-clearing enzymes, such as cystathionine beta-synthase, methionine synthase/methylene tetrahydrofolate reductase,more » and betaine homocysteine methyltransferase, was up-regulated in MCF-7/Adr cells, suggesting that acquiring doxorubicin resistance attenuated methionine-dependence and activated transsulfuration from methionine to cysteine. Homocysteine was similar, which is associated with a balance between the increased expressions of homocysteine-clearing enzymes and decreased extracellular homocysteine. Despite an elevation in cysteine, cellular GSH decreased in MCF-7/Adr cells, which was attributed to over-efflux of GSH into the medium and down-regulation of the GSH synthesis enzyme. Consequently, MCF-7/Adr cells were more sensitive to the oxidative stress induced by bleomycin and menadione than MCF-7 cells. In conclusion, our results suggest that regulating sulfur amino acid metabolism may be a possible therapeutic target for chemoresistant cancer cells. These results warrant further investigations to determine the role of sulfur amino acid metabolism in acquiring anticancer drug resistance in cancer cells using chemical and biological regulators involved in sulfur amino acid metabolism. - Research Highlights: > MCF-7/Adr cells showed decreases in

Identification of human pulmonary alkaline phosphatase isoenzymes.

PubMed

Capelli, A; Cerutti, C G; Lusuardi, M; Donner, C F

1997-04-01

An increase of alkaline phosphatase (ALP) activity has been observed in the bronchoalveolar lavage fluid (BALF) of patients affected by pulmonary fibrosis in chronic interstitial lung disorders. To characterize the ALP isoenzymes in such cases, we used gel filtration, agarose gel electrophoresis, heat and amino acid inhibition assays, wheat-germ agglutinin (WGA) precipitation, and an immunoassay specific for the bone-isoform of ALP. Only one anodic band representing a high-molecular-weight isoform of ALP (Mr approximately 2,000 kDa) was observed on electrophoresis of BALF. The inhibition assay results were consistent for a tissue-nonspecific isoenzyme sensitive to a temperature of 56 degrees C (71.9 +/- 2.5% inhibition) and to homoarginine (65.7 +/- 1.9%), and resistant to L-phenylalanine and L-leucine. Less than 13% of ALP activity was heat-stable. After incubation of BALF specimens with glycosyl-phosphatidylinositol-phospholipase D plus Nonidet P-40, or with phosphatidylinositol-phospholipase C alone, an electrophoretic cathodic band (Mr approximately 220 kDa) appeared near the bone band of a standard serum. With the WGA assay, 84.4 +/- 3.3% of ALP precipitated and the band disappeared. After immunoassay for the bone isoform, a mean of less than 5% enzyme activity was measured. We conclude that the ALP found in BALF is a pulmonary isoform of a tissue nonspecific isoenzyme.

Different designs of kinase-phosphatase interactions and phosphatase sequestration shapes the robustness and signal flow in the MAPK cascade

PubMed Central

2012-01-01

Background The three layer mitogen activated protein kinase (MAPK) signaling cascade exhibits different designs of interactions between its kinases and phosphatases. While the sequential interactions between the three kinases of the cascade are tightly preserved, the phosphatases of the cascade, such as MKP3 and PP2A, exhibit relatively diverse interactions with their substrate kinases. Additionally, the kinases of the MAPK cascade can also sequester their phosphatases. Thus, each topologically distinct interaction design of kinases and phosphatases could exhibit unique signal processing characteristics, and the presence of phosphatase sequestration may lead to further fine tuning of the propagated signal. Results We have built four architecturally distinct types of models of the MAPK cascade, each model with identical kinase-kinase interactions but unique kinases-phosphatases interactions. Our simulations unravelled that MAPK cascade’s robustness to external perturbations is a function of nature of interaction between its kinases and phosphatases. The cascade’s output robustness was enhanced when phosphatases were sequestrated by their target kinases. We uncovered a novel implicit/hidden negative feedback loop from the phosphatase MKP3 to its upstream kinase Raf-1, in a cascade resembling the B cell MAPK cascade. Notably, strength of the feedback loop was reciprocal to the strength of phosphatases’ sequestration and stronger sequestration abolished the feedback loop completely. An experimental method to verify the presence of the feedback loop is also proposed. We further showed, when the models were activated by transient signal, memory (total time taken by the cascade output to reach its unstimulated level after removal of signal) of a cascade was determined by the specific designs of interaction among its kinases and phosphatases. Conclusions Differences in interaction designs among the kinases and phosphatases can differentially shape the robustness and

ERECTA, salicylic acid, abscisic acid, and jasmonic acid modulate quantitative disease resistance of Arabidopsis thaliana to Verticillium longisporum.

PubMed

Häffner, Eva; Karlovsky, Petr; Splivallo, Richard; Traczewska, Anna; Diederichsen, Elke

2014-04-01

Verticillium longisporum is a soil-borne vascular pathogen infecting cruciferous hosts such as oilseed rape. Quantitative disease resistance (QDR) is the major control means, but its molecular basis is poorly understood so far. Quantitative trait locus (QTL) mapping was performed using a new (Bur×Ler) recombinant inbred line (RIL) population of Arabidopsis thaliana. Phytohormone measurements and analyses in defined mutants and near-isogenic lines (NILs) were used to identify genes and signalling pathways that underlie different resistance QTL. QTL for resistance to V. longisporum-induced stunting, systemic colonization by the fungus and for V. longisporum-induced chlorosis were identified. Stunting resistance QTL were contributed by both parents. The strongest stunting resistance QTL was shown to be identical with Erecta. A functional Erecta pathway, which was present in Bur, conferred partial resistance to V. longisporum-induced stunting. Bur showed severe stunting susceptibility in winter. Three stunting resistance QTL of Ler origin, two co-localising with wall-associated kinase-like (Wakl)-genes, were detected in winter. Furthermore, Bur showed a much stronger induction of salicylic acid (SA) by V. longisporum than Ler. Systemic colonization was controlled independently of stunting. The vec1 QTL on chromosome 2 had the strongest effect on systemic colonization. The same chromosomal region controlled the level of abscisic acid (ABA) and jasmonic acid (JA) in response to V. longisporum: The level of ABA was higher in colonization-susceptible Ler than in colonization-resistant Bur after V. longisporum infection. JA was down-regulated in Bur after infection, but not in Ler. These differences were also demonstrated in NILs, varying only in the region containing vec1. All phytohormone responses were shown to be independent of Erecta. Signalling systems with a hitherto unknown role in the QDR of A. thaliana against V. longisporum were identified: Erecta mediated

l-Tartaric acid synthesis from vitamin C in higher plants

PubMed Central

DeBolt, Seth; Cook, Douglas R.; Ford, Christopher M.

2006-01-01

The biosynthetic pathway of l-tartaric acid, the form most commonly encountered in nature, and its catabolic ties to vitamin C, remain a challenge to plant scientists. Vitamin C and l-tartaric acid are plant-derived metabolites with intrinsic human value. In contrast to most fruits during development, grapes accumulate l-tartaric acid, which remains within the berry throughout ripening. Berry taste and the organoleptic properties and aging potential of wines are intimately linked to levels of l-tartaric acid present in the fruit, and those added during vinification. Elucidation of the reactions relating l-tartaric acid to vitamin C catabolism in the Vitaceae showed that they proceed via the oxidation of l-idonic acid, the proposed rate-limiting step in the pathway. Here we report the use of transcript and metabolite profiling to identify candidate cDNAs from genes expressed at developmental times and in tissues appropriate for l-tartaric acid biosynthesis in grape berries. Enzymological analyses of one candidate confirmed its activity in the proposed rate-limiting step of the direct pathway from vitamin C to tartaric acid in higher plants. Surveying organic acid content in Vitis and related genera, we have identified a non-tartrate-forming species in which this gene is deleted. This species accumulates in excess of three times the levels of vitamin C than comparably ripe berries of tartrate-accumulating species, suggesting that modulation of tartaric acid biosynthesis may provide a rational basis for the production of grapes rich in vitamin C. PMID:16567629

Glow discharge plasma pretreatment enhances osteoclast differentiation and survival on titanium plates.

PubMed

Kawai, Hiroyuki; Shibata, Yo; Miyazaki, Takashi

2004-05-01

Despite the fact that several reports have demonstrated osteoclast activity on various bioactive ceramics, osteoclast functions on surface-modified titanium have not come under focus. This study aimed to examine whether the increasing surface energy of glow discharge plasma (GDP) involved in protein adhesion containing the RGD (Arg-Gly-Asp) sequence affects osteoclast responses on titanium plates. We examined osteoclast differentiation and survival rates on titanium plates with and without GDP. The amounts of osteoclasts on titanium plates were not increased by GDP after 1 week. However, osteoclast differentiation was greatly activated by GDP pretreatment, as tartrate-resistant acid phosphatase synthesis significantly increased on the titanium plates with GDP. Additionally, since the presence of osteoclasts was detected only on the titanium plates with GDP, even after 4h cultivation in a coculture test, the osteoclasts survival rate was increased by GDP pretreatment. As osteoclast responses were affected even on surface modified metallic materials, we concluded that novel approaches are needed not only for osteoclastic resorption on ceramic materials but also for osteoclast responses on surface-modified metallic materials.

SH3BP2 is an activator of NFAT Activity and Osteoclastogenesis

PubMed Central

Lietman, Steven A.; Yin, Lihong; Levine, Michael A.

2009-01-01

Heterozygous activating mutations in exon 9 of SH3BP2 have been found in most patients with cherubism, an unusual genetic syndrome characterized by excessive remodeling of the mandible and maxilla due to spontaneous and excessive osteoclastic bone resorption. Osteoclasts differentiate after binding of sRANKL to RANK induces a number of downstream signaling effects, including activation of the calcineurin/NFAT (nuclear factor of activated T cells) pathway. Here we have investigated the functional significance of SH3BP2 protein on osteoclastogenesis in the presence of sRANKL. Our results indicate that SH3BP2 both increases nuclear NFATc1 in sRANKL treated RAW 264.7 preosteoclast cells and enhances expression of tartrate resistant acid phosphatase (TRAP), a specific marker of osteoclast differentiation. Moreover, overexpression of SH3BP2 in RAW 264.7 cells potentiates sRANKL stimulated phosphorylation of PLCγ1 and 2, thus providing a mechanistic pathway for the rapid translocation of NFATc1 into the nucleus and increased osteoclastogenesis in cherubism. PMID:18440306

Retrovirus-mediated conditional immortalization and analysis of established cell lines of osteoclast precursor cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kawata, Shigehisa; Suzuki, Jun; Osaka University Graduate School of Medicine, 2-2 Yamada-oka, Suita, Osaka 565-0871

2006-11-10

Osteoclast precursor cells (OPCs) have previously been established from bone marrow cells of SV40 temperature-sensitive T antigen-expressing transgenic mice. Here, we use retrovirus-mediated gene transfer to conditionally immortalize OPCs by expressing temperature-sensitive large T antigen (tsLT) from wild type bone marrow cells. The immortalized OPCs proliferated at the permissive temperature of 33.5 deg. C, but stopped growing at the non-permissive temperature of 39 deg. C. In the presence of receptor activator of NF{kappa}B ligand (RANKL), the OPCs differentiated into tartrate-resistant acid phosphatase (TRAP)-positive cells and formed multinucleate osteoclasts at 33.5 deg. C. From these OPCs, we cloned two types ofmore » cell lines. Both differentiated into TRAP-positive cells, but one formed multinucleate osteoclasts while the other remained unfused in the presence of RANKL. These results indicate that the established cell lines are useful for analyzing mechanisms of differentiation, particularly multinucleate osteoclast formation. Retrovirus-mediated conditional immortalization should be a useful method to immortalize OPCs from primary bone marrow cells.« less

The effects of binge-pattern alcohol consumption on orthodontic tooth movement

PubMed Central

de Araujo, Cristiano Miranda; Johann, Aline Cristina Batista Rodrigues; Camargo, Elisa Souza; Tanaka, Orlando Motohiro

2014-01-01

OBJECTIVE: This study aimed to assess tissue changes during orthodontic movement after binge-pattern ethanol 20% exposure. METHODS: Male Wistar rats (n = 54) were divided into two groups. The control group (CG) received 0.9% saline solution, while the experimental group (EG) received 20% ethanol in 0.9% saline solution (3 g/kg/day). On the 30th day, a force of 25 cN was applied with a nickel-titanium closed coil spring to move the maxillary right first molar mesially. The groups were further divided into three subgroups (2, 14 and 28 days). Tartrate-resistant acid phosphatase and picrosirius were used to assess bone resorption and neoformation, respectively. Data were compared by two-way ANOVA, Tukey's HSD, Games-Howell and chi-square test. Significance level was set at 5%. RESULTS: There was a decrease in the number of osteoclasts in EG at day 28. The percentage of collagen showed no interaction between group and time. CONCLUSION: Binge-pattern 20% ethanol promoted less bone resorption at the end of tooth movement, thereby suggesting delay in tooth movement. PMID:25628085

Variant hairy cell leukemia following papillary urothelial neoplasm of bladder.

PubMed

Beyan, Cengiz; Kaptan, Kürsat

2014-03-01

A 65 years old man was admitted with multiple lymphadenopathy, weight loss, night sweats and fatigue for 2 months. He had been treated for bladder cancer 2 years ago. Leukocyte count was 37.9 x10(9)/l. Peripheral blood smear had 91% lymphocytes. Lymphocytes had large nuclei with prominent nucleoli, heterogeneous appearance, and large cytoplasm with hairy projections. Flow cytometric immunophenotyping revealed CD20, CD22, CD24, CD45 and HLA-DR positivity. Atypical lymphocytes were stained with tartrate resistant acid phosphatase. Increased metabolic activity was detected in multiple lymph nodes, bone marrow and extremely enlarged spleen with positron emission tomography-computed tomography. Excisional biopsy of the left axillary lymph node revealed infiltration with diffuse B-cell leukemia/lymphoma. Immunohistochemistry showed CD20 positive atypical cells with weak expression of CD11c. The patient was diagnosed as a case of variant hairy cell leukemia and cladribine was administered. A probable second primary malignancy should be kept in mind in cases with a defined malignancy in the presence of unusual symptoms.

Effects of alkylphenols on bone metabolism in vivo and in vitro.

PubMed

Hagiwara, Hiromi; Sugizaki, Toshinori; Tsukamoto, Yu; Senoh, Emi; Goto, Tadashi; Ishihara, Yoko

2008-09-01

Alkylphenols are endocrine disruptors that show estrogen-like effects in various wildlife species. However, little information is available about the action of these chemicals on bone metabolism. We investigated the effects of alkylphenols, such as nonylphenol (NP) and octylphenol (OP), on the formation of bone using several culture systems for osteoclasts and osteoblasts, as well as in vivo experiments. NP and OP dose-dependently inhibited the formation of tartrate-resistant acid phosphatase-positive multinucleated cells (osteoclasts) in cocultures of mouse spleen cells or mouse bone marrow cells with ST2 cells. However, beta-estradiol at 10(-9)M to 10(-6)M did not affect this process. In contrast, neither compound affected the proliferation and differentiation of rat calvarial osteoblast-like cells (ROB cells). When NP or OP (0.1mg/kg body weight) was administered subcutaneously to pregnant mice at 10 days, 12 days and 14 days post-coitus, fetuses at 17.5 days post-coitus showed stimulation of sternebrae bone calcification. Our findings suggest that alkylphenols have critical effects on the formation of bone by non-estrogenic effects.

Effect of an estrogen-deficient state and alendronate therapy on bone loss resulting from experimental periapical lesions in rats.

PubMed

Xiong, Haofei; Peng, Bin; Wei, Lili; Zhang, Xiaolei; Wang, Li

2007-11-01

The aim of the research was to evaluate the impact of an estrogen-deficient state and alendronate (ALD) therapy on bone loss resulting from experimental periapical lesions in rats. Periapical lesions were induced on ovariectomized (OVX) and sham-ovariectomized (Sham) rats. After sample preparation, histologic and radiographic examination for periapical bone loss area and an enzyme histochemical test for tartrate-resistant acid phosphatase (TRAP) were performed. The results showed that OVX significantly increased bone loss resulting from periradicular lesions. After daily subcutaneous injection of ALD, the bone loss area and the number of TRAP-positive cells (osteoclasts) were reduced. These findings suggested that alendronate may protect against increased bone loss from experimental periapical lesions in estrogen-deficient rats. Given recent recognition of adverse effects of bisphosphonates, including an increased risk for osteonecrosis, the findings from this study should not be interpreted as a new indication for ALD treatment. However, they may offer insight into understanding and predicting outcomes in female postmenopausal patients already on ALD therapy for medical indications.

A novel fluorescence biosensor for sensitivity detection of tyrosinase and acid phosphatase based on nitrogen-doped graphene quantum dots.

PubMed

Qu, Zhengyi; Na, Weidan; Liu, Xiaotong; Liu, Hua; Su, Xingguang

2018-01-02

In this paper, we developed a sensitive fluorescence biosensor for tyrosinase (TYR) and acid phosphatase (ACP) activity detection based on nitrogen-doped graphene quantum dots (N-GQDs). Tyrosine could be catalyzed by TYR to generate dopaquinone, which could efficiently quench the fluorescence of N-GQDs, and the degree of fluorescence quenching of N-GQDs was proportional to the concentration of TYR. In the presence of ACP, l-Ascorbic acid-2-phosphate (AAP) was hydrolyzed to generate ascorbic acid (AA), and dopaquinone was reduced to l-dopa, resulting in the fluorescence recovery of the quenched fluorescence by dopaquinone. Thus, a novel fluorescence biosensor for the detection of TYR and ACP activity based on N-GQDs was constructed. Under the optimized experimental conditions, the fluorescence intensity was linearly correlated with the concentration of TYR and ACP in the range of 0.43-3.85 U mL -1 and 0.04-0.7 mU mL -1 with a detection limit of 0.15 U mL -1 and 0.014 mU mL -1 , respectively. The feasibility of the proposed biosensor in real samples assay was also studied and satisfactory results were obtained. Copyright © 2017 Elsevier B.V. All rights reserved.

Hydroxycinnamoyl Glucose and Tartrate Esters and Their Role in the Formation of Ethylphenols in Wine.

PubMed

Hixson, Josh L; Hayasaka, Yoji; Curtin, Christopher D; Sefton, Mark A; Taylor, Dennis K

2016-12-14

Synthesized p-coumaroyl and feruloyl l-tartrate esters were submitted to Brettanomyces bruxellensis strains AWRI 1499, AWRI 1608, and AWRI 1613 to assess their role as precursors to ethylphenols in wine. No evolution of ethylphenols was observed. Additionally, p-coumaroyl and feruloyl glucose were synthesized and submitted to B. bruxellensis AWRI 1499, which yielded both 4-ethylphenol and 4-ethylguaiacol. Unexpected chemical transformations of the hydroxycinnamoyl glucose esters during preparation were investigated to prevent these in subsequent synthetic attempts. Photoisomerization gave an isomeric mixture containing the trans-esters and undesired cis-esters, and acyl migration resulted in a mixture of the desired 1-O-β-ester and two additional migrated forms, the 2-O-α- and 6-O-α-esters. Theoretical studies indicated that the photoisomerization was facilitated by deprotonation of the phenol, and acyl migration is favored during acidic, nonaqueous handling. Preliminary LC-MS/MS studies observed the migrated hydroxycinnamoyl glucose esters in wine and allowed for identification of feruloyl glucose in red wine for the first time.

The leukocyte common antigen (CD45): a putative receptor-linked protein tyrosine phosphatase.

PubMed Central

Charbonneau, H; Tonks, N K; Walsh, K A; Fischer, E H

1988-01-01

A major protein tyrosine phosphatase (PTPase 1B) has been isolated in essentially homogeneous form from the soluble and particulate fractions of human placenta. Unexpectedly, partial amino acid sequences displayed no homology with the primary structures of the protein Ser/Thr phosphatases deduced from cDNA clones. However, the sequence is strikingly similar to the tandem C-terminal homologous domains of the leukocyte common antigen (CD45). A 157-residue segment of PTPase 1B displayed 40% and 33% sequence identity with corresponding regions from cytoplasmic domains I and II of human CD45. Similar degrees of identity have been observed among the catalytic domains of families of regulatory proteins such as protein kinases and cyclic nucleotide phosphodiesterases. On this basis, it is proposed that the CD45 family has protein tyrosine phosphatase activity and may represent a set of cell-surface receptors involved in signal transduction. This suggests that the repertoire of signal transduction mechanisms may include the direct control of an intracellular protein tyrosine phosphatase, offering the possibility of a regulatory balance with those protein tyrosine kinases that act at the internal surface of the membrane. Images PMID:2845400

Stress Degradation Studies on Varenicline Tartrate and Development of a Validated Stability-Indicating HPLC Method

PubMed Central

Pujeri, Sudhakar S.; Khader, Addagadde M. A.; Seetharamappa, Jaldappagari

2012-01-01

A simple, rapid and stability-indicating reversed-phase liquid chromatographic method was developed for the assay of varenicline tartrate (VRT) in the presence of its degradation products generated from forced decomposition studies. The HPLC separation was achieved on a C18 Inertsil column (250 mm × 4.6 mm i.d. particle size is 5 μm) employing a mobile phase consisting of ammonium acetate buffer containing trifluoroacetic acid (0.02M; pH 4) and acetonitrile in gradient program mode with a flow rate of 1.0 mL min−1. The UV detector was operated at 237 nm while column temperature was maintained at 40 °C. The developed method was validated as per ICH guidelines with respect to specificity, linearity, precision, accuracy, robustness and limit of quantification. The method was found to be simple, specific, precise and accurate. Selectivity of the proposed method was validated by subjecting the stock solution of VRT to acidic, basic, photolysis, oxidative and thermal degradation. The calibration curve was found to be linear in the concentration range of 0.1–192 μg mL−1 (R2 = 0.9994). The peaks of degradation products did not interfere with that of pure VRT. The utility of the developed method was examined by analyzing the tablets containing VRT. The results of analysis were subjected to statistical analysis. PMID:22396908

Synchronous delivery of felodipine and metoprolol tartrate using monolithic osmotic pump technology.

PubMed

Zhao, Shiqing; Yu, Fanglin; Liu, Nan; Di, Zhong; Yan, Kun; Liu, Yan; Li, Ying; Zhang, Hui; Yang, Yang; Yang, Zhenbo; Li, Zhiping; Mei, Xingguo

2016-11-01

The synchronous sustained-release of two drugs was desired urgently for patients needing combination therapy in long term. However, sophisticated technologies were used generally to realize the simultaneous delivery of two drugs especially those with different physico-chemical properties. The purpose of this study was to obtain the concurrent release of felodipine and metoprolol tartrate, two drugs with completely different solubilities, in a simple monolithic osmotic pump system (FMOP). Two types of blocking agents were used in monolithic osmotic pump tablets and the synchronous sustained-release of FMOP was acquired in vitro. The tablets were also administered to beagle dogs and the plasma levels of FMOP were determined by HPLC-MS/MS. The pharmacokinetic parameters were calculated using a non-compartmental model. Cmax of both felodipine and metoprolol from the osmotic pump tablets were lower, tmax and mean residence time of both felodipine and metoprolol from the osmotic pump tablets were longer significantly than those from immediate release tablets. These results verified prolonged release of felodipine and metoprolol tartrate from osmotic pump formulations. The similar absorption rate between felodipine and metoprolol in beagles was also obtained by this osmotic pump formulation. Therefore, it could be supposed that the accordant release of two drugs with completely different solubilities may be realized just by using monolithic osmotic pump technology.

Effect of Exogenous Phytase Addition on Soil Phosphatase Activities: a Fluorescence Spectroscopy Study.

PubMed

Yang, Xiao-zhu; Chen, Zhen-hua; Zhang, Yu-lan; Chen, Li-jun

2015-05-01

The utilization of organic phosphorus (P) has directly or indirectly improved after exogenous phytase was added to soil. However, the mechanism by which exogenous phytase affected the soil phosphatases (phosphomonoesterase and phosphodiesterase) activities was not clear. The present work was aimed to study red soil, brown soil and cinnamon soil phosphomonoesterase (acid and alkaline) (AcP and AlP) and phosphodiesterase (PD) activities responding to the addition of exogenous phytase (1 g phytase/50 g air dry soil sample) based on the measurements performed via a fluorescence detection method combined with 96 microplates using a TECAN Infinite 200 Multi-Mode Microplate Reader. The results indicated that the acid phosphomonoesterase activity was significantly enhanced in red soil (p≤0. 01), while it was significantly reduced in cinnamon soil; alkaline phosphomonoesterase activity was significantly enhanced in cinnamon soil (p≤ 0. 01), while it was significantly reduced in red soil; phosphodiesterase activity was increased in three soils but it was significantly increased in brown soil (p≤0. 01) after the addition of exogenous phytase. The activities still remained strong after eight days in different soils, which indicated that exogenous phytase addition could be enhance soil phosphatases activities effectively. This effect was not only related to soil properties, such as pH and phosphorus forms, but might also be related to the excreted enzyme amount of the stimulating microorganism. Using fluorescence spectroscopy to study exogenous phytase addition influence on soil phosphatase activities was the first time at home and abroad. Compared with the conventional spectrophotometric method, the fluorescence microplate method is an accurate, fast and simple to use method to determine the relationships among the soil phosphatases activities.

Influence of exercise mode and osteogenic index on bone biomarker responses during short-term physical training.

PubMed

Lester, Mark E; Urso, Maria L; Evans, Rachel K; Pierce, Joseph R; Spiering, Barry A; Maresh, Carl M; Hatfield, Disa L; Kraemer, William J; Nindl, Bradley C

2009-10-01

Prescribing exercise based on intensity, frequency, and duration of loading may maximize osteogenic responses in bone, but a model of the osteogenic potential of exercise has not been established in humans. In rodents, an osteogenic index (OI) has been used to predict the osteogenic potential of exercise. The current study sought to determine whether aerobic, resistance, or combined aerobic and resistance exercise programs conducted over eight weeks and compared to a control group could produce changes in biochemical markers of bone turnover indicative of bone formation. We further sought to determine whether an OI could be calculated for each of these programs that would reflect observed biochemical changes. We collected serum biomarkers [bone-specific alkaline phosphatase (BAP), osteocalcin, tartrate-resistant acid phosphatase (TRAP), C-terminal telopeptide fragment of type I collagen (CTx), deoxypyridinoline (DPD), 25-hydroxy vitamin D (25(OH)D), and parathyroid hormone (PTH)] in 56 women (20.3+/-1.8 years) before, during and after eight weeks of training. We also measured bone mineral density (BMD) at regional areas of interest using DXA and pQCT. Biomarkers of bone formation (BAP and osteocalcin) increased in the Resistance and Combined groups (p<0.05), while biomarkers of bone resorption (TRAP and DPD) decreased and increased, respectively, after training (p<0.05) in all groups. Small changes in volumetric and areal BMD (p<0.05) were observed in the distal tibia in the Aerobic and Combined groups, respectively. Mean weekly OIs were 16.0+/-1.9, 20.6+/-2.2, and 36.9+/-5.2 for the Resistance, Aerobic, and Combined groups, respectively. The calculated osteogenic potential of our programs did not correlate with the observed changes in biomarkers of bone turnover. The results of the present study demonstrate that participation in an eight week physical training program that incorporates a resistance component by previously inactive young women results in

Associations of erythrocyte fatty acid patterns with insulin resistance

USDA-ARS?s Scientific Manuscript database

Background: Synergistic and/or additive effects on cardiometabolic risk may be missed by examining individual fatty acids (FA). A pattern analysis may be a more useful approach. As well, it remains unclear whether erythrocyte fatty acid composition relates to insulin resistance among Hispanic/Latino...

21 CFR 864.7660 - Leukocyte alkaline phosphatase test.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Leukocyte alkaline phosphatase test. 864.7660... Leukocyte alkaline phosphatase test. (a) Identification. A leukocyte alkaline phosphatase test is a device used to identify the enzyme leukocyte alkaline phosphatase in neutrophilic granulocytes (granular...

21 CFR 864.7660 - Leukocyte alkaline phosphatase test.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Leukocyte alkaline phosphatase test. 864.7660... Leukocyte alkaline phosphatase test. (a) Identification. A leukocyte alkaline phosphatase test is a device used to identify the enzyme leukocyte alkaline phosphatase in neutrophilic granulocytes (granular...

21 CFR 864.7660 - Leukocyte alkaline phosphatase test.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Leukocyte alkaline phosphatase test. 864.7660... Leukocyte alkaline phosphatase test. (a) Identification. A leukocyte alkaline phosphatase test is a device used to identify the enzyme leukocyte alkaline phosphatase in neutrophilic granulocytes (granular...

Phosphatase activity of the voltage-sensing phosphatase, VSP, shows graded dependence on the extent of activation of the voltage sensor

PubMed Central

Sakata, Souhei; Okamura, Yasushi

2014-01-01

The voltage-sensing phosphatase (VSP) consists of a voltage sensor and a cytoplasmic phosphatase region, and the movement of the voltage sensor is coupled to the phosphatase activity. However, its coupling mechanisms still remain unclear. One possible scenario is that the phosphatase is activated only when the voltage sensor is in a fully activated state. Alternatively, the enzymatic activity of single VSP proteins could be graded in distinct activated states of the voltage sensor, and partial activation of the voltage sensor could lead to partial activation of the phosphatase. To distinguish between these two possibilities, we studied a voltage sensor mutant of zebrafish VSP, where the voltage sensor moves in two steps as evidenced by analyses of charge movements of the voltage sensor and voltage clamp fluorometry. Measurements of the phosphatase activity toward phosphatidylinositol 4,5-bisphosphate revealed that both steps of voltage sensor activation are coupled to the tuning of phosphatase activities, consistent with the idea that the phosphatase activity is graded by the magnitude of the movement of the voltage sensor. PMID:24277865