Increase in tartrate-resistant acid phosphatase of bone at the early stage of ascorbic acid deficiency in the ascorbate-requiring Osteogenic Disorder Shionogi (ODS) rat.

PubMed

Goto, A; Tsukamoto, I

2003-08-01

The effect of ascorbic acid deficiency on bone metabolism was evaluated using the ascorbate-requiring Osteogenic Disorder Shionogi (ODS) rat model. Ascorbic acid (Asc)-deficient rats gained body weight in a manner similar to Asc-supplemented rats (control) during 3 weeks, but began to lose weight during the 4th week of Asc deficiency. The tartrate-resistant acid phosphatase (TRAP) activity in serum increased to about 2-fold the control value in the rats fed the Asc-free diet for 2, 3, and 4 weeks (AscD2, AscD3, and AscD4), while a decrease in the alkaline phosphatase (ALP) activity was observed only in AscD4 rats. The serum pyridinoline cross-linked carboxyterminal telopeptide of type I collagen (ICTP) level significantly increased to 1.3-, 1.4-, and 1.9-fold of that in the controls in AscD2, D3, and D4, respectively. The ALP activity in the distal femur was unchanged in AscD1, D2, and D3, but decreased to 50% of the control level in AscD4 rats. The TRAP activity in the distal femur increased to about 2-fold of that in the controls in the AscD2 and D3 and decreased to the control level in the AscD4 rats. The amount of hydroxyproline in the distal femur significantly decreased to about 80%, 70%, and 60% of the control in AscD2, D3, and D4 rats, respectively. These decreases were associated with a similar reduction in the calcium content of the distal femur. Histochemical analysis of the distal femur showed an increase in TRAP-positive cells in AscD2 and AscD3 rats and a decrease in the trabecular bone in AscD2, D3, and D4 rats. These results suggested that a deficiency of Asc stimulated bone resorption at an early stage, followed by a decrease in bone formation in mature ODS rats which already had a well-developed collagen matrix and fully differentiated osteoblasts.

21 CFR 184.1077 - Potassium acid tartrate.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Potassium acid tartrate. 184.1077 Section 184.1077... Listing of Specific Substances Affirmed as GRAS § 184.1077 Potassium acid tartrate. (a) Potassium acid tartrate (C4H5KO6, CAS Reg. No. 868-14-4) is the potassium acid salt of l−(+)−tartaric acid and is also...

21 CFR 184.1077 - Potassium acid tartrate.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Potassium acid tartrate. 184.1077 Section 184.1077... GRAS § 184.1077 Potassium acid tartrate. (a) Potassium acid tartrate (C4H5KO6, CAS Reg. No. 868-14-4) is the potassium acid salt of l−(+)−tartaric acid and is also called potassium bitartrate or cream of...

21 CFR 184.1077 - Potassium acid tartrate.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Potassium acid tartrate. 184.1077 Section 184.1077... Listing of Specific Substances Affirmed as GRAS § 184.1077 Potassium acid tartrate. (a) Potassium acid tartrate (C4H5KO6, CAS Reg. No. 868-14-4) is the potassium acid salt of l−(+)−tartaric acid and is also...

[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.

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.

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.

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.

[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 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

[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.

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.

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.

[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

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

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.

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.

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.

Consumption of vitamin D-and calcium-fortified soft white cheese lowers the biochemical marker of bone resorption TRAP 5b in postmenopausal women at moderate risk of osteoporosis fracture.

PubMed

Bonjour, Jean-Philippe; Benoit, Valérie; Rousseau, Brigitte; Souberbielle, Jean-Claude

2012-04-01

The prevention of increased bone remodeling in postmenopausal women at low 10-y risk of osteoporotic fractures essentially relies on reinforcement of environmental factors known to positively influence bone health, among which nutrition plays an important role. In institutionalized women in their mid-eighties, we previously found that consumption of fortified soft plain cheese increased vitamin D, calcium, and protein intakes, reduced bone resorption biochemical markers, particularly the serum bone specific acid phosphatase tartrate resistant acid phosphatase, isoform 5b (TRAP 5b) that reflects osteoclast activity, and stimulated the serum bone anabolic factor insulin-like growth factor-I (IGF-I). Whether these effects occur in much younger women was tested in a prospective control study. Seventy-one healthy postmenopausal women aged 56.6 ± 3.9 y (mean ± SD) with low spontaneous supply of both Ca and vitamin D were randomized to consume daily (treated, n = 36) or not (controls, n = 35) two servings (2 × 100 g) of skimmed-milk, soft plain cheese for 6 wk. The vitamin D and Ca-fortified dairy product provided daily: 661 kJ, 2.5 μg vitamin D, 400 mg calcium, and 13.8 g protein. At the end of the intervention, the decrease in TRAP 5b and the increase in IGF-I were greater in the treated than in the control group (P < 0.02). The changes in serum carboxy terminal crosslinked telopeptide of type I collagen did not differ significantly between the two groups. In conclusion, like in elderly women, consumption by healthy postmenopausal women of a vitamin D and calcium-fortified dairy product that also increases the protein intake, reduces the serum concentration of the bone resorption biomarker TRAP 5b. This response, combined with the increase in serum IGF-I, is compatible with a nutrition-induced reduction in postmenopausal bone loss rate.

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.

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

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

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.

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

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

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.

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.

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

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.

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

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.

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.

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.

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...

Structural Basis for the Catalytic Activity of Human Serine/Threonine Protein Phosphatase type 5 (PP5)

NASA Technical Reports Server (NTRS)

Swingle, Mark R.; Ciszak, Ewa M.; Honkanen, Richard E.

2004-01-01

Serine/threonine protein phosphatase-5 (PP5) is a member of the PPP-gene family of protein phosphatases that is widely expressed in mammalian tissues and is highly conserved among eukaryotes. PP5 associates with several proteins that affect signal transduction networks, including the glucocorticoid receptor (GR)-heat shock protein-90 (Hsp90)-heterocomplex, the CDC16 and CDC27 subunits of the anaphase-promoting complex, elF2alpha kinase, the A subunit of PP2A, the G12-alpha / G13-alpha subunits of heterotrimeric G proteins and DNA-PK. The catalytic domain of PP5 (PP5c) shares 35-45% sequence identity with the catalytic domains of other PPP-phosphatases, including protein phosphatase-1 (PP1), -2A (PP2A), -2B / calcineurin (PP2B), -4 (PP4), -6 (PP6), and -7 (PP7). Like PP1, PP2A and PP4, PP5 is also sensitive to inhibition by okadaic acid, microcystin, cantharidin, tautomycin, and calyculin A. Here we report the crystal structure of the PP5 catalytic domain (PP5c) at a resolution of 1.6 angstroms. From this structure we propose a mechanism for PP5-mediated hydrolysis of phosphoprotein substrates, which requires the precise positioning of two metal ions within a conserved Asp(sup 271)-M(sub 1):M(sub 2)-W(sup 1)-His(sup 304)-Asp(sup 274) catalytic motif. The structure of PP5c provides a possible structural basis for explaining the exceptional catalytic proficiency of protein phosphatases, which are among the most powerful known catalysts. Resolution of the entire C-terminus revealed a novel subdomain, and the structure of the PP5c should also aid development of type-specific inhibitors.

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

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.

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

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...

BCR-crosslinking induces a transcription of protein phosphatase component G5PR that is required for mature B-cell survival

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

Huq Ronny, Faisal Mahmudul; Igarashi, Hideya; Core Research for Evolutional Science and Technology

2006-02-03

BCR-crosslinking triggers activation-induced cell death (AICD) selectively in the restricted stage of B-cell differentiation. We examined the transcription of a protein phosphatase subunit G5PR in immature and mature B-cells, because absence of this factor augmented cell sensitivity to AICD, associated with increased activation of JNK and Bim. BCR-crosslinking-induced G5pr transcription in AICD-resistant mature splenic IgM{sup lo}IgD{sup hi} B-cells but not in AICD susceptible immature IgM{sup hi}IgD{sup lo} B-cells. Thus, G5pr induction correlated with the prevention of AICD; High in mature splenic CD23{sup hi} B-cells but low in immature B-cells of neonatal mice, sub-lethally irradiated mice, or xid mice. Lack ofmore » G5pr upregulation was associated with the prolonged activation of JNK. The G5pr cDNA transfection protected an immature B-cell line WEHI-231 from BCR-mediated AICD. The differential expression of G5PR might be responsible for the antigen-dependent selection of B-cells.« less

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

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

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.

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...

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.

[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.

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.

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

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.

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.

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.

Isonicotinohydrazones as inhibitors of alkaline phosphatase and ecto-5'-nucleotidase.

PubMed

Channar, Pervaiz Ali; Shah, Syed Jawad Ali; Hassan, Sidra; Nisa, Zaib Un; Lecka, Joanna; Sévigny, Jean; Bajorath, Jürgen; Saeed, Aamer; Iqbal, Jamshed

2017-03-01

A series of isonicotinohydrazide derivatives was synthesized and tested against recombinant human and rat ecto-5'-nucleotidases (h-e5'NT and r-e5'NT) and alkaline phosphatase isozymes including both bovine tissue-non-specific alkaline phosphatase (b-TNAP) and tissue-specific calf intestinal alkaline phosphatase (c-IAP). These enzymes are implicated in vascular calcifications, hypophosphatasia, solid tumors, and cancers, such as colon, lung, breast, pancreas, and ovary. All tested compounds were active against both enzymes. The most potent inhibitor of h-e5'NT was derivative (E)-N'-(1-(3-(4-fluorophenyl)-5-phenyl-4,5-dihydro-1H-pyrazol-1-yl)ethylidene)isonicotinohydrazide (3j), whereas derivative (E)-N'-(4-hydroxy-3-methoxybenzylidene)isonicotinohydrazide (3g) exhibited significant inhibitory activity against r-e5'NT. In addition, the derivative (E)-N'-(4'-chlorobenzylidene)isonicotinohydrazide (3a) was most potent inhibitor against calf intestinal alkaline phosphatase and the derivative (E)-N'-(4-hydroxy-3-methoxybenzylidene)isonicotinohydrazide (3g) was found to be most potent inhibitor of bovine tissue-non-specific alkaline phosphatase. Furthermore, putative binding modes of potent compounds against e5'NT (human and rat e5'NT) and AP (including b-TNAP and c-IAP) were determined computationally. © 2016 John Wiley & Sons A/S.

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

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

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.

2,4-Dihydroxychalcone derivatives as novel potent cell division cycle 25B phosphatase inhibitors and protein tyrosine phosphatase 1B inhibitors.

PubMed

Xie, Chao; Sun, Yuan; Pan, Cheng-Yan; Tang, Li-Ming; Guan, Li-Ping

2014-04-01

Eleven 2,4-dihydroxychalcone compounds were synthesized and identified as reversible and competitive cell division cycle 25 (CDC25) B and protein tyrosine phosphatase (PTP) 1B inhibitors with inhibition values in the micromolar range. The results showed that nine compounds significantly inhibited CDC25B phosphatase, whereas seven compounds inhibited the activity against PTP1B in vitro. Compound 8 had the greatest inhibition activity against CDC25B and PTP1B in vitro, with percentage inhibition values of 97.5% and 96.3% at a dose of 20 microg/mL, respectively. Cytotoxic activity assays revealed that compound 8 was the most potent against HCT116, HeLa, and A549 cells. Furthermore, compound 8 exhibited potent antitumor activity in a colo205 xenograft model.

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.

Docking of oxalyl aryl amino benzoic acid derivatives into PTP1B

PubMed Central

Verma, Neelam; Mittal, Minakshi; Verma, Raman kumar

2008-01-01

Protein Tyrosine Phosphatases (PTPs) that function as negative regulators of the insulin signaling cascade have been identified as novel targets for the therapeutic enhancement of insulin action in insulin resistant disease states. Reducing Protein Tyrosine Phosphatase1B (PTP1B) abundance not only enhances insulin sensitivity and improves glucose metabolism but also protects against obesity induced by high fat feeding. PTP1B inhibitors such as Formylchromone derivatives, 1, 2-Naphthoquinone derivatives and Oxalyl aryl amino benzoic derivatives may eventually find an important clinical role as insulin sensitizers in the management of Type-II Diabetes and metabolic syndrome. We have carried out docking of modified oxalyl aryl amino benzoic acid derivatives into three dimensional structure of PTP1B using BioMed CAChe 6.1. These compounds exhibit good selectivity for PTP1B over most of phosphatases in selectivity panel such as SHP-2, LAR, CD45 and TCPTP found in literature. This series of compounds identified the amino acid residues such as Gly220 and Arg221 are important for achieving specificity via H-bonding interactions. Lipophilic side chain of methionine in modified oxalyl aryl amino benzoic acid derivative [1b (a2, b2, c1, d)] lies in closer vicinity of hydrophobic region of protein consisted of Meth258 and Phe52 in comparison to active ligand. Docking Score in [1b (a2, b2, c1, d)] is -131.740Kcal/mol much better than active ligand score -98.584Kcal/mol. This information can be exploited to design PTP1B specific inhibitors. PMID:19238234

NMDA-induced potentiation of mGluR5 is mediated by activation of protein phosphatase 2B/calcineurin

PubMed Central

Alagarsamy, Sudar; Saugstad, Julie; Warren, Lee; Mansuy, Isabelle M.; Gereau, Robert W.; Conn, P. Jeffrey

2010-01-01

Previous reports have shown that activation of N-methyl-D-aspartate (NMDA) receptors potentiates responses to activation of the group I metabotropic glutamate receptor mGluR5 by reversing PKC-mediated desensitization of this receptor. NMDA-induced reversal of mGluR5 desensitization is dependent on activation of protein phosphatases. However, the specific protein phosphatase involved and the precise mechanism by which NMDA receptor activation reduces mGluR desensitization are not known. We have performed a series of molecular, biochemical, and genetic studies to show that NMDA-induced regulation of mGluR5 is dependent on activation of calcium-dependent protein phosphatase 2B/calcineurin (PP2B/CaN). Furthermore, we report that purified calcineurin directly dephosphorylates the C-terminal tail of mGluR5 at sites that are phosphorylated by PKC. Finally, immunoprecipitation and GST fusion protein pull-down experiments reveal that calcineurin interacts with mGluR5, suggesting that these proteins could be colocalized in a signaling complex. Taken together with previous studies, these data suggest that activation of NMDA receptors leads to activation of calcineurin and that calcineurin modulates mGluR5 function by directly dephosphorylating mGluR5 at PKC sites that are involved in desensitization of this receptor. 2005 Elsevier Ltd. All rights reserved. PMID:16005030

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

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

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.

Jolkinolide B inhibits RANKL-induced osteoclastogenesis by suppressing the activation NF-κB and MAPK signaling pathways.

PubMed

Ma, Xiaojun; Liu, Yupeng; Zhang, Yao; Yu, Xiaobing; Wang, Weiming; Zhao, Dewei

2014-03-07

Osteoclasts together with osteoblasts play pivotal roles in bone remodeling. The unique function and ability of osteoclasts to resorb bone makes them critical in both normal bone homeostasis and pathologic bone diseases such as osteoporosis and rheumatoid arthritis. Thus, new compounds that may inhibit osteoclastogenesis and osteoclast function may be of great value in the treatment of osteoclast-related diseases. In the present study, we examined the effect of jolkinolide B (JB), isolated from the root of Euphorbia fischeriana Steud on receptor activator of NF-κB ligand (RANKL)-induced osteoclast formation. We found that JB inhibited RANKL-induced osteoclast differentiation from bone marrow macrophages (BMMs) without cytotoxicity. Furthermore, the expression of osteoclastic marker genes, such as tartrate-resistant acid phosphatase (TRAP), cathepsin K (CtsK), and calcitonin receptor (CTR), was significantly inhibited. JB inhibited RANKL-induced activation of NF-κB by suppressing RANKL-mediated IκBα degradation. Moreover, JB inhibited RANKL-induced phosphorylation of mitogen-activated protein kinases (p38, JNK, and ERK). This study thus identifies JB as an inhibitor of osteoclast formation and provides evidence that JB might be an alternative medicine for preventing and treating osteolysis. Copyright © 2014 Elsevier Inc. All rights reserved.

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.

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

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

Trivaric acid, a new inhibitor of PTP1b with potent beneficial effect on diabetes.

PubMed

Sun, Wenlong; Zhang, Bowei; Zheng, Haizhou; Zhuang, Chunlin; Li, Xia; Lu, Xinhua; Quan, Chunshan; Dong, Yuesheng; Zheng, Zhihui; Xiu, Zhilong

2017-01-15

To screen a potential PTP1b inhibitor from the microbial origin-based compound library and to investigate the potential anti-diabetic effects of the inhibitor in vivo and determine its primary anti-diabetic mechanism in vitro and in silico. PTP1b inhibitory activity was measured using recombination protein as the enzyme and p-NPP as the substrate. The binding of the inhibitor to PTP1b was analysed by docking in silico and confirmed by ITC experiments. The intracellular signalling pathway was detected by Western blot analysis in HepG2 cells. The anti-diabetic effects were evaluated using a diabetic mice model in vivo. Among 545 microbial origin-based pure compounds tested, trivaric acid, a tridepside, was selected as a PTP1B inhibitor exhibiting strong inhibitory activity with an IC 50 of 173nM. Docking and ITC studies showed that trivaric acid was able to spontaneously bind to PTP1b and may inhibit PTP1b by blocking the catalytic domain of the phosphatase. Trivaric acid also enhanced the ability of insulin to stimulate the IR/IRS/Akt/GLUT2 pathway and increase the glucose consumption in HepG2 cells. In diabetic mice, trivaric acid that had been encapsulated into Eudrgit L100-5.5 showed significant anti-diabetic effects, improving insulin resistance, leptin resistance and lipid profile and weight control at doses of 5mg/kg and 50mg/kg. Trivaric acid is a potential lead compound in the search for anti-diabetic agents targeting PTP1b. Copyright © 2016 Elsevier Inc. All rights reserved.

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

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.

Caffeic Acid Phenethyl Ester (Propolis Extract) Ameliorates Insulin Resistance by Inhibiting JNK and NF-κB Inflammatory Pathways in Diabetic Mice and HepG2 Cell Models.

PubMed

Nie, Jiarui; Chang, Yaning; Li, Yujia; Zhou, Yingjun; Qin, Jiawen; Sun, Zhen; Li, Haibin

2017-10-18

Caffeic acid phenethyl ester (CAPE), extracted from propolis, was evaluated for the ameliorative effects on insulin resistance and the mechanisms were identified, using non-insulin-dependent diabetes mellitus (NIDDM) model mice and insulin resistance (IR) model cells. After 5 weeks of CAPE supplementation, insulin sensitivity, hyperlipidemia, and peroxisome proliferator-activated receptor-α (PPAR-α) levels were improved in mice. Proinflammatory cytokines in serum and the expressions of tumor necrosis factor-alpha (TNF-α) mRNA in tissues were markedly downregulated from CAPE-treated mice. In vitro, CAPE supplement significantly improved glucose consumption, glucose uptake, glycogen content, and oxidative stress and decreased expression of glucose-6-phosphatase (G6Pase) mRNA in cells. Both in vivo and in vitro, CAPE enhanced p-Akt (Ser473) and p-insulin receptor substrate (IRS)-1 (Tyr612), but inhibited p-JNK (Thr183/Tyr185), p-NF-κB p65 (Ser536), and nuclear translocation of p-NF-κB p65 (Ser536). In summary, CAPE can ameliorate insulin resistance through modulation of JNK and NF-κB signaling pathway in mice and HepG2 cells.

"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

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

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

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.

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.

Neogambogic Acid Suppresses Receptor Activator of Nuclear Factor κB Ligand (RANKL)-Induced Osteoclastogenesis by Inhibiting the JNK and NF-κB Pathways in Mouse Bone Marrow-Derived Monocyte/Macrophages.

PubMed

Jin, Gu; Wang, Fang-Fang; Li, Tao; Jia, Dong-Dong; Shen, Yong; Xu, Hai-Chao

2018-04-26

BACKGROUND Neogambogic acid (NGA) is used in traditional Chinese medicine. The aim of this study was to investigate the effects of NGA on gene signaling pathways involved in osteoclastogenesis in mouse bone marrow-derived monocyte/macrophages (BMMs) and on bone resorption in vitro. MATERIAL AND METHODS Primary mouse BMMs were cultured with increasing concentrations of NGA. Real-time polymerase chain reaction was used to study the expression of mRNAs corresponding to gene products specific to receptor activator of NF-κB ligand (RANKL)-induced osteoclast differentiation, including tartrate-resistant acid phosphatase (TRAP), calcitonin receptor (CTR), cathepsin K (CTSK), and nuclear factor of activated T cells c1 (NFATc1). A cell counting kit-8 assay was used to evaluate cell proliferation. Western blotting and confocal immunofluorescence microscopy were used to investigate the signaling pathways. A bone resorption model was used to quantify bone resorption. RESULTS An NGA dose of ≤0.4 μg/ml had no significant effect on the proliferation of mouse BMMs in vitro (P>0.05); concentrations of between 0.1-0.4 μg/ml significantly inhibited RANKL-induced osteoclastogenesis (P<0.01) in a dose-dependent manner. Compared with the control group, NGA significantly reduced RANKL-induced bone resorption in vitro (P <0.01), and downregulated the expression of osteoclast-related mRNAs of TRAP, CTR, CTSK, and NFATc1. NGA suppressed the activation of JNK but not the p38 signaling pathway and significantly reduced NF-κB p65 phosphorylation and the nuclear transport of NF-κB molecules, which inhibited NFATc1 expression. CONCLUSIONS NGA suppressed RANKL-induced osteoclastogenesis by inhibiting the JNK and NF-κB pathways in mouse BMMs in vitro and reduced osteoclastic bone resorption.

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.

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

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.

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.

B56δ-related protein phosphatase 2A dysfunction identified in patients with intellectual disability

PubMed Central

Houge, Gunnar; Haesen, Dorien; Vissers, Lisenka E.L.M.; Mehta, Sarju; Parker, Michael J.; Wright, Michael; Vogt, Julie; McKee, Shane; Tolmie, John L.; Cordeiro, Nuno; Kleefstra, Tjitske; Willemsen, Marjolein H.; Reijnders, Margot R.F.; Berland, Siren; Hayman, Eli; Lahat, Eli; Brilstra, Eva H.; van Gassen, Koen L.I.; Zonneveld-Huijssoon, Evelien; de Bie, Charlotte I.; Hoischen, Alexander; Eichler, Evan E.; Holdhus, Rita; Steen, Vidar M.; Døskeland, Stein Ove; Hurles, Matthew E.; FitzPatrick, David R.; Janssens, Veerle

2015-01-01

Here we report inherited dysregulation of protein phosphatase activity as a cause of intellectual disability (ID). De novo missense mutations in 2 subunits of serine/threonine (Ser/Thr) protein phosphatase 2A (PP2A) were identified in 16 individuals with mild to severe ID, long-lasting hypotonia, epileptic susceptibility, frontal bossing, mild hypertelorism, and downslanting palpebral fissures. PP2A comprises catalytic (C), scaffolding (A), and regulatory (B) subunits that determine subcellular anchoring, substrate specificity, and physiological function. Ten patients had mutations within a highly conserved acidic loop of the PPP2R5D-encoded B56δ regulatory subunit, with the same E198K mutation present in 6 individuals. Five patients had mutations in the PPP2R1A-encoded scaffolding Aα subunit, with the same R182W mutation in 3 individuals. Some Aα cases presented with large ventricles, causing macrocephaly and hydrocephalus suspicion, and all cases exhibited partial or complete corpus callosum agenesis. Functional evaluation revealed that mutant A and B subunits were stable and uncoupled from phosphatase activity. Mutant B56δ was A and C binding–deficient, while mutant Aα subunits bound B56δ well but were unable to bind C or bound a catalytically impaired C, suggesting a dominant-negative effect where mutant subunits hinder dephosphorylation of B56δ-anchored substrates. Moreover, mutant subunit overexpression resulted in hyperphosphorylation of GSK3β, a B56δ-regulated substrate. This effect was in line with clinical observations, supporting a correlation between the ID degree and biochemical disturbance. PMID:26168268

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.

Molecular and Structural Basis for the Roles of Hepatitis C Virus Polymerase NS5B Amino Acids 15, 223, and 321 in Viral Replication and Drug Resistance

PubMed Central

Mosley, Ralph T.; Murakami, Eisuke; Bansal, Shalini; Lugo, Christopher; Bao, Haiying; Otto, Michael J.; Sofia, Michael J.; Furman, Phillip A.

2014-01-01

Resistance to the 2′-F-2′-C-methylguanosine monophosphate nucleotide hepatitis C virus (HCV) inhibitors PSI-352938 and PSI-353661 was associated with a combination of amino acid changes (changes of S to G at position 15 [S15G], C223H, and V321I) within the genotype 2a nonstructural protein 5B (NS5B), an RNA-dependent RNA polymerase. To understand the role of these residues in viral replication, we examined the effects of single and multiple point mutations on replication fitness and inhibition by a series of nucleotide analog inhibitors. An acidic residue at position 15 reduced replicon fitness, consistent with its proximity to the RNA template. A change of the residue at position 223 to an acidic or large residue reduced replicon fitness, consistent with its proposed proximity to the incoming nucleoside triphosphate (NTP). A change of the residue at position 321 to a charged residue was not tolerated, consistent with its position within a hydrophobic cavity. This triple resistance mutation was specific to both genotype 2a virus and 2′-F-2′-C-methylguanosine inhibitors. A crystal structure of the NS5B S15G/C223H/V321I mutant of the JFH-1 isolate exhibited rearrangement to a conformation potentially consistent with short primer-template RNA binding, which could suggest a mechanism of resistance accomplished through a change in the NS5B conformation, which was better tolerated by genotype 2a virus than by viruses of other genotypes. PMID:25182647

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

Protein-Tyrosine Phosphatase-1B Mediates Sleep Fragmentation-Induced Insulin Resistance and Visceral Adipose Tissue Inflammation in Mice.

PubMed

Gozal, David; Khalyfa, Abdelnaby; Qiao, Zhuanghong; Akbarpour, Mahzad; Maccari, Rosanna; Ottanà, Rosaria

2017-09-01

Sleep fragmentation (SF) is highly prevalent and has emerged as an important contributing factor to obesity and metabolic syndrome. We hypothesized that SF-induced increases in protein tyrosine phosphatase-1B (PTP-1B) expression and activity underlie increased food intake, inflammation, and leptin and insulin resistance. Wild-type (WT) and ObR-PTP-1b-/- mice (Tg) were exposed to SF and control sleep (SC), and food intake was monitored. WT mice received a PTP-1B inhibitor (RO-7d; Tx) or vehicle (Veh). Upon completion of exposures, systemic insulin and leptin sensitivity tests were performed as well as assessment of visceral white adipose tissue (vWAT) insulin receptor sensitivity and macrophages (ATM) polarity. SF increased food intake in either untreated or Veh-treated WT mice. Leptin-induced hypothalamic STAT3 phosphorylation was decreased, PTP-1B activity was increased, and reduced insulin sensitivity emerged both systemic and in vWAT, with the latter displaying proinflammatory ATM polarity changes. All of the SF-induced effects were abrogated following PTP-1B inhibitor treatment and in Tg mice. SF induces increased food intake, reduced leptin signaling in hypothalamus, systemic insulin resistance, and reduced vWAT insulin sensitivity and inflammation that are mediated by increased PTP-1B activity. Thus, PTP-1B may represent a viable therapeutic target in the context of SF-induced weight gain and metabolic dysfunction. © Sleep Research Society 2017. Published by Oxford University Press on behalf of the Sleep Research Society. All rights reserved. For permissions, please e-mail journals.permissions@oup.com.

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

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

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

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

In vitro and in vivo synergism between amoxicillin and clavulanic acid against ampicillin-resistant Haemophilus influenzae type b.

PubMed Central

Yogev, R; Melick, C; Kabat, W J

1981-01-01

Eight strans of ampicillin-resistant beta-lactamase-producing Haemophilus influenzae type b were studied in vitro for synergy between amoxicillin and clavulanic acid. The minimal inhibitory concentrations for amoxicillin alone were 6.25 to 12.5 microgram/ml, and for clavulanic acid alone they were 12.5 to 25 microgram/ml. However, seven of eight strains were inhibited by a combination of 0.36 microgram of amoxicillin and 0.36 microgram of clavulanic acid per ml. Infant rat models of bacteremia and meningitis were used to test the efficacy of amoxicillin and clavulanic acid alone and in combination upon four strains of ampicillin-resistant H. influenzae. Neither amoxicillin alone (27 animals) nor clavulanic acid alone (20 animals) sterilized the blood or cerebrospinal fluid of the animals. In contrast, 30 of 33 blood cultures and 29 of 33 cerebrospinal fluid cultures were sterile when a combination of the two drugs in the same dosages was used. The observed in vitro and in vivo synergism between amoxicillin and clavulanic acid suggests that the combination may be effective therapy for invasive infections in humans caused by ampicillin-resistant H. influenzae type b. PMID:6973952

Mutations in the PP2A regulatory subunit B family genes PPP2R5B, PPP2R5C and PPP2R5D cause human overgrowth.

PubMed

Loveday, Chey; Tatton-Brown, Katrina; Clarke, Matthew; Westwood, Isaac; Renwick, Anthony; Ramsay, Emma; Nemeth, Andrea; Campbell, Jennifer; Joss, Shelagh; Gardner, McKinlay; Zachariou, Anna; Elliott, Anna; Ruark, Elise; van Montfort, Rob; Rahman, Nazneen

2015-09-01

Overgrowth syndromes comprise a group of heterogeneous disorders characterised by excessive growth parameters, often in association with intellectual disability. To identify new causes of human overgrowth, we have been undertaking trio-based exome sequencing studies in overgrowth patients and their unaffected parents. Prioritisation of functionally relevant genes with multiple unique de novo mutations revealed four mutations in protein phosphatase 2A (PP2A) regulatory subunit B family genes protein phosphatase 2, regulatory Subunit B', beta (PPP2R5B); protein phosphatase 2, regulatory Subunit B', gamma (PPP2R5C); and protein phosphatase 2, regulatory Subunit B', delta (PPP2R5D). This observation in 3 related genes in 111 individuals with a similar phenotype is greatly in excess of the expected number, as determined from gene-specific de novo mutation rates (P = 1.43 × 10(-10)). Analysis of exome-sequencing data from a follow-up series of overgrowth probands identified a further pathogenic mutation, bringing the total number of affected individuals to 5. Heterozygotes shared similar phenotypic features including increased height, increased head circumference and intellectual disability. The mutations clustered within a region of nine amino acid residues in the aligned protein sequences (P = 1.6 × 10(-5)). We mapped the mutations onto the crystal structure of the PP2A holoenzyme complex to predict their molecular and functional consequences. These studies suggest that the mutations may affect substrate binding, thus perturbing the ability of PP2A to dephosphorylate particular protein substrates. PP2A is a major negative regulator of v-akt murine thymoma viral oncogene homolog 1 (AKT). Thus, our data further expand the list of genes encoding components of the phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K)/AKT signalling cascade that are disrupted in human overgrowth conditions. © The Author 2015. Published by Oxford University Press.

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...

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.

[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.

Crystal structure of human dual specificity phosphatase, JNK stimulatory phosphatase-1, at 1.5 A resolution.

PubMed

Yokota, Takehiro; Nara, Yukinori; Kashima, Akiko; Matsubara, Keiko; Misawa, Satoru; Kato, Ryohei; Sugio, Shigetoshi

2007-02-01

Human JNK stimulatory phosphatase-1 (JSP-1) is a novel member of dual specificity phosphatases. A C-terminus truncated JSP-1 was expressed in Escherichia coli and was crystallized using the sitting-drop vapor diffusion method. Thin-plate crystals obtained at 278 K belong to a monoclinic space group, C2, with unit-cell parameters a = 84.0 A, b = 49.3 A, c = 47.3 A, and beta = 119.5 degrees , and diffract up to 1.5 A resolution at 100 K. The structure of JSP-1 has a single compact (alpha/beta) domain, which consists of six alpha-helices and five beta-strands, and shows a conserved structural scaffold in regard to both DSPs and PTPs. A cleft formed by a PTP-loop at the active site is very shallow, and is occupied by one sulfonate compound, MES, at the bottom. In the binary complex structure of JSP-1 with MES, the conformations of three important segments in regard to the catalytic mechanism are not similar to those in PTP1B. JSP-1 has no loop corresponding to the Lys120-loop of PTP1B, and tryptophan residue corresponding to the substrate-stacking in PTP1B is substituted by alanine residue in JSP-1. Copyright 2006 Wiley-Liss, Inc.

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.

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.

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

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.

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.

Redox regulation of protein tyrosine phosphatase 1B (PTP1B): Importance of steric and electronic effects on the unusual cyclization of the sulfenic acid intermediate to a sulfenyl amide

NASA Astrophysics Data System (ADS)

Sarma, Bani Kanta

2013-09-01

The redox regulation of protein tyrosine phosphatase 1B (PTP1B) via the unusual transformation of its sulfenic acid (PTP1B-SOH) to a cyclic sulfenyl amide intermediate is studied by using small molecule chemical models. These studies suggest that the sulfenic acids derived from the H2O2-mediated reactions o-amido thiophenols do not efficiently cyclize to sulfenyl amides and the sulfenic acids produced in situ can be trapped by using methyl iodide. Theoretical calculations suggest that the most stable conformer of such sulfenic acids are stabilized by nO → σ*S-OH orbital interactions, which force the -OH group to adopt a position trans to the S⋯O interaction, leading to an almost linear arrangement of the O⋯S-O moiety and this may be the reason for the slow cyclization of such sulfenic acids to their corresponding sulfenyl amides. On the other hand, additional substituents at the 6-position of o-amido phenylsulfenic acids that can induce steric environment and alter the electronic properties around the sulfenic acid moiety by S⋯N or S⋯O nonbonded interactions destabilize the sulfenic acids by inducing strain in the molecule. This may lead to efficient the cyclization of such sulfenic acids. This model study suggests that the amino acid residues in the close proximity of the sulfenic acid moiety in PTP1B may play an important role in the cyclization of PTP1B-SOH to produce the corresponding sulfenyl amide.

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

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.

Phosphoinositide 5- and 3-phosphatase activities of a voltage-sensing phosphatase in living cells show identical voltage dependence

PubMed Central

Keum, Dongil; Kim, Dong-Il; Suh, Byung-Chang

2016-01-01

Voltage-sensing phosphatases (VSPs) are homologs of phosphatase and tensin homolog (PTEN), a phosphatidylinositol 3,4-bisphosphate [PI(3,4)P2] and phosphatidylinositol 3,4,5-trisphosphate [PI(3,4,5)P3] 3-phosphatase. However, VSPs have a wider range of substrates, cleaving 3-phosphate from PI(3,4)P2 and probably PI(3,4,5)P3 as well as 5-phosphate from phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] and PI(3,4,5)P3 in response to membrane depolarization. Recent proposals say these reactions have differing voltage dependence. Using Förster resonance energy transfer probes specific for different PIs in living cells with zebrafish VSP, we quantitate both voltage-dependent 5- and 3-phosphatase subreactions against endogenous substrates. These activities become apparent with different voltage thresholds, voltage sensitivities, and catalytic rates. As an analytical tool, we refine a kinetic model that includes the endogenous pools of phosphoinositides, endogenous phosphatase and kinase reactions connecting them, and four exogenous voltage-dependent 5- and 3-phosphatase subreactions of VSP. We show that apparent voltage threshold differences for seeing effects of the 5- and 3-phosphatase activities in cells are not due to different intrinsic voltage dependence of these reactions. Rather, the reactions have a common voltage dependence, and apparent differences arise only because each VSP subreaction has a different absolute catalytic rate that begins to surpass the respective endogenous enzyme activities at different voltages. For zebrafish VSP, our modeling revealed that 3-phosphatase activity against PI(3,4,5)P3 is 55-fold slower than 5-phosphatase activity against PI(4,5)P2; thus, PI(4,5)P2 generated more slowly from dephosphorylating PI(3,4,5)P3 might never accumulate. When 5-phosphatase activity was counteracted by coexpression of a phosphatidylinositol 4-phosphate 5-kinase, there was accumulation of PI(4,5)P2 in parallel to PI(3,4,5)P3 dephosphorylation

Novel Mixed-Type Inhibitors of Protein Tyrosine Phosphatase 1B. Kinetic and Computational Studies.

PubMed

Sarabia-Sánchez, Marie Jazmín; Trejo-Soto, Pedro Josué; Velázquez-López, José Miguel; Carvente-García, Carlos; Castillo, Rafael; Hernández-Campos, Alicia; Avitia-Domínguez, Claudia; Enríquez-Mendiola, Daniel; Sierra-Campos, Erick; Valdez-Solana, Mónica; Salas-Pacheco, José Manuel; Téllez-Valencia, Alfredo

2017-12-20

The Atlas of Diabetes reports 415 million diabetics in the world, a number that has surpassed in half the expected time the twenty year projection. Type 2 diabetes is the most frequent form of the disease; it is characterized by a defect in the secretion of insulin and a resistance in its target organs. In the search for new antidiabetic drugs, one of the principal strategies consists in promoting the action of insulin. In this sense, attention has been centered in the protein tyrosine phosphatase 1B (PTP1B), a protein whose overexpression or increase of its activity has been related in many studies with insulin resistance. In the present work, a chemical library of 250 compounds was evaluated to determine their inhibition capability on the protein PTP1B. Ten molecules inhibited over the 50% of the activity of the PTP1B, the three most potent molecules were selected for its characterization, reporting Ki values of 5.2, 4.2 and 41.3 µM, for compounds 1 , 2 , and 3 , respectively. Docking and molecular dynamics studies revealed that the three inhibitors made interactions with residues at the secondary binding site to phosphate, exclusive for PTP1B. The data reported here support these compounds as hits for the design more potent and selective inhibitors against PTP1B in the search of new antidiabetic treatment.

Involvement of the Eukaryote-Like Kinase-Phosphatase System and a Protein That Interacts with Penicillin-Binding Protein 5 in Emergence of Cephalosporin Resistance in Cephalosporin-Sensitive Class A Penicillin-Binding Protein Mutants in Enterococcus faecium.

PubMed

Desbonnet, Charlene; Tait-Kamradt, Amelia; Garcia-Solache, Monica; Dunman, Paul; Coleman, Jeffrey; Arthur, Michel; Rice, Louis B

2016-04-05

The intrinsic resistance of Enterococcus faecium to ceftriaxone and cefepime (here referred to as "cephalosporins") is reliant on the presence of class A penicillin-binding proteins (Pbps) PbpF and PonA. Mutants lacking these Pbps exhibit cephalosporin susceptibility that is reversible by exposure to penicillin and by selection on cephalosporin-containing medium. We selected two cephalosporin-resistant mutants (Cro1 and Cro2) of class A Pbp-deficient E. faecium CV598. Genome analysis revealed changes in the serine-threonine kinase Stk in Cro1 and a truncation in the associated phosphatase StpA in Cro2 whose respective involvements in resistance were confirmed in separate complementation experiments. In an additional effort to identify proteins linked to cephalosporin resistance, we performed tandem affinity purification using Pbp5 as bait in penicillin-exposed E. faecium; these experiments yielded a protein designated Pbp5-associated protein (P5AP). Transcription of the P5AP gene was increased after exposure to penicillin in wild-type strains and in Cro2 and suppressed in Cro2 complemented with the wild-type stpA Transformation of class A Pbp-deficient strains with the plasmid-carried P5AP gene conferred cephalosporin resistance. These data suggest that Pbp5-associated cephalosporin resistance in E. faecium devoid of typical class A Pbps is related to the presence of P5AP, whose expression is influenced by the activity of the serine-threonine phosphatase/kinase system. β-Lactam antibiotics remain our most effective therapies against susceptible Gram-positive bacteria. The intrinsic resistance of Enterococcus faecium to β-lactams, particularly to cephalosporins, therefore represents a major limitation of therapy. Although the primary mechanism of resistance to β-lactams in E. faecium is the presence of low-affinity monofunctional transpeptidase (class B) penicillin-binding protein Pbp5, the interaction of Pbp5 with other proteins is fundamental to maintain a

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

Naturally occurring resistance mutations within the core and NS5B regions in hepatitis C genotypes, particularly genotype 5a, in South Africa.

PubMed

Prabdial-Sing, N; Blackard, J T; Puren, A J; Mahomed, A; Abuelhassan, W; Mahlangu, J; Vermeulen, M; Bowyer, S M

2016-03-01

Approximately 1 million South Africans are infected with Hepatitis C virus (HCV). The standard of care (SOC) in South Africa is combination therapy (pegylated interferon and ribavirin). HCV genotypes and/or mutations in the core/non-structural regions have been associated with response to therapy and/or disease progression. This study examines mutations in the core (29-280 amino acids, including ∼ 90 E1 amino acids) and NS5B (241-306 amino acids) regions on pre-treatment isolates from patients attending Johannesburg hospitals or asymptomatic South African blood donors. Diversity within known CD4+ and CD8+ T-cell epitopes was also explored. Samples grouped into subtypes 1a(N = 10) 1b(N = 12), 3a(N = 5), 4a(N = 3) and 5a(N = 61). Two mutations, associated with interferon resistance-R70Q and T110N-were present in 29 genotype 5a core sequences. No resistance mutation to NS5B nucleotide inhibitors, sofosbuvir was found. Six putative CD8+ and one CD4+ T-cell epitope sequence in the core region showed binding scores of <300 IC50nM to HLA alleles frequently observed in the South African population. No known CD8+ and CD4+ T-cell epitopes were mapped in the NS5B region. The analysis begs the question whether those infected with genotype 5a will benefit better on interferon-free combination therapies. This study provides new insight into one of the lesser studied HCV genotypes and compares the diversity seen in a large pre-treatment cohort with other subtypes. Copyright © 2015 Elsevier B.V. All rights reserved.

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.

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…

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.

Phosphoinositide 5-phosphatase activities control cell motility in glioblastoma: Two phosphoinositides PI(4,5)P2 and PI(3,4)P2 are involved.

PubMed

Ramos, Ana Raquel; Elong Edimo, William's; Erneux, Christophe

2018-01-01

Inositol polyphosphate 5-phosphatases or phosphoinositide 5-phosphatases (PI 5-phosphatases) are enzymes that can act on soluble inositol phosphates and/or phosphoinositides (PIs). Several PI 5-phosphatases have been linked to human genetic diseases, in particular the Lowe protein or OCRL which is mutated in the Lowe syndrome. There are 10 different members of this family and 9 of them can use PIs as substrate. One of these substrates, PI(3,4,5)P3 binds to specific PH domains and recruits as effectors specific proteins to signaling complexes. Protein kinase B is one target protein and activation of the kinase will have a major impact on cell proliferation, survival and cell metabolism. Two other PIs, PI(4,5)P2 and PI(3,4)P2, are produced or used as substrates of PI 5-phosphatases (OCRL, INPP5B, SHIP1/2, SYNJ1/2, INPP5K, INPP5J, INPP5E). The inositol lipids may influence many aspects of cytoskeletal organization, lamellipodia formation and F-actin polymerization. PI 5-phosphatases have been reported to control cell migration, adhesion, polarity and cell invasion particularly in cancer cells. In glioblastoma, reducing SHIP2 expression can positively or negatively affect the speed of cell migration depending on the glioblastoma cell type. The two PI 5-phosphatases SHIP2 or SKIP could be localized at the plasma membrane and can reduce either PI(3,4,5)P3 or PI(4,5)P2 abundance. In the glioblastoma 1321 N1 cells, SHIP2 controls plasma membrane PI(4,5)P2 thereby participating in the control of cell migration. Copyright © 2017 Elsevier Ltd. All rights reserved.

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.

[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).

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

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

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.

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.

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.

Streptococcus pneumoniae Phosphotyrosine Phosphatase CpsB and Alterations in Capsule Production Resulting from Changes in Oxygen Availability

PubMed Central

Geno, K. Aaron; Hauser, Jocelyn R.; Gupta, Kanupriya

2014-01-01

Streptococcus pneumoniae produces a protective capsular polysaccharide whose production must be modulated for bacterial survival within various host niches. Capsule production is affected in part by a phosphoregulatory system comprised of CpsB, CpsC, and CpsD. Here, we found that growth of serotype 2 strain D39 under conditions of increased oxygen availability resulted in decreased capsule levels concurrent with an ∼5-fold increase in Cps2B-mediated phosphatase activity. The change in Cps2B phosphatase activity did not result from alterations in the levels of either the cps2B transcript or the Cps2B protein. Recombinant Cps2B expressed in Escherichia coli similarly exhibited increased phosphatase activity under conditions of high-oxygen growth. S. pneumoniae D39 derivatives with defined deletion or point mutations in cps2B demonstrated reduced phosphatase activity with corresponding increases in levels of Cps2D tyrosine phosphorylation. There was, however, no correlation between these phenotypes and the level of capsule production. During growth under reduced-oxygen conditions, the Cps2B protein was essential for parental levels of capsule, but phosphatase activity alone could be eliminated without an effect on capsule. Under increased-oxygen conditions, deletion of cps2B did not affect capsule levels. These results indicate that neither Cps2B phosphatase activity nor Cps2D phosphorylation levels per se are determinants of capsule levels, whereas the Cps2B protein is important for capsule production during growth under conditions of reduced but not enhanced oxygen availability. Roles for factors outside the capsule locus, possible interactions between capsule regulatory proteins, and links to other cellular processes are also suggested by the results described in this study. PMID:24659769

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.

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.

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...

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.

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.

Competitive protein tyrosine phosphatase 1B (PTP1B) inhibitors, prenylated caged xanthones from Garcinia hanburyi and their inhibitory mechanism.

PubMed

Tan, Xue Fei; Uddin, Zia; Park, Chanin; Song, Yeong Hun; Son, Minky; Lee, Keun Woo; Park, Ki Hun

2017-04-15

Protein tyrosine phosphatase 1B (PTP1B) plays important role in diabetes, obesity and cancer. The methanol extract of the gum resin of Garcinia hanburyi (G. hanburyi) showed potent PTP1B inhibition at 10µg/ml. The active compounds were identified as prenylated caged xanthones (1-9) which inhibited PTP1B in dose-dependent manner. Carboxybutenyl group within caged motif (A ring) was found to play a critical role in enzyme inhibition such as 1-6 (IC 50 s=0.47-4.69µM), whereas compounds having hydroxymethylbutenyl 7 (IC 50 =70.25µM) and methylbutenyl 8 (IC 50 >200µM) showed less activity. The most potent inhibitor, gambogic acid 1 (IC 50 =0.47µM) showed 30-fold more potency than ursolic acid (IC 50 =15.5µM), a positive control. In kinetic study, all isolated xanthones behaved as competitive inhibitors which were fully demonstrated with K m , V max and K ik /K iv ratio. It was also proved that inhibitor 1 operated under the enzyme isomerization model having k 5 =0.0751µM - 1 S - 1 , k 6 =0.0249µM - 1 S - 1 and K i app =0.499µM. To develop a pharmacophore model, we explored the binding sites of compound 1 and 7 in PTP1B. These modeling results were in agreement with our findings, which revealed that the inhibitory activities are tightly related to caged motif and prenyl group in A ring. Copyright © 2017 Elsevier Ltd. All rights reserved.

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.

TRAF Family Member-Associated NF-κB Activator (TANK) Induced by RANKL Negatively Regulates Osteoclasts Survival and Function

PubMed Central

Wu, Mengrui; Wang, Yiping; Deng, Lianfu; Chen, Wei; Li, Yi-Ping

2012-01-01

Osteoclasts are the principle bone-resorbing cells. Precise control of balanced osteoclast activity is indispensable for bone homeostasis. Osteoclast activation mediated by RANK-TRAF6 axis has been clearly identified. However, a negative regulation-machinery in osteoclast remains unclear. TRAF family member-associated NF-κB activator (TANK) is induced by about 10 folds during osteoclastogenesis, according to a genome-wide analysis of gene expression before and after osteoclast maturation, and confirmed by western blot and quantitative RT-PCR. Bone marrow macrophages (BMMs) transduced with lentivirus carrying tank-shRNA were induced to form osteoclast in the presence of RANKL and M-CSF. Tank expression was downregulated by 90% by Tank-shRNA, which is confirmed by western blot. Compared with wild-type (WT) cells, osteoclastogenesis of Tank-silenced BMMs was increased, according to tartrate-resistant acid phosphatase (TRAP) stain on day 5 and day 7. Number of bone resorption pits by Tank-silenced osteoclasts was increased by 176% compared with WT cells, as shown by wheat germ agglutinin (WGA) stain and scanning electronic microscope (SEM) analysis. Survival rate of Tank-silenced mature osteoclast is also increased. However, acid production of Tank-knockdown cells was not changed compared with control cells. IκBα phosphorylation is increased in tank-silenced cells, indicating that TANK may negatively regulate NF-κB activity in osteoclast. In conclusion, Tank, whose expression is increased during osteoclastogenesis, inhibits osteoclast formation, activity and survival, by regulating NF-κB activity and c-FLIP expression. Tank enrolls itself in a negative feedback loop in bone resorption. These results may provide means for therapeutic intervention in diseases of excessive bone resorption. PMID:23139637

TRAF family member-associated NF-κB activator (TANK) induced by RANKL negatively regulates osteoclasts survival and function.

PubMed

Wu, Mengrui; Wang, Yiping; Deng, Lianfu; Chen, Wei; Li, Yi-Ping

2012-01-01

Osteoclasts are the principle bone-resorbing cells. Precise control of balanced osteoclast activity is indispensable for bone homeostasis. Osteoclast activation mediated by RANK-TRAF6 axis has been clearly identified. However, a negative regulation-machinery in osteoclast remains unclear. TRAF family member-associated NF-κB activator (TANK) is induced by about 10 folds during osteoclastogenesis, according to a genome-wide analysis of gene expression before and after osteoclast maturation, and confirmed by western blot and quantitative RT-PCR. Bone marrow macrophages (BMMs) transduced with lentivirus carrying tank-shRNA were induced to form osteoclast in the presence of RANKL and M-CSF. Tank expression was downregulated by 90% by Tank-shRNA, which is confirmed by western blot. Compared with wild-type (WT) cells, osteoclastogenesis of Tank-silenced BMMs was increased, according to tartrate-resistant acid phosphatase (TRAP) stain on day 5 and day 7. Number of bone resorption pits by Tank-silenced osteoclasts was increased by 176% compared with WT cells, as shown by wheat germ agglutinin (WGA) stain and scanning electronic microscope (SEM) analysis. Survival rate of Tank-silenced mature osteoclast is also increased. However, acid production of Tank-knockdown cells was not changed compared with control cells. IκBα phosphorylation is increased in tank-silenced cells, indicating that TANK may negatively regulate NF-κB activity in osteoclast. In conclusion, Tank, whose expression is increased during osteoclastogenesis, inhibits osteoclast formation, activity and survival, by regulating NF-κB activity and c-FLIP expression. Tank enrolls itself in a negative feedback loop in bone resorption. These results may provide means for therapeutic intervention in diseases of excessive bone resorption.

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

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...

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...

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

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.

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.

Benzene Polyphosphates as Tools for Cell Signalling: Inhibition of Inositol 1,4,5-Trisphosphate 5-Phosphatase and Interaction with the PH Domain of Protein Kinase Bα

PubMed Central

Mills, Stephen J; Vandeput, Fabrice; Trusselle, Melanie N.; Safrany, Stephen T.; Erneux, Christophe; Potter, Barry V. L.

2009-01-01

Novel benzene polyphosphates were synthesised as inositol polyphosphate mimics and evaluated against both type-I inositol 1,4,5-trisphosphate 5-phosphatase, which only binds soluble inositol polyphosphates, and the PH domain of protein kinase Bα (PKBα), which can bind both soluble inositol polyphosphates and inositol phospholipids. The most potent trisphosphate 5-phosphatase inhibitor is benzene 1,2,4-trisphosphate 2, (IC50 of 14 μm) a potential mimic of d-myo-inositol 1,4,5-trisphosphate, and the most potent tetrakisphosphate Ins(1,4,5)P3 5-phosphatase inhibitor is benzene 1,2,4,5-tetrakisphosphate, with an IC50 of 4 μm. Biphenyl 2,3′,4,5′,6-pentakisphosphate 4 was the most potent inhibitor evaluated against type I Ins(1,4,5)P3 5-phosphatase (IC50 of 1 μm). All new benzene polyphosphates are resistant to dephosphorylation by type I Ins(1,4,5)P3 5-phosphatase. Unexpectedly, all benzene polyphosphates studied bind to the PH domain of PKBα with apparent higher affinity than type 1 Ins(1,4,5)P3 5-phosphatase. The most potent ligand for PKBα PH domain is biphenyl 2,3′,4,5′,6-pentakisphosphate 4 (Ki = 27 nm), measured by inhibition of biotinylated diC8-PtdIns(3,4)P2 binding. The ca 80-fold enhancement of binding relative to parent benzene trisphosphate is rationalised by the involvement of a cation–π interaction. These new molecular tools will be of potential use in structural and cell signalling studies. PMID:18574825

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.

Natural products possessing protein tyrosine phosphatase 1B (PTP1B) inhibitory activity found in the last decades

PubMed Central

Jiang, Cheng-shi; Liang, Lin-fu; Guo, Yue-wei

2012-01-01

This article provides an overview of approximately 300 secondary metabolites with inhibitory activity against protein tyrosine phosphatase 1B (PTP1B), which were isolated from various natural sources or derived from synthetic process in the last decades. The structure-activity relationship and the selectivity of some compounds against other protein phosphatases were also discussed. Potential pharmaceutical applications of several PTP1B inhibitors were presented. PMID:22941286

Protein Tyrosine Phosphatase 1B Inhibition and Glucose Uptake Potentials of Mulberrofuran G, Albanol B, and Kuwanon G from Root Bark of Morus alba L. in Insulin-Resistant HepG2 Cells: An In Vitro and In Silico Study.

PubMed

Paudel, Pradeep; Yu, Ting; Seong, Su Hui; Kuk, Eun Bi; Jung, Hyun Ah; Choi, Jae Sue

2018-05-22

Type II diabetes mellitus (T2DM) is the most common form of diabetes and has become a major health problem across the world. The root bark of Morus alba L. is widely used in Traditional Chinese Medicine for treatment and management of diabetes. The aim of the present study was to evaluate the enzyme inhibitory potentials of three principle components, mulberrofuran G ( 1 ), albanol B ( 2 ), and kuwanon G ( 3 ) in M. alba root bark against diabetes, establish their enzyme kinetics, carry out a molecular docking simulation, and demonstrate the glucose uptake activity in insulin-resistant HepG2 cells. Compounds 1 ⁻ 3 showed potent mixed-type enzyme inhibition against protein tyrosine phosphatase 1B (PTP1B) and α-glucosidase. In particular, molecular docking simulations of 1 ⁻ 3 demonstrated negative binding energies in both enzymes. Moreover, 1 ⁻ 3 were non-toxic up to 5 µM concentration in HepG2 cells and enhanced glucose uptake significantly and decreased PTP1B expression in a dose-dependent manner in insulin-resistant HepG2 cells. Our overall results depict 1 ⁻ 3 from M. alba root bark as dual inhibitors of PTP1B and α-glucosidase enzymes, as well as insulin sensitizers. These active constituents in M. alba may potentially be utilized as an effective treatment for T2DM.

Structural Determinants of Substrate Recognition in the HAD Superfamily Member D-glycero-D-manno-Heptose-1,7-bisphosphate Phosphatase (GmhB)

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

Nguyen, H.; Wang, L; Huang, H

2010-01-01

The haloalkanoic acid dehalogenase (HAD) enzyme superfamily is the largest family of phosphohydrolases. In HAD members, the structural elements that provide the binding interactions that support substrate specificity are separated from those that orchestrate catalysis. For most HAD phosphatases, a cap domain functions in substrate recognition. However, for the HAD phosphatases that lack a cap domain, an alternate strategy for substrate selection must be operative. One such HAD phosphatase, GmhB of the HisB subfamily, was selected for structure-function analysis. Herein, the X-ray crystallographic structures of Escherichia coli GmhB in the apo form (1.6 {angstrom} resolution), in a complex with Mg{supmore » 2+} and orthophosphate (1.8 {angstrom} resolution), and in a complex with Mg{sup 2+} and D-glycero-D-manno-heptose 1{beta},7-bisphosphate (2.2 {angstrom} resolution) were determined, in addition to the structure of Bordetella bronchiseptica GmhB bound to Mg{sup 2+} and orthophosphate (1.7 {angstrom} resolution). The structures show that in place of a cap domain, the GmhB catalytic site is elaborated by three peptide inserts or loops that pack to form a concave, semicircular surface around the substrate leaving group. Structure-guided kinetic analysis of site-directed mutants was conducted in parallel with a bioinformatics study of sequence diversification within the HisB subfamily to identify loop residues that serve as substrate recognition elements and that distinguish GmhB from its subfamily counterpart, the histidinol-phosphate phosphatase domain of HisB. We show that GmhB and the histidinol-phosphate phosphatase domain use the same design of three substrate recognition loops inserted into the cap domain yet, through selective residue usage on the loops, have achieved unique substrate specificity and thus novel biochemical function.« less

Selective activators of protein phosphatase 5 target the auto-inhibitory mechanism.

PubMed

Haslbeck, Veronika; Drazic, Adrian; Eckl, Julia M; Alte, Ferdinand; Helmuth, Martin; Popowicz, Grzegorz; Schmidt, Werner; Braun, Frank; Weiwad, Matthias; Fischer, Gunter; Gemmecker, Gerd; Sattler, Michael; Striggow, Frank; Groll, Michael; Richter, Klaus

2015-04-20

Protein phosphatase 5 (PP5) is an evolutionary conserved serine/threonine phosphatase. Its dephosphorylation activity modulates a diverse set of cellular factors including protein kinases and the microtubule-associated tau protein involved in neurodegenerative disorders. It is auto-regulated by its heat-shock protein (Hsp90)-interacting tetratricopeptide repeat (TPR) domain and its C-terminal α-helix. In the present study, we report the identification of five specific PP5 activators [PP5 small-molecule activators (P5SAs)] that enhance the phosphatase activity up to 8-fold. The compounds are allosteric modulators accelerating efficiently the turnover rate of PP5, but do barely affect substrate binding or the interaction between PP5 and the chaperone Hsp90. Enzymatic studies imply that the compounds bind to the phosphatase domain of PP5. For the most promising compound crystallographic comparisons of the apo PP5 and the PP5-P5SA-2 complex indicate a relaxation of the auto-inhibited state of PP5. Residual electron density and mutation analyses in PP5 suggest activator binding to a pocket in the phosphatase/TPR domain interface, which may exert regulatory functions. These compounds thus may expose regulatory mechanisms in the PP5 enzyme and serve to develop optimized activators based on these scaffolds. © 2015 Authors.

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.

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.

Requirement of SH2-containing protein tyrosine phosphatases SHP-1 and SHP-2 for paired immunoglobulin-like receptor B (PIR-B)-mediated inhibitory signal.

PubMed

Maeda, A; Kurosaki, M; Ono, M; Takai, T; Kurosaki, T

1998-04-20

Paired immunoglobulin-like receptor B (PIR-B) (p91) molecule has been proposed to function as an inhibitory receptor in B cells and myeloid lineage cells. We demonstrate here that the cytoplasmic region of PIR-B is capable of inhibiting B cell activation. Mutational analysis of five cytoplasmic tyrosines indicate that tyrosine 771 in the motif VxYxxL plays the most crucial role in mediating the inhibitory signal. PIR-B-mediated inhibition was markedly reduced in the SH2-containing protein tyrosine phosphatases SHP-1 and SHP-2 double-deficient DT40 B cells, whereas this inhibition was unaffected in the inositol polyphosphate 5'-phosphatase SHIP-deficient cells. These data demonstrate that PIR-B can negatively regulate B cell receptor activation and that this PIR-B-mediated inhibition requires redundant functions of SHP-1 and SHP-2.

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.

Protein Tyrosine Phosphatase 1B Regulates Pyruvate Kinase M2 Tyrosine Phosphorylation*

PubMed Central

Bettaieb, Ahmed; Bakke, Jesse; Nagata, Naoto; Matsuo, Kosuke; Xi, Yannan; Liu, Siming; AbouBechara, Daniel; Melhem, Ramzi; Stanhope, Kimber; Cummings, Bethany; Graham, James; Bremer, Andrew; Zhang, Sheng; Lyssiotis, Costas A.; Zhang, Zhong-Yin; Cantley, Lewis C.; Havel, Peter J.; Haj, Fawaz G.

2013-01-01

Protein-tyrosine phosphatase 1B (PTP1B) is a physiological regulator of glucose homeostasis and adiposity and is a drug target for the treatment of obesity and diabetes. Here we identify pyruvate kinase M2 (PKM2) as a novel PTP1B substrate in adipocytes. PTP1B deficiency leads to increased PKM2 total tyrosine and Tyr105 phosphorylation in cultured adipocytes and in vivo. Substrate trapping and mutagenesis studies identify PKM2 Tyr-105 and Tyr-148 as key sites that mediate PTP1B-PKM2 interaction. In addition, in vitro analyses illustrate a direct effect of Tyr-105 phosphorylation on PKM2 activity in adipocytes. Importantly, PTP1B pharmacological inhibition increased PKM2 Tyr-105 phosphorylation and decreased PKM2 activity. Moreover, PKM2 Tyr-105 phosphorylation is regulated nutritionally, decreasing in adipose tissue depots after high-fat feeding. Further, decreased PKM2 Tyr-105 phosphorylation correlates with the development of glucose intolerance and insulin resistance in rodents, non-human primates, and humans. Together, these findings identify PKM2 as a novel substrate of PTP1B and provide new insights into the regulation of adipose PKM2 activity. PMID:23640882

Abscisic acid affects transcription of chloroplast genes via protein phosphatase 2C-dependent activation of nuclear genes: repression by guanosine-3'-5'-bisdiphosphate and activation by sigma factor 5.

PubMed

Yamburenko, Maria V; Zubo, Yan O; Börner, Thomas

2015-06-01

Abscisic acid (ABA) represses the transcriptional activity of chloroplast genes (determined by run-on assays), with the exception of psbD and a few other genes in wild-type Arabidopsis seedlings and mature rosette leaves. Abscisic acid does not influence chloroplast transcription in the mutant lines abi1-1 and abi2-1 with constitutive protein phosphatase 2C (PP2C) activity, suggesting that ABA affects chloroplast gene activity by binding to the pyrabactin resistance (PYR)/PYR1-like or regulatory component of ABA receptor protein family (PYR/PYL/RCAR) and signaling via PP2Cs and sucrose non-fermenting protein-related kinases 2 (SnRK2s). Further we show by quantitative PCR that ABA enhances the transcript levels of RSH2, RSH3, PTF1 and SIG5. RelA/SpoT homolog 2 (RSH2) and RSH3 are known to synthesize guanosine-3'-5'-bisdiphosphate (ppGpp), an inhibitor of the plastid-gene-encoded chloroplast RNA polymerase. We propose, therefore, that ABA leads to an inhibition of chloroplast gene expression via stimulation of ppGpp synthesis. On the other hand, sigma factor 5 (SIG5) and plastid transcription factor 1 (PTF1) are known to be necessary for the transcription of psbD from a specific light- and stress-induced promoter (the blue light responsive promoter, BLRP). We demonstrate that ABA activates the psbD gene by stimulation of transcription initiation at BLRP. Taken together, our data suggest that ABA affects the transcription of chloroplast genes by a PP2C-dependent activation of nuclear genes encoding proteins involved in chloroplast transcription. © 2015 The Authors The Plant Journal © 2015 John Wiley & Sons Ltd.

Protein tyrosine phosphatase 1B as a target for the treatment of impaired glucose tolerance and type II diabetes.

PubMed

Liu, Gang; Trevillyan, James M

2002-11-01

Protein tyrosine phosphatase 1B (PTP1B) is a negative regulator of the insulin signal transduction cascade, initiated when insulin binds to the insulin receptor. PTP1B-deficient mice are more sensitive to insulin, and have improved glycemic control and resistance to diet-induced obesity than wild-type control mice. Diabetic mice treated with PTP1B antisense oligonucleotides intraperitoneally have lower PTP1B protein levels in liver and fat, reduced plasma insulin, blood glucose and hemoglobin A1c (HbA1c) levels. These studies validate PTP1B as a promising drug discovery target for the treatment of insulin resistance, diabetes and obesity. Herein we review the recent advances in the structure-based design of potent and selective small molecule inhibitors of PTP1B, and discuss th e challenge of developing compounds with improved cell permeability and bioavailability.

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.

REDUCED DORMANCY5 Encodes a Protein Phosphatase 2C That Is Required for Seed Dormancy in Arabidopsis[C][W][OPEN

PubMed Central

Xiang, Yong; Nakabayashi, Kazumi; Ding, Jia; He, Fei; Bentsink, Leónie; Soppe, Wim J.J.

2014-01-01

Seed dormancy determines germination timing and contributes to crop production and the adaptation of natural populations to their environment. Our knowledge about its regulation is limited. In a mutagenesis screen of a highly dormant Arabidopsis thaliana line, the reduced dormancy5 (rdo5) mutant was isolated based on its strongly reduced seed dormancy. Cloning of RDO5 showed that it encodes a PP2C phosphatase. Several PP2C phosphatases belonging to clade A are involved in abscisic acid signaling and control seed dormancy. However, RDO5 does not cluster with clade A phosphatases, and abscisic acid levels and sensitivity are unaltered in the rdo5 mutant. RDO5 transcript could only be detected in seeds and was most abundant in dry seeds. RDO5 was found in cells throughout the embryo and is located in the nucleus. A transcriptome analysis revealed that several genes belonging to the conserved PUF family of RNA binding proteins, in particular Arabidopsis PUMILIO9 (APUM9) and APUM11, showed strongly enhanced transcript levels in rdo5 during seed imbibition. Further transgenic analyses indicated that APUM9 reduces seed dormancy. Interestingly, reduction of APUM transcripts by RNA interference complemented the reduced dormancy phenotype of rdo5, indicating that RDO5 functions by suppressing APUM transcript levels. PMID:25415980

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

Antisense protein tyrosine phosphatase 1B reverses activation of p38 mitogen-activated protein kinase in liver of ob/ob mice.

PubMed

Gum, Rebecca J; Gaede, Lori L; Heindel, Matthew A; Waring, Jeffrey F; Trevillyan, James M; Zinker, Bradley A; Stark, Margery E; Wilcox, Denise; Jirousek, Michael R; Rondinone, Cristina M; Ulrich, Roger G

2003-06-01

Phosphorylation of stress-activated kinase p38, a MAPK family member, was increased in liver of ob/ob diabetic mice relative to lean littermates. Treatment of ob/ob mice with protein tyrosine phosphatase 1B (PTP1B) antisense oligonucleotides (ASO) reduced phosphorylation of p38 in liver-to below lean littermate levels-and normalized plasma glucose while reducing plasma insulin. Phosphorylation of ERK, but not JNK, was also decreased in ASO-treated mice. PTP1B ASO decreased TNFalpha protein levels and phosphorylation of the transcription factor cAMP response element binding protein (CREB) in liver, both of which can occur through decreased phosphorylation of p38 and both of which have been implicated in insulin resistance or hyperglycemia. Decreased p38 phosphorylation was not directly due to decreased phosphorylation of the kinases that normally phosphorylate p38-MKK3 and MKK6. Additionally, p38 phosphorylation was not enhanced in liver upon insulin stimulation of ASO-treated ob/ob mice (despite increased activation of other signaling molecules) corroborating that p38 is not directly affected via the insulin receptor. Instead, decreased phosphorylation of p38 may be due to increased expression of MAPK phosphatases, particularly the p38/ERK phosphatase PAC1 (phosphatase of activated cells). This study demonstrates that reduction of PTP1B protein using ASO reduces activation of p38 and its substrates TNFalpha and CREB in liver of diabetic mice, which correlates with decreased hyperglycemia and hyperinsulinemia.

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

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.

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 ...

Effects of 1.8 GHz radiofrequency field on microstructure and bone metabolism of femur in mice.

PubMed

Guo, Ling; Zhang, Jun-Ping; Zhang, Ke-Ying; Wang, Huan-Bo; Wang, Huan; An, Guang-Zhou; Zhou, Yan; Meng, Guo-Lin; Ding, Gui-Rong

2018-04-30

To investigate the effects of 1.8 GHz radiofrequency (RF) field on bone microstructure and metabolism of femur in mice, C57BL/6 mice (male, age 4 weeks) were whole-body exposed or sham exposed to 1.8 GHz RF field. Specific absorption rates of whole body and bone were approximately 2.70 and 1.14 W/kg (6 h/day for 28 days). After exposure, microstructure and morphology of femur were observed by microcomputed tomography (micro-CT), Hematoxylin and Eosin (HE) and Masson staining. Subsequently, bone parameters were calculated directly from the reconstructed images, including structure model index, bone mineral density, trabecular bone volume/total volume, connectivity density, trabecular number, trabecular thickness, and trabecular separation. Biomarkers that reflect bone metabolism, such as serum total alkaline phosphatase (ALP), bone-specific alkaline phosphatase (BALP), and tartrate-resistant acid phosphatase 5b (TRACP-5b), were determined by biochemical assay methods. Micro-CT and histology results showed that there was no significant change in bone microstructure and the above parameters in RF group, compared with sham group. The activity of serum ALP and BALP increased 29.47% and 16.82%, respectively, in RF group, compared with sham group (P < 0.05). In addition, there were no significant differences in the activity of serum TRACP-5b between RF group and sham group. In brief, under present experimental conditions, we did not find support for an effect of 1.8 GHz RF field on bone microstructure; however, it might promote metabolic function of osteoblasts in mice. Bioelectromagnetics. © 2018 Wiley Periodicals, Inc. © 2018 Wiley Periodicals, Inc.

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

Using mass spectrometry to study the photo-affinity labeling of protein tyrosine phosphatase 1B

NASA Astrophysics Data System (ADS)

Leriche, Tammy; Skorey, Kathryn; Roy, Patrick; McKay, Dan; Bateman, Kevin P.

2004-11-01

Protein tyrosine phosphatase 1B (PTP1B) is a potential target for the treatment of Type II diabetes and several companies are developing small molecule inhibitors of this enzyme. Part of the characterization of these compounds as PTP1B inhibitors is the understanding of how they bind in the enzyme active site. The use of photo-activated inhibitors that target the active site can provide such insight. This paper describes the characterization of a photoprobe directed at the active site of PTP1B. Mass spectrometry revealed the specific binding of the probe to the intact protein. Digestion of the labeled protein followed by LC-MS and LC-MS/MS was used to show that the photoprobe binds to a specific active site amino acid. This was confirmed by comparison with the X-ray structure of PTP1B with a PTP1B inhibitor. The probe labels a conserved acidic residue (Asp) that is required for catalytic activity. This photoprobe may prove to be a useful tool for the development of a PTP1B inhibitor or for the study of PTPs in general.

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

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.

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.

Aesculin modulates bone metabolism by suppressing receptor activator of NF-κB ligand (RANKL)-induced osteoclastogenesis and transduction signals.

PubMed

Zhao, Xiao-Li; Chen, Lin-Feng; Wang, Zhen

2017-06-17

Aesculin (AES), a coumarin compound derived from Aesculus hippocasanum L, is reported to exert protective role against inflammatory diseases, gastric disease and cancer. However, direct effect of AES in bone metabolism is deficient. In this study, we examined the effects of AES on osteoclast (OC) differentiation in receptor activator of NF-κB ligand (RANKL)-induced RAW264.7 cells. AES inhibits the OC differentiation in both dose- and time-dependent manner within non-toxic concentrations, as analyzed by Tartrate Resistant Acid Phosphatase (TRAP) staining. The actin ring formation manifesting OC function is also decreased by AES. Moreover, expressions of osteoclastogenesis related genes Trap, Atp6v0d2, Cathepsin K and Mmp-9 are decreased upon AES treatment. Mechanistically, AES attenuates the activation of MAPKs and NF-κB activity upon RANKL induction, thus leading to the reduction of Nfatc1 mRNA expression. Moreover, AES inhibits Rank expression, and RANK overexpression markedly decreases AES's effect on OC differentiation and NF-κB activity. Consistently, AES protects against bone mass loss in the ovariectomized and dexamethasone treated rat osteoporosis model. Taken together, our data demonstrate that AES can modulate bone metabolism by suppressing osteoclastogenesis and related transduction signals. AES therefore could be a promising agent for the treatment of osteoporosis. Copyright © 2017 Elsevier Inc. All rights reserved.

Superoxide anion radicals induce IGF-1 resistance through concomitant activation of PTP1B and PTEN

PubMed Central

Singh, Karmveer; Maity, Pallab; Krug, Linda; Meyer, Patrick; Treiber, Nicolai; Lucas, Tanja; Basu, Abhijit; Kochanek, Stefan; Wlaschek, Meinhard; Geiger, Hartmut; Scharffetter-Kochanek, Karin

2015-01-01

The evolutionarily conserved IGF-1 signalling pathway is associated with longevity, metabolism, tissue homeostasis, and cancer progression. Its regulation relies on the delicate balance between activating kinases and suppressing phosphatases and is still not very well understood. We report here that IGF-1 signalling in vitro and in a murine ageing model in vivo is suppressed in response to accumulation of superoxide anions () in mitochondria, either by chemical inhibition of complex I or by genetic silencing of -dismutating mitochondrial Sod2. The -dependent suppression of IGF-1 signalling resulted in decreased proliferation of murine dermal fibroblasts, affected translation initiation factors and suppressed the expression of α1(I), α1(III), and α2(I) collagen, the hallmarks of skin ageing. Enhanced led to activation of the phosphatases PTP1B and PTEN, which via dephosphorylation of the IGF-1 receptor and phosphatidylinositol 3,4,5-triphosphate dampened IGF-1 signalling. Genetic and pharmacologic inhibition of PTP1B and PTEN abrogated -induced IGF-1 resistance and rescued the ageing skin phenotype. We thus identify previously unreported signature events with , PTP1B, and PTEN as promising targets for drug development to prevent IGF-1 resistance-related pathologies. PMID:25520316

A Loss-of-Function Screen for Phosphatases that Regulate Neurite Outgrowth Identifies PTPN12 as a Negative Regulator of TrkB Tyrosine Phosphorylation

PubMed Central

Ambjørn, Malene; Dubreuil, Véronique; Miozzo, Federico; Nigon, Fabienne; Møller, Bente; Issazadeh-Navikas, Shohreh; Berg, Jacob; Lees, Michael; Sap, Jan

2013-01-01

Alterations in function of the neurotrophin BDNF are associated with neurodegeneration, cognitive decline, and psychiatric disorders. BDNF promotes axonal outgrowth and branching, regulates dendritic tree morphology and is important for axonal regeneration after injury, responses that largely result from activation of its tyrosine kinase receptor TrkB. Although intracellular neurotrophin (NT) signaling presumably reflects the combined action of kinases and phosphatases, little is known about the contributions of the latter to TrkB regulation. The issue is complicated by the fact that phosphatases belong to multiple independently evolved families, which are rarely studied together. We undertook a loss-of-function RNA-interference-based screen of virtually all known (254) human phosphatases to understand their function in BDNF/TrkB-mediated neurite outgrowth in differentiated SH-SY5Y cells. This approach identified phosphatases from diverse families, which either positively or negatively modulate BDNF-TrkB-mediated neurite outgrowth, and most of which have little or no previously established function related to NT signaling. “Classical” protein tyrosine phosphatases (PTPs) accounted for 13% of the candidate regulatory phosphatases. The top classical PTP identified as a negative regulator of BDNF-TrkB-mediated neurite outgrowth was PTPN12 (also called PTP-PEST). Validation and follow-up studies showed that endogenous PTPN12 antagonizes tyrosine phosphorylation of TrkB itself, and the downstream activation of ERK1/2. We also found PTPN12 to negatively regulate phosphorylation of p130cas and FAK, proteins with previously described functions related to cell motility and growth cone behavior. Our data provide the first comprehensive survey of phosphatase function in NT signaling and neurite outgrowth. They reveal the complexity of phosphatase control, with several evolutionarily unrelated phosphatase families cooperating to affect this biological response, and hence

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.

Host Plant Effects on Alkaline Phosphatase Activity in the Whiteflies, Bemisia tabaci Biotype B and Trialeurodes vaporariorum

PubMed Central

Yan, Ying; Peng, Lu; Liu, Wan-Xue; Wan, Fang-Hao; Harris, Marvin K.

2011-01-01

Bemisia tabaci (Gennadius) B-biotype and Trialeurodes vaporariorum (Westwood) (Hemiptera: Aleyrodidae) often coexist on greenhouse-grown vegetable crops in northern China. The recent spread of B. tabaci B-biotype has largely replaced T. vaporariorum, and B-biotype now overlaps with T. vaporariorum where common hosts occur in most invaded areas. The impact of the B-biotype on the agro eco system appears to be widespread, and involves the ability to compete with and perhaps replace other phytophages like T. vaporariorum. An emerging hypothesis is that the B-biotype is physiologically superior due at least in part to an improved ability to metabolically utilize the alkaline phosphatase pathway. To test this hypothesis, alkaline phosphatase activity was studied in the B-biotype and T. vaporariorum after feeding on a number of different hosts for a range of durations, with and without host switching. Alkaline phosphatase activity in T. vaporariorum was 1.45 to 2.53-fold higher than that of the B-biotype when fed on tomato for 4 and 24 h, or switched from tomato to cotton and cabbage for the same durations. However, alkaline phosphatase activity in the B-biotype was 1.40 to 3.35-fold higher than that of T. vaporariorum when the host switching time was ∼72 and ∼120 h on the same plant. Both short-term (4 h) and long-term (72 h) switching of plant hosts can significantly affect the alkaline phosphatase activity in the two species. After ∼120 h, feeding on tomato and cotton alkaline phosphatase activity in the B-biotype was significantly higher than that of T. vaporariorum. It was shown that alkaline phosphatase aids the species feeding on different plant species, and that the B-biotype is physiologically superior to T. vaporariorum in utilizing the enzyme compared to T. vaporariorum over longer periods of feeding. PMID:21521136

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.

Therapeutic effects of systemic vitamin k2 and vitamin d3 on gingival inflammation and alveolar bone in rats with experimentally induced periodontitis.

PubMed

Aral, Kübra; Alkan, Banu Arzu; Saraymen, Recep; Yay, Arzu; Şen, Ahmet; Önder, Gözde Özge

2015-05-01

The synergistic effects of vitamin D3 and vitamin K2 on bone loss prevention have been reported. This study evaluates the effects of vitamin D3 and vitamin K2 supplementation in conjunction with conventional periodontal therapy (scaling and root planing [SRP]) on gingival interleukin (IL)-1β and IL-10, serum bone alkaline phosphatase (B-ALP) and tartrate-resistant acid phosphatase 5b (TRAP-5b), and calcium and alveolar bone levels in rats with experimentally induced periodontitis. Seventy-two rats were divided into the following groups: 1) healthy; 2) periodontitis; 3) SRP; 4) SRP + vitamin D3; 5) SRP + vitamin K2; and 6) SRP + vitamins K2 and D3. Periodontitis was induced by ligature placement for 7 days, and vitamin K2 (30 mg/kg) and/or vitamin D3 (2 μg/kg) were administered for 10 days in the SRP + vitamin D3, SRP + vitamin K2, and SRP + vitamins K2 and D3 groups by oral gavage. On day 18, the animals were sacrificed, serum B-ALP, TRAP-5b, and calcium levels were measured, gingiva specimens were extracted for IL-1β and IL-10 analysis, and distances between the cemento-enamel junction and alveolar bone crest were evaluated. Alveolar bone levels in the periodontitis group were significantly greater than those in the other five groups. No significant differences were found in gingival IL-1β and IL-10, serum B-ALP and TRAP-5b, and calcium and alveolar bone levels between the groups receiving SRP and vitamins and the group receiving SRP alone. Within the limitations of this study, vitamin D3 and K2 alone or in combination did not affect gingival IL-1β and IL-10, serum B-ALP and TRAP-5b levels, or alveolar bone compared with conventional periodontal therapy alone.

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.

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

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.

Synthesis of some novel orcinol based coumarin triazole hybrids with capabilities to inhibit RANKL-induced osteoclastogenesis through NF-κB signaling pathway.

PubMed

Rama Krishna, Boddu; Thummuri, Dinesh; Naidu, V G M; Ramakrishna, Sistla; Venkata Mallavadhani, Uppuluri

2018-08-01

A total of twenty-two novel coumarin triazole hybrids (4a-4k and 6a-6k) were synthesized from orcinol in good to excellent yields of 70-94%. The structures of all the synthesized compounds were elucidated by spectroscopic techniques such as 1 H NMR, 13 C NMR, and HRMS. The anti-inflammatory potential of synthesized compounds was investigated against the proinflammatory cytokine, TNF-α on U937 cell line and compounds 4d, 4j, and 6j were found to exhibit promising anti-inflammatory activity. These three compounds were further screened against TNF-α on LPS-stimulated RAW 264.7 cells, which confirm their anti-inflammatory potential. Furthermore, the above said active compounds were tested for their inhibitory effect on RANKL-induced osteoclastogenesis in RAW 264.7 cells by using tartrate resistant acid phosphatase (TRAP) staining assay at 10 µM. Molecular mechanism studies demonstrated that compound 4d exhibited dose dependent inhibition of RANKL-induced osteoclastogenesis by suppression of the NF-kB pathway. Thus, compound 4d is a promising candidate for further optimization to develop as a potent anti-osteoporotic agent. Copyright © 2018 Elsevier Inc. All rights reserved.

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.

Loss of ephrinB1 in osteogenic progenitor cells impedes endochondral ossification and compromises bone strength integrity during skeletal development.

PubMed

Nguyen, Thao M; Arthur, Agnieszka; Paton, Sharon; Hemming, Sarah; Panagopoulos, Romana; Codrington, John; Walkley, Carl R; Zannettino, Andrew C W; Gronthos, Stan

2016-12-01

The EphB receptor tyrosine kinase family and their ephrinB ligands have been implicated as mediators of skeletal development and bone homeostasis in humans, where mutations in ephrinB1 contribute to frontonasal dysplasia and coronal craniosynostosis. In mouse models, ephrinB1 has been shown to be a critical factor mediating osteoblast function. The present study examined the functional importance of ephrinB1 during endochondral ossification using the Cre recombination system with targeted deletion of ephrinB1 (EfnB1 fl/fl ) in osteogenic progenitor cells, under the control of the osterix (Osx:Cre) promoter. The Osx:EfnB1 -/- mice displayed aberrant bone growth during embryonic and postnatal skeletal development up to 4weeks of age, when compared to the Osx:Cre controls. Furthermore, compared to the Osx:Cre control mice, the Osx:EfnB1 -/- mice exhibited significantly weaker and less rigid bones, with a reduction in trabecular/ cortical bone formation, reduced trabecular architecture and a reduction in the size of the growth plates at the distal end of the femora from newborn through to 4weeks of age. The aberrant bone formation correlated with increased numbers of tartrate resistant acid phosphatase positive osteoclasts and decreased numbers of bone lining osteoblasts in 4week old Osx:EfnB1 -/- mice, compared to Osx:Cre control mice. Taken together, these observations demonstrate the importance of ephrinB1 signalling between cells of the skeleton required for endochondral ossification. Copyright © 2016 Elsevier Inc. All rights reserved.

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.

Randomized, double-blind, placebo-controlled trial of the 5-HT1A receptor antagonist AZD7371 tartrate monohydrate (robalzotan tartrate monohydrate) in patients with irritable bowel syndrome.

PubMed

Drossman, Douglas A; Danilewitz, Mervyn; Naesdal, Jørgen; Hwang, Clara; Adler, John; Silberg, Debra G

2008-10-01

To investigate the efficacy and safety of the 5-hydroxytrypamine 1A (5-HT(1A)) receptor antagonist AZD7371 tartrate monohydrate (robalzotan tartrate monohydrate), termed AZD7371 here, in patients with irritable bowel syndrome (IBS). Patients meeting the Rome II criteria for IBS (N = 402) were randomized to treatment with AZD7371 20 mg or 5 mg or matching placebo tablets twice daily for 12 wk. The patients completed daily and weekly diary assessments, reporting abdominal discomfort or pain and description of bowel movements. They also completed validated symptom and quality-of-life questionnaires. Neither AZD7371 regimen was significantly more effective than placebo in providing adequate relief from IBS symptoms in at least 2 out of 4 wk per month over the 12 wk of treatment. There was also no significant difference between the treatment groups and placebo in the change in score in the validated symptom and quality-of-life questionnaires. Overall, 22.1% of patients experienced adverse events (AEs) attributed to the study medication: 44 of 133 (33.1%) in the 20 mg AZD7371 group, 27 of 131 (20.6%) in the 5 mg AZD7371 group, and 17 of 134 (12.7%) in the placebo group. Also, 31 of 57 (54%) of AEs leading to discontinuation were central nervous system-related. Hallucinations or hallucination-like AEs were reported by eight patients taking AZD7371, and by none of the patients in the placebo group. After these events led to discontinuation in six patients, the study was prematurely terminated. In view of the AE profile and lack of efficacy in IBS, the clinical development of AZD7371 has been stopped.

Markers of Bone Metabolism Are Affected by Renal Function and Growth Hormone Therapy in Children with Chronic Kidney Disease

PubMed Central

Doyon, Anke; Fischer, Dagmar-Christiane; Bayazit, Aysun Karabay; Canpolat, Nur; Duzova, Ali; Sözeri, Betül; Bacchetta, Justine; Balat, Ayse; Büscher, Anja; Candan, Cengiz; Cakar, Nilgun; Donmez, Osman; Dusek, Jiri; Heckel, Martina; Klaus, Günter; Mir, Sevgi; Özcelik, Gül; Sever, Lale; Shroff, Rukshana; Vidal, Enrico; Wühl, Elke; Gondan, Matthias; Melk, Anette; Querfeld, Uwe; Haffner, Dieter; Schaefer, Franz

2015-01-01

Objectives The extent and relevance of altered bone metabolism for statural growth in children with chronic kidney disease is controversial. We analyzed the impact of renal dysfunction and recombinant growth hormone therapy on a panel of serum markers of bone metabolism in a large pediatric chronic kidney disease cohort. Methods Bone alkaline phosphatase (BAP), tartrate-resistant acid phosphatase 5b (TRAP5b), sclerostin and C-terminal FGF-23 (cFGF23) normalized for age and sex were analyzed in 556 children aged 6–18 years with an estimated glomerular filtration rate (eGFR) of 10–60 ml/min/1.73m2. 41 children receiving recombinant growth hormone therapy were compared to an untreated matched control group. Results Standardized levels of BAP, TRAP5b and cFGF-23 were increased whereas sclerostin was reduced. BAP was correlated positively and cFGF-23 inversely with eGFR. Intact serum parathormone was an independent positive predictor of BAP and TRAP5b and negatively associated with sclerostin. BAP and TRAP5B were negatively affected by increased C-reactive protein levels. In children receiving recombinant growth hormone, BAP was higher and TRAP5b lower than in untreated controls. Sclerostin levels were in the normal range and higher than in untreated controls. Serum sclerostin and cFGF-23 independently predicted height standard deviation score, and BAP and TRAP5b the prospective change in height standard deviation score. Conclusion Markers of bone metabolism indicate a high-bone turnover state in children with chronic kidney disease. Growth hormone induces an osteoanabolic pattern and normalizes osteocyte activity. The osteocyte markers cFGF23 and sclerostin are associated with standardized height, and the markers of bone turnover predict height velocity. PMID:25659076

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.

Prevalence of NS5B resistance-associated variants in treatment-naïve Asian patients with chronic hepatitis C.

PubMed

Yang, Song; Xing, Huichun; Feng, Shenghu; Ju, Wei; Liu, Shunai; Wang, Xiaomei; Ou, Weini; Cheng, Jun; Pan, Calvin Q

2018-02-01

There is little information on the association between baseline non-structural protein (NS) 5b resistance-associated variants (RAVs) and treatment failure in hepatitis C patients. This study examined the frequencies of natural hepatitis C virus (HCV) NS5B resistance-associated variants (RAVs) in an Asian cohort. Samples from Asian HCV patients enrolled between October 2009 and September 2014 were analyzed for NS5B RAVs within the region from amino acid 230 to 371. Serum samples were tested by PCR genotyping, with sequence alignment performed using the neighbor-joining method. NS5B was detected by Sanger sequencing followed by Geno2pheno analysis. NS5B RAVs were detected in 80.52% (1199/1489) of patients; 68.4% (1019/1489) and 79.7% (1186/1489) were associated with resistance to sofosbuvir (SOF) and dasabuvir (DSV), respectively. These RAVs were present in 95% (1004/1058) of genotype 1b patients. When genotypes 1b and 2a were compared, SOF-associated RAVs were detected at a higher frequency in genotype 1b (94.8% [1004/1058] vs. 2.9% [9/309]; χ 2 = 1054.433, P < 0.001), C316H/N was more common in genotype 1b (94.7% [1002/1058] vs. 0% [0/309]; χ 2 = 1096.014, P < 0.001), M289F/L/I/W/V had a higher frequency in genotype 2a (0.7% [7/309] vs. 2.3% [7/1058]; χ 2 = 4.589, P = 0.032), DSV-associated RAVs were most often found in genotype 1b (95.0% [1005/1058] vs. 40.1% 124/309]; χ 2 = 500.577, P < 0.001), and frequency of C316Y/H/N/W was higher in genotype 1b (94.7% [1002/1058] vs. 0% [0/309]; χ 2 = 1096.014, P < 0.001). In conclusion, baseline SOF and DSV RAVs are common in Asian HCV patients and predominantly occur in genotype 1b.

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.

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

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...

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

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.

Analysis of in vitro interactions of protein tyrosine phosphatase 1B with insulin receptors.

PubMed

Wang, X Y; Bergdahl, K; Heijbel, A; Liljebris, C; Bleasdale, J E

2001-02-28

One strategy to treat the insulin resistance that is central to type II diabetes mellitus may be to maintain insulin receptors (IR) in the active (tyrosine phosphorylated) form. Because protein tyrosine phosphatase 1B (PTP1B) binds and subsequently dephosphorylates IR, inhibitors of PTP1B-IR binding are potential insulin 'sensitizers.' A Scintillation Proximity Assay (SPA) was developed to characterize and quantitate PTP1B-IR binding. Human IR were solubilized and captured on wheat germ agglutinin (WGA)-coated SPA beads. Subsequent binding of human, catalytically inactive [35S] PTP1B Cys(215)/Ser (PTP1B(C215S)) to the lectin-anchored IR results in scintillation from the SPA beads that can be quantitated. Binding of PTP1B to IR was pH- and divalent cation-sensitive. Ca(2+) and Mn(2+), but not Mg(2+), dramatically attenuated the loss of PTP1B-IR binding observed when pH was raised from 6.2 to 7.8. PTP1B binding to IR from insulin-stimulated cells was much greater than to IR from unstimulated cells and was inhibited by either an antiphosphotyrosine antibody or treatment of IR with alkaline phosphatase, suggesting that tyrosine phosphorylation of IR is required for PTP1B binding. Phosphopeptides modeled after various IR phosphotyrosine domains each only partially inhibited PTP1B-IR binding, indicating that multiple domains of IR are likely involved in binding PTP1B. However, competitive displacement of [35S]PTP1B(C215S) by PTP1B(C215S) fitted best to a single binding site with a K(d) in the range 100-1000 nM, depending upon pH and divalent cations. PNU-200898, a potent and selective inhibitor of PTP1B whose orientation in the active site of PTP1B has been solved, competitively inhibited catalysis and PTP1B-IR binding with equal potency. The results of this novel assay for PTP1B-IR binding suggest that PTP1B binds preferentially to tyrosine phosphorylated IR through its active site and that binding may be susceptible to therapeutic disruption by small molecules.

Protection against gamma-radiation injury by protein tyrosine phosphatase 1B.

PubMed

Mojena, Marina; Pimentel-Santillana, María; Povo-Retana, Adrián; Fernández-García, Victoria; González-Ramos, Silvia; Rada, Patricia; Tejedor, Alberto; Rico, Daniel; Martín-Sanz, Paloma; Valverde, Angela M; Boscá, Lisardo

2018-07-01

Protein tyrosine phosphatase 1B (PTP1B) is widely expressed in mammalian tissues, in particular in immune cells, and plays a pleiotropic role in dephosphorylating many substrates. Moreover, PTP1B expression is enhanced in response to pro-inflammatory stimuli and to different cell stressors. Taking advantage of the use of mice deficient in PTP1B we have investigated the effect of γ-radiation in these animals and found enhanced lethality and decreased respiratory exchange ratio vs. the corresponding wild type animals. Using bone-marrow derived macrophages and mouse embryonic fibroblasts (MEFs) from wild-type and PTP1B-deficient mice, we observed a differential response to various cell stressors. PTP1B-deficient macrophages exhibited an enhanced response to γ-radiation, UV-light, LPS and S-nitroso-glutathione. Macrophages exposed to γ-radiation show DNA damage and fragmentation, increased ROS production, a lack in GSH elevation and enhanced acidic β-galactosidase activity. Interestingly, these differences were not observed in MEFs. Differential gene expression analysis of WT and KO macrophages revealed that the main pathways affected after irradiation were an up-regulation of protein secretion, TGF-β signaling and angiogenesis among other, and downregulation of Myc targets and Hedgehog signaling. These results demonstrate a key role for PTP1B in the protection against the cytotoxicity of irradiation in intact animal and in macrophages, which might be therapeutically relevant. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

Properties of MgB 2 superconductor chemically treated by acetic acid

NASA Astrophysics Data System (ADS)

Hušeková, K.; Hušek, I.; Kováč, P.; Kulich, M.; Dobročka, E.; Štrbík, V.

2010-03-01

Commercial Alfa Aesar MgB 2 powder was chemically treated by acetic acid with the aim of MgO removing. Single-core MgB 2/Fe ex situ wires have been made by powder-in-tube (PIT) process using the powders treated with different acid concentration. All samples were annealed in argon at 950 °C/0.5 h. Differences in transition temperatures and critical currents of acetic acid treated MgB 2 are related to the normal state resistivity, effective carbon substitution from the organic solvent and the active area fraction (grain-connectivity).

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 ...

FAP-1-mediated activation of NF-kappaB induces resistance of head and neck cancer to Fas-induced apoptosis.

PubMed

Wieckowski, Eva; Atarashi, Yoshinari; Stanson, Joanna; Sato, Taka-Aki; Whiteside, Theresa L

2007-01-01

Molecular mechanisms responsible for tumor resistance to apoptosis often involve the Fas/FasL pathway. While squamous cell carcinomas of the head and neck (SCCHN) express both Fas and FasL, their resistance to self-induced apoptosis or apoptosis mediated by Fas agonistic antibody (CH-11Ab) was independent of the level of Fas surface expression or the presence of soluble Fas in supernatants of primary or metastatic SCCHN cell lines. By in vitro immunoselection, using PCI-15A cell line treated with successive cycles of CH-11 Ab, Fas-resistant sublines with the parental genotype were selected. Such sublines failed to cleave caspase-8 upon Fas engagement and were resistant to CH-11 Ab, although they remained sensitive to VP-16 or staurosporin. In the presence of cycloheximide, the selected SCCHN sublines become susceptible to CH-11 Ab, and showed cleavage of caspase-8, suggesting that apoptosis resistance was mediated by an inhibitory protein(s) acting upstream of caspase-8. Overexpression of Fas-associated phosphatase 1 (FAP-1), but not cellular FLICE-inhibitory protein (cFLIP) in SCCHN sublines was documented by Western blots and RT-PCR analyses. The FAP-1+ selected sublines also downregulated cell surface Fas. A high phosphorylation level of IkappaB kappa, NFkappaB activation and upregulation of Bcl-2 expression were observed in the FAP-1+ sublines. Treatment with the phosphatase inhibitor, orthovanadate, or silencing of FAP-1 with siRNA abolished their resistance to apoptosis, suggesting that FAP-1 phosphatase activity could be responsible for NF-kappaB activation and resistance of SCCHN cells to Fas-mediated apoptosis. 2006 Wiley-Liss, Inc.

MiR-20a-5p promotes radio-resistance by targeting Rab27B in nasopharyngeal cancer cells.

PubMed

Huang, Dabing; Bian, Geng; Pan, Yueyin; Han, Xinghua; Sun, Yubei; Wang, Yong; Shen, Guodong; Cheng, Min; Fang, Xiang; Hu, Shilian

2017-01-01

MicroRNAs (miRNAs) was reported to be involved in cancer radio-resistance, which remains a major obstacle for effective cancer therapy. The differently expressed miRNAs were detected by RNA-seq experiment in nasopharyngeal cancer (NPC) cells. MiR-20a-5p was selected as our target, which was subject to finding its target gene Rab27B via bioinformatics analysis. The qRT-PCR, western blot and the luciferase reporter assays were performed to confirm Rab27B as the target of miR-20a-5p. In addition, the roles of miR-20a-5p in NPC radio-resistance were detected by transfection of either miR-20a-5p-mimic or miR-20a-5p-antagomiR. The involvement of Rab27B with NPC radio-resistance was also detected by the experiments with siRNA-mediated repression of Rab27B or over-expression of GFP-Rab27B. Wound healing and invasion assays were performed to detect the roles of both miR-20a-5p and Rab27B. MiR-20a-5p promotes NPC radio-resistance. We identified that its target gene Rab27B negatively correlates with miR-20a-5p-mediated NPC radio-resistance by systematic studies of a radio-sensitive (CNE-2) and resistant (CNE-1) NPC cell lines. Repression of Rab27B by siRNA suppresses cell apoptosis and passivates CNE-2 cells, whereas over-expression of Rab27B triggered cell apoptosis and sensitizes CNE-1 cells. MiR-20a-5p and its target gene Rab27B might be involved in the NPC radio-resistance. Thus the key players and regulators involved in this pathway might be the potential targets for developing effective therapeutic strategies against NPC.

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

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.

Production of Siderophores Increases Resistance to Fusaric Acid in Pseudomonas protegens Pf-5

PubMed Central

Ruiz, Jimena A.; Bernar, Evangelina M.; Jung, Kirsten

2015-01-01

Fusaric acid is produced by pathogenic fungi of the genus Fusarium, and is toxic to plants and rhizobacteria. Many fluorescent pseudomonads can prevent wilt diseases caused by these fungi. This study was undertaken to evaluate the effect of fusaric acid on P. protegens Pf-5 and elucidate the mechanisms that enable the bacterium to survive in the presence of the mycotoxin. The results confirm that fusaric acid negatively affects growth and motility of P. protegens. Moreover, a notable increase in secretion of the siderophore pyoverdine was observed when P. protegens was grown in the presence of fusaric acid. Concomitantly, levels of enzymes involved in the biosynthesis of pyoverdine and enantio-pyochelin, the second siderophore encoded by P. protegens, increased markedly. Moreover, while similar levels of resistance to fusaric acid were observed for P. protegens mutants unable to synthesize either pyoverdine or enanto-pyochelin and the wild type strain, a double mutant unable to synthesize both kinds of siderophores showed a dramatically reduced resistance to this compound. This reduced resistance was not observed when this mutant was grown under conditions of iron excess. Spectrophotometric titrations revealed that fusaric acid binds not only Fe2+ and Fe3+, but also Zn2+, Mn2+ and Cu2+, with high affinity. Our results demonstrate that iron sequestration accounts at least in part for the deleterious effect of the mycotoxin on P. protegens. PMID:25569682

Reduced expression of CD45 Protein-Tyrosine Phosphatase Pr

DTIC Science & Technology

2009-05-08

H S /D T R A on A ugust 19, 2009 w w w .jbc.org D ow nloaded from PTP1B , CD45, TCPTP, LMPTP-A, LMPTP-B, MEG1, MEG2, HePTP, PTP), three belong to...the dual specificity phosphatase VHR or the protein-tyrosine phosphatase PTP1B . Given these FIGURE 5. Mice expressing intermediate CD45 levels survive

Lipoic Acid Decreases the Viability of Breast Cancer Cells and Activity of PTP1B and SHP2.

PubMed

Kuban-Jankowska, Alicja; Gorska-Ponikowska, Magdalena; Wozniak, Michal

2017-06-01

Protein tyrosine phosphatases PTP1B and SHP2 are potential targets for anticancer therapy, because of the essential role they play in the development of tumors. PTP1B and SHP2 are overexpressed in breast cancer cells, thus inhibition of their activity can be potentially effective in breast cancer therapy. Lipoic acid has been previously reported to inhibit the proliferation of colon, breast and thyroid cancer cells. We investigated the effect of alpha-lipoic acid (ALA) and its reduced form of dihydrolipoic acid (DHLA) on the viability of MCF-7 cancer cells and on the enzymatic activity of PTP1B and SHP2 phosphatases. ALA and DHLA decrease the activity of PTP1B and SHP2, and have inhibitory effects on the viability and proliferation of breast cancer cells. ALA and DHLA can be considered as potential agents for the adjunctive treatment of breast cancer. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

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

Saccharomyces cerevisiae sigma 1278b has novel genes of the N-acetyltransferase gene superfamily required for L-proline analogue resistance.

PubMed

Takagi, H; Shichiri, M; Takemura, M; Mohri, M; Nakamori, S

2000-08-01

We discovered on the chromosome of Saccharomyces cerevisiae Sigma 1278b novel genes involved in L-proline analogue L-azetidine-2-carboxylic acid resistance which are not present in the standard laboratory strains. The 5.4 kb-DNA fragment was cloned from the genomic library of the L-azetidine-2-carboxylic acid-resistant mutant derived from a cross between S. cerevisiae strains S288C and Sigma 1278b. The nucleotide sequence of a 4.5-kb segment exhibited no identity with the sequence in the genome project involving strain S288C. Deletion analysis indicated that one open reading frame encoding a predicted protein of 229 amino acids is indispensable for L-azetidine-2-carboxylic acid resistance. The protein sequence was found to be a member of the N-acetyltransferase superfamily. Genomic Southern analysis and gene disruption showed that two copies of the novel gene with one amino acid change at position 85 required for L-azetidine-2-carboxylic acid resistance were present on chromosomes X and XIV of Sigma 1278b background strains. When this novel MPR1 or MPR2 gene (sigma 1278b gene for L-proline analogue resistance) was introduced into the other S. cerevisiae strains, all of the recombinants were resistant to L-azetidine-2-carboxylic acid, indicating that both MPR1 and MPR2 are expressed and have a global function in S. cerevisiae.

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

Protein tyrosine phosphatase 1B (PTP1B) is dispensable for IgE-mediated cutaneous reaction in vivo.

PubMed

Yang, Ting; Xie, Zhongping; Li, Hua; Yue, Lei; Pang, Zheng; MacNeil, Adam J; Tremblay, Michel L; Tang, Jin-Tian; Lin, Tong-Jun

2016-01-01

Mast cells play a critical role in allergic reactions. The cross-linking of FcεRI-bound IgE with multivalent antigen initiates a cascade of signaling events leading to mast cell activation. It has been well-recognized that cross linking of FcεRI mediates tyrosine phosphorylation. However, the mechanism involved in tyrosine dephosphorylation in mast cells is less clear. Here we demonstrated that protein tyrosine phosphatase 1B (PTP1B)-deficient mast cells showed increased IgE-mediated phosphorylation of the signal transducer and activator of transcription 5 (STAT5) and enhanced production of CCL9 (MIP-1γ) and IL-6 in IgE-mediated mast cells activation in vitro. However, IgE-mediated calcium mobilization, β-hexaosaminidase release (degranulation), and phosphorylation of IκB and MAP kinases were not affected by PTP1B deficiency. Furthermore, PTP1B deficient mice showed normal IgE-dependent passive cutaneous anaphylaxis and late phase cutaneous reactions in vivo. Thus, PTP1B specifically regulates IgE-mediated STAT5 pathway, but is redundant in influencing mast cell function in vivo. Copyright © 2016 Elsevier Inc. All rights reserved.

[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.

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.

Sulfone-stabilized carbanions for the reversible covalent capture of a posttranslationally-generated cysteine oxoform found in protein tyrosine phosphatase 1B (PTP1B).

PubMed

Parsons, Zachary D; Ruddraraju, Kasi Viswanatharaju; Santo, Nicholas; Gates, Kent S

2016-06-15

Redox regulation of protein tyrosine phosphatase 1B (PTP1B) involves oxidative conversion of the active site cysteine thiolate into an electrophilic sulfenyl amide residue. Reduction of the sulfenyl amide by biological thiols regenerates the native cysteine residue. Here we explored fundamental chemical reactions that may enable covalent capture of the sulfenyl amide residue in oxidized PTP1B. Various sulfone-containing carbon acids were found to react readily with a model peptide sulfenyl amide via attack of the sulfonyl carbanion on the electrophilic sulfur center in the sulfenyl amide. Both the products and the rates of these reactions were characterized. The results suggest that capture of a peptide sulfenyl amide residue by sulfone-stabilized carbanions can slow, but not completely prevent, thiol-mediated generation of the corresponding cysteine-containing peptide. Sulfone-containing carbon acids may be useful components in the construction of agents that knock down PTP1B activity in cells via transient covalent capture of the sulfenyl amide oxoform generated during insulin signaling processes. Copyright © 2016 Elsevier Ltd. All rights reserved.

Structural Basis for Selective Inhibition of Mycobacterium tuberculosis Protein Tyrosine Phosphatase PtpB

PubMed Central

Grundner, Christoph; Perrin, Dominique; van Huijsduijnen, Rob Hooft; Swinnen, Dominique; Gonzalez, Jérome; Gee, Christine L.; Wells, Timothy N.; Alber, Tom

2007-01-01

Tyrosine kinases and phosphatases establish the crucial balance of tyrosine phosphorylation in cellular signaling, but creating specific inhibitors of protein Tyr phosphatases (PTPs) remains a challenge. Here we report the development of a potent, selective inhibitor of Mycobacterium tuberculosis PtpB, a bacterial PTP that is secreted into host cells where it disrupts unidentified signaling pathways. The inhibitor, (oxalylamino-methylene)-thiophene sulfonamide (OMTS), showed an IC50 of 440 +/− 50 nM and >60-fold specificity for PtpB over six human PTPs. The 2-Å resolution crystal structure of PtpB in complex with OMTS revealed a large rearrangement of the enzyme, with some residues shifting >27 Å relative to the PtpB:PO4 complex. Extensive contacts with the catalytic loop provide a potential basis for inhibitor selectivity. Two OMTS molecules bound adjacent to each other, raising the possibility of a second substrate phosphotyrosine binding site in PtpB. The PtpB:OMTS structure provides an unanticipated framework to guide inhibitor improvement. PMID:17437721

Biotechnological approach for systemic delivery of membrane Receptor Activator of NF-κB Ligand (RANKL) active domain into the circulation

PubMed Central

Cappariello, Alfredo; Paone, Riccardo; Maurizi, Antonio; Capulli, Mattia; Rucci, Nadia; Muraca, Maurizio; Teti, Anna

2015-01-01

Deficiency of Receptor Activator of NF-κB Ligand (RANKL) prevents osteoclast formation causing osteopetrosis. RANKL is a membrane-bound protein cleaved into active soluble (s)RANKL by metalloproteinase 14 (MMP14). We created a bio-device that harbors primary osteoblasts, cultured on 3D hydroxyapatite scaffolds carrying immobilized MMP14 catalytic domain. Scaffolds were sealed in diffusion chambers and implanted in RANKL-deficient mice. Mice received 1 or 2 diffusion chambers, once or twice and were sacrificed after 1 or 2 months from implants. A progressive increase of body weight was observed in the implanted groups. Histological sections of tibias of non-implanted mice were negative for the osteoclast marker Tartrate-Resistant Acid Phosphatase (TRAcP), consistent with the lack of osteoclasts. In contrast, tibias excised from implanted mice showed TRAcP-positive cells in the bone marrow and on the bone surface, these latter morphologically similar to mature osteoclasts. In mice implanted with 4 diffusion chambers total, we noted the highest number and size of TRAcP-positive cells, with quantifiable eroded bone surface and significant reduction of trabecular bone volume. These data demonstrate that our bio-device delivers effective sRANKL, inducing osteoclastogenesis in RANKL-deficient mice, supporting the feasibility of an innovative experimental strategy to treat systemic cytokine deficiencies. PMID:25678116

Structure and functions of the placenta in common minke (Balaenoptera acutorostrata), Bryde’s (B. brydei) and sei (B. borealis) whales

PubMed Central

KITAYAMA, Chiyo; SASAKI, Motoki; ISHIKAWA, Hajime; MOGOE, Toshihiro; OHSUMI, Seiji; FUKUI, Yutaka; BUDIPITOJO, Teguh; KONDOH, Daisuke; KITAMURA, Nobuo

2015-01-01

The structure and functions of placentas were examined in 3 species of rorqual whales, common minke (Balaenoptera acutorostrata), Bryde’s (B. brydei) and sei (B. borealis) whales, with the aim of confirming the structural characteristics of the chorion, including the presence of the areolar part, and clarifying steroidogenic activities and fetomaternal interactions in the placentas of these whales. Placentas were collected from the second phase of the Japanese Whale Research Program under Special Permit in the North Pacific (JARPN II). Histological and ultrastructural examinations revealed that these whale placentas were epitheliochorial placentas with the interdigitation of chorionic villi lined by monolayer uninucleate cells (trophoblast cells) and endometrial crypts as well as folded placentation by fold-like chorionic villi. Moreover, well-developed pouch-like areolae were observed in the placentas, and active absorption was suggested in the chorionic epithelial cells of the areolar part (areolar trophoblast cells). Berlin blue staining showed the presence of ferric ions (Fe3+) in the uterine glandular epithelial cells and within the stroma of chorionic villi in the areolar part. An immunohistochemical examination revealed tartrate-resistant acid phosphatase (TRAP; known as uteroferrin in uteri) in the cytoplasm of glandular cells and areolar trophoblast cells. This result suggested that, in cetaceans, uteroferrin is used to supply iron to the fetus. Furthermore, immunoreactivity for P450scc and P450arom was detected in trophoblast cells, but not in areolar trophoblast cells, suggesting that trophoblast cells synthesize estrogen in whale placentas. Therefore, we herein immunohistochemically revealed the localization of aromatase and uteroferrin in cetacean placentas during pregnancy for the first time. PMID:26096685

Protein phosphatase 5 promotes hepatocarcinogenesis through interaction with AMP-activated protein kinase.

PubMed

Chen, Yao-Li; Hung, Man-Hsin; Chu, Pei-Yi; Chao, Tzu-I; Tsai, Ming-Hsien; Chen, Li-Ju; Hsiao, Yung-Jen; Shih, Chih-Ting; Hsieh, Feng-Shu; Chen, Kuen-Feng

2017-08-15

The serine-threonine protein phosphatase family members are known as critical regulators of various cellular functions, such as survival and transformation. Growing evidence suggests that pharmacological manipulation of phosphatase activity exhibits therapeutic benefits. Ser/Thr protein phosphatase 5 (PP5) is known to participate in glucocorticoid receptor (GR) and stress-induced signaling cascades that regulate cell growth and apoptosis, and has been shown to be overexpressed in various human malignant diseases. However, the role of PP5 in hepatocellular carcinoma (HCC) and whether PP5 may be a viable therapeutic target for HCC treatment are unknown. Here, by analyzing HCC clinical samples obtained from 215 patients, we found that overexpression of PP5 is tumor specific and associated with worse clinical outcomes. We further characterized the oncogenic properties of PP5 in HCC cells. Importantly, both silencing of PP5 with lentiviral-mediated short hairpin RNA (shRNA) and chemical inhibition of PP5 phosphatase activity using the natural compound cantharidin/norcantharidin markedly suppressed the growth of HCC cells and tumors in vitro and in vivo. Moreover, we identified AMP-activated protein kinase (AMPK) as a novel downstream target of oncogenic PP5 and demonstrated that the antitumor mechanisms underlying PP5 inhibition involve activation of AMPK signaling. Overall, our results establish a pathological function of PP5 in hepatocarcinogenesis via affecting AMPK signaling and suggest that PP5 inhibition is an attractive therapeutic approach for HCC. Copyright © 2017 Elsevier Inc. All rights reserved.

Asperentin B, a New Inhibitor of the Protein Tyrosine Phosphatase 1B.

PubMed

Wiese, Jutta; Aldemir, Hülya; Schmaljohann, Rolf; Gulder, Tobias A M; Imhoff, Johannes F

2017-06-21

In the frame of studies on secondary metabolites produced by fungi from deep-sea environments we have investigated inhibitors of enzymes playing key roles in signaling cascades of biochemical pathways relevant for the treatment of diseases. Here we report on a new inhibitor of the human protein tyrosine phosphatase 1B (PTP1B), a target in the signaling pathway of insulin. A new asperentin analog is produced by an Aspergillus sydowii strain isolated from the sediment of the deep Mediterranean Sea. Asperentin B ( 1 ) contains an additional phenolic hydroxy function at C-6 and exhibits an IC 50 value against PTP1B of 2 μM in vitro, which is six times stronger than the positive control, suramin. Interestingly, asperentin ( 2 ) did not show any inhibition of this enzymatic activity. Asperentin B ( 1 ) is discussed as possible therapeutic agents for type 2 diabetes and sleeping sickness.

Asperentin B, a New Inhibitor of the Protein Tyrosine Phosphatase 1B

PubMed Central

Wiese, Jutta; Aldemir, Hülya; Schmaljohann, Rolf; Gulder, Tobias A. M.; Imhoff, Johannes F.

2017-01-01

In the frame of studies on secondary metabolites produced by fungi from deep-sea environments we have investigated inhibitors of enzymes playing key roles in signaling cascades of biochemical pathways relevant for the treatment of diseases. Here we report on a new inhibitor of the human protein tyrosine phosphatase 1B (PTP1B), a target in the signaling pathway of insulin. A new asperentin analog is produced by an Aspergillus sydowii strain isolated from the sediment of the deep Mediterranean Sea. Asperentin B (1) contains an additional phenolic hydroxy function at C-6 and exhibits an IC50 value against PTP1B of 2 μM in vitro, which is six times stronger than the positive control, suramin. Interestingly, asperentin (2) did not show any inhibition of this enzymatic activity. Asperentin B (1) is discussed as possible therapeutic agents for type 2 diabetes and sleeping sickness. PMID:28635658

High bone turnover elevates the risk of denosumab-induced hypocalcemia in women with postmenopausal osteoporosis

PubMed Central

Ishikawa, Koji; Nagai, Takashi; Sakamoto, Keizo; Ohara, Kenji; Eguro, Takeshi; Ito, Hiroshi; Toyoshima, Yoichi; Kokaze, Akatsuki; Toyone, Tomoaki; Inagaki, Katsunori

2016-01-01

Hypocalcemia is the most common major adverse event in patients with osteoporosis receiving the bone resorption inhibitor denosumab; however, limited information is available regarding risk factors of hypocalcemia. Therefore, this study aimed to identify the risk factors of hypocalcemia induced by denosumab treatment for osteoporosis. We retrospectively reviewed the records of patients who had received initial denosumab supplemented with activated vitamin D for osteoporosis. Serum levels of the following bone turnover markers (BTMs) were measured at baseline: bone-specific alkaline phosphatase (BAP), total N-terminal propeptide of type 1 procollagen (P1NP), tartrate-resistant acid phosphatase 5b (TRACP-5b), and urinary cross-linked N-telopeptide of type 1 collagen (NTX). Of the 85 denosumab-treated patients with osteoporosis studied, 22 (25.9%) developed hypocalcemia. Baseline serum total P1NP, TRACP-5b, and urinary NTX were significantly higher in patients with hypocalcemia than in those with normocalcemia following denosumab administration (all P<0.01). Multivariate logistic regression analysis revealed that patients with total P1NP >76.5 μg/L, TRACP-5b >474 mU/dL, or urinary NTX >49.5 nmol bone collagen equivalent/mmol creatinine had a higher risk of hypocalcemia (P<0.01). Our study suggests that denosumab may have a greater impact on serum calcium levels in patients with postmenopausal osteoporosis with higher baseline bone turnover than in patients with postmenopausal osteoporosis with normal baseline bone turnover, because maintenance of normal serum calcium in this subgroup is more dependent on bone resorption. Close monitoring of serum calcium levels is strongly recommended for denosumab-treated patients with high bone turnover, despite supplementation with activated vitamin D and oral calcium. PMID:27980413

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

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

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.

Caged xanthones displaying protein tyrosine phosphatase 1B (PTP1B) inhibition from Cratoxylum cochinchinense.

PubMed

Li, Zuo Peng; Lee, Hyeong-Hwan; Uddin, Zia; Song, Yeong Hun; Park, Ki Hun

2018-08-01

Four new caged xanthones (1-4) and two known compounds (5, 6) were isolated from the roots of Cratoxylum cochinchinense, a polyphenol rich plant, collected in China. The structures of the isolated compounds (1-6) were characterized by obtaining their detailed spectroscopic data. In particular, compounds 1 and 6 were fully identified by X-ray crystallographic data. The isolated compounds (1-6) were evaluated against protein tyrosine phosphatase 1B (PTP1B), which plays an important role in diabetes, obesity, and cancer. Among these compounds, 3, 4, and 6 displayed significant inhibition with IC 50 values of 76.3, 43.2, and 6.6 µM, respectively. A detailed kinetic study was conducted by determining K m , V max , and the ratio of K ik and K iv , which revealed that all the compounds behaved as competitive inhibitors. Copyright © 2018 Elsevier Inc. All rights reserved.

Inhibition of CDC25B Phosphatase Through Disruption of Protein–Protein Interaction

DOE PAGES

Lund, George; Dudkin, Sergii; Borkin, Dmitry; ...

2014-11-25

CDC25 phosphatases are key cell cycle regulators and represent very attractive but challenging targets for anticancer drug discovery. Here in this paper, we explored whether fragment-based screening represents a valid approach to identify inhibitors of CDC25B. This resulted in identification of 2-fluoro-4-hydroxybenzonitrile, which directly binds to the catalytic domain of CDC25B. Interestingly, NMR data and the crystal structure demonstrate that this compound binds to the pocket distant from the active site and adjacent to the protein–protein interaction interface with CDK2/Cyclin A substrate. Furthermore, we developed a more potent analogue that disrupts CDC25B interaction with CDK2/Cyclin A and inhibits dephosphorylation ofmore » CDK2. Based on these studies, we provide a proof of concept that targeting CDC25 phosphatases by inhibiting their protein–protein interactions with CDK2/Cyclin A substrate represents a novel, viable opportunity to target this important class of enzymes.« less

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.

Covalent Allosteric Inactivation of Protein Tyrosine Phosphatase 1B (PTP1B) by an Inhibitor-Electrophile Conjugate.

PubMed

Punthasee, Puminan; Laciak, Adrian R; Cummings, Andrea H; Ruddraraju, Kasi Viswanatharaju; Lewis, Sarah M; Hillebrand, Roman; Singh, Harkewal; Tanner, John J; Gates, Kent S

2017-04-11

Protein tyrosine phosphatase 1B (PTP1B) is a validated drug target, but it has proven difficult to develop medicinally useful, reversible inhibitors of this enzyme. Here we explored covalent strategies for the inactivation of PTP1B using a conjugate composed of an active site-directed 5-aryl-1,2,5-thiadiazolidin-3-one 1,1-dioxide inhibitor connected via a short linker to an electrophilic α-bromoacetamide moiety. Inhibitor-electrophile conjugate 5a caused time-dependent loss of PTP1B activity consistent with a covalent inactivation mechanism. The inactivation occurred with a second-order rate constant of (1.7 ± 0.3) × 10 2 M -1 min -1 . Mass spectrometric analysis of the inactivated enzyme indicated that the primary site of modification was C121, a residue distant from the active site. Previous work provided evidence that covalent modification of the allosteric residue C121 can cause inactivation of PTP1B [Hansen, S. K., Cancilla, M. T., Shiau, T. P., Kung, J., Chen, T., and Erlanson, D. A. (2005) Biochemistry 44, 7704-7712]. Overall, our results are consistent with an unusual enzyme inactivation process in which noncovalent binding of the inhibitor-electrophile conjugate to the active site of PTP1B protects the nucleophilic catalytic C215 residue from covalent modification, thus allowing inactivation of the enzyme via selective modification of allosteric residue C121.

Increased PTP1B expression and phosphatase activity in colorectal cancer results in a more invasive phenotype and worse patient outcome.

PubMed

Hoekstra, Elmer; Das, Asha M; Swets, Marloes; Cao, Wanlu; van der Woude, C Janneke; Bruno, Marco J; Peppelenbosch, Maikel P; Kuppen, Peter J K; Ten Hagen, Timo L M; Fuhler, Gwenny M

2016-04-19

Cell signaling is dependent on the balance between phosphorylation of proteins by kinases and dephosphorylation by phosphatases. This balance if often disrupted in colorectal cancer (CRC), leading to increased cell proliferation and invasion. For many years research has focused on the role of kinases as potential oncogenes in cancer, while phosphatases were commonly assumed to be tumor suppressive. However, this dogma is currently changing as phosphatases have also been shown to induce cancer growth. One of these phosphatases is protein tyrosine phosphatase 1B (PTP1B). Here we report that the expression of PTP1B is increased in colorectal cancer as compared to normal tissue, and that the intrinsic enzymatic activity of the protein is also enhanced. This suggests a role for PTP1B phosphatase activity in CRC formation and progression. Furthermore, we found that increased PTP1B expression is correlated to a worse patient survival and is an independent prognostic marker for overall survival and disease free survival. Knocking down PTP1B in CRC cell lines results in a less invasive phenotype with lower adhesion, migration and proliferation capabilities. Together, these results suggest that inhibition of PTP1B activity is a promising new target in the treatment of colorectal cancer and the prevention of metastasis.

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.

Varic acid analogues from fungus as PTP1B inhibitors: Biological evaluation and structure-activity relationships.

PubMed

Sun, Wenlong; Zhuang, Chunlin; Li, Xia; Zhang, Bowei; Lu, Xinhua; Zheng, Zhihui; Dong, Yuesheng

2017-08-01

Protein tyrosine phosphatase 1B (PTP1B) inhibitors as potential therapies for diabetes and obesity have attracted much attention in recent years. Six varic acid analogues were isolated from two strains of fungi and evaluated for PTP1B inhibition activities. The structure-activity relationships were also characterized and predicted by molecular modeling. Further kinetic studies indicated the reversible and competitive inhibition manner of varic acid analogues. Trivaric acid showed insulin-sensitizing effect not only in vitro but also in vivo, representing a promising lead compound for further optimization. Copyright © 2017 Elsevier Ltd. All rights reserved.

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.

Salvianolic acid B reverses multidrug resistance in nude mice bearing human colon cancer stem cells.

PubMed

Guo, Piaoting; Wang, Jianchao; Gao, Wencang; Liu, Xia; Wu, Shaofei; Wan, Boshun; Xu, Lei; Li, Yanhua

2018-05-29

Salvianolic acid B (SalB) is a water‑soluble phenolic compound, extractable from Salvia miltiorrhiza, and has previously been demonstrated to reverse tumor multidrug resistance (MDR) in colon cancer cells. Cancer stem cells (CSCs) are closely associated with drug resistance. Therefore, establishing a nude mouse model bearing human colon CSCs is important for the study of the mechanisms underlying colon cancer drug resistance as well as the reversal of drug resistance. The present study aimed to establish a nude mouse model bearing human colon CSCs and to investigate the effects of SalB on the drug resistance exhibited by the nude mouse model as well as determine its underlying mechanism. Cells from two colon cancer cell lines (LoVo and HCT‑116) were cultured in serum‑free medium to obtain CSCs‑enriched spheroid cells. Following this, nude mice were transplanted with LoVo and HCT‑116 colon CSCs to establish the CSC nude mouse model, which was subsequently demonstrated to exhibit MDR. The results of the present study revealed that following treatment with SalB, the chemotherapeutic drug resistance of xenografts was reversed to a certain extent. Western blot analysis was performed to investigate the expression levels of cluster of differentiation (CD)44, CD133, transcription factor sox‑2 (SOX2) and ATP‑binding cassette sub‑family G member 2 (ABCG2) proteins, and the results demonstrated that treatment with SalB downregulated the expression of CD44, SOX2 and ABCG2 proteins in both LoVo and HCT‑116 colon CSCs xenografts. In conclusion, the results of the present study revealed that a serum‑free suspension method can be performed to successfully isolate colon CSCs. In addition, a nude mice bearing colon CSCs animal model was successfully established, and associated tumors were confirmed to exhibit MDR. Furthermore, SalB was demonstrated to successfully reverse MDR in nude mice bearing LoVo and HCT‑116 colon CSCs, as well as suppress the expression

Fucosterol activates the insulin signaling pathway in insulin resistant HepG2 cells via inhibiting PTP1B.

PubMed

Jung, Hyun Ah; Bhakta, Himanshu Kumar; Min, Byung-Sun; Choi, Jae Sue

2016-10-01

Insulin resistance is a characteristic feature of type 2 diabetes mellitus (T2DM) and is characterized by defects in insulin signaling. This study investigated the modulatory effects of fucosterol on the insulin signaling pathway in insulin-resistant HepG2 cells by inhibiting protein tyrosine phosphatase 1B (PTP1B). In addition, molecular docking simulation studies were performed to predict binding energies, the specific binding site of fucosterol to PTP1B, and to identify interacting residues using Autodock 4.2 software. Glucose uptake was determined using a fluorescent D-glucose analogue and the glucose tracer 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxyglucose, and the signaling pathway was detected by Western blot analysis. We found that fucosterol enhanced insulin-provoked glucose uptake and conjointly decreased PTP1B expression level in insulin-resistant HepG2 cells. Moreover, fucosterol significantly reduced insulin-stimulated serine (Ser307) phosphorylation of insulin receptor substrate 1 (IRS1) and increased phosphorylation of Akt, phosphatidylinositol-3-kinase, and extracellular signal- regulated kinase 1 at concentrations of 12.5, 25, and 50 µM in insulin-resistant HepG2 cells. Fucosterol inhibited caspase-3 activation and nuclear factor kappa B in insulin-resistant hepatocytes. These results suggest that fucosterol stimulates glucose uptake and improves insulin resistance by downregulating expression of PTP1B and activating the insulin signaling pathway. Thus, fucosterol has potential for development as an anti-diabetic agent.

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.

Lnk adaptor suppresses radiation resistance and radiation-induced B-cell malignancies by inhibiting IL-11 signaling

PubMed Central

Louria-Hayon, Igal; Frelin, Catherine; Ruston, Julie; Gish, Gerald; Jin, Jing; Kofler, Michael M.; Lambert, Jean-Philippe; Adissu, Hibret A.; Milyavsky, Michael; Herrington, Robert; Minden, Mark D.; Dick, John E.; Gingras, Anne-Claude; Iscove, Norman N.; Pawson, Tony

2013-01-01

The Lnk (Sh2b3) adaptor protein dampens the response of hematopoietic stem cells and progenitors (HSPCs) to a variety of cytokines by inhibiting JAK2 signaling. As a consequence, Lnk−/− mice develop hematopoietic hyperplasia, which progresses to a phenotype resembling the nonacute phase of myeloproliferative neoplasm. In addition, Lnk mutations have been identified in human myeloproliferative neoplasms and acute leukemia. We find that Lnk suppresses the development of radiation-induced acute B-cell malignancies in mice. Lnk-deficient HSPCs recover more effectively from irradiation than their wild-type counterparts, and this resistance of Lnk−/− HSPCs to radiation underlies the subsequent emergence of leukemia. A search for the mechanism responsible for radiation resistance identified the cytokine IL-11 as being critical for the ability of Lnk−/− HSPCs to recover from irradiation and subsequently become leukemic. In IL-11 signaling, wild-type Lnk suppresses tyrosine phosphorylation of the Src homology region 2 domain-containing phosphatase-2/protein tyrosine phosphatase nonreceptor type 11 and its association with the growth factor receptor-bound protein 2, as well as activation of the Erk MAP kinase pathway. Indeed, Src homology region 2 domain-containing phosphatase-2 has a binding motif for the Lnk Src Homology 2 domain that is phosphorylated in response to IL-11 stimulation. IL-11 therefore drives a pathway that enhances HSPC radioresistance and radiation-induced B-cell malignancies, but is normally attenuated by the inhibitory adaptor Lnk. PMID:24297922

Cloning and sequence analysis of a full-length cDNA of SmPP1cb encoding turbot protein phosphatase 1 beta catalytic subunit

NASA Astrophysics Data System (ADS)

Qi, Fei; Guo, Huarong; Wang, Jian

2008-02-01

Reversible protein phosphorylation, catalyzed by protein kinases and phosphatases, is an important and versatile mechanism by which eukaryotic cells regulate almost all the signaling processes. Protein phosphatase 1 (PP1) is the first and well-characterized member of the protein serine/threonine phosphatase family. In the present study, a full-length cDNA encoding the beta isoform of the catalytic subunit of protein phosphatase 1(PP1cb), was for the first time isolated and sequenced from the skin tissue of flatfish turbot Scophthalmus maximus, designated SmPP1cb, by the rapid amplification of cDNA ends (RACE) technique. The cDNA sequence of SmPP1cb we obtained contains a 984 bp open reading frame (ORF), flanked by a complete 39 bp 5' untranslated region and 462 bp 3' untranslated region. The ORF encodes a putative 327 amino acid protein, and the N-terminal section of this protein is highly acidic, Met-Ala-Glu-Gly-Glu-Leu-Asp-Val-Asp, a common feature for PP1 catalytic subunit but absent in protein phosphatase 2B (PP2B). And its calculated molecular mass is 37 193 Da and pI 5.8. Sequence analysis indicated that, SmPP1cb is extremely conserved in both amino acid and nucleotide acid levels compared with the PP1cb of other vertebrates and invertebrates, and its Kozak motif contained in the 5'UTR around ATG start codon is GXXAXXGXX ATGG, which is different from mammalian in two positions A-6 and G-3, indicating the possibility of different initiation of translation in turbot, and also the 3'UTR of SmPP1cb is highly diverse in the sequence similarity and length compared with other animals, especially zebrafish. The cloning and sequencing of SmPP1cb gene lays a good foundation for the future work on the biological functions of PP1 in the flatfish turbot.

Inhibition of protein tyrosine phosphatase 1B by flavonoids: A structure - activity relationship study.

PubMed

Proença, Carina; Freitas, Marisa; Ribeiro, Daniela; Sousa, Joana L C; Carvalho, Félix; Silva, Artur M S; Fernandes, Pedro A; Fernandes, Eduarda

2018-01-01

The classical non-transmembrane protein tyrosine phosphatase 1B (PTP1B) has emerged as a key negative regulator of insulin signaling pathways that leads to insulin resistance, turning this enzyme a promising therapeutic target in the management of type 2 diabetes mellitus (T2DM). In the present work, the in vitro inhibitory activity of a panel of structurally related flavonoids, for recombinant human PTP1B was studied and the type of inhibition of the most active compounds further evaluated. The majority of the studied flavonoids was tested in this work for the first time, including flavonoid C13, which was the most potent inhibitor. It was observed that the ability to inhibit PTP1B depends on the nature, position and number of substituents in the flavonoid structure, as the presence of both 7- and 8-OBn groups in the A ring, together with the presence of both 3' and 4'-OMe groups in the B ring and the 3-OH group in the C ring; these substituents increase the flavonoids' ability to inhibit PTP1B. In conclusion, some of the tested flavonoids seem to be promising PTP1B inhibitors and potential effective agents in the management of T2DM, by increasing insulin sensitivity. Copyright © 2017 Elsevier Ltd. All rights reserved.

Inhibitory Activity of Iron Chelators ATA and DFO on MCF-7 Breast Cancer Cells and Phosphatases PTP1B and SHP2.

PubMed

Kuban-Jankowska, Alicja; Sahu, Kamlesh K; Gorska-Ponikowska, Magdalena; Tuszynski, Jack A; Wozniak, Michal

2017-09-01

Rapidly-dividing cancer cells have higher requirement for iron compared to non-transformed cells, making iron chelating a potential anticancer strategy. In the present study we compared the anticancer activity of uncommon iron chelator aurintricarboxylic acid (ATA) with the known deferoxamine (DFO). We investigated the impact of ATA and DFO on the viability and proliferation of MCF-7 cancer cells. Moreover we performed enzymatic activity assays and computational analysis of the ATA and DFO effects on pro-oncogenic phosphatases PTP1B and SHP2. ATA and DFO decrease the viability and proliferation of breast cancer cells, but only ATA considerably reduces the activity of PTP1B and SHP2 phosphatases. Our studies indicated that ATA strongly inactivates and binds in the PTP1B and SHP2 active site, interacting with arginine residue essential for enzyme activity. We confirmed that iron chelating can be considered as a potential strategy for the adjunctive treatment of breast cancer. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

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.

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.

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.

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

Spondyloenchondrodysplasia Due to Mutations in ACP5: A Comprehensive Survey.

PubMed

Briggs, Tracy A; Rice, Gillian I; Adib, Navid; Ades, Lesley; Barete, Stephane; Baskar, Kannan; Baudouin, Veronique; Cebeci, Ayse N; Clapuyt, Philippe; Coman, David; De Somer, Lien; Finezilber, Yael; Frydman, Moshe; Guven, Ayla; Heritier, Sébastien; Karall, Daniela; Kulkarni, Muralidhar L; Lebon, Pierre; Levitt, David; Le Merrer, Martine; Linglart, Agnes; Livingston, John H; Navarro, Vincent; Okenfuss, Ericka; Puel, Anne; Revencu, Nicole; Scholl-Bürgi, Sabine; Vivarelli, Marina; Wouters, Carine; Bader-Meunier, Brigitte; Crow, Yanick J

2016-04-01

Spondyloenchondrodysplasia is a rare immuno-osseous dysplasia caused by biallelic mutations in ACP5. We aimed to provide a survey of the skeletal, neurological and immune manifestations of this disease in a cohort of molecularly confirmed cases. We compiled clinical, genetic and serological data from a total of 26 patients from 18 pedigrees, all with biallelic ACP5 mutations. We observed a variability in skeletal, neurological and immune phenotypes, which was sometimes marked even between affected siblings. In total, 22 of 26 patients manifested autoimmune disease, most frequently autoimmune thrombocytopenia and systemic lupus erythematosus. Four patients were considered to demonstrate no clinical autoimmune disease, although two were positive for autoantibodies. In the majority of patients tested we detected upregulated expression of interferon-stimulated genes (ISGs), in keeping with the autoimmune phenotype and the likely immune-regulatory function of the deficient protein tartrate resistant acid phosphatase (TRAP). Two mutation positive patients did not demonstrate an upregulation of ISGs, including one patient with significant autoimmune disease controlled by immunosuppressive therapy. Our data expand the known phenotype of SPENCD. We propose that the OMIM differentiation between spondyloenchondrodysplasia and spondyloenchondrodysplasia with immune dysregulation is no longer appropriate, since the molecular evidence that we provide suggests that these phenotypes represent a continuum of the same disorder. In addition, the absence of an interferon signature following immunomodulatory treatments in a patient with significant autoimmune disease may indicate a therapeutic response important for the immune manifestations of spondyloenchondrodysplasia.

Differential effects of protein phosphatases in the recycling of metabotropic glutamate receptor 5.

PubMed

Mahato, P K; Pandey, S; Bhattacharyya, S

2015-10-15

The major excitatory neurotransmitter Glutamate acts on both ionotropic and metabotropic glutamate receptors (mGluRs) in the central nervous system. mGluR5, a member of the group I mGluR family is widely expressed throughout the brain and plays important roles in a variety of neuronal processes including various forms of synaptic plasticity. This receptor is also involved in various neuropsychiatric disorders, viz., Fragile X syndrome, autism etc. It has been reported that mGluR5 undergoes desensitization and subsequently internalization on ligand exposure in various cell types. However, the downstream events after the internalization and the molecular players involved in the post-endocytic events of this receptor have not been studied. In the present study, we find that subsequent to internalization mGluR5 enters the recycling compartment. After that the receptor recycles back to the cell surface. We also show here that the recycling of mGluR5 is dependent on protein phosphatases. Our data suggest that mGluR5 recycling is completely dependent on the activity of PP2A whereas, PP2B has partial effect on this process. Thus our study suggests that mGluR5 recycles back to the cell surface after ligand-dependent internalization and protein phosphatases that have been implicated in various forms of synaptic plasticity have differential effects on the recycling of mGluR5. Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.

Contribution of glutamate decarboxylase in Lactobacillus reuteri to acid resistance and persistence in sourdough fermentation

PubMed Central

2011-01-01

Background Acid stress impacts the persistence of lactobacilli in industrial sourdough fermentations, and in intestinal ecosystems. However, the contribution of glutamate to acid resistance in lactobacilli has not been demonstrated experimentally, and evidence for the contribution of acid resistance to the competitiveness of lactobacilli in sourdough is lacking. It was therefore the aim of this study to investigate the ecological role of glutamate decarboxylase in L. reuteri. Results A gene coding for a putative glutamate decarboxylase, gadB, was identified in the genome of L. reuteri 100-23. Different from the organization of genetic loci coding for glutamate decarboxylase in other lactic acid bacteria, gadB was located adjacent to a putative glutaminase gene, gls3. An isogenic deletion mutant, L. reuteri ∆gadB, was generated by a double crossover method. L. reuteri 100-23 but not L. reuteri ∆gadB converted glutamate to γ-aminobutyrate (GABA) in phosphate butter (pH 2.5). In sourdough, both strains converted glutamine to glutamate but only L. reuteri 100-23 accumulated GABA. Glutamate addition to phosphate buffer, pH 2.5, improved survival of L. reuteri 100-23 100-fold. However, survival of L. reuteri ∆gadB remained essentially unchanged. The disruption of gadB did not affect growth of L. reuteri in mMRS or in sourdough. However, the wild type strain L. reuteri 100-23 displaced L. reuteri ∆gadB after 5 cycles of fermentation in back-slopped sourdough fermentations. Conclusions The conversion of glutamate to GABA by L. reuteri 100-23 contributes to acid resistance and to competitiveness in industrial sourdough fermentations. The organization of the gene cluster for glutamate conversion, and the availability of amino acids in cereals imply that glutamine rather than glutamate functions as the substrate for GABA formation. The exceptional coupling of glutamine deamidation to glutamate decarboxylation in L. reuteri likely reflects adaptation to cereal

5-LO inhibition ameliorates palmitic acid-induced ER stress, oxidative stress and insulin resistance via AMPK activation in murine myotubes.

PubMed

Kwak, Hyun Jeong; Choi, Hye-Eun; Cheon, Hyae Gyeong

2017-07-10

Leukotriene B4 (LTB4) production via the 5-lipoxygenase (5-LO) pathway contributes to the development of insulin resistance in adipose and hepatic tissues, but the role of LTB4 in skeletal muscle is relatively unknown. Here, the authors investigated the role of LTB4 in C2C12 myotubes in palmitic acid (PA)-induced ER stress, inflammation and insulin resistance. PA (750 μM) evoked lipotoxicity (ER stress, oxidative stress, inflammation and insulin resistance) in association with LTB4 production. 5-LO inhibition reduced all the lipotoxic effects induced by PA. On the other hand, PA did not induce cysteinyl leukotrienes (CysLTs), which themselves had no effect on ER stress and inflammation. The beneficial effects of 5-LO suppression from PA-induced lipotoxicity were related with AMPK activation. In ob/ob mice, once daily oral administration of zileuton (50, 100 mg/kg) for 5 weeks improved insulin resistance, increased AMPK phosphorylation, and reduced LTB4 and ER stress marker expression in skeletal muscle. These results show that 5-LO inhibition by either zileuton or 5-LO siRNA protects C2C12 myotubes from PA-induced lipotoxicity, at least partly via AMPK activation, and suggest that the in vivo insulin-sensitizing effects of zileuton are in part attributable to its direct action on skeletal muscle via LTB4 downregulation followed by AMPK activation.

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

[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.

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.

Dendrobium moniliforme Exerts Inhibitory Effects on Both Receptor Activator of Nuclear Factor Kappa-B Ligand-Mediated Osteoclast Differentiation in Vitro and Lipopolysaccharide-Induced Bone Erosion in Vivo.

PubMed

Baek, Jong Min; Kim, Ju-Young; Ahn, Sung-Jun; Cheon, Yoon-Hee; Yang, Miyoung; Oh, Jaemin; Choi, Min Kyu

2016-03-01

Dendrobium moniliforme (DM) is a well-known plant-derived extract that is widely used in Oriental medicine. DM and its chemical constituents have been reported to have a variety of pharmacological effects, including anti-oxidative, anti-inflammatory, and anti-tumor activities; however, no reports discuss the beneficial effects of DM on bone diseases such as osteoporosis. Thus, we investigated the relationship between DM and osteoclasts, cells that function in bone resorption. We found that DM significantly reduced receptor activator of nuclear factor kappa-B ligand (RANKL)-induced tartrate-resistant acid phosphatase (TRAP)-positive osteoclast formation; DM directly induced the down-regulation of c-Fos and nuclear factor of activated T cells c1 (NFATc1) without affecting other RANKL-dependent transduction pathways. In the later stages of osteoclast maturation, DM negatively regulated the organization of filamentous actin (F-actin), resulting in impaired bone-resorbing activity by the mature osteoclasts. In addition, micro-computed tomography (μ-CT) analysis of the murine model revealed that DM had a beneficial effect on lipopolysaccharide (LPS)-mediated bone erosion. Histological analysis showed that DM attenuated the degradation of trabecular bone matrix and formation of TRAP-positive osteoclasts in bone tissues. These results suggest that DM is a potential candidate for the treatment of metabolic bone disorders such as osteoporosis.

Phosphatase of regenerating liver-3 is expressed in acute lymphoblastic leukemia and mediates leukemic cell adhesion, migration and drug resistance

PubMed Central

Hjort, Magnus A.; Abdollahi, Pegah; Vandsemb, Esten N.; Fenstad, Mona H.; Lund, Bendik; Slørdahl, Tobias S.; Børset, Magne; Rø, Torstein B.

2018-01-01

Phosphatase of regenerating liver-3 (PRL-3/PTP4A3) is upregulated in multiple cancers, including BCR-ABL1- and ETV6-RUNX-positive acute lymphoblastic leukemia (ALL). With this study, we aim to characterize the biological role of PRL-3 in B cell ALL (B-ALL). Here, we demonstrate that PRL-3 expression at mRNA and protein level was higher in B-ALL cells than in normal cells, as measured by qRT-PCR or flow cytometry. Further, we demonstrate that inhibition of PRL-3 using shRNA or a small molecular inhibitor reduced cell migration towards an SDF-1α gradient in the preB-ALL cell lines Reh and MHH-CALL-4. Knockdown of PRL-3 also reduced cell adhesion towards fibronectin in Reh cells. Mechanistically, PRL-3 mediated SDF-1α stimulated calcium release, and activated focal adhesion kinase (FAK) and Src, important effectors of migration and adhesion. Finally, PRL-3 expression made Reh cells more resistance to cytarabine treatment. In conclusion, the expression level of PRL-3 was higher in B-ALL cells than in normal cells. PRL-3 promoted adhesion, migration and resistance to cytarabine. PRL-3 may represent a novel target in the treatment of B-ALL. PMID:29423065

A Novel Inositol Pyrophosphate Phosphatase in Saccharomyces cerevisiae: Siw14 PROTEIN SELECTIVELY CLEAVES THE β-PHOSPHATE FROM 5-DIPHOSPHOINOSITOL PENTAKISPHOSPHATE (5PP-IP5).

PubMed

Steidle, Elizabeth A; Chong, Lucy S; Wu, Mingxuan; Crooke, Elliott; Fiedler, Dorothea; Resnick, Adam C; Rolfes, Ronda J

2016-03-25

Inositol pyrophosphates are high energy signaling molecules involved in cellular processes, such as energetic metabolism, telomere maintenance, stress responses, and vesicle trafficking, and can mediate protein phosphorylation. Although the inositol kinases underlying inositol pyrophosphate biosynthesis are well characterized, the phosphatases that selectively regulate their cellular pools are not fully described. The diphosphoinositol phosphate phosphohydrolase enzymes of the Nudix protein family have been demonstrated to dephosphorylate inositol pyrophosphates; however, theSaccharomyces cerevisiaehomolog Ddp1 prefers inorganic polyphosphate over inositol pyrophosphates. We identified a novel phosphatase of the recently discovered atypical dual specificity phosphatase family as a physiological inositol pyrophosphate phosphatase. Purified recombinant Siw14 hydrolyzes the β-phosphate from 5-diphosphoinositol pentakisphosphate (5PP-IP5or IP7)in vitro. In vivo,siw14Δ yeast mutants possess increased IP7levels, whereas heterologousSIW14overexpression eliminates IP7from cells. IP7levels increased proportionately whensiw14Δ was combined withddp1Δ orvip1Δ, indicating independent activity by the enzymes encoded by these genes. We conclude that Siw14 is a physiological phosphatase that modulates inositol pyrophosphate metabolism by dephosphorylating the IP7isoform 5PP-IP5to IP6. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Bone markers during acute burn care: Relevance to clinical practice?

PubMed

Rousseau, Anne-Françoise; Damas, Pierre; Delanaye, Pierre; Cavalier, Etienne

2017-02-01

Bone changes are increasingly described after burn. How bone markers could help to detect early bone changes or to screen burn patients at higher risk of demineralization is still not made clear. We performed an observational study assessing the changes in serum bone markers after moderate burn. Adults admitted in the first 24h following burn extended on >10% body surface area were included. Serum levels of collagen type 1 cross-linked C-telopeptide (CTX), tartrate-resistant acid phosphatase 5b (TRAP), type 1 procollagen N-terminal (P1NP) and bone alkaline phosphatase (b-ALP) were measured at admission and every week during the first month. Data are expressed as median [min-max]. Bone markers were measured in 20 patients: 18 men, 2 women (including one post-menopausal). Age was 46 [19-86] years old, burn surface area reached 15 [7-85] %. Twelve patients completed the study. All biomarkers mainly remained into normal ranges during evolution. A huge variability was observed regarding biomarkers evolution. Patient's evolution was not linear and could fluctuate from a decrease to an increase of blood concentrations. There was not necessarily a consistency between the two formation or the two resorption markers. Variations observed between two consecutive measurements were lesser than the accepted critical difference in almost one third of the cases. Considering available data, role and interest of bone markers in management of burn related bone disease remain unclear. Copyright © 2016 Elsevier Ltd and ISBI. All rights reserved.

Tanzawaic acid derivatives from a marine isolate of Penicillium sp. (SF-6013) with anti-inflammatory and PTP1B inhibitory activities.

PubMed

Quang, Tran Hong; Ngan, Nguyen Thi Thanh; Ko, Wonmin; Kim, Dong-Cheol; Yoon, Chi-Su; Sohn, Jae Hak; Yim, Joung Han; Kim, Youn-Chul; Oh, Hyuncheol

2014-12-15

Chemical investigation of a marine-derived fungus Penicillium sp. SF-6013 resulted in the discovery of a new tanzawaic acid derivative, 2E,4Z-tanzawaic acid D (1), together with four known analogues, tanzawaic acids A (2) and D (3), a salt form of tanzawaic acid E (4), and tanzawaic acid B (5). Their structures were mainly determined by analysis of NMR and MS data, along with chemical methods. Preliminary screening for anti-inflammatory effects in lipopolysaccharide (LPS)-activated microglial BV-2 cells showed that compounds 1, 2, and 5 inhibited the production of nitric oxide (NO) with IC50 values of 37.8, 7.1, and 42.5 μM, respectively. Compound 2 also inhibited NO production in LPS-stimulated RAW264.7 murine macrophages with an IC50 value of 27.0 μM. Moreover, these inhibitory effects correlated with the suppressive effect of compound 2 on inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expression in LPS-stimulated RAW264.7 and BV2 cells. In addition, compounds 2 and 5 significantly inhibited the activity of protein tyrosine phosphatase 1B (PTP1B) with the same IC50 value (8.2 μM). Copyright © 2014 Elsevier Ltd. All rights reserved.

Arthritogenic alphaviral infection perturbs osteoblast function and triggers pathologic bone loss

PubMed Central

Chen, Weiqiang; Foo, Suan-Sin; Rulli, Nestor E.; Taylor, Adam; Sheng, Kuo-Ching; Herrero, Lara J.; Herring, Belinda L.; Lidbury, Brett A.; Li, Rachel W.; Walsh, Nicole C.; Sims, Natalie A.; Smith, Paul N.; Mahalingam, Suresh

2014-01-01

Arthritogenic alphaviruses including Ross River virus (RRV), Sindbis virus, and chikungunya virus cause worldwide outbreaks of musculoskeletal disease. The ability of alphaviruses to induce bone pathologies remains poorly defined. Here we show that primary human osteoblasts (hOBs) can be productively infected by RRV. RRV-infected hOBs produced high levels of inflammatory cytokine including IL-6. The RANKL/OPG ratio was disrupted in the synovial fluid of RRV patients, and this was accompanied by an increase in serum Tartrate-resistant acid phosphatase 5b (TRAP5b) levels. Infection of bone cells with RRV was validated using an established RRV murine model. In wild-type mice, infectious virus was detected in the femur, tibia, patella, and foot, together with reduced bone volume in the tibial epiphysis and vertebrae detected by microcomputed tomographic (µCT) analysis. The RANKL/OPG ratio was also disrupted in mice infected with RRV; both this effect and the bone loss were blocked by treatment with an IL-6 neutralizing antibody. Collectively, these findings provide previously unidentified evidence that alphavirus infection induces bone loss and that OBs are capable of producing proinflammatory mediators during alphavirus-induced arthralgia. The perturbed RANKL/OPG ratio in RRV-infected OBs may therefore contribute to bone loss in alphavirus infection. PMID:24733914

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.

Transgenic Arabidopsis Plants Expressing the Type 1 Inositol 5-Phosphatase Exhibit Increased Drought Tolerance and Altered Abscisic Acid Signaling[W

PubMed Central

Perera, Imara Y.; Hung, Chiu-Yueh; Moore, Candace D.; Stevenson-Paulik, Jill; Boss, Wendy F.

2008-01-01

The phosphoinositide pathway and inositol-1,4,5-trisphosphate (InsP3) are implicated in plant responses to stress. To determine the downstream consequences of altered InsP3-mediated signaling, we generated transgenic Arabidopsis thaliana plants expressing the mammalian type I inositol polyphosphate 5-phosphatase (InsP 5-ptase), which specifically hydrolyzes soluble inositol phosphates and terminates the signal. Rapid transient Ca2+ responses to a cold or salt stimulus were reduced by ∼30% in these transgenic plants. Drought stress studies revealed, surprisingly, that the InsP 5-ptase plants lost less water and exhibited increased drought tolerance. The onset of the drought stress was delayed in the transgenic plants, and abscisic acid (ABA) levels increased less than in the wild-type plants. Stomatal bioassays showed that transgenic guard cells were less responsive to the inhibition of opening by ABA but showed an increased sensitivity to ABA-induced closure. Transcript profiling revealed that the drought-inducible ABA-independent transcription factor DREB2A and a subset of DREB2A-regulated genes were basally upregulated in the InsP 5-ptase plants, suggesting that InsP3 is a negative regulator of these DREB2A-regulated genes. These results indicate that the drought tolerance of the InsP 5-ptase plants is mediated in part via a DREB2A-dependent pathway and that constitutive dampening of the InsP3 signal reveals unanticipated interconnections between signaling pathways. PMID:18849493

Effects of light and the regulatory B-subunit composition of protein phosphatase 2A on the susceptibility of Arabidopsis thaliana to aphid (Myzus persicae) infestation

PubMed Central

Rasool, Brwa; Karpinska, Barbara; Konert, Grzegorz; Durian, Guido; Denessiouk, Konstantin; Kangasjärvi, Saijaliisa; Foyer, Christine H.

2014-01-01

The interactions between biotic and abiotic stress signaling pathways are complex and poorly understood but protein kinase/phosphatase cascades are potentially important components. Aphid fecundity and susceptibility to Pseudomonas syringae infection were determined in the low light-grown Arabidopsis thaliana wild type and in mutant lines defective in either the protein phosphatase (PP)2A regulatory subunit B'γ (gamma; pp2a-b'γ) or B'ζ (zeta; pp2a-b'ζ1-1 and pp2a-b'ζ 1-2) and in gamma zeta double mutants (pp2a-b'γζ) lacking both subunits. All the mutants except for pp2a-b'ζ 1-1 had significantly lower leaf areas than the wild type. Susceptibility to P. syringae was similar in all genotypes. In contrast, aphid fecundity was significantly decreased in the pp2a-b'γ mutant relative to the wild type but not in the pp2a-b'γζ double mutant. A high light pre-treatment, which led to a significant increase in rosette growth in all mutant lines but not in the wild type, led to a significant decrease in aphid fecundity in all genotypes. The high light pre-treatment abolished the differences in aphid resistance observed in the pp2a-b'γ mutant relative to the wild type. The light and CO2 response curves for photosynthesis were changed in response to the high light pre-treatment, but the high light effects were similar in all genotypes. These data demonstrate that a pre-exposure to high light and the composition of B-subunits on the trimeric PP2A holoenzymes are important in regulating plant resistance to aphids. The functional specificity for the individual regulatory B-subunits may therefore limit aphid colonization, depending on the prevailing abiotic stress environment. PMID:25191331

Peptide Nucleic Acid Array for Detection of Point Mutations in Hepatitis B Virus Associated with Antiviral Resistance ▿ †

PubMed Central

Jang, Hyunjung; Kim, Jihyun; Choi, Jae-jin; Son, Yeojin; Park, Heekyung

2010-01-01

The detection of antiviral-resistant hepatitis B virus (HBV) mutations is important for monitoring the response to treatment and for effective treatment decisions. We have developed an array using peptide nucleic acid (PNA) probes to detect point mutations in HBV associated with antiviral resistance. PNA probes were designed to detect mutations associated with resistance to lamivudine, adefovir, and entecavir. The PNA array assay was sensitive enough to detect 102 copies/ml. The PNA array assay was able to detect mutants present in more than 5% of the virus population when the total HBV DNA concentration was greater than 104 copies/ml. We analyzed a total of 68 clinical samples by this assay and validated its usefulness by comparing results to those of the sequencing method. The PNA array correctly identified viral mutants and has high concordance (98.3%) with direct sequencing in detecting antiviral-resistant mutations. Our results showed that the PNA array is a rapid, sensitive, and easily applicable assay for the detection of antiviral-resistant mutation in HBV. Thus, the PNA array is a useful and powerful diagnostic tool for the detection of point mutations or polymorphisms. PMID:20573874

Structure of a Protein Phosphatase 2A Holoenzyme: Insights into B55-Mediated Tau Dephosphorylation

PubMed Central

Xu, Yanhui; Chen, Yu; Zhang, Ping; Jeffrey, Philip D.; Shi, Yigong

2009-01-01

Summary Protein phosphatase 2A (PP2A) regulates many essential aspects of cellular physiology. Members of the regulatory B/B55/PR55 family are thought to play a key role in the dephosphorylation of Tau, whose hyperphosphorylation contributes to Alzheimer's disease. The underlying mechanisms of the PP2A-Tau connection remain largely enigmatic. Here, we report the complete reconstitution of a Tau dephosphorylation assay and the crystal structure of a heterotrimeric PP2A holoenzyme involving the regulatory subunit Bα. We show that Bα specifically and markedly facilitates dephosphorylation of the phosphorylated Tau in our reconstituted assay. The Bα subunit comprises a seven-bladed β propeller, with an acidic, substrate-binding groove located in the center of the propeller. The β propeller latches onto the ridge of the PP2A scaffold subunit with the help of a protruding β hairpin arm. Structure-guided mutagenesis studies revealed the underpinnings of PP2A-mediated dephosphorylation of Tau. PMID:18922469

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.

Biotechnological approach for systemic delivery of membrane Receptor Activator of NF-κB Ligand (RANKL) active domain into the circulation.

PubMed

Cappariello, Alfredo; Paone, Riccardo; Maurizi, Antonio; Capulli, Mattia; Rucci, Nadia; Muraca, Maurizio; Teti, Anna

2015-04-01

Deficiency of Receptor Activator of NF-κB Ligand (RANKL) prevents osteoclast formation causing osteopetrosis. RANKL is a membrane-bound protein cleaved into active soluble (s)RANKL by metalloproteinase 14 (MMP14). We created a bio-device that harbors primary osteoblasts, cultured on 3D hydroxyapatite scaffolds carrying immobilized MMP14 catalytic domain. Scaffolds were sealed in diffusion chambers and implanted in RANKL-deficient mice. Mice received 1 or 2 diffusion chambers, once or twice and were sacrificed after 1 or 2 months from implants. A progressive increase of body weight was observed in the implanted groups. Histological sections of tibias of non-implanted mice were negative for the osteoclast marker Tartrate-Resistant Acid Phosphatase (TRAcP), consistent with the lack of osteoclasts. In contrast, tibias excised from implanted mice showed TRAcP-positive cells in the bone marrow and on the bone surface, these latter morphologically similar to mature osteoclasts. In mice implanted with 4 diffusion chambers total, we noted the highest number and size of TRAcP-positive cells, with quantifiable eroded bone surface and significant reduction of trabecular bone volume. These data demonstrate that our bio-device delivers effective sRANKL, inducing osteoclastogenesis in RANKL-deficient mice, supporting the feasibility of an innovative experimental strategy to treat systemic cytokine deficiencies. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

Emodin suppresses cadmium-induced osteoporosis by inhibiting osteoclast formation.

PubMed

Chen, Xiao; Ren, Shuai; Zhu, Guoying; Wang, Zhongqiu; Wen, Xiaolin

2017-09-01

Environmental level of cadmium (Cd) exposure can induce bone loss. Emodin, a naturally compound found in Asian herbal medicines, could influence osteoblast/osteoclast differentiation. However, the effects of emodin on Cd-induced bone damage are not clarified. The aim of this study was to investigate the role of emodin on Cd-induced osteoporosis. Sprague-Dawley male rats were divided into three groups which were given 0mg/L, 50mg Cd/L and 50mg Cd/L plus emodin (50mg/kg body weight). Bone histological investigation, microCT analysis, metabolic biomarker determination and immunohistochemical staining were performed at the 12th week. The bone mass and bone microstructure index of rats treated with Cd were obviously lower than in control. Cd markedly enhanced the osteoclast formation compared with control. Emodin significantly abolished the Cd-induced bone microstructure damage (p<0.05), osteoclast formation and increase of tartrate-resistant acid phosphatase 5b level (p<0.05). Our data further showed that emodin attenuated the Cd-induced inhibition of osteoprotegerin expression and stimulation of receptor activator for nuclear factor-κ B ligand expression. Our data show that emodin suppresses the Cd-induced osteoporosis by inhibiting osteoclast formation. Copyright © 2017 Elsevier B.V. All rights reserved.

Potential effects of valproate and oxcarbazepine on growth velocity and bone metabolism in epileptic children- a medical center experience.

PubMed

Lin, Chien-Ming; Fan, Hueng-Chuen; Chao, Tsu-Yi; Chu, Der-Ming; Lai, Chi-Chieh; Wang, Chih-Chien; Chen, Shyi-Jou

2016-05-03

Children with longstanding use of antiepileptic drugs (AEDs) are susceptible to developing low bone mineral density and an increased fracture risk. However, the literature regarding the effects of AEDs on growth in epileptic children is limited. The aim of this study was to investigate the potential effects of valproate (VPA) and/or oxcarbazepine (OXC) therapy on growth velocity and bone metabolism. Seventy-three ambulatory children (40 boys and 33 girls) with epilepsy, aged between 1 and 18 years (mean age 9.8 ± 4.1 years), were evaluated for growth velocity before and for 1 year after VPA and/or OXC treatment. The bone resorption marker serum tartrate-resistant acid phosphatase 5b (TRAcP5b) and the bone formation marker serum bone-specific alkaline phosphatase (BAP) were measured post-AEDs therapy for 1 year. The difference in growth velocity (ΔHt) and body weight change (ΔWt) between pre- and post-AEDs treatment were -1.0 ± 2.8 cm/year (P < 0.05) and 0.1 ± 3.9 kg/year (P = 0.84), respectively. The study population had serum TRAcP5b-SDS of -1.6 ± 1.2 and BAP-SDS of 1.7 ± 3.7 compared with sex- and age-matched healthy children. Significant correlation between serum TRAcP 5b and BAP activities was noted (r = 0.60, p < 0.001). There was a positive correlation between growth velocity and serum TRAcP 5b activity after AED treatment (r = 0.42, p < 0.01). No correlation was found between ΔHt, ΔWt, serum TRAcP 5b, BAP activity and types of AEDs. Growth velocity was significantly decreased in epileptic children after 1 year of VPA and/or OXC treatment. The effect of VPA and/or OXC therapy on dysregulation of bone metabolism might play a crucial role in physical growth.

Disruption of protein-tyrosine phosphatase 1B expression in the pancreas affects β-cell function.

PubMed

Liu, Siming; Xi, Yannan; Bettaieb, Ahmed; Matsuo, Kosuke; Matsuo, Izumi; Kulkarni, Rohit N; Haj, Fawaz G

2014-09-01

Protein-tyrosine phosphatase 1B (PTP1B) is a physiological regulator of glucose homeostasis and energy balance. However, the role of PTP1B in pancreatic endocrine function remains largely unknown. To investigate the metabolic role of pancreatic PTP1B, we generated mice with pancreas PTP1B deletion (panc-PTP1B KO). Mice were fed regular chow or a high-fat diet, and metabolic parameters, insulin secretion and glucose tolerance were determined. On regular chow, panc-PTP1B KO and control mice exhibited comparable glucose tolerance whereas aged panc-PTP1B KO exhibited mild glucose intolerance. Furthermore, high-fat feeding promoted earlier impairment of glucose tolerance and attenuated glucose-stimulated insulin secretion in panc-PTP1B KO mice. The secretory defect in glucose-stimulated insulin secretion was recapitulated in primary islets ex vivo, suggesting that the effects were likely cell-autonomous. At the molecular level, PTP1B deficiency in vivo enhanced basal and glucose-stimulated tyrosyl phosphorylation of EphA5 in islets. Consistently, PTP1B overexpression in the glucose-responsive MIN6 β-cell line attenuated EphA5 tyrosyl phosphorylation, and substrate trapping identified EphA5 as a PTP1B substrate. In summary, these studies identify a novel role for PTP1B in pancreatic endocrine function.

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

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.

HF diets increase hypothalamic PTP1B and induce leptin resistance through both leptin-dependent and -independent mechanisms

PubMed Central

White, Christy L.; Whittington, Amy; Barnes, Maria J.; Wang, Zhong; Bray, George A.; Morrison, Christopher D.

2009-01-01

Protein tyrosine phosphatase 1B (PTP1B) contributes to leptin resistance by inhibiting intracellular leptin receptor signaling. Mice with whole body or neuron-specific deletion of PTP1B are hypersensitive to leptin and resistant to diet-induced obesity. Here we report a significant increase in PTP1B protein levels in the mediobasal hypothalamus (P = 0.003) and a concomitant reduction in leptin sensitivity following 28 days of high-fat (HF) feeding in rats. A significant increase in PTP1B mRNA levels was also observed in rats chronically infused with leptin (3 μg/day icv) for 14 days (P = 0.01) and in leptin-deficient ob/ob mice infused with leptin (5 μg/day sc for 14 days; P = 0.003). When saline-infused ob/ob mice were placed on a HF diet for 14 days, an increase in hypothalamic PTP1B mRNA expression was detected (P = 0.001) despite the absence of circulating leptin. In addition, although ob/ob mice were much more sensitive to leptin on a low-fat (LF) diet, a reduction in this sensitivity was still observed following exposure to a HF diet. Taken together, these data indicate that hypothalamic PTP1B is specifically increased during HF diet-induced leptin resistance. This increase in PTP1B is due in part to chronic hyperleptinemia, suggesting that hyperleptinemia is one mechanism contributing to the development of leptin resistance. However, these data also indicate that leptin is not required for the increase in hypothalamic PTP1B or the development of leptin resistance. Therefore, additional, leptin-independent mechanisms must exist that increase hypothalamic PTP1B and contribute to leptin resistance. PMID:19017730

Role of 5-HT5A and 5-HT1B/1D receptors in the antinociception produced by ergotamine and valerenic acid in the rat formalin test.

PubMed

Vidal-Cantú, Guadalupe C; Jiménez-Hernández, Mildred; Rocha-González, Héctor I; Villalón, Carlos M; Granados-Soto, Vinicio; Muñoz-Islas, Enriqueta

2016-06-15

Sumatriptan, dihydroergotamine and methysergide inhibit 1% formalin-induced nociception by activation of peripheral 5-HT1B/1D receptors. This study set out to investigate the pharmacological profile of the antinociception produced by intrathecal and intraplantar administration of ergotamine (a 5-HT1B/1D and 5-HT5A/5B receptor agonist) and valerenic acid (a partial agonist at 5-HT5A receptors). Intraplantar injection of 1% formalin in the right hind paw resulted in spontaneous flinching behavior of the injected hindpaw of female Wistar rats. Intrathecal ergotamine (15nmol) or valerenic acid (1 nmol) blocked in a dose dependent manner formalin-induced nociception. The antinociception by intrathecal ergotamine (15nmol) or valerenic acid (1nmol) was partly or completely blocked by intrathecal administration of the antagonists: (i) methiothepin (non-selective 5-HT5A/5B; 0.01-0.1nmol); (ii) SB-699551 (selective 5-HT5A; up to 10nmol); (iii) anti-5-HT5A antibody; (iv) SB-224289 (selective 5-HT1B; 0.1-1nmol); or (v) BRL-15572 (selective 5-HT1D; 0.1-1nmol). Likewise, antinociception by intraplantar ergotamine (15nmol) and valerenic acid (10nmol) was: (i) partially blocked by methiothepin (1nmol), SB-699551 (10nmol) or SB-224289 (1nmol); and (ii) abolished by BRL-15572 (1nmol). The above doses of antagonists (which did not affect per se the formalin-induced nociception) were high enough to completely block their respective receptors. Our results suggest that ergotamine and valerenic acid produce antinociception via 5-HT5A and 5-HT1B/1D receptors located at both spinal and peripheral sites. This provides new evidence for understanding the modulation of nociceptive pathways in inflammatory pain. Copyright © 2016 Elsevier B.V. All rights reserved.

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.

Regulated binding of PTP1B-like phosphatase to N-cadherin: control of cadherin-mediated adhesion by dephosphorylation of beta-catenin

PubMed Central

1996-01-01

Cadherins are a family of cell-cell adhesion molecules which play a central role in controlling morphogenetic movements during development. Cadherin function is regulated by its association with the actin containing cytoskeleton, an association mediated by a complex of cytoplasmic proteins, the catenins: alpha, beta, and gamma. Phosphorylated tyrosine residues on beta-catenin are correlated with loss of cadherin function. Consistent with this, we find that only nontyrosine phosphorylated beta-catenin is associated with N-cadherin in E10 chick retina tissue. Moreover, we demonstrate that a PTP1B-like tyrosine phosphatase associates with N-cadherin and may function as a regulatory switch controlling cadherin function by dephosphorylating beta-catenin, thereby maintaining cells in an adhesion-competent state. The PTP1B-like phosphatase is itself tyrosine phosphorylated. Moreover, both direct binding experiments performed with phosphorylated and dephosphorylated molecules, and treatment of cells with tyrosine kinase inhibitors indicate that the interaction of the PTP1B-like phosphatase with N-cadherin depends on its tyrosine phosphorylation. Concomitant with the tyrosine kinase inhibitor-induced loss of the PTP1B-like phosphatase from its association with N-cadherin, phosphorylated tyrosine residues are retained on beta-catenin, the association of N- cadherin with the actin containing cytoskeleton is lost and N-cadherin- mediated cell adhesion is prevented. Tyrosine phosphatase inhibitors also result in the accumulation of phosphorylated tyrosine residues on beta-catenin, loss of the association of N-cadherin with the actin- containing cytoskeleton, and prevent N-cadherin mediated adhesion, presumably by directly blocking the function of the PTP1B-like phosphatase. We previously showed that the binding of two ligands to the cell surface N-acetylgalactosaminylphosphotransferase (GalNAcPTase), the monoclonal antibody 1B11 and a proteoglycan with a 250-kD core protein

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.

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

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.

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.

Cloning and Characterization of Purple Acid Phosphatase Phytases from Wheat, Barley, Maize, and Rice[W][OA

PubMed Central

Dionisio, Giuseppe; Madsen, Claus K.; Holm, Preben B.; Welinder, Karen G.; Jørgensen, Malene; Stoger, Eva; Arcalis, Elsa; Brinch-Pedersen, Henrik

2011-01-01

Barley (Hordeum vulgare) and wheat (Triticum aestivum) possess significant phytase activity in the mature grains. Maize (Zea mays) and rice (Oryza sativa) possess little or virtually no preformed phytase activity in the mature grain and depend fully on de novo synthesis during germination. Here, it is demonstrated that wheat, barley, maize, and rice all possess purple acid phosphatase (PAP) genes that, expressed in Pichia pastoris, give fully functional phytases (PAPhys) with very similar enzyme kinetics. Preformed wheat PAPhy was localized to the protein crystalloid of the aleurone vacuole. Phylogenetic analyses indicated that PAPhys possess four conserved domains unique to the PAPhys. In barley and wheat, the PAPhy genes can be grouped as PAPhy_a or PAPhy_b isogenes (barley, HvPAPhy_a, HvPAPhy_b1, and HvPAPhy_b2; wheat, TaPAPhy_a1, TaPAPhy_a2, TaPAPhy_b1, and TaPAPhy_b2). In rice and maize, only the b type (OsPAPhy_b and ZmPAPhy_b, respectively) were identified. HvPAPhy_a and HvPAPhy_b1/b2 share 86% and TaPAPhya1/a2 and TaPAPhyb1/b2 share up to 90% (TaPAPhy_a2 and TaPAPhy_b2) identical amino acid sequences. despite of this, PAPhy_a and PAPhy_b isogenes are differentially expressed during grain development and germination. In wheat, it was demonstrated that a and b isogene expression is driven by different promoters (approximately 31% identity). TaPAPhy_a/b promoter reporter gene expression in transgenic grains and peptide mapping of TaPAPhy purified from wheat bran and germinating grains confirmed that the PAPhy_a isogene set present in wheat/barley but not in rice/maize is the origin of high phytase activity in mature grains. PMID:21220762

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.

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

Inhibition effects of chlorogenic acid on benign prostatic hyperplasia in mice.

PubMed

Huang, Ya; Chen, Huaguo; Zhou, Xin; Wu, Xingdong; Hu, Enming; Jiang, Zhengmeng

2017-08-15

This study aimed to evaluate the inhibitory effects and explore mechanisms of chlorogenic acid against testosterone-induced benign prostatic hyperplasia (BPH) in mice. Benign prostatic hyperplasia model was induced in experimental groups by daily subcutaneous injections of testosterone propionate (7.5mg/kg/d) consecutively for 14 d. A total of 60 mice were randomly divided into six groups: (Group 1) normal control group, (Group 2) benign prostatic hyperplasia model control group, (Group 3) benign prostatic hyperplasia mice treated with finasteride at a dose of 1mg/kg, (Group 4) benign prostatic hyperplasia mice treated with chlorogenic acid at dose levels of 0.8mg/kg (low dose group), (Group 5) benign prostatic hyperplasia mice treated with chlorogenic acid at dose levels of 1.6mg/kg (medium dose group) and (Group 6) benign prostatic hyperplasia mice treated with chlorogenic acid at dose levels of 3.2mg/kg (high dose group). Animals were sacrificed on the scheduled termination, pick out the eyeball to get blood, then prostates were weighed and prostatic index were determined. Then the serum acid phosphatase (ACP), prostatic acid phosphatase (PACP) and typeⅡ5-alpha-reductase (SRD5A2) levels were measured and observed morphological changes of the prostate. Comparing with benign prostatic hyperplasia model group, the high and medium dose of chlorogenic acid could significantly reduce prostate index and levels of acid phosphatase, prostatic acid phosphatase and typeⅡ5-alpha-reductase (P<0.05 or P<0.01). These findings were supported by histopathological observations of prostate tissues. Histopathological examination also indicated that chlorogenic acid treatment at the high and medium doses inhibited testosterone-induced prostatic hyperplasia. The results indicated that chlorogenic acid exhibited restraining effect on benign prostatic hyperplasia model animals, and its mechanism might be related to inhibit typeⅡ5-alpha reductase activity. Copyright © 2017

Pharmacological inhibition of protein tyrosine phosphatase 1B: a promising strategy for the treatment of obesity and type 2 diabetes mellitus.

PubMed

Panzhinskiy, E; Ren, J; Nair, S

2013-01-01

Obesity and metabolic syndrome represent major public health problems, and are the biggest preventable causes of death worldwide. Obesity is the leading risk factor for type 2 diabetes mellitus (T2DM), cardiovascular diseases and non-alcoholic fatty liver disease. Obesity-associated insulin resistance, which is characterized by reduced uptake and utilization of glucose in muscle, adipose and liver tissues, is a key predictor of metabolic syndrome and T2DM. With increasing prevalence of obesity in adults and children, the need to identify and characterize potential targets for treating metabolic syndrome is imminent. Emerging evidence from animal models, clinical studies and cell lines studies suggest that protein tyrosine phosphatase 1B (PTP1B), an enzyme that negatively regulates insulin signaling, is likely to be involved in the pathways leading to insulin resistance. PTP1B is tethered to the cytosolic surface of endoplasmic reticulum (ER), an organelle that is responsible for folding, modification, and trafficking of proteins. Recent evidence links the disruption of ER homeostasis, referred to as ER stress, to the pathogenesis of obesity and T2DM. PTP1B has been recognized as an important player linking ER stress and insulin resistance in obese subjects. This review highlights recent advances in the research related to the role of PTP1B in signal transduction processes implicated in pathophysiology of obesity and type 2 diabetes, and focuses on the potential therapeutic exploitation of PTP1B inhibitors for the management of these conditions.

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.

The M Phase Kinase Greatwall (Gwl) Promotes Inactivation of PP2A/B55δ, a Phosphatase Directed Against CDK Phosphosites

PubMed Central

Castilho, Priscila V.; Williams, Byron C.; Mochida, Satoru; Zhao, Yong

2009-01-01

We have previously shown that Greatwall kinase (Gwl) is required for M phase entry and maintenance in Xenopus egg extracts. Here, we demonstrate that Gwl plays a crucial role in a novel biochemical pathway that inactivates, specifically during M phase, “antimitotic” phosphatases directed against phosphorylations catalyzed by cyclin-dependent kinases (CDKs). A major component of this phosphatase activity is heterotrimeric PP2A containing the B55δ regulatory subunit. Gwl is activated during M phase by Cdk1/cyclin B (MPF), but once activated, Gwl promotes PP2A/B55δ inhibition with no further requirement for MPF. In the absence of Gwl, PP2A/B55δ remains active even when MPF levels are high. The removal of PP2A/B55δ corrects the inability of Gwl-depleted extracts to enter M phase. These findings support the hypothesis that M phase requires not only high levels of MPF function, but also the suppression, through a Gwl-dependent mechanism, of phosphatase(s) that would otherwise remove MPF-driven phosphorylations. PMID:19793917

Relationship between hydroxycinnamic acids and the resistance of apple cultivars to rosy apple aphid.

PubMed

Berrueta, Luis A; Sasía-Arriba, Andrea; Miñarro, Marcos; Antón, María J; Alonso-Salces, Rosa M; Micheletti, Diego; Gallo, Blanca; Dapena, Enrique

2018-09-01

The phenolic profiles of apple cultivars from the SERIDA Asturian cider apple breeding program, including parents and progenies, were determined by ultrahigh-performance liquid chromatography-diode array detector-electrospray ionization-quadrupole time of flight/mass spectrometer in order to study the relationship between phenols and the resistance of apple tree cultivars to rosy apple aphid (RAA). A pattern recognition technique named partial least square discriminant analysis (PLS-DA) was used to classify apple cultivars based on resistance to RAA, resistant and susceptible, reaching scores with accuracy higher than 97% and 91% respectively. Hydroxycinnamic acids, particularly 4-caffeoylquinic acid (4-CQA) and 4-p-coumaroylquinic acid (4-pCoQA), were identified as the major player in RAA resistance by the PLS-DA model. Indeed, the isomerisation 5-CQA → 4-CQA is favoured in resistant cultivars, whereas the isomerisation 5-pCoQA → 4-pCoQA is favoured in susceptible cultivars. As a result, resistant cultivars accumulate higher amounts of 4-CQA than susceptible ones, and the opposite occurs for 4-pCoQA. Also, minor isomerisations of 5-CQA to 1-CQA or 3-CQA show opposite behaviour for resistant and susceptible cultivars. Cultivar resistance to RAA is concluded to be related with the phenylpropanoid pathway, the isomerisation reactions being the key metabolic reaction for a cultivar to be resistant or susceptible to RAA. Copyright © 2018 Elsevier B.V. All rights reserved.

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+.

3-Phenylpropanoic acid-based phosphotyrosine (pTyr) mimetics: hit evolution to a novel orally active protein tyrosine phosphatase 1B (PTP1B) inhibitor.

PubMed

Tang, Yan-Bo; Liu, Jun-Zheng; Zhang, Shu-En; Du, Xin; Nie, Feilin; Tian, Jin-Ying; Ye, Fei; Huang, Kai; Hu, Jin-Ping; Li, Yan; Xiao, Zhiyan

2014-05-01

Protein tyrosine phosphatase 1B (PTP1B) is a promising therapeutic target for type 2 diabetes. Herein, we report the evolution of a previously identified 3-phenylpropanoic acid-based PTP1B inhibitor to an orally active lead compound. A series of 3-phenylpropanoic acid-based PTP1B inhibitors were synthesized, and three of them, 3-(4-(9H-carbazol-9-yl)phenyl)-5-(3,5-di-tert-butyl-4-methoxyphenyl)-5-oxopentanoic acid (9), 3-(4-(9H-carbazol-9-yl)phenyl)-5-(4'-bromo-[1,1'-biphenyl]-4-yl)-5-oxopentanoic acid (10) and 3-(4-(9H-carbazol-9-yl)-2-fluorophenyl)-5-(4-cyclohexylphenyl)-5-oxopentanoic acid (16), showed IC50 values at sub-micromolar level. Further in vivo evaluation indicated the sodium salt of 9 not only exhibited significant insulin-sensitizing and hypoglycemia effects, but also decreased the serum levels of triglyceride and total cholesterol in high-fat-diet-induced insulin resistance model mice. Preliminary in vivo pharmacokinetic studies on the sodium salt of 9 revealed its pharmacokinetic profile after oral administration in rats. These results provide proof-of-concept for the dual effects of PTP1B inhibitors on both glucose and lipid metabolisms. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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

Mutations in the inositol polyphosphate-5-phosphatase E gene link phosphatidyl inositol signaling to the ciliopathies

PubMed Central

Bielas, Stephanie L.; Silhavy, Jennifer L.; Brancati, Francesco; Kisseleva, Marina V.; Al-Gazali, Lihadh; Sztriha, Laszlo; Bayoumi, Riad A.; Zaki, Maha S.; Abdel-Aleem, Alice; Rosti, Ozgur; Kayserili, Hulya; Swistun, Dominika; Scott, Lesley C.; Bertini, Enrico; Boltshauser, Eugen; Fazzi, Elisa; Travaglini, Lorena; Field, Seth J.; Gayral, Stephanie; Jacoby, Monique; Schurmans, Stephane; Dallapiccola, Bruno; Majerus, Philip W.; Valente, Enza Maria; Gleeson, Joseph G.

2009-01-01

Phosphotidylinositol (PtdIns) signaling is tightly regulated, both spatially and temporally, by subcellularly localized PtdIns kinases and phosphatases that dynamically alter downstream signaling events 1. Joubert Syndrome (JS) characterized by a specific midbrain-hindbrain malformation (“molar tooth sign”) and variably associated retinal dystrophy, nephronophthisis, liver fibrosis and polydactyly 2, and is included in the newly emerging group of “ciliopathies”. In patients linking to JBTS1, we identified mutations in the INPP5E gene, encoding inositol polyphosphate-5-phosphatase E, which hydrolyzes the 5-phosphate of PtdIns(3,4,5)P3 and PtdIns(4,5)P2. Mutations clustered in the phosphatase domain and impaired 5-phosphatase activity, resulting in altered cellular PtdIns ratios. INPP5E localized to cilia in major organs affected in JS, and mutations promoted premature destabilization of cilia in response to stimulation. Thus, these data links PtdIns signaling to the primary cilium, a cellular structure that is becoming increasingly appreciated for its role in mediating cell signals and neuronal function. PMID:19668216

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.

Low bone mineral density and vitamin d deficiency correlated with genetics and other bone markers in female Turkish immigrants in Germany.

PubMed

Tastan, Yasemin; Kann, Peter Herbert; Tinneberg, Hans-Rudolf; Hadji, Peyman; Müller-Ladner, Ulf; Lange, Uwe

2016-11-01

Patients with osteoporosis have a low bone mass resulting in an increased risk for bone fractures, morbidity and mortality. One hundred thirty-one female pre-menopausal participants (98 Turkish immigrants living in Germany in comparison with 33 age-matched healthy Germans) were recruited for this study which explored vitamin D deficiency and specific genetic modifications of bone metabolism. The subjects were investigated for their femoral and lumbar bone mineral density (BMD) by dual-energy X-ray absorptiometry (DEXA) of the right total femur and the lumbar spine. Serum levels of osteologic parameters were determined: parathormone (PTH), calcium (Ca), osteocalcin (OC), phosphate (P), alkaline phosphatase (AP), beta-crossLaps (CL), tartrate-resistant acid phosphatase isoform 5b (TRAP5b), and 25-vitamin D 3 (25-OH D 3 ). The Bsml- and Fokl-polymorphisms of the vitamin D receptor (VDR) gene and the collagen type I alpha 1 (COLIA1)-gene polymorphism were also genotyped. An extremely high prevalence of vitamin D deficiency could be found in the immigrant cohort (87.8 %). Osteoporosis but not osteopenia was more prevalent in this group. Among immigrants with osteoporosis, TRAP5b was elevated in 42.9 % and beta-CL in 28.6 %. Only the Fokl FF-genotype of the VDR polymorphism was significantly more prevalent among the Turkish women, Ff-genotyped immigrants showed significantly decreased BMD. A significant correlation between the COLIA1-gene polymorphism and BMD could not be identified in the two groups. Vitamin D deficiency and osteoporosis appear to be dominant and unrecognized problem among female Turkish immigrants in Germany. Therefore, in this population, osteologic parameters and BMD should be routinely analyzed and deficiencies be treated immediately.

Effects of Kalsis, A Dietary Supplement, on Bone Metabolism in the Ovariectomized Rats

PubMed Central

Montero, Mercedes; Díaz-Curiel, Manuel; Guede, David; Caeiro, Jose Ramón; Martín-Fernández, Marta; Rubert, Mercedes; Navarro, Daisy; de la Piedra, Concepción

2012-01-01

We studied the ability of Kalsis, a food supplement that contains selenium, citric acid, and vitamin E, to prevent the effects of ovariectomy on bone loss. Six-month-old, Wistar female rats were studied. Groups (n = 12): SHAM: sham-operated rats; OVX: ovariectomized rats, treated with vehicle; OVX + Kalsis: ovariectomized rats treated with Kalsis (25 mg/kg/day) for 3 months. Bone mineral density (BMD) was determined by DXA in lumbar spine and femur. Computerized microtomography (μCT) in femur and serum osteocalcin (BGP), aminoterminal propeptide of procollagen I (PINP), β-isomer of carboxyterminal telopeptide of collagen I (CTX), and 5b isoenzyme of tartrate-resistant acid phosphatase (TRAP) were performed. Treatment with Kalsis prevented BMD loss in OVX group. μCT showed a decrease in BV/TV, and trabecular number, and an increase in trabecular separation in OVX rats. Kalsis administration attenuated partially bone loss observed by μCT due to ovariectomy. BGP, PINP, and the resorption index (CTX/TRAP) were increased in OVX group. Treatment with Kalsis maintained this increase. The mechanism of action of this supplement is not through a decrease in bone remodelling rate. The antioxidant action of this food supplement, due to the synergism of all its components, as a cause of its beneficial effect is suggested. PMID:23094197

Effects of kalsis, a dietary supplement, on bone metabolism in the ovariectomized rats.

PubMed

Montero, Mercedes; Díaz-Curiel, Manuel; Guede, David; Caeiro, Jose Ramón; Martín-Fernández, Marta; Rubert, Mercedes; Navarro, Daisy; de la Piedra, Concepción

2012-01-01

We studied the ability of Kalsis, a food supplement that contains selenium, citric acid, and vitamin E, to prevent the effects of ovariectomy on bone loss. Six-month-old, Wistar female rats were studied. Groups (n = 12): SHAM: sham-operated rats; OVX: ovariectomized rats, treated with vehicle; OVX + Kalsis: ovariectomized rats treated with Kalsis (25 mg/kg/day) for 3 months. Bone mineral density (BMD) was determined by DXA in lumbar spine and femur. Computerized microtomography (μCT) in femur and serum osteocalcin (BGP), aminoterminal propeptide of procollagen I (PINP), β-isomer of carboxyterminal telopeptide of collagen I (CTX), and 5b isoenzyme of tartrate-resistant acid phosphatase (TRAP) were performed. Treatment with Kalsis prevented BMD loss in OVX group. μCT showed a decrease in BV/TV, and trabecular number, and an increase in trabecular separation in OVX rats. Kalsis administration attenuated partially bone loss observed by μCT due to ovariectomy. BGP, PINP, and the resorption index (CTX/TRAP) were increased in OVX group. Treatment with Kalsis maintained this increase. The mechanism of action of this supplement is not through a decrease in bone remodelling rate. The antioxidant action of this food supplement, due to the synergism of all its components, as a cause of its beneficial effect is suggested.

Prediction of Bacillus weihenstephanensis acid resistance: the use of gene expression patterns to select potential biomarkers.

PubMed

Desriac, N; Postollec, F; Coroller, L; Sohier, D; Abee, T; den Besten, H M W

2013-10-01

Exposure to mild stress conditions can activate stress adaptation mechanisms and provide cross-resistance towards otherwise lethal stresses. In this study, an approach was followed to select molecular biomarkers (quantitative gene expressions) to predict induced acid resistance after exposure to various mild stresses, i.e. exposure to sublethal concentrations of salt, acid and hydrogen peroxide during 5 min to 60 min. Gene expression patterns of unstressed and mildly stressed cells of Bacillus weihenstephanensis were correlated to their acid resistance (3D value) which was estimated after exposure to lethal acid conditions. Among the twenty-nine candidate biomarkers, 12 genes showed expression patterns that were correlated either linearly or non-linearly to acid resistance, while for the 17 other genes the correlation remains to be determined. The selected genes represented two types of biomarkers, (i) four direct biomarker genes (lexA, spxA, narL, bkdR) for which expression patterns upon mild stress treatment were linearly correlated to induced acid resistance; and (ii) nine long-acting biomarker genes (spxA, BcerKBAB4_0325, katA, trxB, codY, lacI, BcerKBAB4_1716, BcerKBAB4_2108, relA) which were transiently up-regulated during mild stress exposure and correlated to increased acid resistance over time. Our results highlight that mild stress induced transcripts can be linearly or non-linearly correlated to induced acid resistance and both approaches can be used to find relevant biomarkers. This quantitative and systematic approach opens avenues to select cellular biomarkers that could be incremented in mathematical models to predict microbial behaviour. Copyright © 2013 Elsevier B.V. All rights reserved.

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

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.

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.

Comparative stability of repackaged metoprolol tartrate tablets.

PubMed

Yang, Yongsheng; Gupta, Abhay; Carlin, Alan S; Faustino, Patrick J; Lyon, Robbe C; Ellison, Christopher D; Rothman, Barry; Khan, Mansoor A

2010-01-29

The stability of metoprolol tartrate tablets packaged in original high density polyethylene containers and repackaged in USP Class A unit-dose blister packs was investigated. Studies were conducted at 25 degrees C/60% relative humidity (RH) for 52 weeks and at 40 degrees C/75% RH for 13 weeks. The potency, dissolution, water content, loss on drying and hardness of the drug products were analyzed. Results indicated no differences in the stability between the tablets in both packages stored under 25 degrees C/60% RH. No difference in potency was found in both packages under either condition. However, a significant weight increase due to moisture uptake was observed for the repackaged tablets stored under 40 degrees C/75% RH. The weight increase was accompanied by a decrease in tablet hardness (6.5-0 kp) and a increase in dissolution rate (51-92%) in 5 min. Near-infrared (NIR) chemical imaging also monitored moisture uptake of the tablet non-invasively through the package. The observed changes in product stability may adversely affect the products bioavailability profile, even though the potency of the active drug remained within USP specification range of 90-110%. Study results suggest product quality can be negatively impacted even when using USP Class A repackaging materials. Published by Elsevier B.V.

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.

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

Alternative translation initiation of Caveolin-2 desensitizes insulin signaling through dephosphorylation of insulin receptor by PTP1B and causes insulin resistance.

PubMed

Kwon, Hayeong; Jang, Donghwan; Choi, Moonjeong; Lee, Jaewoong; Jeong, Kyuho; Pak, Yunbae

2018-06-01

Insulin resistance, defined as attenuated sensitivity responding to insulin, impairs insulin action. Direct causes and molecular mechanisms of insulin resistance have thus far remained elusive. Here we show that alternative translation initiation (ATI) of Caveolin-2 (Cav-2) regulates insulin sensitivity. Cav-2β isoform yielded by ATI desensitizes insulin receptor (IR) via dephosphorylation by protein-tyrosine phosphatase 1B (PTP1B), and subsequent endocytosis and lysosomal degradation of IR, causing insulin resistance. Blockage of Cav-2 ATI protects against insulin resistance by preventing Cav-2β-PTP1B-directed IR desensitization, thereby normalizing insulin sensitivity and glucose uptake. Our findings show that Cav-2β is a negative regulator of IR signaling, and identify a mechanism causing insulin resistance through control of insulin sensitivity via Cav-2 ATI. Copyright © 2018 Elsevier B.V. All rights reserved.

Evolutionary Silence of the Acid Chaperone Protein HdeB in Enterohemorrhagic Escherichia coli O157:H7

PubMed Central

Louie, Jacqueline W.; Fagerquist, Clifton K.; Sultan, Omar; Miller, William G.; Mandrell, Robert E.

2012-01-01

The periplasmic chaperones HdeA and HdeB are known to be important for cell survival at low pH (pH < 3) in Escherichia coli and Shigella spp. Here we investigated the roles of HdeA and HdeB in the survival of various enterohemorrhagic E. coli (EHEC) following exposure to pH 2.0. Similar to K-12 strains, the acid protections conferred by HdeA and HdeB in EHEC O145 were significant: loss of HdeA and HdeB led to over 100- to 1,000-fold reductions in acid survival, depending on the growth condition of prechallenge cells. However, this protection was much less in E. coli O157:H7 strains. Deletion of hdeB did not affect the acid survival of cells, and deletion of hdeA led to less than a 5-fold decrease in survival. Sequence analysis of the hdeAB operon revealed a point mutation at the putative start codon of the hdeB gene in all 26 E. coli O157:H7 strains analyzed, which shifted the ATG start codon to ATA. This mutation correlated with the lack of HdeB in E. coli O157:H7; however, the plasmid-borne O157-hdeB was able to restore partially the acid resistance in an E. coli O145ΔhdeAB mutant, suggesting the potential function of O157-HdeB as an acid chaperone. We conclude that E. coli O157:H7 strains have evolved acid survival strategies independent of the HdeA/B chaperones and are more acid resistant than nonpathogenic K-12 for cells grown under nonfavorable culturing conditions such as in Luria-Bertani no-salt broth at 28°C. These results suggest a divergent evolution of acid resistance mechanisms within E. coli. PMID:22179243

Overexpression of PP2A-C5 that encodes the catalytic subunit 5 of protein phosphatase 2A in Arabidopsis confers better root and shoot development under salt conditions

USDA-ARS?s Scientific Manuscript database

Protein phosphatase 2A (PP2A) is an enzyme consisting of three subunits: a scaffolding A subunit, a regulatory B subunit and a catalytic C subunit. PP2As were shown to play diverse roles in eukaryotes. In this study, the function of the Arabidopsis PP2A-C5 gene that encodes the catalytic subunit 5 o...

Membrane interaction and functional plasticity of inositol polyphosphate 5-phosphatases.

PubMed

Braun, Werner; Schein, Catherine H

2014-05-06

In this issue of Structure, Trésaugues and colleagues determined the interaction of membrane-bound phosphoinositides with three clinically significant human inositol polyphosphate 5-phosphatases (I5Ps). A comparison to the structures determined with soluble substrates revealed differences in the binding mode and suggested how the I5Ps and apurinic endonuclease (APE1) activities evolved from the same metal-binding active center. Copyright © 2014 Elsevier Ltd. All rights reserved.

Relationship between fluoride exposure and osteoclast markers during RANKL-induced osteoclast differentiation.

PubMed

Junrui, Pei; Bingyun, Li; Yanhui, Gao; Xu, Jiaxun; Darko, Gottfried M; Dianjun, Sun

2016-09-01

Skeletal fluorosis is a metabolic bone disease caused by excessive accumulation of fluoride. Although the cause of this disease is known, the mechanism by which fluoride accumulates on the bone has not been clearly defined, thus there are no markers that can be used for screening skeletal fluorosis in epidemiology. In this study, osteoclasts were formed from bone marrow cells of C57BL/6 mice-treated with macrophage colony stimulating factor and receptor activator of nuclear factor kappa-B ligand. The mRNA expression of tartrate-resistant acid phosphatase 5b (TRAP5b), osteoclast-associated receptor (OSCAR), calcitonin receptor (CTR), matrix metalloproteinase 9 (MMP9) and cathepsin K (CK) were detected using real-time PCR (RT-PCR). Results showed that fluoride between 0.5 and 8mg/l had no effect on osteoclast formation. However fluoride at 0.5mg/l level significantly decreased the activity of osteoclast bone resorption. Fluoride concentration was negatively correlated with the activity of osteoclast bone resorption. On day 5 of osteoclast differentiation maturity, MMP9 and CK mRNA expression were not only negatively correlated with fluoride concentration, but directly correlated with the activity of osteoclast bone resorption. TRAP5b, CTR and OSCAR mRNA expression were positively correlated with the number of osteoclast and they had no correlation with the activity of osteoclast bone resorption. Thus, it can be seen that MMP9 and CK may reflect the change of activity of bone resorption as well the degree of fluoride exposure. TRAP5b, CTR and OSCAR can represent the change of number of osteoclast formed. Copyright © 2016 Elsevier B.V. All rights reserved.

A novel function of twins, B subunit of protein phosphatase 2A, in regulating actin polymerization.

PubMed

Yeh, Po-An; Chang, Ching-Jin

2017-01-01

Actin is an important component of the cytoskeleton and its polymerization is delicately regulated by several kinases and phosphatases. Heterotrimeric protein phosphatase 2A (PP2A) is a potent phosphatase that is crucial for cell proliferation, apoptosis, tumorigenesis, signal transduction, cytoskeleton arrangement, and neurodegeneration. To facilitate these varied functions, different regulators determine the different targets of PP2A. Among these regulators of PP2A, the B subunits in particular may be involved in cytoskeleton arrangement. However, little is known about the role of PP2A in actin polymerization in vivo. Using sophisticated fly genetics, we demonstrated a novel function for the fly B subunit, twins, to promote actin polymerization in varied tissue types, suggesting a broad and conserved effect. Furthermore, our genetic data suggest that twins may act upstream of the actin-polymerized-proteins, Moesin and Myosin-light-chain, and downstream of Rho to promote actin polymerization. This work opens a new avenue for exploring the biological functions of a PP2A regulator, twins, in cytoskeleton regulation.

A novel function of twins, B subunit of protein phosphatase 2A, in regulating actin polymerization

PubMed Central

Chang, Ching-Jin

2017-01-01

Actin is an important component of the cytoskeleton and its polymerization is delicately regulated by several kinases and phosphatases. Heterotrimeric protein phosphatase 2A (PP2A) is a potent phosphatase that is crucial for cell proliferation, apoptosis, tumorigenesis, signal transduction, cytoskeleton arrangement, and neurodegeneration. To facilitate these varied functions, different regulators determine the different targets of PP2A. Among these regulators of PP2A, the B subunits in particular may be involved in cytoskeleton arrangement. However, little is known about the role of PP2A in actin polymerization in vivo. Using sophisticated fly genetics, we demonstrated a novel function for the fly B subunit, twins, to promote actin polymerization in varied tissue types, suggesting a broad and conserved effect. Furthermore, our genetic data suggest that twins may act upstream of the actin-polymerized-proteins, Moesin and Myosin-light-chain, and downstream of Rho to promote actin polymerization. This work opens a new avenue for exploring the biological functions of a PP2A regulator, twins, in cytoskeleton regulation. PMID:28977036

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.