Aniline-induced nitrosative stress in rat spleen: Proteomic identification of nitrated proteins

International Nuclear Information System (INIS)

Fan Xiuzhen; Wang Jianling; Soman, Kizhake V.; Ansari, G.A.S.; Khan, M. Firoze

2011-01-01

Aniline exposure is associated with toxicity to the spleen which is characterized by splenomegaly, hyperplasia, fibrosis, and a variety of sarcomas on chronic exposure in rats. However, mechanisms by which aniline elicits splenotoxic responses are not well understood. Earlier we have shown that aniline exposure leads to increased nitration of proteins in the spleen. However, nitrated proteins remain to be characterized. Therefore, in the current study using proteomic approaches, we focused on characterizing the nitrated proteins in the spleen of aniline-exposed rats. Aniline exposure led to increased tyrosine nitration of proteins, as determined by 2D Western blotting with anti-3-nitrotyrosine specific antibody, compared to the controls. The analyzed nitrated proteins were found in the molecular weight range of 27.7 to 123.6 kDa. A total of 37 nitrated proteins were identified in aniline-treated and control spleens. Among them, 25 were found only in aniline-treated rats, 11 were present in both aniline-treated and control rats, while one was found in controls only. The nitrated proteins identified mainly represent skeletal proteins, chaperones, ferric iron transporter, enzymes, nucleic acids binding protein, and signaling and protein synthesis pathways. Furthermore, aniline exposure led to significantly increased iNOS mRNA and protein expression in the spleen, suggesting its role in increased reactive nitrogen species formation and contribution to increased nitrated proteins. The identified nitrated proteins provide a global map to further investigate alterations in their structural and functional properties, which will lead to a better understanding of the role of protein nitration in aniline-mediated splenic toxicity. - Highlights: → Proteomic approaches are used to identify nitrated proteins in the spleen. → Twenty five nitrated proteins were found only in the spleen of aniline-treated rats. → Aniline exposure led to increased iNOS mRNA and protein

Atomic structure of nitrate-binding protein crucial for photosynthetic productivity

Energy Technology Data Exchange (ETDEWEB)

Koropatkin, Nicole M.; Pakrasi, Himadri B.; Smith, Thomas J.

2006-06-27

Cyanobacteria, blue-green algae, are the most abundant autotrophs in aquatic environments and form the base of all aquatic food chains by fixing carbon and nitrogen into cellular biomass. The single most important nutrient for photosynthesis and growth is nitrate, which is severely limiting in many aquatic environments particularly the open ocean (1, 2). It is therefore not surprising that NrtA, the solute-binding component of the high-affinity nitrate ABC transporter, is the single-most abundant protein in the plasma membrane of these bacteria (3). Here we describe the first structure of a nitratespecific receptor, NrtA from Synechocystis sp. PCC 6803, complexed with nitrate and determined to a resolution of 1.5Å. NrtA is significantly larger than other oxyanionbinding proteins, representing a new class of transport proteins. From sequence alignments, the only other solute-binding protein in this class is CmpA, a bicarbonatebinding protein. Therefore, these organisms created a novel solute-binding protein for two of the most important nutrients; inorganic nitrogen and carbon. The electrostatic charge distribution of NrtA appears to force the protein off of the membrane while the flexible tether facilitates the delivery of nitrate to the membrane pore. The structure not only details the determinants for nitrate selectivity in NrtA, but also the bicarbonate specificity in CmpA. Nitrate and bicarbonate transport are regulated by the cytoplasmic proteins NrtC and CmpC, respectively. Interestingly, the residues lining the ligand binding pockets suggest that they both bind nitrate. This implies that the nitrogen and carbon uptake pathways are synchronized by intracellular nitrate and nitrite.3 The nitrate ABC transporter of cyanobacteria is composed of four polypeptides (Figure 1): a high-affinity periplasmic solute-binding lipoprotein (NrtA), an integral membrane permease (NrtB), a cytoplasmic ATPase (NrtD), and a unique ATPase/solute-binding fusion protein (Nrt

Induction of the Nitrate Assimilation nirA Operon and Protein-Protein Interactions in the Maturation of Nitrate and Nitrite Reductases in the Cyanobacterium Anabaena sp. Strain PCC 7120.

Science.gov (United States)

Frías, José E; Flores, Enrique

2015-07-01

Nitrate is widely used as a nitrogen source by cyanobacteria, in which the nitrate assimilation structural genes frequently constitute the so-called nirA operon. This operon contains the genes encoding nitrite reductase (nirA), a nitrate/nitrite transporter (frequently an ABC-type transporter; nrtABCD), and nitrate reductase (narB). In the model filamentous cyanobacterium Anabaena sp. strain PCC 7120, which can fix N2 in specialized cells termed heterocysts, the nirA operon is expressed at high levels only in media containing nitrate or nitrite and lacking ammonium, a preferred nitrogen source. Here we examined the genes downstream of the nirA operon in Anabaena and found that a small open reading frame of unknown function, alr0613, can be cotranscribed with the operon. The next gene in the genome, alr0614 (narM), showed an expression pattern similar to that of the nirA operon, implying correlated expression of narM and the operon. A mutant of narM with an insertion mutation failed to produce nitrate reductase activity, consistent with the idea that NarM is required for the maturation of NarB. Both narM and narB mutants were impaired in the nitrate-dependent induction of the nirA operon, suggesting that nitrite is an inducer of the operon in Anabaena. It has previously been shown that the nitrite reductase protein NirA requires NirB, a protein likely involved in protein-protein interactions, to attain maximum activity. Bacterial two-hybrid analysis confirmed possible NirA-NirB and NarB-NarM interactions, suggesting that the development of both nitrite reductase and nitrate reductase activities in cyanobacteria involves physical interaction of the corresponding enzymes with their cognate partners, NirB and NarM, respectively. Nitrate is an important source of nitrogen for many microorganisms that is utilized through the nitrate assimilation system, which includes nitrate/nitrite membrane transporters and the nitrate and nitrite reductases. Many cyanobacteria

Protein Tyrosine Nitration: Biochemical Mechanisms and Structural Basis of its Functional Effects

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Radi, Rafael

2012-01-01

CONSPECTUS The nitration of protein tyrosine residues to 3-nitrotyrosine represents an oxidative postranslational modification that unveils the disruption of nitric oxide (•NO) signaling and metabolism towards pro-oxidant processes. Indeed, excess levels of reactive oxygen species in the presence of •NO or •NO-derived metabolites lead to the formation of nitrating species such as peroxynitrite. Thus, protein 3-nitrotyrosine has been established as a biomarker of cell, tissue and systemic “nitroxidative stress”. Moreover, tyrosine nitration modifies key properties of the amino acid (i.e. phenol group pKa, redox potential, hydrophobicity and volume). Thus, the incorporation of a nitro group (−NO2) to protein tyrosines can lead to profound structural and functional changes, some of which contribute to altered cell and tissue homeostasis. In this Account, I describe our current efforts to define 1) biologically-relevant mechanisms of protein tyrosine nitration and 2) how this modification can cause changes in protein structure and function at the molecular level. First, the relevance of protein tyrosine nitration via free radical-mediated reactions (in both peroxynitrite-dependent or independent pathways) involving the intermediacy of tyrosyl radical (Tyr•) will be underscored. This feature of the nitration process becomes critical as Tyr• can take variable fates, including the formation of 3-nitrotyrosine. Fast kinetic techniques, electron paramagnetic resonance (EPR) studies, bioanalytical methods and kinetic simulations have altogether assisted to characterize and fingerprint the reactions of tyrosine with peroxynitrite and one-electron oxidants and its further evolution to 3-nitrotyrosine. Recent findings show that nitration of tyrosines in proteins associated to biomembranes is linked to the lipid peroxidation process via a connecting reaction that involves the one-electron oxidation of tyrosine by lipid peroxyl radicals (LOO•). Second

A reassessment of synchronous fluorescence in the separation of Trp and Tyr contributions in protein emission and in the determination of conformational changes

DEFF Research Database (Denmark)

Bobone, Sara; van de Weert, Marco; Stella, Lorenzo

2014-01-01

solvents, as well as a real protein (bovine serum albumin). Unfortunately, synchronous spectra were found to be unreliable in the separation of Trp and Tyr emission components in proteins. A simple alternative approach based on the deconvolution of emission spectra is presented. In addition, an equation...

Differential nitrate accumulation, nitrate reduction, nitrate reductase ...

African Journals Online (AJOL)

However, the effects of potassium nitrate were higher than sodium nitrate, which was due to the positive effects of potassium on the enzyme activity, sugars transport, water and nutrient transport, protein synthesis and carbohydrate metabolism. In conclusion, potassium nitrate has better effect on the nitrate assimilatory ...

Fundamentals on the biochemistry of peroxynitrite and protein tyrosine nitration

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Silvina Bartesaghi

2018-04-01

Full Text Available In this review we provide an analysis of the biochemistry of peroxynitrite and tyrosine nitration. Peroxynitrite is the product of the diffusion-controlled reaction between superoxide (O2•- and nitric oxide (•NO. This process is in competition with the enzymatic dismutation of O2•- and the diffusion of •NO across cells and tissues and its reaction with molecular targets (e.g. guanylate cyclase. Understanding the kinetics and compartmentalization of the O2•- / •NO interplay is critical to rationalize the shift of •NO from a physiological mediator to a cytotoxic intermediate. Once formed, peroxynitrite (ONOO- and ONOOH; pKa = 6,8 behaves as a strong one and two-electron oxidant towards a series of biomolecules including transition metal centers and thiols. In addition, peroxynitrite anion can secondarily evolve to secondary radicals either via its fast reaction with CO2 or through proton-catalyzed homolysis. Thus, peroxynitrite can participate in direct (bimolecular and indirect (through secondary radical intermediates oxidation reactions; through these processes peroxynitrite can participate as cytotoxic effector molecule against invading pathogens and/or as an endogenous pathogenic mediator. Peroxynitrite can cause protein tyrosine nitration in vitro and in vivo. Indeed, tyrosine nitration is a hallmark of the reactions of •NO-derived oxidants in cells and tissues and serves as a biomarker of oxidative damage. Protein tyrosine nitration can mediate changes in protein structure and function that affect cell homeostasis. Tyrosine nitration in biological systems is a free radical process that can be promoted either by peroxynitrite-derived radicals or by other related •NO-dependent oxidative processes. Recently, mechanisms responsible of tyrosine nitration in hydrophobic biostructures such as membranes and lipoproteins have been assessed and involve the parallel occurrence and connection with lipid

Hemin-Graphene Derivatives with Increased Peroxidase Activities Restrain Protein Tyrosine Nitration.

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Xu, Huan; Yang, Zhen; Li, Hailing; Gao, Zhonghong

2017-12-14

Protein tyrosine nitration is implicated in the occurrence and progression of pathological conditions involving free radical reactions. It is well recognized that hemin can catalyze protein tyrosine nitration in the presence of nitrite and hydrogen peroxide. Generally, the catalytic efficiency is positively correlated to its peroxidase activity. In this study, however, it is found that the efficiency of hemin in catalyzing protein tyrosine nitration is largely suppressed after functionalization with graphene derivatives, even though its peroxidase-like activity is more than quadrupled. Further studies show that the oxidation of tyrosine is still observed for these composites; dityrosine formation, however, is greatly inhibited. Furthermore, these composites also exhibit strong effects on the oxidation of nitrite into nitrate. Therefore, we propose a mechanism in which hemin-graphene derivatives facilitate the oxidation of tyrosine and nitrite to produce tyrosyl radicals and nitrogen dioxide radicals in the presence of hydrogen peroxide, but graphene interlayers serve as barriers that hinder radical-radical coupling reactions; consequently, protein tyrosine nitration is restrained. This property of hemin-graphene derivatives, by which they catalyze substrate oxidation but suppress radical-radical coupling reactions, shows their great potential in selective oxidation procedures for byproduct removal. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Electrophoretic Detection and Confocal Microscopic Imaging of Tyrosine Nitrated Proteins in Plant Tissue.

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Arora, Dhara; Singh, Neha; Bhatla, Satish C

2018-01-01

Tyrosine nitrated proteins can be detected in plant cells electrophoretically and their distribution can be monitored by confocal laser scanning microscopy (CLSM) imaging. One-dimensional polyacrylamide gel electrophoresis (1D PAGE) followed by Western blotting using polyclonal antibody against 3-nitrotyrosine residues enables detection of tyrosine nitrated proteins in plant cells. Here we describe detection of tyrosine nitrated proteins in the homogenates derived from sunflower (Helianthus annuus L.) seedling cotyledons. Total soluble proteins obtained from tissue homogenates are resolved using vertical gel electrophoresis followed by their electrophoretic transfer on to a microporous membrane support for immunodetection. Spatial distribution of tyrosine nitrated proteins can be visualized using an antibody against 3-nitrotyrosine residues. Immunofluorescent localization is performed by cutting 7 μm thick wax sections of tissue followed by incubation in primary anti-nitrotyrosine antibody (dilution 1:200) and secondary Cy-3 labeled anti-rabbit IgG antibody (dilution 1:1500). Confocal laser scanning microscopy analysis is undertaken using argon lasers (ex: 530-550 nm and em: 570 nm) at pinhole 1. Modulation in the abundance and spatial localization of tyrosine nitrated proteins in plant tissues can be monitored using these techniques.

Protein-tyrosine phosphatase SHP2 contributes to GDNF neurotrophic activity through direct binding to phospho-Tyr687 in the RET receptor tyrosine kinase.

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Perrinjaquet, Maurice; Vilar, Marçal; Ibáñez, Carlos F

2010-10-08

The signaling mechanisms by which neurotrophic receptors regulate neuronal survival and axonal growth are still incompletely understood. In the receptor tyrosine kinase RET, a receptor for GDNF (glial cell line-derived neurotrophic factor), the functions of the majority of tyrosine residues that become phosphorylated are still unknown. Here we have identified the protein-tyrosine phosphatase SHP2 as a novel direct interactor of RET and the first effector known to bind to phosphorylated Tyr(687) in the juxtamembrane region of the receptor. We show that SHP2 is recruited to RET upon ligand binding in a cooperative fashion, such that both interaction with Tyr(687) and association with components of the Tyr(1062) signaling complex are required for stable recruitment of SHP2 to the receptor. SHP2 recruitment contributes to the ability of RET to activate the PI3K/AKT pathway and promote survival and neurite outgrowth in primary neurons. Furthermore, we find that activation of protein kinase A (PKA) by forskolin reduces the recruitment of SHP2 to RET and negatively affects ligand-mediated neurite outgrowth. In agreement with this, mutation of Ser(696), a known PKA phosphorylation site in RET, enhances SHP2 binding to the receptor and eliminates the effect of forskolin on ligand-induced outgrowth. Together, these findings establish SHP2 as a novel positive regulator of the neurotrophic activities of RET and reveal Tyr(687) as a critical platform for integration of RET and PKA signals. We anticipate that several other phosphotyrosines of unknown function in neuronal receptor tyrosine kinases will also support similar regulatory functions.

The protonation state around TyrD/TyrD• in photosystem II is reflected in its biphasic oxidation kinetics.

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Sjöholm, Johannes; Ho, Felix; Ahmadova, Nigar; Brinkert, Katharina; Hammarström, Leif; Mamedov, Fikret; Styring, Stenbjörn

2017-02-01

The tyrosine residue D2-Tyr160 (Tyr D ) in photosystem II (PSII) can be oxidized through charge equilibrium with the oxygen evolving complex in PSII. The kinetics of the electron transfer from Tyr D has been followed using time-resolved EPR spectroscopy after triggering the oxidation of pre-reduced Tyr D by a short laser flash. After its oxidation Tyr D is observed as a neutral radical (Tyr D • ) indicating that the oxidation is coupled to a deprotonation event. The redox state of Tyr D was reported to be determined by the two water positions identified in the crystal structure of PSII [Saito et al. (2013) Proc. Natl. Acad. Sci. USA 110, 7690]. To assess the mechanism of the proton coupled electron transfer of Tyr D the oxidation kinetics has been followed in the presence of deuterated buffers, thereby resolving the kinetic isotope effect (KIE) of Tyr D oxidation at different H/D concentrations. Two kinetic phases of Tyr D oxidation - the fast phase (msec-sec time range) and the slow phase (tens of seconds time range) were resolved as was previously reported [Vass and Styring (1991) Biochemistry 30, 830]. In the presence of deuterated buffers the kinetics was significantly slower compared to normal buffers. Furthermore, although the kinetics were faster at both high pH and pD values the observed KIE was found to be similar (~2.4) over the whole pL range investigated. We assign the fast and slow oxidation phases to two populations of PSII centers with different water positions, proximal and distal respectively, and discuss possible deprotonation events in the vicinity of Tyr D . Copyright © 2016 Elsevier B.V. All rights reserved.

Protein-tyrosine Phosphatase SHP2 Contributes to GDNF Neurotrophic Activity through Direct Binding to Phospho-Tyr687 in the RET Receptor Tyrosine Kinase*

Science.gov (United States)

Perrinjaquet, Maurice; Vilar, Marçal; Ibáñez, Carlos F.

2010-01-01

The signaling mechanisms by which neurotrophic receptors regulate neuronal survival and axonal growth are still incompletely understood. In the receptor tyrosine kinase RET, a receptor for GDNF (glial cell line-derived neurotrophic factor), the functions of the majority of tyrosine residues that become phosphorylated are still unknown. Here we have identified the protein-tyrosine phosphatase SHP2 as a novel direct interactor of RET and the first effector known to bind to phosphorylated Tyr687 in the juxtamembrane region of the receptor. We show that SHP2 is recruited to RET upon ligand binding in a cooperative fashion, such that both interaction with Tyr687 and association with components of the Tyr1062 signaling complex are required for stable recruitment of SHP2 to the receptor. SHP2 recruitment contributes to the ability of RET to activate the PI3K/AKT pathway and promote survival and neurite outgrowth in primary neurons. Furthermore, we find that activation of protein kinase A (PKA) by forskolin reduces the recruitment of SHP2 to RET and negatively affects ligand-mediated neurite outgrowth. In agreement with this, mutation of Ser696, a known PKA phosphorylation site in RET, enhances SHP2 binding to the receptor and eliminates the effect of forskolin on ligand-induced outgrowth. Together, these findings establish SHP2 as a novel positive regulator of the neurotrophic activities of RET and reveal Tyr687 as a critical platform for integration of RET and PKA signals. We anticipate that several other phosphotyrosines of unknown function in neuronal receptor tyrosine kinases will also support similar regulatory functions. PMID:20682772

Nitrate-induced changes in protein synthesis and translation of RNA in maize roots

International Nuclear Information System (INIS)

McClure, P.R.; Omholt, T.E.; Pace, G.M.; Bouthyette, P.Y.

1987-01-01

Nitrate regulation of protein synthesis and RNA translation in maize (Zea mays L. var B73) roots was examined, using in vivo labeling with [ 35 S]methionine and in vitro translation. Nitrate enhanced the synthesis of a 31 kilodalton membrane polypeptide which was localized in a fraction enriched in tonoplast and/or endoplasmic reticulum membrane vesicles. The nitrate-enhanced synthesis was correlated with an acceleration of net nitrate uptake by seedlings during initial exposure to nitrate. Nitrate did not consistently enhance protein synthesis in other membrane fractions. Synthesis of up to four soluble polypeptides (21, 40, 90, and 168 kilodaltons) was also enhanced by nitrate. The most consistent enhancement was that of the 40 kilodalton polypeptide. No consistent nitrate-induced changes were noted in the organellar fraction (14,000g pellet of root homogenates). When roots were treated with nitrate, the amount of [ 35 S]methionine increased in six in vitro translation products (21, 24, 41, 56, 66, and 90 kilodaltons). Nitrate treatment did not enhance accumulation of label in translation products with a molecular weight of 31,000 (corresponding to the identified nitrate-inducible membrane polypeptide). Incubation of in vitro translation products with root membranes caused changes in the SDS-PAGE profiles in the vecinity of 31 kilodaltons. The results suggest that the nitrate-inducible, 31 kilodalton polypeptide from a fraction enriched in tonoplast and/or endoplasmic reticulum may be involved in regulating nitrate accumulation by maize roots

PTP1B Regulates Cortactin Tyrosine Phosphorylation by Targeting Tyr446*S⃞

Science.gov (United States)

Stuible, Matthew; Dubé, Nadia; Tremblay, Michel L.

2008-01-01

The emergence of protein-tyrosine phosphatase 1B (PTP1B) as a potential drug target for treatment of diabetes, obesity, and cancer underlies the importance of understanding its full range of cellular functions. Here, we have identified cortactin, a central regulator of actin cytoskeletal dynamics, as a substrate of PTP1B. A trapping mutant of PTP1B binds cortactin at the phosphorylation site Tyr446, the regulation and function of which have not previously been characterized. We show that phosphorylation of cortactin Tyr446 is induced by hyperosmolarity and potentiates apoptotic signaling during prolonged hyperosmotic stress. This study advances the importance of Tyr446 in the regulation of cortactin and provides a potential mechanism to explain the effects of PTP1B on processes including cell adhesion, migration, and tumorigenesis. PMID:18387954

Impaired mitochondrial respiration and protein nitration in the rat hippocampus after acute inhalation of combustion smoke

International Nuclear Information System (INIS)

Lee, Heung M.; Reed, Jason; Greeley, George H.; Englander, Ella W.

2009-01-01

Survivors of massive inhalation of combustion smoke endure critical injuries, including lasting neurological complications. We have previously reported that acute inhalation of combustion smoke disrupts the nitric oxide homeostasis in the rat brain. In this study, we extend our findings and report that a 30-minute exposure of awake rats to ambient wood combustion smoke induces protein nitration in the rat hippocampus and that mitochondrial proteins are a sensitive nitration target in this setting. Mitochondria are central to energy metabolism and cellular signaling and are critical to proper cell function. Here, analyses of the mitochondrial proteome showed elevated protein nitration in the course of a 24-hour recovery following exposure to smoke. Mass spectrometry identification of several significantly nitrated mitochondrial proteins revealed diverse functions and involvement in central aspects of mitochondrial physiology. The nitrated proteins include the ubiquitous mitochondrial creatine kinase, F1-ATP synthase α subunit, dihydrolipoamide dehydrogenase (E3), succinate dehydrogenase Fp subunit, and voltage-dependent anion channel (VDAC1) protein. Furthermore, acute exposure to combustion smoke significantly compromised the respiratory capacity of hippocampal mitochondria. Importantly, elevated protein nitration and reduced mitochondrial respiration in the hippocampus persisted beyond the time required for restoration of normal oxygen and carboxyhemoglobin blood levels after the cessation of exposure to smoke. Thus, the time frame for intensification of the various smoke-induced effects differs between blood and brain tissues. Taken together, our findings suggest that nitration of essential mitochondrial proteins may contribute to the reduction in mitochondrial respiratory capacity and underlie, in part, the brain pathophysiology after acute inhalation of combustion smoke

Nitration of soluble proteins in organotypic culture models of Parkinson's disease

DEFF Research Database (Denmark)

Larsen, Trine R; Söderling, Ann-Sofi; Caidahl, Kenneth

2008-01-01

Protein nitration due to oxidative and nitrative stress has been linked to the pathogenesis of Parkinson's disease (PD), but its relationship to the loss of dopamine (DA) or tyrosine hydroxylase (TH) activity is not clear. Here we quantified protein-bound 3-nitrotyrosine (3-NT) by a novel gas...... chromatography/negative chemical ionization tandem mass spectrometry technique and DA and 3,4-dihydroxyphenylalanine (DOPA) by HPLC in tissues or medium of organotypic, mouse mesencephalon cultures after acute or chronic treatments with the peroxynitrite donor 3-morpholino-sydnonimine (SIN-1), the dopaminergic...

Exposure of tropoelastin to peroxynitrous acid gives high yields of nitrated tyrosine residues, di-tyrosine cross-links and altered protein structure and function

DEFF Research Database (Denmark)

Degendorfer, Georg; Chuang, Christine Yu-Nung; Mariotti, Michele

2018-01-01

Elastin is an abundant extracellular matrix protein in elastic tissues, including the lungs, skin and arteries, and comprises 30–57% of the aorta by dry mass. The monomeric precursor, tropoelastin (TE), undergoes complex processing during elastogenesis to form mature elastic fibres. Peroxynitrous......-protein di-tyrosine cross-links have been characterized by mass spectrometry. Examination of human atherosclerotic lesions shows colocalization of 3-nitroTyr with elastin epitopes, consistent with TE or elastin modification in vivo, and also an association of 3-nitroTyr containing proteins and elastin...

The structure of Haemophilus influenzae prephenate dehydrogenase suggests unique features of bifunctional TyrA enzymes

International Nuclear Information System (INIS)

Chiu, Hsiu-Ju; Abdubek, Polat; Astakhova, Tamara; Axelrod, Herbert L.; Carlton, Dennis; Clayton, Thomas; Das, Debanu; Deller, Marc C.; Duan, Lian; Feuerhelm, Julie; Grant, Joanna C.; Grzechnik, Anna; Han, Gye Won; Jaroszewski, Lukasz; Jin, Kevin K.; Klock, Heath E.; Knuth, Mark W.; Kozbial, Piotr; Krishna, S. Sri; Kumar, Abhinav; Marciano, David; McMullan, Daniel; Miller, Mitchell D.; Morse, Andrew T.; Nigoghossian, Edward; Okach, Linda; Reyes, Ron; Tien, Henry J.; Trame, Christine B.; Bedem, Henry van den; Weekes, Dana; Xu, Qingping; Hodgson, Keith O.; Wooley, John; Elsliger, Marc-André; Deacon, Ashley M.; Godzik, Adam; Lesley, Scott A.; Wilson, Ian A.

2010-01-01

The crystal structure of the prephenate dehydrogenase component of the bifunctional H. influenzae TyrA reveals unique structural differences between bifunctional and monofunctional TyrA enzymes. Chorismate mutase/prephenate dehydrogenase from Haemophilus influenzae Rd KW20 is a bifunctional enzyme that catalyzes the rearrangement of chorismate to prephenate and the NAD(P) + -dependent oxidative decarboxylation of prephenate to 4-hydroxyphenylpyruvate in tyrosine biosynthesis. The crystal structure of the prephenate dehydrogenase component (HinfPDH) of the TyrA protein from H. influenzae Rd KW20 in complex with the inhibitor tyrosine and cofactor NAD + has been determined to 2.0 Å resolution. HinfPDH is a dimeric enzyme, with each monomer consisting of an N-terminal α/β dinucleotide-binding domain and a C-terminal α-helical dimerization domain. The structure reveals key active-site residues at the domain interface, including His200, Arg297 and Ser179 that are involved in catalysis and/or ligand binding and are highly conserved in TyrA proteins from all three kingdoms of life. Tyrosine is bound directly at the catalytic site, suggesting that it is a competitive inhibitor of HinfPDH. Comparisons with its structural homologues reveal important differences around the active site, including the absence of an α–β motif in HinfPDH that is present in other TyrA proteins, such as Synechocystis sp. arogenate dehydrogenase. Residues from this motif are involved in discrimination between NADP + and NAD + . The loop between β5 and β6 in the N-terminal domain is much shorter in HinfPDH and an extra helix is present at the C-terminus. Furthermore, HinfPDH adopts a more closed conformation compared with TyrA proteins that do not have tyrosine bound. This conformational change brings the substrate, cofactor and active-site residues into close proximity for catalysis. An ionic network consisting of Arg297 (a key residue for tyrosine binding), a water molecule, Asp206 (from

Nano titanium dioxide photocatalytic protein tyrosine nitration: A potential hazard of TiO2 on skin

International Nuclear Information System (INIS)

Lu, Naihao; Zhu Zhening; Zhao Xuqi; Tao Ran; Yang Xiangliang; Gao Zhonghong

2008-01-01

Protein tyrosine nitration is a prevalent post-translational modification which occurs as a result of oxidative and nitrative stress, it may be directly involved in the onset and/or progression of diseases. Considering the existence of nano titanium dioxide (TiO 2 ) in environment and sunscreen products along with the high content of nitrite in sweat, the UV-exposed skin may be a significant target for the photosensitized damage. In this paper, tyrosine nitration of bovine serum albumin (BSA) was initiated in the UV-irradiated reaction mixture containing 0.2-3.0 mg/ml of three commercially nano TiO 2 products and 0.25-1.0 mM NO 2 - . It was found that anatase TiO 2 and Degussa P25 TiO 2 showed prominent photocatalytic activity on promoting the formation of protein tyrosine nitration, and the optimum condition for the reaction was around physiological pH. Meanwhile, the photocatalytic effect of rutile on protein tyrosine nitration was subtle. The potential physiological significance of nano TiO 2 -photocatalytic protein nitration was also demonstrated in mouse skin homogenate. Although the relationship between photocatalytic protein tyrosine nitration and chronic cutaneous diseases needs further study, the toxicity of nano TiO 2 to the skin disease should be paid more attention in the production and utilization process

Biosynthetically directed fractional 13C labeling facilitates identification of Phe and Tyr aromatic signals in proteins

International Nuclear Information System (INIS)

Jacob, Jaison; Louis, John M.; Nesheiwat, Issa; Torchia, Dennis A.

2002-01-01

Analysis of 2D [ 13 C, 1 H]-HSQC spectra of biosynthetic fractionally 13 C labeled proteins is a reliable, straightforward means to obtain stereospecific assignments of Val and Leu methyl sites in proteins. Herein we show that the same fractionally labeled protein sample facilitates observation and identification of Phe and Tyr aromatic signals. This is the case, in part, because the fractional 13 C labeling yields aromatic rings in which some of the 13 C- 13 C J-couplings, present in uniformly labeled samples, are absent. Also, the number of homonuclear J-coupling partners differs for the δ-, ε- and ζ-carbons. This enabled us to vary their signal intensities in distinctly different ways by appropriately setting the 13 C constant-time period in 2D [ 13 C, 1 H]-HSQC spectra. We illustrate the application of this approach to an 18 kDa protein, c-VIAF, a modulator of apoptosis. In addition, we show that cancellation of the aromatic 13 C CSA and 13 C- 1 H dipolar interactions can be fruitfully utilized in the case of the fractionally labeled sample to obtain high resolution 13 C constant-time spectra with good sensitivity

Post-anthesis nitrate uptake is critical to yield and grain protein content in Sorghum bicolor.

Science.gov (United States)

Worland, Belinda; Robinson, Nicole; Jordan, David; Schmidt, Susanne; Godwin, Ian

2017-09-01

Crops only use ∼50% of applied nitrogen (N) fertilizer creating N losses and pollution. Plants need to efficiently uptake and utilize N to meet growing global food demands. Here we investigate how the supply and timing of nitrate affects N status and yield in Sorghum bicolor (sorghum). Sorghum was grown in pots with either 10mM (High) or 1mM (Low) nitrate supply. Shortly before anthesis the nitrate supply was either maintained, increased 10-fold or eliminated. Leaf sheaths of sorghum grown with High nitrate accumulated nitrate in concentrations >3-times higher than leaves. Removal of nitrate supply pre-anthesis resulted in the rapid reduction of stored nitrate in all organs. Plants receiving a 10-fold increase in nitrate supply pre-anthesis achieved similar grain yield and protein content and 29% larger grains than those maintained on High nitrate, despite receiving 24% less nitrate over the whole growth period. In sorghum, plant available N is important throughout development, particularly anthesis and grain filling, for grain yield and grain protein content. Nitrate accumulation in leaf sheaths presents opportunities for the genetic analysis of mechanisms behind nitrate storage and remobilization in sorghum to improve N use efficiency. Copyright © 2017 Elsevier GmbH. All rights reserved.

Nitrosative stress and nitrated proteins in trichloroethene-mediated autoimmunity.

Directory of Open Access Journals (Sweden)

Gangduo Wang

Full Text Available Exposure to trichloroethene (TCE, a ubiquitous environmental contaminant, has been linked to a variety of autoimmune diseases (ADs including SLE, scleroderma and hepatitis. Mechanisms involved in the pathogenesis of ADs are largely unknown. Earlier studies from our laboratory in MRL+/+ mice suggested the contribution of oxidative/nitrosative stress in TCE-induced autoimmunity, and N-acetylcysteine (NAC supplementation provided protection by attenuating oxidative stress. This study was undertaken to further evaluate the contribution of nitrosative stress in TCE-mediated autoimmunity and to identify proteins susceptible to nitrosative stress. Groups of female MRL +/+ mice were given TCE, NAC or TCE + NAC for 6 weeks (TCE, 10 mmol/kg, i.p., every 4th day; NAC, ∼ 250 mg/kg/day via drinking water. TCE exposure led to significant increases in serum anti-nuclear and anti-histone antibodies together with significant induction of iNOS and increased formation of nitrotyrosine (NT in sera and livers. Proteomic analysis identified 14 additional nitrated proteins in the livers of TCE-treated mice. Furthermore, TCE exposure led to decreased GSH levels and increased activation of NF-κB. Remarkably, NAC supplementation not only ameliorated TCE-induced nitrosative stress as evident from decreased iNOS, NT, nitrated proteins, NF-κB p65 activation and increased GSH levels, but also the markers of autoimmunity, as evident from decreased levels of autoantibodies in the sera. These findings provide support to the role of nitrosative stress in TCE-mediated autoimmune response and identify specific nitrated proteins which could have autoimmune potential. Attenuation of TCE-induced autoimmunity in mice by NAC provides an approach for designing therapeutic strategies.

Regulation by nitrate of protein synthesis and translation of RNA in maize roots

International Nuclear Information System (INIS)

McClure, P.R.; Bouthyette, P.Y.

1986-01-01

Roots of maize seedlings were exposed to 35 S-methionine in the presence or absence of nitrate. Using SDS-PAGE, nitrate-induced changes in labeled polypeptides were noted in the soluble (at 92, 63 and 21kD) and organellar(at 14kD) fractions, as well as in a membrane fraction of putative tonoplast origin (at 31kD). No nitrate-induced changes were noted in a plasmamembrane-enriched fraction or in a membrane fraction of mixed origin. Total RNA from nitrate-treated and control roots was translated in a rabbit reticulocyte system. Five translation products (94, 63, 41, 39 and 21kD) were identified as nitrate-inducible by comparative gel electrophoresis. Changes in protein synthesis and translation of mRNA were apparent within 2-3 h after introduction of nitrate. Within 4-6 h after removal of nitrate, the level of nitrate-inducible translation products diminished to that of control roots. In contrast, the 31kD tonoplast polypeptide was still labeled 26 h after removal of external nitrate and 35 S-methionine. The results will be discussed in relation to the nitrate induction of nitrate reductase, nitrite reductase, and the nitrate uptake system

A novel method for the biosynthesis of deuterated proteins with selective protonation at the aromatic rings of Phe, Tyr and Trp

International Nuclear Information System (INIS)

Rajesh, Sundaresan; Nietlispach, Daniel; Nakayama, Hiroshi; Takio, Koji; Laue, Ernest D.; Shibata, Takehiko; Ito, Yutaka

2003-01-01

A novel biosynthetic strategy is described for the preparation of deuterated proteins containing protons at the ring carbons of Phe, Tyr and Trp, using the aromatic amino acid precursor shikimic acid. Specific protonation at aromatic side chains, with complete deuteration at C α/β positions was achieved in proteins overexpressed in bacteria grown in shikimate-supplemented D 2 O medium. Co-expression of a shikimate transporter in prototrophic bacteria resulted in protonation levels of 62-79%, whereas complete labeling was accomplished using shikimate auxotrophic bacteria. Our labeling protocol permits the measurement of important aromatic side chain derived distance restraints in perdeuterated proteins that could be utilized to enhance the accuracy of NMR structures calculated using low densities of NOEs from methyl selectively protonated samples

Tyr51: Key Determinant of the Low Thermostability of the Colwellia psychrerythraea Cold-Shock Protein.

Science.gov (United States)

Lee, Yeongjoon; Kwak, Chulhee; Jeong, Ki-Woong; Durai, Prasannavenkatesh; Ryu, Kyoung-Seok; Kim, Eun-Hee; Cheong, Chaejoon; Ahn, Hee-Chul; Kim, Hak Jun; Kim, Yangmee

2018-05-18

Cold-shock proteins (Csps) are expressed at lower-than-optimum temperatures, and they function as RNA chaperones; however, no structural studies on psychrophilic Csps have been reported. Here, we aimed to investigate the structure and dynamics of the Csp of psychrophile Colwellia psychrerythraea 34H, ( Cp-Csp). Although Cp-Csp shares sequence homology, common folding patterns, and motifs, including a five β-stranded barrel, with its thermophilic counterparts, its thermostability (37 °C) was markedly lower than those of other Csps. Cp-Csp binds heptathymidine with an affinity of 10 -7 M, thereby increasing its thermostability to 50 °C. Nuclear magnetic resonance spectroscopic analysis of the Cp-Csp structure and backbone dynamics revealed a flexible structure with only one salt bridge and 10 residues in the hydrophobic cavity. Notably, Cp-Csp contains Tyr51 instead of the conserved Phe in the hydrophobic core, and its phenolic hydroxyl group projects toward the surface. The Y51F mutation increased the stability of hydrophobic packing and may have allowed for the formation of a K3-E21 salt bridge, thereby increasing its thermostability to 43 °C. Cp-Csp exhibited conformational exchanges in its ribonucleoprotein motifs 1 and 2 (754 and 642 s -1 ), and heptathymidine binding markedly decreased these motions. Cp-Csp lacks salt bridges and has longer flexible loops and a less compact hydrophobic cavity resulting from Tyr51 compared to mesophilic and thermophilic Csps. These might explain the low thermostability of Cp-Csp. The conformational flexibility of Cp-Csp facilitates its accommodation of nucleic acids at low temperatures in polar oceans and its function as an RNA chaperone for cold adaptation.

Cloning and nitrate induction of nitrate reductase mRNA

OpenAIRE

Cheng, Chi-Lien; Dewdney, Julia; Kleinhofs, Andris; Goodman, Howard M.

1986-01-01

Nitrate is the major source of nitrogen taken from the soil by higher plants but requires reduction to ammonia prior to incorporation into amino acids. The first enzyme in the reducing pathway is a nitrate-inducible enzyme, nitrate reductase (EC 1.6.6.1). A specific polyclonal antiserum raised against purified barley nitrate reductase has been used to immunoprecipitate in vivo labeled protein and in vitro translation products, demonstrating that nitrate induction increases nitrate reductase p...

Glucose Elevates NITRATE TRANSPORTER2.1 Protein Levels and Nitrate Transport Activity Independently of Its HEXOKINASE1-Mediated Stimulation of NITRATE TRANSPORTER2.1 Expression1[W][OPEN

Science.gov (United States)

de Jong, Femke; Thodey, Kate; Lejay, Laurence V.; Bevan, Michael W.

2014-01-01

Mineral nutrient uptake and assimilation is closely coordinated with the production of photosynthate to supply nutrients for growth. In Arabidopsis (Arabidopsis thaliana), nitrate uptake from the soil is mediated by genes encoding high- and low-affinity transporters that are transcriptionally regulated by both nitrate and photosynthate availability. In this study, we have studied the interactions of nitrate and glucose (Glc) on gene expression, nitrate transport, and growth using glucose-insensitive2-1 (gin2-1), which is defective in sugar responses. We confirm and extend previous work by showing that HEXOKINASE1-mediated oxidative pentose phosphate pathway (OPPP) metabolism is required for Glc-mediated NITRATE TRANSPORTER2.1 (NRT2.1) expression. Treatment with pyruvate and shikimate, two products derived from intermediates of the OPPP that are destined for amino acid production, restores wild-type levels of NRT2.1 expression, suggesting that metabolites derived from OPPP metabolism can, together with Glc, directly stimulate high levels of NRT2.1 expression. Nitrate-mediated NRT2.1 expression is not influenced by gin2-1, showing that Glc does not influence NRT2.1 expression through nitrate-mediated mechanisms. We also show that Glc stimulates NRT2.1 protein levels and transport activity independently of its HEXOKINASE1-mediated stimulation of NRT2.1 expression, demonstrating another possible posttranscriptional mechanism influencing nitrate uptake. In gin2-1 plants, nitrate-responsive biomass growth was strongly reduced, showing that the supply of OPPP metabolites is essential for assimilating nitrate for growth. PMID:24272701

An adventitious interaction of filamin A with RhoGDI2(Tyr153Glu)

International Nuclear Information System (INIS)

Song, Mia; He, Qianjing; Berk, Benjamin-Andreas; Hartwig, John H.; Stossel, Thomas P.; Nakamura, Fumihiko

2016-01-01

Filamin A (FLNA) is an actin filament crosslinking protein with multiple intracellular binding partners. Mechanical force exposes cryptic FLNA binding sites for some of these ligands. To identify new force-dependent binding interactions, we used a fusion construct composed of two FLNA domains, one of which was previously identified as containing a force-dependent binding site as a bait in a yeast two-hybrid system and identified the Rho dissociation inhibitor 2 (RhoGDI2) as a potential interacting partner. A RhoGDI2 truncate with 81 N-terminal amino acid residues and a phosphomimetic mutant, RhoGDI(Tyr153Glu) interacted with the FLNA construct. However, neither wild-type or full-length RhoGDI2 phosphorylated at Y153 interacted with FLNA. Our interpretation of these contradictions is that truncation and/or mutation of RhoGDI2 perturbs its conformation to expose a site that adventitiously binds FLNA and is not a bona–fide interaction. Therefore, previous studies reporting that a RhoGDI(Y153E) mutant suppresses the metastasis of human bladder cancer cells must be reinvestigated in light of artificial interaction of this point mutant with FLNA. - Highlights: • RhoGDI2 is identified as a potential filamin A (FLNA)-binding partner. • Phosphomimetic mutant, RhoGDI2(Tyr153Glu) interacts with FLNA. • RhoGDI2 phosphorylated (Tyr153) by src kinase does not interact with FLNA. • Mutation of Tyr-153 to Glu of RhoGDI2 does not mimic phosphorylation. • RhoGDI2(Tyr153Glu) provokes an adventitious interaction with FLNA.

An adventitious interaction of filamin A with RhoGDI2(Tyr153Glu)

Energy Technology Data Exchange (ETDEWEB)

Song, Mia; He, Qianjing [Hematology Division, Department of Medicine, Brigham and Women' s Hospital, Harvard Medical School, Boston MA (United States); Berk, Benjamin-Andreas [Faculty of Veterinary Medicine and Faculty of Biosciences and Pharmacy, University of Leipzig, Leipzig (Germany); Hartwig, John H.; Stossel, Thomas P. [Hematology Division, Department of Medicine, Brigham and Women' s Hospital, Harvard Medical School, Boston MA (United States); Nakamura, Fumihiko, E-mail: fnakamura@partners.org [Hematology Division, Department of Medicine, Brigham and Women' s Hospital, Harvard Medical School, Boston MA (United States)

2016-01-15

Filamin A (FLNA) is an actin filament crosslinking protein with multiple intracellular binding partners. Mechanical force exposes cryptic FLNA binding sites for some of these ligands. To identify new force-dependent binding interactions, we used a fusion construct composed of two FLNA domains, one of which was previously identified as containing a force-dependent binding site as a bait in a yeast two-hybrid system and identified the Rho dissociation inhibitor 2 (RhoGDI2) as a potential interacting partner. A RhoGDI2 truncate with 81 N-terminal amino acid residues and a phosphomimetic mutant, RhoGDI(Tyr153Glu) interacted with the FLNA construct. However, neither wild-type or full-length RhoGDI2 phosphorylated at Y153 interacted with FLNA. Our interpretation of these contradictions is that truncation and/or mutation of RhoGDI2 perturbs its conformation to expose a site that adventitiously binds FLNA and is not a bona–fide interaction. Therefore, previous studies reporting that a RhoGDI(Y153E) mutant suppresses the metastasis of human bladder cancer cells must be reinvestigated in light of artificial interaction of this point mutant with FLNA. - Highlights: • RhoGDI2 is identified as a potential filamin A (FLNA)-binding partner. • Phosphomimetic mutant, RhoGDI2(Tyr153Glu) interacts with FLNA. • RhoGDI2 phosphorylated (Tyr153) by src kinase does not interact with FLNA. • Mutation of Tyr-153 to Glu of RhoGDI2 does not mimic phosphorylation. • RhoGDI2(Tyr153Glu) provokes an adventitious interaction with FLNA.

Phosphoproteome analysis of E-coli reveals evolutionary conservation of bacterial Ser/Thr/Tyr phosphorylation

DEFF Research Database (Denmark)

Macek, B.; Gnad, F.; Soufi, Boumediene

2008-01-01

Protein phosphorylation on serine, threonine, and tyrosine (Ser/Thr/Tyr) is generally considered the major regulatory posttranslational modification in eukaryotic cells. Increasing evidence at the genome and proteome level shows that this modification is also present and functional in prokaryotes...

Different zinc sensitivity of Brassica organs is accompanied by distinct responses in protein nitration level and pattern.

Science.gov (United States)

Feigl, Gábor; Kolbert, Zsuzsanna; Lehotai, Nóra; Molnár, Árpád; Ördög, Attila; Bordé, Ádám; Laskay, Gábor; Erdei, László

2016-03-01

Zinc is an essential microelement, but its excess exerts toxic effects in plants. Heavy metal stress can alter the metabolism of reactive oxygen (ROS) and nitrogen species (RNS) leading to oxidative and nitrosative damages; although the participation of these processes in Zn toxicity and tolerance is not yet known. Therefore this study aimed to evaluate the zinc tolerance of Brassica organs and the putative correspondence of it with protein nitration as a relevant marker for nitrosative stress. Both examined Brassica species (B. juncea and B. napus) proved to be moderate Zn accumulators; however B. napus accumulated more from this metal in its organs. The zinc-induced damages (growth diminution, altered morphology, necrosis, chlorosis, and the decrease of photosynthetic activity) were slighter in the shoot system of B. napus than in B. juncea. The relative zinc tolerance of B. napus shoot was accompanied by moderate changes of the nitration pattern. In contrast, the root system of B. napus suffered more severe damages (growth reduction, altered morphology, viability loss) and slighter increase in nitration level compared to B. juncea. Based on these, the organs of Brassica species reacted differentially to excess zinc, since in the shoot system modification of the nitration pattern occurred (with newly appeared nitrated protein bands), while in the roots, a general increment in the nitroproteome could be observed (the intensification of the same protein bands being present in the control samples). It can be assumed that the significant alteration of nitration pattern is coupled with enhanced zinc sensitivity of the Brassica shoot system and the general intensification of protein nitration in the roots is attached to relative zinc endurance. Copyright © 2015 Elsevier Inc. All rights reserved.

Long-term toxicity of [177Lu-DOTA0,Tyr3]octreotate in rats

International Nuclear Information System (INIS)

Rolleman, Edgar J.; Krenning, Eric P.; Bernard, Bert F.; Visser, Monique de; Bijster, Magda; Jong, Marion de; Visser, Theo J.; Vermeij, Marcel; Lindemans, Jan

2007-01-01

Studies on peptide receptor radionuclide therapy (PRRT) using radiolabelled somatostatin analogues have shown promising results with regard to tumour control. The efficacy of PRRT is limited by uptake and retention in the proximal tubules of the kidney, which might lead to radiation nephropathy. We investigated the long-term renal toxicity after different doses of [ 177 Lu-DOTA 0 ,Tyr 3 ]octreotate and the effects of dose fractionation and lysine co-injection in two tumour-bearing rat models. Significant renal toxicity was detected beyond 100 days after start of treatment as shown by elevated serum creatinine and proteinuria. Microscopically, tubules were strongly dilated with flat epithelium, containing protein cylinders. Creatinine levels rose significantly after 555 MBq [ 177 Lu-DOTA 0 ,Tyr 3 ]octreotate, but were significantly lower after 278 MBq (single injection) or two weekly doses of 278 MBq. Renal damage scores were maximal after 555 MBq and significantly lower in the 278 and 2 x 278 MBq groups. Three doses of 185 MBq [ 177 Lu-DOTA 0 ,Tyr 3 ]octreotate with intervals of a day, a week or a month significantly influenced serum creatinine (469±18, 134±70 and 65±15 μmol/l, respectively; p 177 Lu-DOTA 0 ,Tyr 3 ]octreotate resulted in severe renal damage in rats as indicated by proteinuria, elevated serum creatinine and histological damage. This damage was dose dependent and became overt between 100 and 200 days after treatment. Dose fractionation had significant beneficial effects on kidney function. Also, lysine co-injection successfully prevented functional damage. (orig.)

Protein phosphorylation during coconut zygotic embryo development

International Nuclear Information System (INIS)

Islas-Flores, I.; Oropeza, C.; Hernandez-Sotomayor, S.M.T.

1998-01-01

Evidence was obtained on the occurrence of protein threonine, serine, and tyrosine (Tyr) kinases in developing coconut (Cocos nucifera L.) zygotic embryos, based on in vitro phosphorylation of proteins in the presence of [gamma-32P]ATP, alkaline treatment, and thin-layer chromatography analysis, which showed the presence of [32P]phosphoserine, [32P]phosphothreonine, and [32P]phosphotyrosine in [32P]-labeled protein hydrolyzates. Tyr kinase activity was further confirmed in extracts of embryos at different stages of development using antiphosphotyrosine monoclonal antibodies and the synthetic peptide derived from the amino acid sequence surrounding the phosphorylation site in pp60src (RR-SRC), which is specific for Tyr kinases. Anti-phosphotyrosine western blotting revealed a changing profile of Tyr-phosphorylated proteins during embryo development. Tyr kinase activity, as assayed using RR-SRC, also changed during embryo development, showing two peaks of activity, one during early and another during late embryo development. In addition, the use of genistein, a Tyr kinase inhibitor, diminished the ability of extracts to phosphorylate RR-SRC. Results presented here show the occurrence of threonine, serine, and Tyr kinases in developing coconut zygotic embryos, and suggest that protein phosphorylation, and the possible inference of Tyr phosphorylation in particular, may play a role in the coordination of the development of embryos in this species

The levels of nitrite and nitrate, proline and protein profiles in tomato plants infected with pseudomonas syringae

International Nuclear Information System (INIS)

Berber, I.; Onlu, H.

2012-01-01

In this study, the contents of nitrite-nitrate and free L-proline, and pathogenesis-related (PR) proteins in tomato plants following inoculation with Pseudomonas syringae pv. tomato strain were examined. The results of the nitrite and nitrate indicated that there was a reduction in the levels of nitrate in the infected tomato plants through 1-8 study days, compared with the healthy plants. On the other hands, when the nitrite amounts increased in the first and second days, the nitrite concentrations reduced in infected plants at subsequent time periods, compared with uninfected plants. The accumulation of free proline increased in the infected plants, according to control plants. The whole-cell protein profiles displayed that the levels of the protein bands of molecular masses 204.6 kDa and 69.9 kDa significantly increased in infected and uninfected plants during 2-10 study days. In additionally, in the quantities of the protein bands of molecular weights 90.3 and 79.4 kDa were observed an increase in the infected and healthy plants after the fourth day. However, the protein band of molecular weight 54.3 kDa was visible only in uninfected plants for the fourth and eighth days. Finally, the study suggest that there were the sophisticate relationships among the proline accumulation, the conversion of nitrate to nitrite and the induction of PR protein genes in the regulation of defense mechanisms toward microbial invaders. Our results also indicated that the increases in nitrite and proline contents might be useful indicator for the response toward pathogen attacks. (author)

Sensitive radioimmunoassay for somatostatin using N-(/sup 125/I)-tyr-somatostatin as labelled antigen

Energy Technology Data Exchange (ETDEWEB)

Dupont, A [CHUL, Quebec (Canada). Lab. of Molecular Endocrinology and Service of Gastro-Enterology; Coy, D H; Alvarado-Urbina, G; Cote, J; Meyers, C A; McManus, J; Barden, N; De Lean, A; Labrie, F

1979-01-01

A sensitive radioimmunoassay for somatostatin using N-(/sup 125/I)-Tyr-somatostatin is described and compared with using (/sup 125/I)-Tyr/sup 1/-somatostatin. The minimum detectable amount of somatostatin using N-(/sup 125/I)-Tyr-somatostatin as tracer was 0.1 to 0.5 pg, which is approximately 10-fold lower detection limit of the RIA using (/sup 125/I)-Tyr/sup 1/-somatostatin. Moreover, it was found that the shelf-life of N-(/sup 125/I)-Tyr-somatostatin was prolonged in comparison with labelled Tyr/sup 1/-somatostatin. Human pancreatic and gastric extracts displayed immunological similarity to synthetic somatostatin tetradecapeptide.

Nitrate as a probe of cytochrome c surface: crystallographic identification of crucial "hot spots" for protein-protein recognition.

Science.gov (United States)

De March, Matteo; Demitri, Nicola; De Zorzi, Rita; Casini, Angela; Gabbiani, Chiara; Guerri, Annalisa; Messori, Luigi; Geremia, Silvano

2014-06-01

The electrostatic surface of cytochrome c and its changes with the iron oxidation state are involved in the docking and undocking processes of this protein to its biological partners in the mitochondrial respiratory pathway. To investigate the subtle mechanisms of formation of productive macromolecular complexes and of their breakage following the electron transfer process, the X-ray structures of horse heart ferri-cytochrome c (trigonal form) and ferro-cytochrome c (monoclinic form) were obtained using nitrate ions both as a crystallizing agent and an anionic probe for mapping the electrostatic surface changes. Both crystal forms contain three protein molecules in the asymmetric unit. In addition, a total of 21.5 and 18 crystallographically independent nitrate ions were identified for the trigonal and monoclinic forms, respectively. By matching all the six crystallographically independent protein molecules, 26 different anion-protein interaction sites were identified on the surfaces of cytochrome c, 10 of which were found in both forms, 8 present only in the oxidized and 8 only in the reduced form. The structural analysis of the electron transfer complexes, based on this new information, suggests a specific exit strategy for cytochrome c after formation of productive protein-protein complexes: a directional sliding mechanism for the electron shuttle on the surface of the redox partner is proposed to take place after the electron transfer process has occurred. Copyright © 2014 Elsevier Inc. All rights reserved.

Temporal pattern changes in duodenal protein tyrosine nitration events in response to Eimeria acervulina infection in chickens

Science.gov (United States)

Intracellular generation of nitric oxide (NO) and superoxide anion (SOA) can result in the formation of 3'-nitrotyrosine proteins (NTp). Nitrated proteins usually are associated with significant perturbation in protein function, apoptosis, and cell death. We undertook the present study to establis...

Nitrate Activation of Cytosolic Protein Kinases Diverts Photosynthetic Carbon from Sucrose to Amino Acid Biosynthesis

Science.gov (United States)

Champigny, Marie-Louise; Foyer, Christine

1992-01-01

The regulation of carbon partitioning between carbohydrates (principally sucrose) and amino acids has been only poorly characterized in higher plants. The hypothesis that the pathway of sucrose and amino acid biosynthesis compete for carbon skeletons and energy is widely accepted. In this review, we suggest a mechanism involving the regulation of cytosolic protein kinases whereby the flow of carbon is regulated at the level of partitioning between the pathways of carbohydrate and nitrogen metabolism via the covalent modulation of component enzymes. The addition of nitrate to wheat seedlings (Triticum aestivum) grown in the absence of exogenous nitrogen has a dramatic, if transient, impact on sucrose formation and on the activities of sucrose phosphate synthase (which is inactivated) and phosphoenolpyruvate carboxylase (which is activated). The activities of these two enzymes are modulated by protein phosphorylation in response to the addition of nitrate, but they respond in an inverse fashion. Sucrose phosphate synthase in inactivated and phosphoenolpyruvate carboxylase is activated. Nitrate functions as a signal metabolite activating the cytosolic protein kinase, thereby modulating the activities of at least two of the key enzymes in assimilate partitioning and redirecting the flow of carbon away from sucrose biosynthesis toward amino acid synthesis. PMID:16653003

Albinism in the domestic cat (Felis catus) is associated with a tyrosinase (TYR) mutation

Science.gov (United States)

Imes, DL; Geary, LA; Grahn, RA; Lyons, LA

2006-01-01

Summary Albino phenotypes are documented in a variety of species including the domestic cat. As albino phenotypes in other species are associated with tyrosinase (TYR) mutations, TYR was proposed as a candidate gene for albinism in cats. An Oriental and Colourpoint Shorthair cat pedigree segregating for albinism was analysed for association with TYR by linkage and sequence analyses. Microsatellite FCA931, which is closely linked to TYR and TYR sequence variants were tested for segregation with the albinism phenotype. Sequence analysis of genomic DNA from wild-type and albino cats identified a cytosine deletion in TYR at position 975 in exon 2, which causes a frame shift resulting in a premature stop codon nine residues downstream from the mutation. The deletion mutation in TYR and an allele of FCA931 segregated concordantly with the albino phenotype. Taken together, our results suggest that the TYR gene corresponds to the colour locus in cats and its alleles, from dominant to recessive, are as follows: C (full colour) > cb (burmese) ≥ cs (siamese) > c (albino). PMID:16573534

Autoradiographic localization of (125I-Tyr4)bombesin-binding sites in rat brain

International Nuclear Information System (INIS)

Zarbin, M.A.; Kuhar, M.J.; O'Donohue, T.L.; Wolf, S.S.; Moody, T.W.

1985-01-01

The binding of ( 125 I-Tyr 4 )bombesin to rat brain slices was investigated. Radiolabeled (Tyr 4 )bombesin bound with high affinity (K/sub d/ . 4 nM) to a single class of sites (B/sub max/ . 130 fmol/mg of protein); the ratio of specific to nonspecific binding was 6/1. Also, pharmacology studies indicated that the C-terminal of bombesin was important for the high affinity binding activity. Autoradiographic studies indicated that the ( 125 I-Tyr4)bombesin-binding sites were discretely distributed in certain gray but not white matter regions of rat brain. Highest grain densities were present in the olfactory bulb and tubercle, nucleus accumbens, suprachiasmatic and periventricular nuclei of the hypothalamus, central medial thalamic nucleus, medial amygdaloid nucleus, hippocampus, dentate gyrus, subiculum, nucleus of the solitary tract, and substantia gelatinosa. Moderate grain densities were present in the parietal cortex, deep layers of the neocortex, rhinal cortex, caudate putamen, stria terminalis, locus ceruleus, parabrachial nucleus, and facial nucleus. Low grain densities were present in the globus pallidus, lateral thalamus, and midbrain. Negligible grain densities were present in the cerebellum, corpus callosum, and all regions treated with 1 microM unlabeled bombesin. The discrete regional distribution of binding suggests that endogenous bombesin-like peptides may function as important regulatory agents in certain brain loci

Potent μ-Opioid Receptor Agonists from Cyclic Peptides Tyr-c[D-Lys-Xxx-Tyr-Gly]: Synthesis, Biological, and Structural Evaluation.

Science.gov (United States)

Li, Yangmei; Cazares, Margret; Wu, Jinhua; Houghten, Richard A; Toll, Laurence; Dooley, Colette

2016-02-11

To optimize the structure of a μ-opioid receptor ligand, analogs H-Tyr-c[D-Lys-Xxx-Tyr-Gly] were synthesized and their biological activity was tested. The analog containing a Phe(3) was identified as not only exhibiting binding affinity 14-fold higher than the original hit but also producing agonist activity 3-fold more potent than morphine. NMR study suggested that a trans conformation at D-Lys(2)-Xxx(3) is crucial for these cyclic peptides to maintain high affinity, selectivity, and functional activity toward the μ-opioid receptor.

Singlet oxygen-mediated protein oxidation

DEFF Research Database (Denmark)

Wright, Adam; Bubb, William A; Hawkins, Clare Louise

2002-01-01

Singlet oxygen (1O2) is generated by a number of enzymes as well as by UV or visible light in the presence of a sensitizer and has been proposed as a damaging agent in a number of pathologies including cataract, sunburn, and skin cancers. Proteins, and Cys, Met, Trp, Tyr and His side chains...... in particular, are major targets for 1O2 as a result of their abundance and high rate constants for reaction. In this study it is shown that long-lived peroxides are formed on free Tyr, Tyr residues in peptides and proteins, and model compounds on exposure to 1O2 generated by both photochemical and chemical....... These studies demonstrate that long-lived Tyr-derived peroxides are formed on proteins exposed to 1O2 and that these may promote damage to other targets via further radical generation....

Electroencephalographic changes in the lateral septum complex following systemic administration of DES-TYR1 -[alpha]-endorphin, Des-Tyr1-[gamma]-endorphin and haloperidol in rats

NARCIS (Netherlands)

Urban, I.J.A.; Wied, D. de

1982-01-01

The influence of systemically administered Des-Tyr1-α-endorphin (DTαE), Des-Tyr1-γ-endorphin (DTγE) and haloperidol on electroencephalographic (EEG) activity of the lateral septum complex (LSC) and the frontal cortex was studied in male rats. DTαE (2 μg) significantly increased whereas DTγE (10 μg)

Increased nitration and carbonylation of proteins in MRL +/+ mice exposed to trichloroethene: Potential role of protein oxidation in autoimmunity

International Nuclear Information System (INIS)

Wang Gangduo; Wang Jianling; Ma Huaxian; Khan, M. Firoze

2009-01-01

Even though reactive oxygen and nitrogen species (RONS) are implicated as mediators of autoimmune diseases (ADs), little is known about contribution of protein oxidation (carbonylation and nitration) in the pathogenesis of such diseases. The focus of this study was, therefore, to establish a link between protein oxidation and induction and/or exacerbation of autoimmunity. To achieve this, female MRL +/+ mice were treated with trichloroethene (TCE), an environmental contaminant known to induce autoimmune response, for 6 or 12 weeks (10 mmol/kg, i.p., every 4 th day). TCE treatment resulted in significantly increased formation of nitrotyrosine (NT) and induction of iNOS in the serum at both 6 and 12 weeks of treatment, but the response was greater at 12 weeks. Likewise, TCE treatment led to greater NT formation, and iNOS protein and mRNA expression in the livers and kidneys. Moreover, TCE treatment also caused significant increases (∼3 fold) in serum protein carbonyls (a marker of protein oxidation) at both 6 and 12 weeks. Significantly increased protein carbonyls were also observed in the livers and kidneys (2.1 and 1.3 fold, respectively) at 6 weeks, and to a greater extent at 12 weeks (3.5 and 2.1 fold, respectively) following TCE treatment. The increases in TCE-induced protein oxidation (carbonylation and nitration) were associated with significant increases in Th1 specific cytokine (IL-2, IFN-γ) release into splenocyte cultures. These results suggest an association between protein oxidation and induction/exacerbation of autoimmune response. The results present a potential mechanism by which oxidatively modified proteins could contribute to TCE-induced autoimmune response and necessitates further investigations for clearly establishing the role of protein oxidation in the pathogenesis of ADs.

Stereospecific transport of Tyr-MIF-1 across the blood-brain barrier by peptide transport system-1

Energy Technology Data Exchange (ETDEWEB)

Banks, W.A.; Kastin, A.J.; Michals, E.A.; Barrera, C.M. (Veterans Affairs Medical Center, New Orleans, LA (USA))

1990-10-01

Previous studies have suggested that peptide transport system-1 (PTS-1), the saturable system that transports Tyr-MIF-1, the enkephalins, and related peptides out of the central nervous system (CNS), exhibits stereospecificity. In the present studies, we showed that {sup 125}I-L-Tyr-MIF-1, but not {sup 131}I-D-Tyr-MIF-1, was cleared from the CNS more rapidly than could be accounted for by nonspecific mechanisms. Such clearance was inhibited by a 1.0 nmol dose of L-Tyr-MIF-1, but not by D-Tyr-MIF-1. Neither L- nor D-Tyr-MIF-1 altered the much lower clearance of I-D-Tyr-MIF-1 from the brain. Radioactivity recovered from the vascular space after the injection of {sup 125}I-Tyr-MIF-1 into the lateral ventricle of the brain eluted by HPLC primarily as intact peptide, demonstrating that most of the Tyr-MIF-1 was not degraded during transport. By contrast, the nonsaturable unidirectional influx of Tyr-MIF-1 into the CNS did not distinguish between the isomers. These studies confirm and extend the observations that Tyr-MIF-1 is transported out of the CNS by a saturable, stereospecific transport system as an intact peptide while the influx into the CNS is by a nonsaturable mechanism that does not distinguish between the isomers.

Tyrosine and carboxyl protonation changes in the bacteriorhodopsin photocycle. 2. Tyrosine-26 and -64

International Nuclear Information System (INIS)

Roepe, P.; Scherrer, P.; Ahl, P.L.; Gupta, S.K.D.; Bogomolni, R.A.; Herzfeld, J.; Rothschild, K.J.

1987-01-01

Low-temperature Fourier transform infrared (FTIR) and UV difference spectroscopies combined with selective tyrosine nitration and tyrosine isotopic labeling have been used to investigate the participation of tyrosines-26 and -64 in the bacteriorhodopsin (bR) photocycle. Nitration of Tyr-26 has no detectable effect on the FTIR or UV difference spectra of the BR 570 → K 630 or BR 570 → M 412 transitions. In contrast, nitration of Tyr-64 causes changes in both the FTIR and UV spectra of these transitions. However, this nitration does not alter tyrosine peaks in the FTIR difference spectra which have previously been associated with the protonation of a tyrosinate by K 630 and the deprotonation of a tyrosine by M 412 . Instead, Tyr-64 nitration appears to affect other tyrosine peaks. These results and changes in UV difference spectra upon Tyr-64 nitration are consistent with the deprotonation of Tyr-64 by M 412 as concluded previously. Effects on chromophore vibrations caused by Tyr-64 nitration are unaltered upon reducing the nitrotyrosine to aminotyrosine with sodium dithionite. Finally, nitro-Tyr-64 causes a shift in the frequency of a positive peak at 1739 cm -1 in the BR 570 → M 412 FTIR difference spectrum which reflects the protonation of a carboxyl-containing residue. The shift does not occur for samples containing amino-Tyr-64. These data suggest that Tyr-64 may interact with this carboxyl group

Tyr66 acts as a conformational switch in the closed-to-open transition of the SHP-2 N-SH2-domain phosphotyrosine-peptide binding cleft

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MacKerell Alexander D

2007-03-01

Full Text Available Abstract Background The N-terminal SH2 domain (N-SH2 of the non-receptor tyrosine phosphatase SHP-2 is involved both in localization of SHP-2 by recognition of phosphotyrosine (pY peptides and self-inhibition of SHP-2 phosphatase activity through the formation of a protein – protein interface with the phosphatase domain. Mutations that disrupt this interface break the coupling between pY-peptide binding cleft conformation and self-inhibition, thereby increasing both SHP-2 phosphatase activity and pY-peptide binding affinity, and are associated with the congenital condition Noonan syndrome and various pediatric leukemias. To better characterize the molecular process involved in N-SH2 pY-dependent binding, we have applied explicit-solvent molecular dynamics simulations to study the closed-to-open transition of the N-SH2 pY-peptide binding cleft. Results The existence of stable conformations in the left-handed helical and the extended regions of Tyr66 φ/ψ space prevent rapid interconversion of the backbone and create a conformational switch such that Tyr66 in a left-handed helical backbone conformation results in an open cleft and in an extended backbone conformation results in a closed cleft. The stable conformations arise from deep, well-localized free-energy minima in the left-handed helical and extended regions of the Tyr66 φ/ψ map. Changing the Tyr66 backbone conformation from extended to left-handed helical induces a closed-to-open transition in the cleft, and the reverse change in backbone conformation induces the reverse, open-to-closed transition. In the open-cleft state, weak solvent-exposed interactions involving the sidechains of Tyr66, Asp40, Lys55, and Gln57 serve to anchor the Tyr66 sidechain to the surface of the protein and away from the binding cleft entrance, thereby facilitating pY-peptide access to the binding cleft. Conclusion The simulations point to a regulatory role for Tyr66 and surrounding residues in SHP-2 function

The dual effects of nitrite on hemoglobin-dependent redox reactions.

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Lu, Naihao; Chen, Chao; He, Yingjie; Tian, Rong; Xiao, Qiang; Peng, Yi-Yuan

2014-08-31

Evidence to support the role of heme proteins-dependent reactions as major inducers of oxidative damage is increasingly present. Nitrite (NO2(-)) is one of the major end products of NO metabolism, and from the daily consumption. Although the biological significance of heme proteins/NO2(-)-mediated protein tyrosine nitration is a subject of great interest, the important roles of NO2(-) on heme proteins-dependent redox reactions have been greatly underestimated. In this study, we investigated the influence of NO2(-) on met-hemoglobin (Hb)-dependent oxidative and nitrative stress. It was found that NO2(-) effectively reduced cytotoxic ferryl intermediate back to ferric Hb in a biphasic kinetic reaction. However, the presence of NO2(-) surprisingly exerted pro-oxidant effect on Hb-H2O2-induced protein (bovine serum albumin, enolase) oxidation at low concentrations and enhanced the loss of HepG2 cell viability. In the reduction of ferryl Hb to ferric state, NO2(-) was decreased and oxidized to a nitrating agent NO2, Tyr12 and Tyr191 in enolase were subsequently nitrated. In contrast to the frequently inhibitive effect of nitrotyrosine, NO2(-)-triggered tyrosine nitration might play an important role in enolase activation. These data provided novel evidence that the dietary intake and potential therapeutic application of NO2(-) would possess anti- and pro-oxidant activities through interfering in hemoglobin-dependent redox reactions. Besides the classic role in protein tyrosine nitration, the dual effects on hemoglobin-triggered oxidative stress may provide new insights into the physiological and toxicological implications of NO2(-) with heme proteins. Copyright © 2014 Elsevier Inc. All rights reserved.

Inorganic Nitrate Promotes the Browning of White Adipose Tissue through the Nitrate-Nitrite-Nitric Oxide Pathway

Science.gov (United States)

Roberts, Lee D; Ashmore, Tom; Kotwica, Aleksandra O; Murfitt, Steven A; Fernandez, Bernadette O; Feelisch, Martin; Griffin, Julian L

2015-01-01

Inorganic nitrate was once considered an oxidation end-product of nitric oxide metabolism with little biological activity. However, recent studies have demonstrated that dietary nitrate can modulate mitochondrial function in man and is effective in reversing features of the metabolic syndrome in mice. Using a combined histological, metabolomics, and transcriptional and protein analysis approach we mechanistically define that nitrate not only increases the expression of thermogenic genes in brown-adipose tissue but also induces the expression of brown adipocyte-specific genes and proteins in white adipose tissue, substantially increasing oxygen consumption and fatty acid β-oxidation in adipocytes. Nitrate induces these phenotypic changes through a mechanism distinct from known physiological small molecule activators of browning, the recently identified nitrate-nitrite-nitric oxide pathway. The nitrate-induced browning effect was enhanced in hypoxia, a serious co-morbidity affecting white adipose tissue in obese individuals, and corrected impaired brown adipocyte-specific gene expression in white adipose tissue in a murine model of obesity. Since resulting beige/brite cells exhibit anti-obesity and anti-diabetic effects, nitrate may be an effective means of inducing the browning response in adipose tissue to treat the metabolic syndrome. PMID:25249574

Mineral, protein and nitrate contents in leaves of Pereskia aculeata subjected to nitrogen fertilization

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Maria Regina de Miranda Souza

2016-03-01

Full Text Available Considering that nitrogen is directly related to leaf protein content, the nitrogen fertilization in Pereskia aculeata plants may affect the protein content and increase its nutritional potential. This study aimed at assessing the effect of nitrogen fertilization on mineral, protein and nitrate contents, as well as the yield of P. aculeata leaves. A randomized blocks design was used, with three replications and five treatments, consisting of increasing topdressing nitrogen doses (0-400 kg ha-1, in soil with organic matter content of 4.0 dag kg-1. Three harvests were performed for leaf analysis. No significant effect was observed for mineral and protein content or leaf fresh mass yield. The mean values for mineral composition were: 3.52 dag kg-1 of N, 0.47 dag kg-1 of P, 4.65 dag kg-1 of Ca, 0.71 dag kg-1 of Mg, 0.25 dag kg-1 of S, 36.64 mg kg-1 of Zn and 174.13 mg kg-1 of Fe. The mean content for protein was 21.86 % and the leaf fresh mass yield was 0.971 kg plant-1. K levels decreased from 50 kg ha-1 of N. Nitrate increased linearly with the nitrogen fertilization, reaching a maximum value of 78.2 mg kg-1 of fresh mass, well below the health risk threshold. It was concluded that a soil with high organic matter content does not require nitrogen fertilization. However, doses up to 400 kg ha-1 of nitrogen ensure adequate leaf yield and protein and mineral contents within the desired range for the species, being a food rich in proteins, iron and calcium.

Tyrosine phosphorylation switching of a G protein.

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Li, Bo; Tunc-Ozdemir, Meral; Urano, Daisuke; Jia, Haiyan; Werth, Emily G; Mowrey, David D; Hicks, Leslie M; Dokholyan, Nikolay V; Torres, Matthew P; Jones, Alan M

2018-03-30

Heterotrimeric G protein complexes are molecular switches relaying extracellular signals sensed by G protein-coupled receptors (GPCRs) to downstream targets in the cytoplasm, which effect cellular responses. In the plant heterotrimeric GTPase cycle, GTP hydrolysis, rather than nucleotide exchange, is the rate-limiting reaction and is accelerated by a receptor-like regulator of G signaling (RGS) protein. We hypothesized that posttranslational modification of the Gα subunit in the G protein complex regulates the RGS-dependent GTPase cycle. Our structural analyses identified an invariant phosphorylated tyrosine residue (Tyr 166 in the Arabidopsis Gα subunit AtGPA1) located in the intramolecular domain interface where nucleotide binding and hydrolysis occur. We also identified a receptor-like kinase that phosphorylates AtGPA1 in a Tyr 166 -dependent manner. Discrete molecular dynamics simulations predicted that phosphorylated Tyr 166 forms a salt bridge in this interface and potentially affects the RGS protein-accelerated GTPase cycle. Using a Tyr 166 phosphomimetic substitution, we found that the cognate RGS protein binds more tightly to the GDP-bound Gα substrate, consequently reducing its ability to accelerate GTPase activity. In conclusion, we propose that phosphorylation of Tyr 166 in AtGPA1 changes the binding pattern with AtRGS1 and thereby attenuates the steady-state rate of the GTPase cycle. We coin this newly identified mechanism "substrate phosphoswitching." © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Biotic and environmental stress induces nitration and changes in structure and function of the sea urchin major yolk protein toposome.

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Castellano, Immacolata; Migliaccio, Oriana; Ferraro, Giarita; Maffioli, Elisa; Marasco, Daniela; Merlino, Antonello; Zingone, Adriana; Tedeschi, Gabriella; Palumbo, Anna

2018-03-15

The major yolk protein toposome plays crucial roles during gametogenesis and development of sea urchins. We previously found that nitration of toposome increases in the gonads of a Paracentrotus lividus population living in a marine protected area affected by toxic blooms of Ostreospsis cf. ovata, compared to control populations. This modification is associated with ovatoxin accumulation, high levels of nitric oxide in the gonads, and a remarkable impairment of progeny development. However, nothing is known about the environmental-mediated-regulation of the structure and biological function of toposome. Here, we characterize through wide-ranging biochemical and structural analyses the nitrated toposome of sea urchins exposed to the bloom, and subsequently detoxified. The increased number of nitrated tyrosines in toposome of sea urchins collected during algal bloom induced structural changes and improvement of the Ca 2+ -binding affinity of the protein. After 3 months' detoxification, ovatoxin was undetectable, and the number of nitric oxide-modified tyrosines was reduced. However, the nitration of specific residues was irreversible and occurred also in embryos treated with metals, used as a proxy of environmental pollutants. The structural and functional changes of toposome caused by nitration under adverse environmental conditions may be related to the defective development of sea urchins' progeny.

Substitution of Tyr254 with Phe at the active site of flavocytochrome b2: consequences on catalysis of lactate dehydrogenation

International Nuclear Information System (INIS)

Dubois, J.; Chapman, S.K.; Mathews, F.S.; Reid, G.A.; Lederer, F.

1990-01-01

A role for Tyr254 in L-lactate dehydrogenation catalyzed by flavocytochrome b2 has recently been proposed on the basis of the known active-site structure and of studies that had suggested a mechanism involving the initial formation of a lactate carbanion. This role is now examined after replacement of Tyr254 with phenylalanine. The kcat is decreased about 40-fold, Km for lactate appears unchanged, and the mainly rate-limiting step is still alpha-hydrogen abstraction, as judged from the steady-state deuterium isotope effect. Modeling studies with lactate introduced into the active site indicate two possible substrate conformations with different hydrogen-bonding partners for the substrate hydroxyl. If the hydrogen bond is formed with Tyr254, as was initially postulated, the mechanism must involve removal by His373 of the C2 hydrogen, with carbanion formation. If, in the absence of the Tyr254 phenol group, the hydrogen bond is formed with His373 N3, the substrate is positioned in such a way that the reaction must proceed by hydride transfer. Therefore the mechanism of the Y254F enzyme was investigated so as to distinguish between the two mechanistic possibilities. 2-Hydroxy-3-butynoate behaves with the mutant as a suicide reagent, as with the wild-type enzyme. Similarly, the mutant protein also catalyzes the reduction and the dehydrohalogenation of bromopyruvate under transhydrogenation conditions

The albinism of the feral Asinara white donkeys (Equus asinus) is determined by a missense mutation in a highly conserved position of the tyrosinase (TYR) gene deduced protein.

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Utzeri, V J; Bertolini, F; Ribani, A; Schiavo, G; Dall'Olio, S; Fontanesi, L

2016-02-01

A feral donkey population (Equus asinus), living in the Asinara National Park (an island north-west of Sardinia, Italy), includes a unique white albino donkey subpopulation or colour morph that is a major attraction of this park. Disrupting mutations in the tyrosinase (TYR) gene are known to cause recessive albinisms in humans (oculocutaneous albinism Type 1; OCA1) and other species. In this study, we analysed the donkey TYR gene as a strong candidate to identify the causative mutation of the albinism of these donkeys. The TYR gene was sequenced from 13 donkeys (seven Asinara white albino and six coloured animals). Seven single nucleotide polymorphisms were identified. A missense mutation (c.604C>G; p.His202Asp) in a highly conserved amino acid position (even across kingdoms), which disrupts the first copper-binding site (CuA) of functional protein, was identified in the homozygous condition (G/G or D/D) in all Asinara white albino donkeys and in the albino son of a trio (the grey parents had genotype C/G or H/D), supporting the recessive mode of inheritance of this mutation. Genotyping 82 donkeys confirmed that Asinara albino donkeys had genotype G/G whereas all other coloured donkeys had genotype C/C or C/G. Across-population association between the c.604C>G genotypes and the albino coat colour was highly significant (P = 6.17E-18). The identification of the causative mutation of the albinism in the Asinara white donkeys might open new perspectives to study the dynamics of this putative deleterious allele in a feral population and to manage this interesting animal genetic resource. © 2015 Stichting International Foundation for Animal Genetics.

Long-term toxicity of [{sup 177}Lu-DOTA{sup 0},Tyr{sup 3}]octreotate in rats

Energy Technology Data Exchange (ETDEWEB)

Rolleman, Edgar J.; Krenning, Eric P.; Bernard, Bert F.; Visser, Monique de; Bijster, Magda; Jong, Marion de [Erasmus MC Rotterdam, Department of Nuclear Medicine, Rotterdam (Netherlands); Visser, Theo J. [Erasmus MC Rotterdam, Department of Internal Medicine, Rotterdam (Netherlands); Vermeij, Marcel [Erasmus MC Rotterdam, Department of Pathology, Rotterdam (Netherlands); Lindemans, Jan [Erasmus MC Rotterdam, Department of Clinical Chemistry, Rotterdam (Netherlands)

2007-02-15

Studies on peptide receptor radionuclide therapy (PRRT) using radiolabelled somatostatin analogues have shown promising results with regard to tumour control. The efficacy of PRRT is limited by uptake and retention in the proximal tubules of the kidney, which might lead to radiation nephropathy. We investigated the long-term renal toxicity after different doses of [{sup 177}Lu-DOTA{sup 0},Tyr{sup 3}]octreotate and the effects of dose fractionation and lysine co-injection in two tumour-bearing rat models. Significant renal toxicity was detected beyond 100 days after start of treatment as shown by elevated serum creatinine and proteinuria. Microscopically, tubules were strongly dilated with flat epithelium, containing protein cylinders. Creatinine levels rose significantly after 555 MBq [{sup 177}Lu-DOTA{sup 0},Tyr{sup 3}]octreotate, but were significantly lower after 278 MBq (single injection) or two weekly doses of 278 MBq. Renal damage scores were maximal after 555 MBq and significantly lower in the 278 and 2 x 278 MBq groups. Three doses of 185 MBq [{sup 177}Lu-DOTA{sup 0},Tyr{sup 3}]octreotate with intervals of a day, a week or a month significantly influenced serum creatinine (469{+-}18, 134{+-}70 and 65{+-}15 {mu}mol/l, respectively; p<0.001). Renal histological damage scores were not significantly influenced by dose fractionation. Lysine co-administration with three weekly treatments of 185 MBq significantly lowered serum creatinine and proteinuria. Injection of high doses of [{sup 177}Lu-DOTA{sup 0},Tyr{sup 3}]octreotate resulted in severe renal damage in rats as indicated by proteinuria, elevated serum creatinine and histological damage. This damage was dose dependent and became overt between 100 and 200 days after treatment. Dose fractionation had significant beneficial effects on kidney function. Also, lysine co-injection successfully prevented functional damage. (orig.)

Dityrosine, 3,4-Dihydroxyphenylalanine (DOPA), and radical formation from tyrosine residues on milk proteins with globular and flexible structures as a result of riboflavin-mediated photo-oxidation

DEFF Research Database (Denmark)

Dalsgaard, Trine Kastrup; Nielsen, Jacob Holm; Brown, Bronwyn

2011-01-01

Riboflavin-mediated photo-oxidative damage to protein Tyr residues has been examined to determine whether protein structure influences competing protein oxidation pathways in single proteins and protein mixtures. EPR studies resulted in the detection of Tyr-derived o-semiquione radicals, with thi......Riboflavin-mediated photo-oxidative damage to protein Tyr residues has been examined to determine whether protein structure influences competing protein oxidation pathways in single proteins and protein mixtures. EPR studies resulted in the detection of Tyr-derived o-semiquione radicals...

Mussel glue protein has an open conformation.

Science.gov (United States)

Williams, T; Marumo, K; Waite, J H; Henkens, R W

1989-03-01

Both native glue protein from marine mussels and a synthetic nonhydroxylated analog were analyzed by far-uv CD under a variety of conditions. Analysis of the CD spectra using various models strongly suggest a primarily random coil structure for both forms of the protein, a fact also supported by the absence of spectral change for the glue protein upon dilution into 6 M guanidine hydrochloride. The nonhydroxylated analog, which consists of 20 repeats of the peptide sequence Ala-Lys-Pro-Ser-Tyr-Pro-Pro-Thr-Tyr-Lys, was further characterized by enzyme modification using mushroom tyrosinase. Enzymatic hydroxylation of tyrosines was found to be best fit by a model containing two rate constants, 5.6 (+/- 0.6) X 10(-3) and 7.2 (+/- 0.3) X 10(-2) min-1. At equilibrium, HPLC analysis of digests showed nearly 100% conversion of Tyr-9 and only 15 to 35% conversion of Tyr-5. The Chou and Fasman rules for predicting structure were applied to the repeat sequence listed above. The rules predict the absence of alpha helix and beta pleated sheets in the structure of this peptide. On the other hand, beta turns are predicted to be present with Tyr-5 being in the region of highest probability. These data suggest that the protein in solution has only a small amount of secondary structure.

Protein tyrosine nitration in chronic intramuscular parasitism: immunohistochemical evaluation of relationships between nitration, and fiber type-specific responses to infection

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Ted H. Elsasser

2011-05-01

Full Text Available The present study was conducted to determine whether preferential muscle catabolism [psoas major (PM > rectus femoris (RF] observed during the chronic intramuscular stage of Sarcocystis cruzi infection could be associated with the pathological consequences of increased protein tyrosine nitration in fibers characteristically more metabolically active due to higher mitochondrial density. Holstein calves were assigned to control (C, or S. cruzi-infected (I groups, n=5/group. Calves were euthanized on day 63 of infection. Samples of RF and PM were prepared for metabolic fiber typing (MFT: slow oxidative, SO – Type I; fast oxidative glycolytic, FOG - Type IIa; fast glycolytic, FG – Type IIb, fiber area, and immunohistochemical localization of fast myosin heavy chain 2a and 2b, nitrotyrosine (NT, and mitochondrial Complex V ATP-synthase. MFT analysis documented that PM contained twice the number of SO fibers compared to RF (32 v 16%, P<0.002. SO and FOG fibers (Both higher in mitochondrial density than FG fibers in both PM and RF were significantly smaller in area in I calves with mean FG areas not different between C and I. Muscle NT content (Western blot of myofibrillar protein fraction increased with infection; NT was immunohistochemically localized into three distinct patterns in fibers: i sparse fiber staining, ii dense punctuate intrafiber staining, and iii pericystic staining. By image analysis, the greatest punctuate intrafiber pixel density of NT was associated with SO fibers from I calves with the NT colocalizing with mitochondrial Complex V – F1F0 ATP synthase. More fibers were positive for the colocalization in PM than RF (P<0.04. The data are consistent with the concept that fibers rich in mitochondria possessing more inherent oxidative energy capacity generate more nitrated proteins than glycolytic fibers and as such are more affected by the proinflammatory response to infections like Sarcocystosis.

Cloning and Functional Characterization of Octβ2-Receptor and Tyr1-Receptor in the Chagas Disease Vector, Rhodnius prolixus

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Sam Hana

2017-09-01

Full Text Available Octopamine and tyramine, both biogenic amines, are bioactive chemicals important in diverse physiological processes in invertebrates. In insects, octopamine and tyramine operate analogously to epinephrine and norepinephrine in the vertebrates. Octopamine and tyramine bind to G-protein coupled receptors (GPCRs leading to changes in second messenger levels and thereby modifying the function in target tissues and insect behavior. In this paper, we report the cDNA sequences of two GPCRs, RhoprOctβ2-R, and RhoprTyr1-R, have been cloned and functionally characterized from Rhodnius prolixus. Octopamine and tyramine each activate RhoprOctβ2-R and RhoprTyr1-R in a dose-dependent manner. Octopamine is one order of magnitude more potent than tyramine in activating RhoprOctβ2-R. Tyramine is two orders of magnitude more potent than octopamine in activating RhoprTyr1-R. Phentolamine and gramine significantly antagonize RhoprOctβ2-R, whereas yohimbine and phenoxybenzamine are effective blockers of RhoprTyr1-R. The transcripts of both receptors are enriched in the central nervous system (CNS and are expressed throughout the adult female reproductive system. It has been shown in other insects that Octβ2-R is essential for processes such as ovulation and fertilization. We previously reported that octopamine and tyramine modulate oviducts and bursa contractions in R. prolixus. Our data confirm the importance of octopamine and tyramine signaling in the reproductive system of R. prolixus.

A NIN-LIKE PROTEIN mediates nitrate-induced control of root nodule symbiosis in Lotus japonicus.

Science.gov (United States)

Nishida, Hanna; Tanaka, Sachiko; Handa, Yoshihiro; Ito, Momoyo; Sakamoto, Yuki; Matsunaga, Sachihiro; Betsuyaku, Shigeyuki; Miura, Kenji; Soyano, Takashi; Kawaguchi, Masayoshi; Suzaki, Takuya

2018-02-05

Legumes and rhizobia establish symbiosis in root nodules. To balance the gains and costs associated with the symbiosis, plants have developed two strategies for adapting to nitrogen availability in the soil: plants can regulate nodule number and/or stop the development or function of nodules. Although the former is accounted for by autoregulation of nodulation, a form of systemic long-range signaling, the latter strategy remains largely enigmatic. Here, we show that the Lotus japonicus NITRATE UNRESPONSIVE SYMBIOSIS 1 (NRSYM1) gene encoding a NIN-LIKE PROTEIN transcription factor acts as a key regulator in the nitrate-induced pleiotropic control of root nodule symbiosis. NRSYM1 accumulates in the nucleus in response to nitrate and directly regulates the production of CLE-RS2, a root-derived mobile peptide that acts as a negative regulator of nodule number. Our data provide the genetic basis for how plants respond to the nitrogen environment and control symbiosis to achieve proper plant growth.

Contribution of TyrB26 to the Function and Stability of Insulin

Science.gov (United States)

Pandyarajan, Vijay; Phillips, Nelson B.; Rege, Nischay; Lawrence, Michael C.; Whittaker, Jonathan; Weiss, Michael A.

2016-01-01

Crystallographic studies of insulin bound to receptor domains have defined the primary hormone-receptor interface. We investigated the role of TyrB26, a conserved aromatic residue at this interface. To probe the evolutionary basis for such conservation, we constructed 18 variants at B26. Surprisingly, non-aromatic polar or charged side chains (such as Glu, Ser, or ornithine (Orn)) conferred high activity, whereas the weakest-binding analogs contained Val, Ile, and Leu substitutions. Modeling of variant complexes suggested that the B26 side chains pack within a shallow depression at the solvent-exposed periphery of the interface. This interface would disfavor large aliphatic side chains. The analogs with highest activity exhibited reduced thermodynamic stability and heightened susceptibility to fibrillation. Perturbed self-assembly was also demonstrated in studies of the charged variants (Orn and Glu); indeed, the GluB26 analog exhibited aberrant aggregation in either the presence or absence of zinc ions. Thus, although TyrB26 is part of insulin's receptor-binding surface, our results suggest that its conservation has been enjoined by the aromatic ring's contributions to native stability and self-assembly. We envisage that such classical structural relationships reflect the implicit threat of toxic misfolding (rather than hormonal function at the receptor level) as a general evolutionary determinant of extant protein sequences. PMID:27129279

Evaluation of some feedstuffs with special emphasis on the effect of nitrate and other non-protein nitrogen fractions on ruminal microbial metabolism

International Nuclear Information System (INIS)

Nikolic, J.A.

1988-01-01

Green herbage grown under widely different conditions was examined for nitrate content. The mean value was 0.14% (range 0.01-0.32%) NO 3 -N in the dry matter (DM). It was concluded that the cut green fodder would induce high levels of blood methaemoglobin when fed to ruminants in mixed diets. Nevertheless, experiments in vitro indicated that relatively small amounts of dietary nitrate induced considerable microbial nitrate reducing activity in rumen contents. Moreover, nitrate reduction to ammonia was accompanied by alterations in microbial sulphur and energy metabolism. A technique for determining organic matter digestibility (OMD) in vitro was standardized and used to evaluate the potential nutritive value of some fruit wastes. Studies on apple pomace silage indicated that added urea did not significantly increase the silage protein content but remained as a potential source of non-protein nitrogen for rumen microorganisms. Apple pomace ensiled alone or preferably with other ingredients was acceptable to sheep and cattle, particularly as a supplement to their winter rations. (author). 27 refs, 2 figs, 5 tabs

Nitrates and nitrites intoxications’ management

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Alexandra Trif

2007-12-01

Full Text Available The study pointed out the major sources for clinical and subclinical intoxications with nitrates/nitrites (drinking water and nitrates containing fertilizers, circumstances that determine fertilizers to became sources of intoxication (excessive fertilization/consecutive high level of nitrates in fodders, free access of animals to the fertilizers, administration into the diet instead of natrium chloride, factors that determine high nitrates accumulation in fodders despite optimal fertilization (factors related to the plants, soil, clime, harvest methods, storage, agrotechnical measures, nitrates/nitrites toxicity (over 45 ppm nitrates in drinking water, over 0.5 g nitrate/100 g D.M fodder/diet, the factors that influence nitrates/nitrites toxicity ( species, age, rate of feeding, diet balance especially energetically, pathological effects and symptoms (irritation and congestions on digestive tract, resulting diarrhoea, transformation of hemoglobin into methemoglobin determining severe respiratory insufficiency, vascular collapse, low blood pressure inthe acute nitrates intoxication; hypotiroidism, hypovitaminosis A, reproductive disturbances(abortion, low rate of fertility, dead born offspring, diarrhoea and/or respiratory insufficiency in new born e.g. calves, immunosuppression, decrease of milk production in chronic intoxication. There were presented some suggestions concerning management practices to limit nitrate intoxication (analyze of nitrates/nitrites in water and fodders, good management of the situation of risk ,e .g. dilution of the diet with low nitrate content fodders, feeding with balanced diet in energy, protein, minerals and vitamins, accommodation to high nitrate level diet, avoid grazing one week after a frost period, avoid feeding chop green fodders stored a couple of days, monitoring of health status of animals fed with fodders containing nitrates at risk level, a.o..

Recombinant Brucella abortus gene expressing immunogenic protein

Energy Technology Data Exchange (ETDEWEB)

Mayfield, J.E.; Tabatabai, L.B.

1991-06-11

This patent describes a synthetic recombinant DNA molecule containing a DNA sequence. It comprises a gene of Brucella abortus encoding an immunogenic protein having a molecular weight of approximately 31,000 daltons as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis under denaturing conditions, the protein having an isoelectric point around 4.9, and containing a twenty-five amino acid sequence from its amino terminal end consisting of Gln-Ala-Pro-Thr-Phe-Phe-Arg-Ile-Gly-Thr-Gly-Gly-Thr-Ala-Gly-Thr-Tyr-Tyr-Pro-Ile-Gly-Gly-Leu-Ile-Ala, wherein Gln, Ala, Pro, Thr, Phe, Arg, Ile, Gly, Tyr, and Leu, respectively, represent glutamine, alanine, proline, threonine, phenylalanine, arginine, isolecuine, glycine, tyrosine, and leucine.

Nitrate contamination of groundwater and its countermeasures

Energy Technology Data Exchange (ETDEWEB)

Mitamura, Hisayoshi [Japan Atomic Energy Research Inst., Tokai, Ibaraki (Japan). Tokai Research Establishment

2003-03-01

The inevitable increases of food production and energy consumption with an increase in world population become main causes of an increase of nitrate load to the environment. Although nitrogen is essential for the growth of animal and plant as a constituent element of protein, excessive nitrate load to the environment contaminates groundwater resources used as drinking water and leads to seriously adverse effects on the health of man and livestock. In order to clarify the problem of nitrate contamination of groundwater and search a new trend of technology development from the viewpoint of environment remediation and protection, the present paper has reviewed adverse effects of nitrate on human health, the actual state of nitrogen cycle, several kinds of nitrate sources, measures for reducing nitrate level, etc. (author)

Nitrate Enhanced Microbial Cr(VI) Reduction-Final Report

Energy Technology Data Exchange (ETDEWEB)

John F. Stolz

2011-06-15

A major challenge for the bioremediation of radionuclides (i.e., uranium, technetium) and metals (i.e., Cr(VI), Hg) is the co-occurrence of nitrate as it can inhibit metal transformation. Denitrification (nitrate reduction to dinitrogen gas) is considered the most important ecological process. For many metal and metalloid reducing bacteria, however, ammonia is the end product through respiratory nitrate reduction (RNRA). The focus of this work was to determine how RNRA impacts Cr(VI) transformation. The goal was to elucidate the specific mechanism(s) that limits Cr(VI) reduction in the presence of nitrate and to use this information to develop strategies that enhance Cr(VI) reduction (and thus detoxification). Our central hypothesis is that nitrate impacts the biotransformation of metals and metalloids in three ways 1) as a competitive alternative electron acceptor (inhibiting transformation), 2) as a co-metabolite (i.e., concomitant reduction, stimulating transformation), and 3) as an inducer of specific proteins and pathways involved in oxidation/reduction reactions (stimulating transformation). We have identified three model organisms, Geobacter metallireducens (mechanism 1), Sulfurospirillum barnesii, (mechasism 2), and Desulfovibrio desulfuricans (mechanisms 3). Our specific aims were to 1) investigate the role of Cr(VI) concentration on the kinetics of both growth and reduction of nitrate, nitrite, and Cr(VI) in these three organisms; 2) develop a profile of bacterial enzymes involved in nitrate transformation (e.g., oxidoreductases) using a proteomic approach; 3) investigate the function of periplasmic nitrite reductase (Nrf) as a chromate reductase; and 4) develop a strategy to maximize microbial chromium reduction in the presence of nitrate. We found that growth on nitrate by G. metallireducens was inhibited by Cr(VI). Over 240 proteins were identified by LC/MS-MS. Redox active proteins, outer membrane heavy metal efflux proteins, and chemotaxis sensory

Screening of TYR, OCA2, GPR143, and MC1R in patients with congenital nystagmus, macular hypoplasia, and fundus hypopigmentation indicating albinism

Science.gov (United States)

Preising, Markus N.; Gonser, Miriam; Lorenz, Birgit

2011-01-01

Background A broad spectrum of pigmentation of the skin and hair is found among patients diagnosed with ocular albinism (OA) and oculocutaneous albinism (OCA). Even though complexion is variable, three ocular features, i.e., hypopigmentation of the fundus, hypoplasia of the macula, and nystagmus, are classical pathological findings in these patients. We screened 172 index patients with a clinical diagnosis of OA or OCA based on the classical findings, to evaluate the frequency of sequence variants in tyrosinase (TYR), P-gene, P-protein (OCA2), and the G-protein-coupled receptor 143 gene, OA1 (GPR143). In addition, we investigated the association of sequence variants in the melanocortin receptor 1 gene (MC1R) and OCA2. Methods Pigmentation of the hair, skin, iris, and fundus were included in the evaluation of OCA and OA. Male OA patients showing X-linked inheritance were screened for GPR143. Females showing OA without family history were regarded as representing autosomal recessive OA (OA3). Direct sequencing was applied to PCR products showing aberrant single-strand conformation polymorphism–banding patterns. Results Fifty-seven male index patients were screened for OA. We identified 16 potentially pathogenic sequence variations in GPR143 (10 novel) in 22 males. In TYR, we identified 23 (7 novel), and in OCA2 28 (11 novel) possibly pathogenic variants. Variants on both alleles were identified in TYR or OCA2 in 29/79 OCA patients and 14/71 OA patients. Sequence changes in TYR were identified almost exclusively in OCA patients, while sequence changes in OCA2 occurred in OCA and OA patients. MC1R sequencing was performed in 47 patients carrying mutations in OCA2 and revealed MC1R mutations in 42 of them. Conclusions TYR gene mutations have a more severe effect on pigmentation than mutations in OCA2 and the GPR143 gene. Nevertheless, mutations in these genes affect the development of visual function either directly or by interaction with other genes like MC1R, which

Reaction between protein radicals and other biomolecules

DEFF Research Database (Denmark)

Østdal, H.; Davies, M.J.; Andersen, Henrik Jørgen

2002-01-01

The present study investigates the reactivity of bovine serum albumin (BSA) radicals towards different biomolecules (urate, linoleic acid, and a polypeptide, poly(Glu-Ala-Tyr)). The BSA radical was formed at room temperature through a direct protein-to-protein radical transfer from H(2)O(2....... Subsequent analysis showed a decrease in the concentration of urate upon reaction with the BSA radical, while the BSA radical in the presence of poly(Glu-Ala-Tyr) resulted in increased formation of the characteristic protein oxidation product, dityrosine. Reaction between the BSA radical and a linoleic acid...

Laboratoire de Chimie Bactérienne C.N.R.S., Marsielle, France.

Science.gov (United States)

Chippaux, M; Giudici, D; Abou-Jaoudé, A; Casse, F; Pascal, M C

1978-04-06

Mutants of E. coli, completely devoid of nitrite reductase activity with glucose or formate as donor were studied. Biochemical analysis indicates that they are simultaneously affected in nitrate reductase, nitrite reductase, fumarate reductase and hydrogenase activities as well as in cytochrome C552 biosynthesis. The use of an antiserum specific for nitrate reductase shows that the nitrate reductase protein is probably missing. A single mutation is responsible for this phenotype: the gene affected, nir R, is located close to tyr R i.e. at 29 min on the chromosomal map.

[An overview of oculocutaneous albinism: TYR gene mutations in five Colombian individuals].

Science.gov (United States)

Sanabria, Diana; Groot, Helena; Guzmán, Julio; Lattig, María Claudia

2012-06-01

Oculocutaneus albinism is a pigment-related inherited disorder characterized by hypopigmentation of the skin, hair and eyes, foveal hypoplasia and low vision. To date, 230 mutations in the TYR gene have been reported as responsible for oculocutaneus albinism type 1 worldwide. TYR gene encodes the enzyme tyrosinase involved in the metabolic pathway of melanin synthesis. Mutations were identified in the TYR gene as responsible for oculocutaneous albinism type 1 in five Colombian individuals, and a new ophthalmic system was tested that corrected visual defects and symptoms in a patient with oculocutaneous albinism. Samples were taken from 5 individuals, four of whom belong to a single family, along with a fifth individual not related to the family. Five exons in the TYR gene were sequenced to search for the gene carriers in the family and in the non-related individual. In addition, clinical ophthalmological evaluation and implementation of an new oculo-visual system was undertaken. A G47D and 1379delTT mutation was identified in the family. The unrelated individual carried a compound heterozygote for the G47D and D42N mutations. The oculo-visual corrective system was able to increase visual acuity and to diminish the nystagmus and photophobia. This is the first study in Colombia where albinism mutations are reported. The methods developed will enable future molecular screening studies in Colombian populations.

Tumoral fibrosis effect on the radiation absorbed dose of 177Lu–Tyr3-octreotate and 177Lu–Tyr3-octreotate conjugated to gold nanoparticles

International Nuclear Information System (INIS)

Azorín-Vega, E.P.; Zambrano-Ramírez, O.D.; Rojas-Calderón, E.L.; Ocampo-García, B.E.; Ferro-Flores, G.

2015-01-01

The aim of this work was to evaluate the tumoral fibrosis effect on the radiation absorbed dose of the radiopharmaceuticals 177 Lu–Tyr 3 -octreotate (monomeric) and 177 Lu–Tyr 3 -octreotate–gold nanoparticles (multimeric) using an experimental HeLa cells tumoral model and the Monte Carlo PENELOPE code. Experimental and computer micro-environment models with or without fibrosis were constructed. Results showed that fibrosis increases up to 33% the tumor radiation absorbed dose, although the major effect on the dose was produced by the type of radiopharmaceutical (112 Gy-multimeric vs. 43 Gy-monomeric). - Highlights: • Fibrosis increases the radiation absorbed dose to the tumor. • Fibrosis increases the radiopharmaceutical residence time in the tumor. • The multimeric nature of the radiopharmaceuticals enhances the radiopharmaceutical retention

Effects of nitrate and phosphate availabilities on growth, photosynthesis and pigment and protein contents in colour strains of Hypnea musciformis (Wulfen in Jacqu. J.V. Lamour. (Gigartinales, Rhodophyta

Directory of Open Access Journals (Sweden)

Aline P. Martins

2011-05-01

Full Text Available In Brazil, Hypnea musciformis is the main raw material for carrageenan production and the knowledge of nitrogen and phosphorus metabolism in algae is critical for the success of cultivation because these elements can limit seaweed productivity. Thus, the objective of this study was to evaluate the effects of nitrate (zero to 100 μM and nitrate plus phosphate (zero to 25 μM availabilities on the growth, the contents of photosynthetic pigments (phycobiliproteins and chlorophyll a and proteins, and the photosynthesis and respiration of the brown (BR and light green (LG strains of H. musciformis. The results revealed metabolic differences between the colour strains of H. musciformis for nitrogen metabolism: upon nitrate addition, the LG strain stored nitrogen mainly as proteins, while the BR strain stored it as proteins and pigments. Moreover, the respiration of the LG strain and the photosynthesis of the BR strain increased with nitrate concentrations, indicating that the BR strain fixed more photosynthetic carbon than the LG strain.

Effects of nitrate and phosphate availabilities on growth, photosynthesis and pigment and protein contents in colour strains of Hypnea musciformis (Wulfen in Jacqu. J.V. Lamour. (Gigartinales, Rhodophyta

Directory of Open Access Journals (Sweden)

Aline P. Martins

2011-04-01

Full Text Available In Brazil, Hypnea musciformis is the main raw material for carrageenan production and the knowledge of nitrogen and phosphorus metabolism in algae is critical for the success of cultivation because these elements can limit seaweed productivity. Thus, the objective of this study was to evaluate the effects of nitrate (zero to 100 μM and nitrate plus phosphate (zero to 25 μM availabilities on the growth, the contents of photosynthetic pigments (phycobiliproteins and chlorophyll a and proteins, and the photosynthesis and respiration of the brown (BR and light green (LG strains of H. musciformis. The results revealed metabolic differences between the colour strains of H. musciformis for nitrogen metabolism: upon nitrate addition, the LG strain stored nitrogen mainly as proteins, while the BR strain stored it as proteins and pigments. Moreover, the respiration of the LG strain and the photosynthesis of the BR strain increased with nitrate concentrations, indicating that the BR strain fixed more photosynthetic carbon than the LG strain.

Mutational Analysis of the TYR and OCA2 Genes in Four Chinese Families with Oculocutaneous Albinism.

Science.gov (United States)

Wang, Yun; Wang, Zhi; Chen, Mengping; Fan, Ning; Yang, Jie; Liu, Lu; Wang, Ying; Liu, Xuyang

2015-01-01

Oculocutaneous albinism (OCA) is an autosomal recessive disorder. The most common type OCA1 and OCA2 are caused by homozygous or compound heterozygous mutations in the tyrosinase gene (TYR) and OCA2 gene, respectively. The purpose of this study was to evaluate the molecular basis of oculocutaneous albinism in four Chinese families. Four non-consanguineous OCA families were included in the study. The TYR and OCA2 genes of all individuals were amplified by polymerase chain reaction (PCR), sequenced and compared with a reference database. Four patients with a diagnosis of oculocutaneous albinism, presented with milky skin, white or light brown hair and nystagmus. Genetic analyses demonstrated that patient A was compound heterozygous for c.1037-7T.A, c.1037-10_11delTT and c.1114delG mutations in the TYR gene; patient B was heterozygous for c.593C>T and c.1426A>G mutations in the OCA2 gene, patients C and D were compound heterozygous mutations in the TYR gene (c.549_550delGT and c.896G>A, c.832C>T and c.985T>C, respectively). The heterozygous c.549_550delGT and c.1114delG alleles in the TYR gene were two novel mutations. Interestingly, heterozygous members in these pedigrees who carried c.1114delG mutations in the TYR gene or c.1426A>G mutations in the OCA2 gene presented with blond or brown hair and pale skin, but no ocular disorders when they were born; the skin of these patients accumulated pigment over time and with sun exposure. This study expands the mutation spectrum of oculocutaneous albinism. It is the first time, to the best of our knowledge, to report that c.549_550delGT and c.1114delG mutations in the TYR gene were associated with OCA. The two mutations (c.1114delG in the TYR gene and c.1426A>G in the OCA2 gene) may be responsible for partial clinical manifestations of OCA.

Hormonogenic donor Tyr2522 of bovine thyroglobulin. Insight into preferential T3 formation at thyroglobulin carboxyl terminus at low iodination level

Energy Technology Data Exchange (ETDEWEB)

Cetrangolo, Giovanni Paolo, E-mail: giovanni.cetrangolo@unimol.it [Dipartimento di Medicina e Scienze della Salute, Università del Molise, Via De Sanctis, snc, Campobasso 86100 (Italy); Arcaro, Alessia, E-mail: alessia.arcaro@unimol.it [Dipartimento di Medicina e Scienze della Salute, Università del Molise, Via De Sanctis, snc, Campobasso 86100 (Italy); Lepore, Alessio, E-mail: alessiolepore@alice.it [Dipartimento di Medicina Molecolare e Biotecnologie Mediche, Università di Napoli Federico II, Via S. Pansini, 5, Napoli 80131 (Italy); Graf, Maria, E-mail: mariagraf@tiscali.it [Dipartimento di Medicina Molecolare e Biotecnologie Mediche, Università di Napoli Federico II, Via S. Pansini, 5, Napoli 80131 (Italy); Mamone, Gianfranco, E-mail: mamone@isa.cnr.it [Centro di Spettrometria di Massa Proteomica e Biomolecolare, ISA-CNR, Via Roma 52 a, Avellino 83100 (Italy); Ferranti, Pasquale, E-mail: ferranti@unina.it [Dipartimento di Agraria, Università di Napoli “Federico II”, Parco Gussone, Portici (Napoli) 80055 (Italy); Palumbo, Giuseppe, E-mail: palumbo@unina.it [Dipartimento di Medicina Molecolare e Biotecnologie Mediche, Università di Napoli Federico II, Via S. Pansini, 5, Napoli 80131 (Italy); Gentile, Fabrizio, E-mail: gentilefabrizio@unimol.it [Dipartimento di Medicina e Scienze della Salute, Università del Molise, Via De Sanctis, snc, Campobasso 86100 (Italy)

2014-07-18

Highlights: • A carboxy-terminal fragment (residues 2515–2750) was isolated from a low-iodine bTg. • Post-translational status of 8 tyrosines in bTg region 2515–2750 was assessed by MS. • Tyr2522 of bovine Tg is an interspecifically conserved hormonogenic donor site. • Propensities of Tyr residues to mono or diiodination optimize T3 yield from Tyr2748. - Abstract: A tryptic fragment (b5{sub TR,NR}), encompassing residues 2515–2750, was isolated from a low-iodine (0.26% by mass) bovine thyroglobulin, by limited proteolysis with trypsin and preparative, continuous-elution SDS–PAGE. The fragment was digested with Asp-N endoproteinase and analyzed by reverse-phase HPLC electrospray ionization quadrupole time-of-flight mass spectrometry, revealing the formation of: 3-monoiodotyrosine and dehydroalanine from Tyr2522; 3-monoiodotyrosine from Tyr2555 and Tyr2569; 3-monoiodotyrosine and 3,5-diiodotyrosine from Tyr2748. The data presented document, by direct mass spectrometric identifications, efficient iodophenoxyl ring transfer from monoiodinated hormonogenic donor Tyr2522 and efficient mono- and diiodination of hormonogenic acceptor Tyr2748, under conditions which permitted only limited iodination of Tyr2555 and Tyr2569, in low-iodine bovine thyroglobulin. The present study thereby provides: (1) a rationale for the preferential synthesis of T3 at the carboxy-terminal end of thyroglobulin, at low iodination level; (2) confirmation for the presence of an interspecifically conserved hormonogenic donor site in the carboxy-terminal domain of thyroglobulin; (3) solution for a previous uncertainty, concerning the precise location of such donor site in bovine thyroglobulin.

Hormonogenic donor Tyr2522 of bovine thyroglobulin. Insight into preferential T3 formation at thyroglobulin carboxyl terminus at low iodination level

International Nuclear Information System (INIS)

Cetrangolo, Giovanni Paolo; Arcaro, Alessia; Lepore, Alessio; Graf, Maria; Mamone, Gianfranco; Ferranti, Pasquale; Palumbo, Giuseppe; Gentile, Fabrizio

2014-01-01

Highlights: • A carboxy-terminal fragment (residues 2515–2750) was isolated from a low-iodine bTg. • Post-translational status of 8 tyrosines in bTg region 2515–2750 was assessed by MS. • Tyr2522 of bovine Tg is an interspecifically conserved hormonogenic donor site. • Propensities of Tyr residues to mono or diiodination optimize T3 yield from Tyr2748. - Abstract: A tryptic fragment (b5 TR,NR ), encompassing residues 2515–2750, was isolated from a low-iodine (0.26% by mass) bovine thyroglobulin, by limited proteolysis with trypsin and preparative, continuous-elution SDS–PAGE. The fragment was digested with Asp-N endoproteinase and analyzed by reverse-phase HPLC electrospray ionization quadrupole time-of-flight mass spectrometry, revealing the formation of: 3-monoiodotyrosine and dehydroalanine from Tyr2522; 3-monoiodotyrosine from Tyr2555 and Tyr2569; 3-monoiodotyrosine and 3,5-diiodotyrosine from Tyr2748. The data presented document, by direct mass spectrometric identifications, efficient iodophenoxyl ring transfer from monoiodinated hormonogenic donor Tyr2522 and efficient mono- and diiodination of hormonogenic acceptor Tyr2748, under conditions which permitted only limited iodination of Tyr2555 and Tyr2569, in low-iodine bovine thyroglobulin. The present study thereby provides: (1) a rationale for the preferential synthesis of T3 at the carboxy-terminal end of thyroglobulin, at low iodination level; (2) confirmation for the presence of an interspecifically conserved hormonogenic donor site in the carboxy-terminal domain of thyroglobulin; (3) solution for a previous uncertainty, concerning the precise location of such donor site in bovine thyroglobulin

A comparison of the effect of lead nitrate on rat liver chromatin, DNA and histone proteins in solution.

Science.gov (United States)

Rabbani-Chadegani, Azra; Abdosamadi, Sayeh; Fani, Nesa; Mohammadian, Shayesteh

2009-06-01

Although lead is widely recognized as a toxic substance in the environment and directly damage DNA, no studies are available on lead interaction with chromatin and histone proteins. In this work, we have examined the effect of lead nitrate on EDTA-soluble chromatin (SE chromatin), DNA and histones in solution using absorption and fluorescence spectroscopy, thermal denaturation and gel electrophoresis techniques. The results demonstrate that lead nitrate binds with higher affinity to chromatin than to DNA and produces an insoluble complex as monitored at 400 nm. Binding of lead to DNA decreases its Tm, increases its fluorescence intensity and exhibits hypochromicity at 210 nm which reveal that both DNA bases and the backbone participate in the lead-DNA interaction. Lead also binds strongly to histone proteins in the absence of DNA. The results suggest that although lead destabilizes DNA structure, in the chromatin, the binding of lead introduces some sort of compaction and aggregation, and the histone proteins play a key role in this aspect. This chromatin condensation, upon lead exposure, in turn may decrease fidelity of DNA, and inhibits DNA and RNA synthesis, the process that introduces lead toxicity at the chromatin level.

In vivo binding properties of SH2 domains from GTPase-activating protein and phosphatidylinositol 3-kinase.

Science.gov (United States)

Cooper, J A; Kashishian, A

1993-01-01

We have used a transient expression system and mutant platelet-derived growth factor (PDGF) receptors to study the binding specificities of the Src homology 2 (SH2) regions of the Ras GTPase-activator protein (GAP) and the p85 alpha subunit of phosphatidylinositol 3-kinase (PI3 kinase). A number of fusion proteins, each tagged with an epitope allowing recognition by a monoclonal antibody, were expressed at levels comparable to those of endogenous GAP. Fusion proteins containing the central SH2-SH3-SH2 region of GAP or the C-terminal region of p85 alpha, which includes two SH2 domains, bound to PDGF receptors in response to PDGF stimulation. Both fusion proteins showed the same requirements for tyrosine phosphorylation sites in the PDGF receptor as the full-length proteins from which they were derived, i.e., binding of the GAP fusion protein was reduced by mutation of Tyr-771, and binding of the p85 fusion protein was reduced by mutation of Tyr-740, Tyr-751, or both residues. Fusion proteins containing single SH2 domains from either GAP or p85 alpha did not bind detectably to PDGF receptors in this system, suggesting that two SH2 domains in a single polypeptide cooperate to raise the affinity of binding. The sequence specificities of individual SH2 domains were deduced from the binding properties of fusion proteins containing one SH2 domain from GAP and another from p85. The results suggest that the C-terminal GAP SH2 domain specifies binding to Tyr-771, the C-terminal p85 alpha SH2 domain binds to either Tyr-740 or Tyr-751, and each protein's N-terminal SH2 domain binds to unidentified phosphorylation sites.(ABSTRACT TRUNCATED AT 250 WORDS) Images PMID:8382774

Protein Ser/Thr/Tyr phosphorylation in the Archaea.

Science.gov (United States)

Kennelly, Peter J

2014-04-04

The third domain of life, the Archaea (formerly Archaebacteria), is populated by a physiologically diverse set of microorganisms, many of which reside at the ecological extremes of our global environment. Although ostensibly prokaryotic in morphology, the Archaea share much closer evolutionary ties with the Eukarya than with the superficially more similar Bacteria. Initial genomic, proteomic, and biochemical analyses have revealed the presence of "eukaryotic" protein kinases and phosphatases and an intriguing set of serine-, threonine-, and tyrosine-phosphorylated proteins in the Archaea that may offer new insights into this important regulatory mechanism.

Ammonium nitrate-potassium nitrate system

Energy Technology Data Exchange (ETDEWEB)

Cady, H.H.

1981-01-01

A portion of the binary phase diagram for the system ammonium nitrate-potassium nitrate has been determined from -55/sup 0/C to 185/sup 0/C. Results are presented for the ammonium-nitrate-rich end of the system up to 30 wt% potassium nitrate.

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

Science.gov (United States)

Cheng, H F; Tao, M

1989-10-19

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

Disruption of the rice nitrate transporter OsNPF2.2 hinders root-to-shoot nitrate transport and vascular development

Science.gov (United States)

Li, Yuge; Ouyang, Jie; Wang, Ya-Yun; Hu, Rui; Xia, Kuaifei; Duan, Jun; Wang, Yaqin; Tsay, Yi-Fang; Zhang, Mingyong

2015-01-01

Plants have evolved to express some members of the nitrate transporter 1/peptide transporter family (NPF) to uptake and transport nitrate. However, little is known of the physiological and functional roles of this family in rice (Oryza sativa L.). Here, we characterized the vascular specific transporter OsNPF2.2. Functional analysis using cDNA-injected Xenopus laevis oocytes revealed that OsNPF2.2 is a low-affinity, pH-dependent nitrate transporter. Use of a green fluorescent protein tagged OsNPF2.2 showed that the transporter is located in the plasma membrane in the rice protoplast. Expression analysis showed that OsNPF2.2 is nitrate inducible and is mainly expressed in parenchyma cells around the xylem. Disruption of OsNPF2.2 increased nitrate concentration in the shoot xylem exudate when nitrate was supplied after a deprivation period; this result suggests that OsNPF2.2 may participate in unloading nitrate from the xylem. Under steady-state nitrate supply, the osnpf2.2 mutants maintained high levels of nitrate in the roots and low shoot:root nitrate ratios; this observation suggests that OsNPF2.2 is involved in root-to-shoot nitrate transport. Mutation of OsNPF2.2 also caused abnormal vasculature and retarded plant growth and development. Our findings demonstrate that OsNPF2.2 can unload nitrate from the xylem to affect the root-to-shoot nitrate transport and plant development. PMID:25923512

Application of SAIL phenylalanine and tyrosine with alternative isotope-labeling patterns for protein structure determination

Energy Technology Data Exchange (ETDEWEB)

Takeda, Mitsuhiro [Nagoya University, Structural Biology Research Center, Graduate School of Science (Japan); Ono, Akira M.; Terauchi, Tsutomu [SAIL Technologies Co., Inc. (Japan); Kainosho, Masatsune, E-mail: kainosho@nmr.chem.metro-u.ac.j [Nagoya University, Structural Biology Research Center, Graduate School of Science (Japan)

2010-01-15

The extensive collection of NOE constraint data involving the aromatic ring signals is essential for accurate protein structure determination, although it is often hampered in practice by the pervasive signal overlapping and tight spin couplings for aromatic rings. We have prepared various types of stereo-array isotope labeled phenylalanines ({epsilon}- and {zeta}-SAIL Phe) and tyrosine ({epsilon}-SAIL Tyr) to overcome these problems (Torizawa et al. 2005), and proven that these SAIL amino acids provide dramatic spectral simplification and sensitivity enhancement for the aromatic ring NMR signals. In addition to these SAIL aromatic amino acids, we recently synthesized {delta}-SAIL Phe and {delta}-SAIL Tyr, which allow us to observe and assign {delta}-{sup 13}C/{sup 1}H signals very efficiently. Each of the various types of SAIL Phe and SAIL Tyr yields well-resolved resonances for the {delta}-, {epsilon}- or {zeta}-{sup 13}C/{sup 1}H signals, respectively, which can readily be assigned by simple and robust pulse sequences. Since the {delta}-, {epsilon}-, and {zeta}-proton signals of Phe/Tyr residues give rise to complementary NOE constraints, the concomitant use of various types of SAIL-Phe and SAIL-Tyr would generate more accurate protein structures, as compared to those obtained by using conventional uniformly {sup 13}C, {sup 15}N-double labeled proteins. We illustrated this with the case of an 18.2 kDa protein, Escherichia coli peptidyl-prolyl cis-trans isomerase b (EPPIb), and concluded that the combined use of {zeta}-SAIL Phe and {epsilon}-SAIL Tyr would be practically the best choice for protein structural determinations.

Application of SAIL phenylalanine and tyrosine with alternative isotope-labeling patterns for protein structure determination.

Science.gov (United States)

Takeda, Mitsuhiro; Ono, Akira M; Terauchi, Tsutomu; Kainosho, Masatsune

2010-01-01

The extensive collection of NOE constraint data involving the aromatic ring signals is essential for accurate protein structure determination, although it is often hampered in practice by the pervasive signal overlapping and tight spin couplings for aromatic rings. We have prepared various types of stereo-array isotope labeled phenylalanines (epsilon- and zeta-SAIL Phe) and tyrosine (epsilon-SAIL Tyr) to overcome these problems (Torizawa et al. 2005), and proven that these SAIL amino acids provide dramatic spectral simplification and sensitivity enhancement for the aromatic ring NMR signals. In addition to these SAIL aromatic amino acids, we recently synthesized delta-SAIL Phe and delta-SAIL Tyr, which allow us to observe and assign delta-(13)C/(1)H signals very efficiently. Each of the various types of SAIL Phe and SAIL Tyr yields well-resolved resonances for the delta-, epsilon- or zeta-(13)C/(1)H signals, respectively, which can readily be assigned by simple and robust pulse sequences. Since the delta-, epsilon-, and zeta-proton signals of Phe/Tyr residues give rise to complementary NOE constraints, the concomitant use of various types of SAIL-Phe and SAIL-Tyr would generate more accurate protein structures, as compared to those obtained by using conventional uniformly (13)C, (15)N-double labeled proteins. We illustrated this with the case of an 18.2 kDa protein, Escherichia coli peptidyl-prolyl cis-trans isomerase b (EPPIb), and concluded that the combined use of zeta-SAIL Phe and epsilon-SAIL Tyr would be practically the best choice for protein structural determinations.

Application of SAIL phenylalanine and tyrosine with alternative isotope-labeling patterns for protein structure determination

International Nuclear Information System (INIS)

Takeda, Mitsuhiro; Ono, Akira M.; Terauchi, Tsutomu; Kainosho, Masatsune

2010-01-01

The extensive collection of NOE constraint data involving the aromatic ring signals is essential for accurate protein structure determination, although it is often hampered in practice by the pervasive signal overlapping and tight spin couplings for aromatic rings. We have prepared various types of stereo-array isotope labeled phenylalanines (ε- and ζ-SAIL Phe) and tyrosine (ε-SAIL Tyr) to overcome these problems (Torizawa et al. 2005), and proven that these SAIL amino acids provide dramatic spectral simplification and sensitivity enhancement for the aromatic ring NMR signals. In addition to these SAIL aromatic amino acids, we recently synthesized δ-SAIL Phe and δ-SAIL Tyr, which allow us to observe and assign δ- 13 C/ 1 H signals very efficiently. Each of the various types of SAIL Phe and SAIL Tyr yields well-resolved resonances for the δ-, ε- or ζ- 13 C/ 1 H signals, respectively, which can readily be assigned by simple and robust pulse sequences. Since the δ-, ε-, and ζ-proton signals of Phe/Tyr residues give rise to complementary NOE constraints, the concomitant use of various types of SAIL-Phe and SAIL-Tyr would generate more accurate protein structures, as compared to those obtained by using conventional uniformly 13 C, 15 N-double labeled proteins. We illustrated this with the case of an 18.2 kDa protein, Escherichia coli peptidyl-prolyl cis-trans isomerase b (EPPIb), and concluded that the combined use of ζ-SAIL Phe and ε-SAIL Tyr would be practically the best choice for protein structural determinations.

Extending Halogen-based Medicinal Chemistry to Proteins: IODO-INSULIN AS A CASE STUDY.

Science.gov (United States)

El Hage, Krystel; Pandyarajan, Vijay; Phillips, Nelson B; Smith, Brian J; Menting, John G; Whittaker, Jonathan; Lawrence, Michael C; Meuwly, Markus; Weiss, Michael A

2016-12-30

Insulin, a protein critical for metabolic homeostasis, provides a classical model for protein design with application to human health. Recent efforts to improve its pharmaceutical formulation demonstrated that iodination of a conserved tyrosine (Tyr B26 ) enhances key properties of a rapid-acting clinical analog. Moreover, the broad utility of halogens in medicinal chemistry has motivated the use of hybrid quantum- and molecular-mechanical methods to study proteins. Here, we (i) undertook quantitative atomistic simulations of 3-[iodo-Tyr B26 ]insulin to predict its structural features, and (ii) tested these predictions by X-ray crystallography. Using an electrostatic model of the modified aromatic ring based on quantum chemistry, the calculations suggested that the analog, as a dimer and hexamer, exhibits subtle differences in aromatic-aromatic interactions at the dimer interface. Aromatic rings (Tyr B16 , Phe B24 , Phe B25 , 3-I-Tyr B26 , and their symmetry-related mates) at this interface adjust to enable packing of the hydrophobic iodine atoms within the core of each monomer. Strikingly, these features were observed in the crystal structure of a 3-[iodo-Tyr B26 ]insulin analog (determined as an R 6 zinc hexamer). Given that residues B24-B30 detach from the core on receptor binding, the environment of 3-I-Tyr B26 in a receptor complex must differ from that in the free hormone. Based on the recent structure of a "micro-receptor" complex, we predict that 3-I-Tyr B26 engages the receptor via directional halogen bonding and halogen-directed hydrogen bonding as follows: favorable electrostatic interactions exploiting, respectively, the halogen's electron-deficient σ-hole and electronegative equatorial band. Inspired by quantum chemistry and molecular dynamics, such "halogen engineering" promises to extend principles of medicinal chemistry to proteins. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Resolution of two native monomeric 90 kDa nitrate reductase active proteins from Shewanella gelidimarina and the sequence of two napA genes

International Nuclear Information System (INIS)

Simpson, Philippa J.L.; McKinzie, Audra A.; Codd, Rachel

2010-01-01

Research highlights: → Two monomeric 90 kDa nitrate reductase active proteins from Shewanella gelidimarina. → Sequence of napA from napEDABC-type operon and napA from NapDAGHB-type operon. → Isolation of NAP as NapA or NapAB correlated with NapA P47E amino acid substitution. -- Abstract: The reduction of nitrate to nitrite in the bacterial periplasm occurs in the 90 kDa NapA subunit of the periplasmic nitrate reductase (NAP) system. Most Shewanella genomes contain two nap operons: napEDABC and napDAGHB, which is an unusual feature of this genus. Two native, monomeric, 90 kDa nitrate reductase active proteins were resolved by hydrophobic interaction chromatography from aerobic cultures of Shewanella gelidimarina replete with reduced nitrogen compounds. The 90 kDa protein obtained in higher yield was characterized as NapA by electronic absorption and electron paramagnetic resonance spectroscopies and was identified by LC/MS/MS and MALDI-TOF/TOF MS as NapA from the napEDABC-type operon. The other 90 kDa protein, which was unstable and produced in low yields, was posited as NapA from the napDAGHB-type operon. Two napA genes have been sequenced from the napEDABC-type and napDAGHB-type operons of S. gelidimarina. Native NAP from S. putrefaciens was resolved as one NapA monomer and one NapAB heterodimer. Two amino acid substitutions in NapA correlated with the isolation of NAP as a NapA monomer or a NapAB heterodimer. The resolution of native, redox-active NapA isoforms in Shewanella provides new insight into the respiratory versatility of this genus, which has implications in bioremediation and the assembly of microbial fuel cells.

Nitration of β-Lactoglobulin but Not of Ovomucoid Enhances Anaphylactic Responses in Food Allergic Mice.

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Susanne C Diesner

Full Text Available We revealed in previous studies that nitration of food proteins reduces the risk of de novo sensitization in a murine food allergy model. In contrast, in situations with preformed specific IgE antibodies, in vitro experiments suggested an increased capacity of effector cell activation by nitrated food proteins.The aim of this study was to investigate the influence of protein nitration on the effector phase of food allergy.BALB/c mice were immunized intraperitoneally (i.p. with the milk allergen β-lactoglobulin (BLG or the egg allergen ovomucoid (OVM, followed by intragastric (i.g. gavages to induce a strong local inflammatory response and allergen-specific antibodies. Subsequently, naïve and allergic mice were intravenously (i.v. challenged with untreated, sham-nitrated or nitrated BLG or OVM. Anaphylaxis was monitored by measuring core body temperature and determination of mouse mast cell protease-1 (mMCP-1 levels in blood.A significant drop of body temperature accompanied with significantly elevated concentrations of the anaphylaxis marker mMCP-1 were only observed in BLG allergic animals challenged with nitrated BLG and not in OVM allergic mice challenged with nitrated OVM. SDS-PAGE and circular dichroism analysis of the differentially modified allergens revealed an effect of nitration on the secondary protein structure exclusively for BLG together with enhanced protein aggregation.Our data suggest that nitration affects differently the food allergens BLG and OVM. In the case of BLG, structural changes favored dimerization possibly explaining the increased anaphylactic reactivity in BLG allergic animals.

Effects of forest fertilization on nitrate and crude protein content in some important reindeer forage species

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Gustaf Åhman

1984-05-01

Full Text Available When forests are fertilized with ammonia nitrate it is possible that grazing reindeer ingest ammonia nitrate by eating grains of fertilizer from the ground or by drinking contaminated water. They can also get nitrate through plants that have absorbed and disposed nitrate. This latter factor is studied in this report. In addition the effect of fertilization on crude protein content in forage plants is investigated. Fertilizing trials were done within two different areas. One was a dry scotch pine forest and the other a humid scotch pine forest. Both were situated 10 to 15 km north west of Lycksele (northern Sweden. Three different rations (75, 150 and 250 kg N/ha of ammonianitrate and one (150 kg N/ha of urea was used. Fertilization was done at two occations, in June and in July. To investigate the effect of fertilization on nitrate and crude protein content in reindeer forage plants, samples were taken of reindeer lichens (Cladina spp., heather {Calluna vulgaris, crowberry (Empetrum spp., cowberry (Vaccinium vitis ideae, blueberry (Vaccinium myrtillus and hair-grass (Deschampsia flexuosa at different times after fertilization. In this trial we could not find any higher degree of contamination of nitrate in lichens. The highest value was 0.013% nitrate-N in dry matter (table 1. Nitrate accumulation was low in shrubs and grass (table 2. The highest value (0.05% was found in heather. The concentrations were definitly below the level that could be considered as injurious to the reindeer. The effect of fertilization on crude protein content in reindeer forage plants was obvious. It was most evident in hair-grass. Four weeks after fertilization with 150 kg N/ha, crude protein content was more than doubled and reached 20% in dry matter (figure 1 and 2. In withered hair-grass in the autumn the effect was very small. One year after fertilization a small rise in crude protein was registered in both grass and shrubs (table 3. Some effect still remained

Protein nitration and nitrosylation by NO-donating aspirin in colon cancer cells: Relevance to its mechanism of action

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Williams, Jennie L.; Ji, Ping; Ouyang, Nengtai; Kopelovich, Levy; Rigas, Basil

2011-01-01

Nitric oxide-donating aspirin (NO-ASA) is a promising agent for cancer prevention. Although studied extensively, its molecular targets and mechanism of action are still unclear. S-nitrosylation of signaling proteins is emerging as an important regulatory mechanism by NO. Here, we examined whether S-nitrosylation of the NF-κB, p53, and Wnt signaling proteins by NO-ASA might explain, in part, its mechanism of action in colon cancer. NO-ASA releases significant amounts of NO detected intracellularly in HCT116 and HT-29 colon cells. Using a modified biotin switch assay we demonstrated that NO-ASA S-nitrosylates the signaling proteins p53, β-catenin, and NF-κB, in colon cancer cells in a time- and concentration-dependent manner. NO-ASA suppresses NF-κB binding to its cognate DNA oligonucleotide, which occurs without changes in the nuclear levels of the NF-κB subunits p65 and p50 and is reversed by dithiothreitol that reduces -S-NO to -SH. In addition to S-nitrosylation, we documented both in vitro and in vivo widespread nitration of tyrosine residues of cellular proteins in response to NO-ASA. Our results suggest that the increased intracellular NO levels following treatment with NO-ASA modulate cell signaling by chemically modifying key protein members of signaling cascades. We speculate that S-nitrosylation and tyrosine nitration are responsible, at least in part, for the inhibitory growth effect of NO-ASA on cancer cell growth and that this may represent a general mechanism of action of NO-releasing agents.

Protein nitration and nitrosylation by NO-donating aspirin in colon cancer cells: Relevance to its mechanism of action

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Williams, Jennie L.; Ji, Ping; Ouyang, Nengtai [Division of Cancer Prevention, Stony Brook University, HSC, T17-080, Stony Brook, NY 11794-8173 (United States); Kopelovich, Levy [Division of Cancer Prevention NCI, NIH, Bethesda, MD (United States); Rigas, Basil, E-mail: basil.rigas@stonybrook.edu [Division of Cancer Prevention, Stony Brook University, HSC, T17-080, Stony Brook, NY 11794-8173 (United States)

2011-06-10

Nitric oxide-donating aspirin (NO-ASA) is a promising agent for cancer prevention. Although studied extensively, its molecular targets and mechanism of action are still unclear. S-nitrosylation of signaling proteins is emerging as an important regulatory mechanism by NO. Here, we examined whether S-nitrosylation of the NF-{kappa}B, p53, and Wnt signaling proteins by NO-ASA might explain, in part, its mechanism of action in colon cancer. NO-ASA releases significant amounts of NO detected intracellularly in HCT116 and HT-29 colon cells. Using a modified biotin switch assay we demonstrated that NO-ASA S-nitrosylates the signaling proteins p53, {beta}-catenin, and NF-{kappa}B, in colon cancer cells in a time- and concentration-dependent manner. NO-ASA suppresses NF-{kappa}B binding to its cognate DNA oligonucleotide, which occurs without changes in the nuclear levels of the NF-{kappa}B subunits p65 and p50 and is reversed by dithiothreitol that reduces -S-NO to -SH. In addition to S-nitrosylation, we documented both in vitro and in vivo widespread nitration of tyrosine residues of cellular proteins in response to NO-ASA. Our results suggest that the increased intracellular NO levels following treatment with NO-ASA modulate cell signaling by chemically modifying key protein members of signaling cascades. We speculate that S-nitrosylation and tyrosine nitration are responsible, at least in part, for the inhibitory growth effect of NO-ASA on cancer cell growth and that this may represent a general mechanism of action of NO-releasing agents.

Engineered P450 biocatalysts show improved activity and regio-promiscuity in aromatic nitration.

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Zuo, Ran; Zhang, Yi; Jiang, Chao; Hackett, John C; Loria, Rosemary; Bruner, Steven D; Ding, Yousong

2017-04-12

Nitroaromatics are among the most important and commonly used chemicals but their production often suffers from multiple unsolved challenges. We have previously described the development of biocatalytic nitration processes driven by an engineered P450 TxtE fusion construct. Herein we report the creation of improved nitration biocatalysts through constructing and characterizing fusion proteins of TxtE with the reductase domain of CYP102A1 (P450BM3, BM3R). The majority of constructs contained variable linker length while one was rationally designed for optimizing protein-protein interactions. Detailed biochemical characterization identified multiple active chimeras that showed improved nitration activity, increased coupling efficiency and higher total turnover numbers compared with TxtE. Substrate promiscuity of the most active chimera was further assessed with a substrate library. Finally, a biocatalytic nitration process was developed to nitrate 4-Me-DL-Trp. The production of both 4-Me-5-NO 2 -L-Trp and 4-Me-7-NO 2 -L-Trp uncovered remarkable regio-promiscuity of nitration biocatalysts.

The Biogenic Amine Tyramine and its Receptor (AmTyr1 in Olfactory Neuropils in the Honey Bee (Apis mellifera Brain

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Irina T. Sinakevitch

2017-10-01

Full Text Available This article describes the cellular sources for tyramine and the cellular targets of tyramine via the Tyramine Receptor 1 (AmTyr1 in the olfactory learning and memory neuropils of the honey bee brain. Clusters of approximately 160 tyramine immunoreactive neurons are the source of tyraminergic fibers with small varicosities in the optic lobes, antennal lobes, lateral protocerebrum, mushroom body (calyces and gamma lobes, tritocerebrum and subesophageal ganglion (SEG. Our tyramine mapping study shows that the primary sources of tyramine in the antennal lobe and calyx of the mushroom body are from at least two Ventral Unpaired Median neurons (VUMmd and VUMmx with cell bodies in the SEG. To reveal AmTyr1 receptors in the brain, we used newly characterized anti-AmTyr1 antibodies. Immunolocalization studies in the antennal lobe with anti-AmTyr1 antibodies showed that the AmTyr1 expression pattern is mostly in the presynaptic sites of olfactory receptor neurons (ORNs. In the mushroom body calyx, anti-AmTyr1 mapped the presynaptic sites of uniglomerular Projection Neurons (PNs located primarily in the microglomeruli of the lip and basal ring calyx area. Release of tyramine/octopamine from VUM (md and mx neurons in the antennal lobe and mushroom body calyx would target AmTyr1 expressed on ORN and uniglomerular PN presynaptic terminals. The presynaptic location of AmTyr1, its structural similarity with vertebrate alpha-2 adrenergic receptors, and previous pharmacological evidence suggests that it has an important role in the presynaptic inhibitory control of neurotransmitter release.

Hydrogen exchange rate of tyrosine hydroxyl groups in proteins as studied by the deuterium isotope effect on C(zeta) chemical shifts.

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Takeda, Mitsuhiro; Jee, Jungoo; Ono, Akira Mei; Terauchi, Tsutomu; Kainosho, Masatsune

2009-12-30

We describe a new NMR method for monitoring the individual hydrogen exchange rates of the hydroxyl groups of tyrosine (Tyr) residues in proteins. The method utilizes (2S,3R)-[beta(2),epsilon(1,2)-(2)H(3);0,alpha,beta,zeta-(13)C(4);(15)N]-Tyr, zeta-SAIL Tyr, to detect and assign the (13)C(zeta) signals of Tyr rings efficiently, either by indirect (1)H-detection through 7-8 Hz (1)H(delta)-(13)C(zeta) spin couplings or by direct (13)C(zeta) observation. A comparison of the (13)C(zeta) chemical shifts of three Tyr residues of an 18.2 kDa protein, EPPIb, dissolved in H(2)O and D(2)O, revealed that all three (13)C(zeta) signals in D(2)O appeared at approximately 0.13 ppm ( approximately 20 Hz at 150.9 MHz) higher than those in H(2)O. In a H(2)O/D(2)O (1:1) mixture, however, one of the three signals for (13)C(zeta) appeared as a single peak at the averaged chemical shifts, and the other two appeared as double peaks at exactly the same chemical shifts in H(2)O and D(2)O, in 50 mM phosphate buffer (pH 6.6) at 40 degrees C. These three peaks were assigned to Tyr-36, Tyr-120, and Tyr-30, from the lower to higher chemical shifts, respectively. The results indicate that the hydroxyl proton of Tyr-120 exchanges faster than a few milliseconds, whereas those of Tyr-30 and Tyr-36 exchange more slowly. The exchange rate of the Tyr-30 hydroxyl proton, k(ex), under these conditions was determined by (13)C NMR exchange spectroscopy (EXSY) to be 9.2 +/- 1.1 s(-1). The Tyr-36 hydroxyl proton, however, exchanges too slowly to be determined by EXSY. These profound differences among the hydroxyl proton exchange rates are closely related to their relative solvent accessibility and the hydrogen bonds associated with the Tyr hydroxyl groups in proteins.

Discovery of nitrate-CPK-NLP signalling in central nutrient-growth networks

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Liu, Kun-hsiang; Niu, Yajie; Konishi, Mineko; Wu, Yue; Du, Hao; Sun Chung, Hoo; Li, Lei; Boudsocq, Marie; McCormack, Matthew; Maekawa, Shugo; Ishida, Tetsuya; Zhang, Chao; Shokat, Kevan; Yanagisawa, Shuichi; Sheen, Jen

2018-01-01

Nutrient signalling integrates and coordinates gene expression, metabolism and growth. However, its primary molecular mechanisms remain incompletely understood in plants and animals. Here we report novel Ca2+ signalling triggered by nitrate with live imaging of an ultrasensitive biosensor in Arabidopsis leaves and roots. A nitrate-sensitized and targeted functional genomic screen identifies subgroup III Ca2+-sensor protein kinases (CPKs) as master regulators orchestrating primary nitrate responses. A chemical switch with the engineered CPK10(M141G) kinase enables conditional analyses of cpk10,30,32 to define comprehensive nitrate-associated regulatory and developmental programs, circumventing embryo lethality. Nitrate-CPK signalling phosphorylates conserved NIN-LIKE PROTEIN (NLP) transcription factors (TFs) to specify reprogramming of gene sets for downstream TFs, transporters, N-assimilation, C/N-metabolism, redox, signalling, hormones, and proliferation. Conditional cpk10,30,32 and nlp7 similarly impair nitrate-stimulated system-wide shoot growth and root establishment. The nutrient-coupled Ca2+ signalling network integrates transcriptome and cellular metabolism with shoot-root coordination and developmental plasticity in shaping organ biomass and architecture. PMID:28489820

Aminoethyl nitrate – the novel super nitrate?

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Bauersachs, Johann

2009-01-01

Long-term use of most organic nitrates is limited by development of tolerance, induction of oxidative stress and endothelial dysfunction. In this issue of the BJP, Schuhmacher et al. characterized a novel class of organic nitrates with amino moieties (aminoalkyl nitrates). Aminoethyl nitrate was identified as a novel organic mononitrate with high potency but devoid of induction of mitochondrial oxidative stress. Cross-tolerance to nitroglycerin or the endothelium-dependent agonist acetylcholine after in vivo treatment was not observed. Like all nitrates, aminoethyl nitrate induced vasorelaxation by activation of soluble guanylate cyclase. Thus, in contrast to the prevailing view, high potency in an organic nitrate is not necessarily accompanied by induction of oxidative stress or endothelial dysfunction. This work from Daiber's group is an important step forward in the understanding of nitrate bioactivation, tolerance phenomena and towards the development of better organic nitrates for clinical use. PMID:19732062

Study of Nitrate Stress in Desulfovibrio vulgaris Hildenborough Using iTRAQ Proteomics

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Redding, A.M.; Mukhopadhyay, A.; Joyner, D.; Hazen, T.C.; Keasling, J.D.

2006-10-12

The response of Desulfovibrio vulgaris Hildenborough (DvH),a sulphate-reducing bacterium, to nitrate stress was examined usingquantitative proteomic analysis. DvH was stressed with 105 m M sodiumnitrate(NaNO3), a level that caused a 50 percent inhibition in growth.The protein profile of stressed cells was compared with that of cellsgrown in the absence of nitrate using the iTRAQ peptide labellingstrategy and tandem liquid chromatography separation coupled with massspectrometry (quadrupoletime-of-flight) detection. A total of 737 uniqueproteins were identified by two or more peptides, representing 22 percentof the total DvH proteome and spanning every functional category. Theresults indicate that this was a mild stress, as proteins involved incentral metabolism and the sulphate reduction pathway were unperturbed.Proteins involved in the nitrate reduction pathway increased. Increasesseen in transport systems for proline, glycine^ betaineandglutamateindicate that the NaNO3 exposure led to both salt stress and nitratestress.Up-regulation observed in oxidative stress response proteins (Rbr,RbO, etc.) and a large number of ABC transport systems as well as in iron^ sulphur -cluster-containing proteins, however, appear to be specific tonitrate exposure. Finally, a number of hypothetical proteins were amongthe most significant changers, indicating that there may be unknownmechanisms initiated upon nitrate stress in DvH.

Synthesis of (/sup 3/H-Tyr/sup B26/)-human insulin by enzymic semisynthesis

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Haensicke, A; Kaufmann, K D; Beyermann, M; Oehlke, J; Kertscher, U; Bienert, M; Niedrich, H; Mittag, E; Bespalova, Zh D; Titov, M I

1988-11-01

A procedure is described for tritium labelling of human insulin in position Tyr/sup B26/ by means of trypsin catalyzed condensation of DiBoc-DOI with (N/sup /epsilon//-Boc, /sup 3/H-Tyr/sup B26/)-IOP, subsequent deprotection and purification by HPLC. The tritium labelling of the octapeptide was accomplished by dehalotritiation of the corresponding Dit/sup B26/-octapeptide which was obtained both by iodination of N/sup /epsilon//-Boc-IOP and by total synthesis. (author). 2 figs., 1 tab., 17 refs.

Tyr721 regulates specific binding of the CSF-1 receptor kinase insert to PI 3'-kinase SH2 domains: a model for SH2-mediated receptor-target interactions.

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Reedijk, M; Liu, X; van der Geer, P; Letwin, K; Waterfield, M D; Hunter, T; Pawson, T

1992-01-01

Efficient binding of active phosphatidylinositol (PI) 3'-kinase to the autophosphorylated macrophage colony stimulating factor receptor (CSF-1R) requires the noncatalytic kinase insert (KI) region of the receptor. To test whether this region could function independently to bind PI 3'-kinase, the isolated CSF-1R KI was expressed in Escherichia coli, and was inducibly phosphorylated on tyrosine. The tyrosine phosphorylated form of the CSF-1R KI bound PI 3'-kinase in vitro, whereas the unphosphorylated form had no binding activity. The p85 alpha subunit of PI 3'-kinase contains two Src homology (SH)2 domains, which are implicated in the interactions of signalling proteins with activated receptors. Bacterially expressed p85 alpha SH2 domains complexed in vitro with the tyrosine phosphorylated CSF-1R KI. Binding of the CSF-1R KI to PI 3'-kinase activity, and to the p85 alpha SH2 domains, required phosphorylation of Tyr721 within the KI domain, but was independent of phosphorylation at Tyr697 and Tyr706. Tyr721 was also critical for the association of activated CSF-1R with PI 3'-kinase in mammalian cells. Complex formation between the CSF-1R and PI 3'-kinase can therefore be reconstructed in vitro in a specific interaction involving the phosphorylated receptor KI and the SH2 domains of p85 alpha. Images PMID:1314163

A hot-spot mutation in CDC42 (p.Tyr64Cys) and novel phenotypes in the third patient with Takenouchi-Kosaki syndrome.

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Motokawa, Midori; Watanabe, Satoshi; Nakatomi, Akiko; Kondoh, Tatsuro; Matsumoto, Tadashi; Morifuji, Kanako; Sawada, Hirotake; Nishimura, Toyoki; Nunoi, Hiroyuki; Yoshiura, Koh-Ichiro; Moriuchi, Hiroyuki; Dateki, Sumito

2018-03-01

Takenouchi-Kosaki syndrome (TKS) is a congenital malformation syndrome characterized by severe developmental delay, macrothrombocytopenia, camptodactyly, sensorineural hearing loss, and dysmorphic facial features. Recently, a heterozygous de novo mutation (p.Tyr64Cys) in the CDC42 gene, which encodes a key small GTP-binding protein of the Rho-subfamily, was identified in two unrelated patients with TKS. We herein report a third patient with TKS who had the same heterozygous CDC42 mutation. The phenotype of the patient was very similar to those of the two previously reported patients with TKS; however, she also demonstrated novel clinical manifestations, such as congenital hypothyroidism and immunological disturbance. Thus, despite the heterozygous mutation of CDC42 (p.Tyr64Cys) likely being a hot-spot mutation for TKS, its phenotype may be variable. Further studies and the accumulation of patients with CDC42 mutations are needed to clarify the phenotype in patients with TKS and the pathophysiological roles of the CDC42 mutation.

Amifostine protects rat kidneys during peptide receptor radionuclide therapy with [177Lu-DOTA0,Tyr3]octreotate

International Nuclear Information System (INIS)

Rolleman, Edgar J.; Forrer, Flavio; Bernard, Bert; Bijster, Magda; Valkema, Roelf; Krenning, Eric P.; Jong, Marion de; Vermeij, Marcel

2007-01-01

In peptide receptor radionuclide therapy (PRRT) using radiolabelled somatostatin analogues, the kidneys are the major dose-limiting organs, because of tubular reabsorption and retention of radioactivity. Preventing renal uptake or toxicity will allow for higher tumour radiation doses. We tested the cytoprotective drug amifostine, which selectively protects healthy tissue during chemo- and radiotherapy, for its renoprotective capacities after PRRT with high-dose [ 177 Lu-DOTA 0 ,Tyr 3 ]octreotate. Male Lewis rats were injected with 278 or 555 MBq [ 177 Lu-DOTA 0 ,Tyr 3 ]octreotate to create renal damage and were followed up for 130 days. For renoprotection, rats received either amifostine or co-injection with lysine. Kidneys, blood and urine were collected for toxicity measurements. At 130 days after PRRT, a single-photon emission computed tomography (SPECT) scan was performed to quantify tubular uptake of 99m Tc-dimercaptosuccinic acid (DMSA), a measure of tubular function. Treatment with 555 MBq [ 177 Lu-DOTA 0 ,Tyr 3 ]octreotate resulted in body weight loss, elevated creatinine and proteinuria. Amifostine and lysine treatment significantly prevented this rise in creatinine and the level of proteinuria, but did not improve the histological damage. In contrast, after 278 MBq [ 177 Lu-DOTA 0 ,Tyr 3 ]octreotate, creatinine values were slightly, but not significantly, elevated compared with the control rats. Proteinuria and histological damage were different from controls and were significantly improved by amifostine treatment. Quantification of 99m Tc-DMSA SPECT scintigrams at 130 days after [ 177 Lu-DOTA 0 ,Tyr 3 ]octreotate therapy correlated well with 1/creatinine (r 2 = 0.772, p 177 Lu-DOTA 0 ,Tyr 3 ]octreotate. Besides lysine, amifostine might be used in clinical PRRT as well as to maximise anti-tumour efficacy. (orig.)

Genetic control of nitrate assimilation in Klebsiella oxytoca. Final technical report; FINAL

International Nuclear Information System (INIS)

Stewart, Valley J.

2001-01-01

Some microorganisms can use nitrate as the sole source of nitrogen for biosynthesis. This project focused on the bacterium Klebsiella oxytoca, an enterobacterium found in soil and water. Mutagenesis and molecular cloning identified the nasFEDCBA operon encoding enzymes for the uptake and reduction of nitrate and nitrite to ammonium, and the adjacent nasR regulatory gene. Analysis of nasF operon expression revealed that transcription is activated by the Ntr (general nitrogen regulation ) system in response to nitrogen limitation. Transcription antitermination control in response to nitrate and nitrite is mediated by the NasR protein. Additional work established that the NasR protein is an RNA-binding protein that interacts with nasF operon leader RNA to control transcription readthrough

Tumoral fibrosis effect on the radiation absorbed dose of 177Lu-Tyr3-octreotate-gold nanoparticles and 177Lu-Tyr3-octreotate radiopharmaceuticals

International Nuclear Information System (INIS)

Zambrano R, O. D.

2015-01-01

In this work was comparatively evaluated the effect of tumoral fibrosis in the radiation absorbed dose of the radiopharmaceutical 177 Lu-Tyr 3 -octreotate with and without gold nanoparticles. For this, was used an experimental array of tumoral fibrosis and computer models based on Monte Carlo calculations to simulate tumoral micro environments without fibrosis and with fibrosis. The computer simulation code Penelope (Penetration Energy Loss of Positron and Electrons) and MCNP (Monte Carlo N-particle Transport Code System) which are based on the Monte Carlo methodology were used to create the computer models for the simulation of the transport of particles (emitted by 177 Lu) in the micro environments (without fibrosis and with fibrosis) with the purpose of calculating the radiation absorbed dose in the interstitial space and in the nucleus of cancer cells. The first computational model consisted of multiple concentric spheres (as onion shells) with the radioactive source homogeneously distributed in the shell between 5 and 10 μm in diameter which represents the internalization of the radioactive source into the cell cytoplasm as it occurs in target specific radiotherapy. The concentric spheres were useful to calculate the radiation absorbed dose in depth in the models without fibrosis and with fibrosis. Furthermore, there were constructed other computer models using two different codes that simulate the transport of radiation (Penelope and MCNP). These models consist of seven spheres that represent cancer cells (HeLa cells) of 10 μm in diameter and each one of them contain another smaller sphere in the center that represents the cell nucleus. A comparison was done of the radiation absorbed dose in the nucleus of the cells, calculated with both codes, Penelope and MCNP. The radioactive source ( 177 Lu) used for the simulations was given to the codes by means of a convoluted spectrum of the most important beta particles (high percentage emission). To this spectrum

Induction of nitrate transport in maize roots, and kinetics of influx, measured with nitrogen-13

International Nuclear Information System (INIS)

Hole, D.J.; Drew, M.C.; Emran, A.M.; Fares, Y.

1990-01-01

Unlike phosphate or potassium transport, uptake of nitrate by roots is induced, in part, by contact with the substrate ion. Plasmalemma influx of 13 N-labeled nitrate in maize roots was studied in relation to induction of the uptake system, and the influence of short-term N starvation. Maize (Zea mays) roots not previously exposed to nitrate had a constitutive transport system (state 1), but influx increased 250% during six hours of contact with 100 micromolar nitrate, by which time the transport mechanism appeared to be fully synthesized (state 2). A three-day period of N starvation prior to induction and measurement of nitrate influx resulted in a greater capacity to transport nitrate than in unstarved controls, but this was fully expressed only if roots were kept in contact with nitrate for the six hours needed for full induction (state 2E). A kinetic analysis indicated a 160% increase in maximum influx in N-starved, induced roots with a small decrease in K m . The inducible component to nitrate influx was induced only by contact with nitrate. Full expression of the nitrate inducible transport system was dependent upon mRNA synthesis. An inhibitor of cytoplasmic protein synthesis (cycloheximide) eliminated the formation of the transport system while inhibition by chloramphenicol of mitochondrial- or plastid-coded protein synthesis had no effect. Poisoning of membrane-bound proteins effectively disabled both the constitutive and induced transport systems

Immunological comparison of the NADH:nitrate reductase from different cucumber tissues

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Jolanta Marciniak

2014-01-01

Full Text Available Soluble nitrate reductase from cucumber roots (Cucumis sativus L. was isolated and purified with blue-Sepharose 4B. Specific antibodies against the NR protein were raised by immunization of a goat. Using polyclonal antibodies anti-NR properties of the nitrate reductase from various cucumber tissues were examined. Experiments showed difference in immuno-logical properties of nitrate reductase (NR from cotyledon roots and leaves.

Toxic effects of lead and nickel nitrate on rat liver chromatin components.

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Rabbani-Chadegani Iii, Azra; Fani, Nesa; Abdossamadi, Sayeh; Shahmir, Nosrat

2011-01-01

The biological activity of heavy metals is related to their physicochemical interaction with biological receptors. In the present study, the effect of low concentrations of nickel nitrate and lead nitrate (lead nitrate to chromatin compared to nickel nitrate. Also, the binding affinity of lead nitrate to histone proteins free in solution was higher than nickel. On the basis of the results, it is concluded that lead reacts with chromatin components even at very low concentrations and induce chromatin aggregation through histone-DNA cross-links. Whereas, nickel nitrate is less effective on chromatin at low concentrations, suggesting higher toxicity of lead nitrate on chromatin compared to nickel. Copyright © 2010 Wiley Periodicals, Inc.

Cathepsin X Cleaves Profilin 1 C-Terminal Tyr139 and Influences Clathrin-Mediated Endocytosis.

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Urša Pečar Fonović

Full Text Available Cathepsin X, a cysteine carboxypeptidase, is upregulated in several types of cancer. Its molecular target in tumor cells is profilin 1, a known tumor suppressor and regulator of actin cytoskeleton dynamics. Cathepsin X cleaves off the C-terminal Tyr139 of profilin 1, affecting binding of poly-L-proline ligands and, consequently, tumor cell migration and invasion. Profilin 1 with mutations at the C-terminus, transiently expressed in prostate cancer cells PC-3, showed that Tyr139 is important for proper function of profilin 1 as a tumor suppressor. Cleaving off Tyr139 prevents the binding of clathrin, a poly-L-proline ligand involved in endocytosis. More profilin 1-clathrin complexes were present in PC-3 cells when cathepsin X was inhibited by its specific inhibitor AMS36 or silenced by siRNA. As a consequence, the endocytosis of FITC-labeled dextran and transferrin conjugate was significantly increased. These results constitute the first report of the regulation of clathrin-mediated endocytosis in tumor cells through proteolytic processing of profilin 1.

Cathepsin X Cleaves Profilin 1 C-Terminal Tyr139 and Influences Clathrin-Mediated Endocytosis

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Pečar Fonović, Urša; Kos, Janko

2015-01-01

Cathepsin X, a cysteine carboxypeptidase, is upregulated in several types of cancer. Its molecular target in tumor cells is profilin 1, a known tumor suppressor and regulator of actin cytoskeleton dynamics. Cathepsin X cleaves off the C-terminal Tyr139 of profilin 1, affecting binding of poly-L-proline ligands and, consequently, tumor cell migration and invasion. Profilin 1 with mutations at the C-terminus, transiently expressed in prostate cancer cells PC-3, showed that Tyr139 is important for proper function of profilin 1 as a tumor suppressor. Cleaving off Tyr139 prevents the binding of clathrin, a poly-L-proline ligand involved in endocytosis. More profilin 1—clathrin complexes were present in PC-3 cells when cathepsin X was inhibited by its specific inhibitor AMS36 or silenced by siRNA. As a consequence, the endocytosis of FITC-labeled dextran and transferrin conjugate was significantly increased. These results constitute the first report of the regulation of clathrin-mediated endocytosis in tumor cells through proteolytic processing of profilin 1. PMID:26325675

Phase extraction equilibria in systems rare earth (3) nitrates-ammonium nitrate-water-trialkylmethylammonium nitrate

International Nuclear Information System (INIS)

Pyartman, A.K.; Kopyrin, A.A.; Puzikov, E.A.

1995-01-01

The distribution of rare earth metals (3) between aqueous and organic phases in the systems rare earth metal (3) (praseodymium-lutetium (3), yttrium (3)) nitrate-ammonium nitrate-water-trialkylmethylammonium (kerosene diluent nitrate has been studied. It is shown that in organic phase di- and trisolvates of metals (3) with tralkylmethylammonium nitrate are formed. The influence of concentration of rare earth metal (3) nitrate and ammonium nitrate on the values of extraction concentrational constants has been ascertained: they decrease with increase in the ordinal number of lanthanide (3). 11 refs., 4 figs. 1 tab

21 CFR 181.33 - Sodium nitrate and potassium nitrate.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Sodium nitrate and potassium nitrate. 181.33...-Sanctioned Food Ingredients § 181.33 Sodium nitrate and potassium nitrate. Sodium nitrate and potassium nitrate are subject to prior sanctions issued by the U.S. Department of Agriculture for use as sources of...

Systems of cerium(3) nitrate-dimethyl amine nitrate-water and cerium(3) nitrate-dimethyl amine nitrate-water

International Nuclear Information System (INIS)

Mininkov, N.E.; Zhuravlev, E.F.

1976-01-01

Solubility of solid phases in the systems cerium(3)nitrate-water-dimethyl amine nitrate and cerium(3)nitrate-water-dimethyl amine nitrate has been st ed by the method of isothermal sections at 25 and 50 deo. C. It has been shown that one anhydrous compound is formed in each system with a ratio of cerium(3) nitrate to amine nitrate 1:5. The compounds formed in the systems have been separated from the corresponding solutions and studied by microcrystalloscopic, X-ray phase, thermal and infrared spectroscopic methods. On the basis of spectroscopic studies the following formula has been assigned to the compound: [(CH 3 ) 2 NH 2 + ] 5 x[Ce(NO 3 ) 8 ]. The thermal analysis of the compound has shown that its melting point is 106 deg C. The solubility isotherms in the system Ce(NO 3 ) 3 -H 2 O-(C 2 H 5 ) 2 NHxHNO 3 consist of three branches which intersect in two eutonic points

CD6 and Linker of Activated T Cells are Potential Interaction Partners for T Cell-Specific Adaptor Protein.

Science.gov (United States)

Hem, C D; Ekornhol, M; Granum, S; Sundvold-Gjerstad, V; Spurkland, A

2017-02-01

The T cell-specific adaptor protein (TSAd) contains several protein interaction domains, and is merging as a modulator of T cell activation. Several interaction partners for the TSAd proline-rich region and phosphotyrosines have been identified, including the Src and Tec family kinases lymphocyte-specific protein tyrosine kinase and interleukin 2-inducible T cell kinase. Via its Src homology 2 (SH2) domain, TSAd may thus function as a link between these enzymes and other signalling molecules. However, few binding partners to the TSAd SH2 domain in T cells are hitherto known. Through the use of in silico ligand prediction, peptide spot arrays, pull-down and immunoprecipitation experiments, we here report novel interactions between the TSAd SH2 domain and CD6 phosphotyrosine (pTyr) 629 and linker of activated T cells (LAT) pTyr 171 , pTyr 191 and pTyr 226 . © 2016 The Foundation for the Scandinavian Journal of Immunology.

Nitrate biosensors and biological methods for nitrate determination.

Science.gov (United States)

Sohail, Manzar; Adeloju, Samuel B

2016-06-01

The inorganic nitrate (NO3‾) anion is present under a variety of both natural and artificial environmental conditions. Nitrate is ubiquitous within the environment, food, industrial and physiological systems and is mostly present as hydrated anion of a corresponding dissolved salt. Due to the significant environmental and toxicological effects of nitrate, its determination and monitoring in environmental and industrial waters are often necessary. A wide range of analytical techniques are available for nitrate determination in various sample matrices. This review discusses biosensors available for nitrate determination using the enzyme nitrate reductase (NaR). We conclude that nitrate determination using biosensors is an excellent non-toxic alternative to all other available analytical methods. Over the last fifteen years biosensing technology for nitrate analysis has progressed very well, however, there is a need to expedite the development of nitrate biosensors as a suitable alternative to non-enzymatic techniques through the use of different polymers, nanostructures, mediators and strategies to overcome oxygen interference. Copyright © 2016 Elsevier B.V. All rights reserved.

Purification of nitrate reductase from Nicotiana plumbaginifolia by affinity chromatography using 5'AMP-sepharose and monoclonal antibodies.

Science.gov (United States)

Moureaux, T; Leydecker, M T; Meyer, C

1989-02-15

Nitrate reductase was purified from leaves of Nicotiana plumbaginifolia using either 5'AMP-Sepharose chromatography or two steps of immunoaffinity chromatography involving monoclonal antibodies directed against nitrate reductase from maize and against ribulose-1,5-bisphosphate carboxylase from N. plumbaginifolia. Nitrate reductase obtained by the first method was purified 1000-fold to a specific activity of 9 units/mg protein. The second method produced an homogenous enzyme, purified 21,000-fold to a specific activity of 80 units/mg protein. SDS/PAGE of nitrate reductase always resulted in two bands of 107 and 99.5 kDa. The 107-kDa band was the nitrate reductase subunit of N. plumbaginifolia; the smaller one of 99.5 kDa is thought, as commonly reported, to result from proteolysis of the larger protein. The molecular mass of 107 kDa is close to the values calculated from the coding sequences of the two nitrate reductase genes recently cloned from tobacco (Nicotiana tabacum cv Xanthi).

Neurotensin analogs [D-TYR11] and [D-PHE11]neurotensin resist degradation by brain peptidases in vitro and in vivo

International Nuclear Information System (INIS)

Checler, F.; Vincent, J.P.; Kitabgi, P.

1983-01-01

The present study was designed to compare the susceptibility of neurotensin (NT), [ 3 H]NT, [D-Tyr11]NT and [D-Phe11]NT to degradation by 1) rat brain synaptic membranes in vitro and 2) after i.c.v. administration in the rat in vivo. Degradation was assessed by purifying the peptides using reverse phase high-performance liquid chromatography and by measuring the amount of radioactive or absorbing (OD 230) material under each peptide peak. In contrast to NT, [D-Tyr11]NT and [D-Phe11]NT were resistant to degradation by brain synaptic peptidases in vitro. Furthermore, NT was rapidly metabolized in brain tissues after i.c.v. administration, whereas [D-Tyr11]NT was metabolically stable. The present data confirm the central role of NT residue Tyr11 in the mechanisms of NT inactivation by brain synaptic peptidases. They account for the higher in vivo potency of [D-Tyr11]NT as compared with its in vitro potency. Finally, they explain, at least in part, the need to administer large doses of NT in the brain in order to observe neurobehavioral and neuropharmacological effects

Formation of Shc-Grb2 complexes is necessary to induce neoplastic transformation by overexpression of Shc proteins

DEFF Research Database (Denmark)

Salcini, A E; McGlade, J; Pelicci, G

1994-01-01

The mammalian SHC gene encodes three overlapping proteins which all contain a carboxy-terminal SH2 domain. Shc proteins are phosphorylated on tyrosine by a variety of receptor and cytoplasmic tyrosine kinases. Phosphorylated Shc proteins form a complex with the SH2-SH3 containing Grb2 protein which...... of Grb2 to Shc proteins requires phosphorylation of Shc at Tyr317, which lies within the high affinity binding motif for the Grb2 SH2 domain, pYVNV, where Asn at the +2 position is crucial for complex formation. In vivo, Tyr317 is the major, but not the only, site for Shc phosphorylation, and is the sole...... aminoterminal deletion, but retain the Tyr317 site and the SH2 domain conserve the capacity to be phosphorylated, to bind to Grb2 and to induce cell transformation. These data indicate that the formation of the Shc-Grb2 complex is a crucial event in the transformation induced by overexpression of Shc...

Identification of tyrosine-phosphorylated proteins associated with metastasis and functional analysis of FER in human hepatocellular carcinoma cells

International Nuclear Information System (INIS)

Li, Haiyu; Ren, Zhenggang; Kang, Xiaonan; Zhang, Lan; Li, Xuefei; Wang, Yan; Xue, Tongchun; Shen, Yuefang; Liu, Yinkun

2009-01-01

Aberrant activity of tyrosine-phosphorylated proteins is commonly associated with HCC metastasis. Cell signaling events driven by these proteins are implicated in numerous processes that alter cancer cell behavior. Exploring the activities and signaling pathways of these proteins in HCC metastasis may help in identifying new candidate molecules for HCC-targeted therapy. Hep3B (a nonmetastatic HCC cell line) and MHCC97H (a highly metastatic HCC cell line) were used in this study, and the tyrosine-phosphorylated proteins expressed in these cell lines were profiled by a phosphoproteomics technique based on LC-MS/MS. Protein-protein interaction and functional clustering analyses were performed to determine the activities of the identified proteins and the signaling pathways closely related to HCC metastasis. In both cell lines, a total of 247 phosphotyrosine (pTyr) proteins containing 281 pTyr sites were identified without any stimulation. The involvement of almost 30% of these in liver or liver cancer has not been reported previously. Biological process clustering analysis indicated that pTyr proteins involved in cell motility, migration, protein autophosphorylation, cell-cell communication, and antiapoptosis functions were overexpressed during metastasis. Pathway clustering analysis revealed that signaling pathways such as those involved in EGFR signaling, cytokine- and chemokine-mediated signal transduction, and the PI3K and JAK-STAT cascades were significantly activated during HCC metastasis. Moreover, noncanonical regulation of the JNK cascade might also provide new targets for HCC metastasis. After comparing the pTyr proteins that were differentially expressed during HCC cell metastasis, we selected FER, a nonreceptor tyrosine kinase, and validated its role in terms of both expression and function. The data confirmed that FER might play a critical role in the invasion and metastasis of HCC. The identification of pTyr proteins and signaling pathways associated

Over-expression of a tobacco nitrate reductase gene in wheat (Triticum aestivum L.) increases seed protein content and weight without augmenting nitrogen supplying.

Science.gov (United States)

Zhao, Xiao-Qiang; Nie, Xuan-Li; Xiao, Xing-Guo

2013-01-01

Heavy nitrogen (N) application to gain higher yield of wheat (Triticum aestivum L.) resulted in increased production cost and environment pollution. How to diminish the N supply without losing yield and/or quality remains a challenge. To meet the challenge, we integrated and expressed a tobacco nitrate reductase gene (NR) in transgenic wheat. The 35S-NR gene was transferred into two winter cultivars, "Nongda146" and "Jimai6358", by Agrobacterium-mediation. Over-expression of the transgene remarkably enhanced T1 foliar NR activity and significantly augmented T2 seed protein content and 1000-grain weight in 63.8% and 68.1% of T1 offspring (total 67 individuals analyzed), respectively. Our results suggest that constitutive expression of foreign nitrate reductase gene(s) in wheat might improve nitrogen use efficiency and thus make it possible to increase seed protein content and weight without augmenting N supplying.

Over-expression of a tobacco nitrate reductase gene in wheat (Triticum aestivum L. increases seed protein content and weight without augmenting nitrogen supplying.

Directory of Open Access Journals (Sweden)

Xiao-Qiang Zhao

Full Text Available Heavy nitrogen (N application to gain higher yield of wheat (Triticum aestivum L. resulted in increased production cost and environment pollution. How to diminish the N supply without losing yield and/or quality remains a challenge. To meet the challenge, we integrated and expressed a tobacco nitrate reductase gene (NR in transgenic wheat. The 35S-NR gene was transferred into two winter cultivars, "Nongda146" and "Jimai6358", by Agrobacterium-mediation. Over-expression of the transgene remarkably enhanced T1 foliar NR activity and significantly augmented T2 seed protein content and 1000-grain weight in 63.8% and 68.1% of T1 offspring (total 67 individuals analyzed, respectively. Our results suggest that constitutive expression of foreign nitrate reductase gene(s in wheat might improve nitrogen use efficiency and thus make it possible to increase seed protein content and weight without augmenting N supplying.

NarK is a nitrite-extrusion system involved in anaerobic nitrate respiration by Escherichia coli

NARCIS (Netherlands)

Rowe, John J.; Ubbink-Kok, Trees; Molenaar, Douwe; Konings, Wilhelmus; Driessen, Arnold J.M.

Escherichia coli can use nitrate as a terminal electron acceptor for anaerobic respiration. A polytopic membrane protein, termed NarK, has been implicated in nitrate uptake and nitrite excretion and is thought to function as a nitrate/nitrite antiporter. The longest-lived radioactive isotope of

Radical transfer between proteins: role of tyrosine, tryptophan and protein peroxyl radicals

International Nuclear Information System (INIS)

Irwin, J.A.; Ostdal, H.; Davies, M.J.

1998-01-01

Reaction of the Fe(III) forms of the heme proteins myoglobin (Mb) and horseradish peroxidase (HRP) with H 2 O 2 gives rise to high-oxidation-state heme-derived species which can be described as a Fe(IV)-oxo porphyrin radical-cation ('Compound 1'). In the case of Mb, the Fe(IV)-oxo porphyrin radical-cation undergoes rapid electron transfer with the surrounding protein to give protein (globin)-derived radicals and an Fe(lV)-oxo species ('Compound 2'). The globin-derived radicals have been shown to be located at two (or more) sites: Tyr-103 or Trp-14, with the latter radical known to react with oxygen to give a Trp-derived peroxyl radical (Mb-Trp-OO*). With HRP, the Fe(lV)-oxo porphyrin radical-cation carries out two successive one-electron oxidation reactions at the exposed heme edge to give firstly 'Compound 2' [the Fe(lV)oxo species] and then the resting Fe(III) state of the enzyme. n this study we have investigated whether the Trp-14 peroxyl radical from Mb and the Compound 1 and 2 species from HRP (in the absence and presence of free Tyr) can oxidise amino acids, peptides and proteins. Such reactions constitute intermolecular protein-to-protein radical transfer reactions and hence protein chain-oxidation. We have also examined whether these oxidants react with antioxidants. Reaction of these heme-protein derived oxidants with amino acids, proteins and antioxidants has been carried out at room temperature for defined periods of time before freeze-quenching to 77K to halt reaction. The radical species present in the reaction system at the time of freezing were subsequently examined by EPR spectroscopy at 77K. Three free amino acids, Tyr, Trp and Cys (with Cys the least efficient) have been shown to react rapidly with Mb-Trp-OO*, as evidenced by the loss of the characteristic EPR features of Mb-Trp-OO* on inclusion of increasing concentrations of the amino acids. All other amino acids are much less reactive. Evidence has also been obtained for (inefficient) hydrogen

Density and electrical conductivity of molten salts. Comparative study of binary mixtures of alkali nitrates with silver nitrate and with thallium nitrate; Densite et conductibilite de sels fondus. Etude comparative des melanges binaires nitrates alcalins-nitrate d'argent et nitrates alcalins-nitrate de thallium

Energy Technology Data Exchange (ETDEWEB)

Brillant, S [Commissariat a l' Energie Atomique Saclay (France). Centre d' Etudes Nucleaires

1967-10-01

The choice of methods and the number of measurements made enable us to give results on the density and electrical conductivity of molten binary mixtures, alkali nitrate and silver nitrate, and alkali nitrate and thallium nitrate, in the form of equations. The deviations from linearity of the volume and the molar conductivity are determined by calculating the corresponding excess values whose variations are analyzed as a function of the Tobolsky parameter. The absence of any relationship in the sign of the entropy and the excess volume is justified. It is shown that the silver and thallium nitrates, in contrast to the thermodynamic properties, behave as the alkali nitrates in so far as the excess conductivity is concerned. This result is confirmed by the study of changes in the activation enthalpy for the partial molar conductivity; this study also shows the particular behaviour of lithium nitrate. (author) [French] Le choix des methodes et le nombre de mesures effectuees nous permettent de donner les resultats de densite et de conductibilite electrique des melanges fondus binaires nitrate alcalin-nitrate d'argent et nitrate alcalin-nitrate de thallium sous forme d'equations. Les ecarts a la linearite des isothermes de volume et de conductibilite molaire sont precises en calculant les grandeurs d'exces correspondantes dont les variations sont analysees en fonction du parametre de Tobolsky. Nous justifions l'absence de relation de signe entre l'entropie et le volume d'exces. Nous montrons que les nitrates d'argent et de thallium, vis-a-vis de la conductibilite d'exces, contrairement aux proprietes thermodynamiques, se conduisent comme les nitrates alcalins. Ce resultat est confirme par l'etude des variations des enthalpies d'activation de conductibilite partielle molaire qui met d'autre part en evidence le comportement particulier du nitrate de lithium. (auteur)

A bioinformatics-based overview of protein Lys-Ne-acetylation

Science.gov (United States)

Among posttranslational modifications, there are some conceptual similarities between Lys-N'-acetylation and Ser/Thr/Tyr O-phosphorylation. Herein we present a bioinformatics-based overview of reversible protein Lys-acetylation, including some comparisons with reversible protein phosphorylation. T...

Structure of the red fluorescent protein from a lancelet (Branchiostoma lanceolatum): a novel GYG chromophore covalently bound to a nearby tyrosine

Energy Technology Data Exchange (ETDEWEB)

Pletnev, Vladimir Z., E-mail: vzpletnev@gmail.com; Pletneva, Nadya V.; Lukyanov, Konstantin A.; Souslova, Ekaterina A.; Fradkov, Arkady F.; Chudakov, Dmitry M.; Chepurnykh, Tatyana; Yampolsky, Ilia V. [Russian Academy of Sciences, Moscow (Russian Federation); Wlodawer, Alexander [National Cancer Institute, Frederick, MD 21702 (United States); Dauter, Zbigniew [National Cancer Institute, Argonne, IL 60439 (United States); Pletnev, Sergei, E-mail: vzpletnev@gmail.com [National Cancer Institute, Argonne, IL 60439 (United States); SAIC-Frederick, Argonne, IL 60439 (United States); Russian Academy of Sciences, Moscow (Russian Federation)

2013-09-01

The crystal structure of the novel red emitting fluorescent protein from lancelet Branchiostoma lanceolatum (Chordata) revealed an unusual five residues cyclic unit comprising Gly58-Tyr59-Gly60 chromophore, the following Phe61 and Tyr62 covalently bound to chromophore Tyr59. A key property of proteins of the green fluorescent protein (GFP) family is their ability to form a chromophore group by post-translational modifications of internal amino acids, e.g. Ser65-Tyr66-Gly67 in GFP from the jellyfish Aequorea victoria (Cnidaria). Numerous structural studies have demonstrated that the green GFP-like chromophore represents the ‘core’ structure, which can be extended in red-shifted proteins owing to modifications of the protein backbone at the first chromophore-forming position. Here, the three-dimensional structures of green laGFP (λ{sub ex}/λ{sub em} = 502/511 nm) and red laRFP (λ{sub ex}/λ{sub em} ≃ 521/592 nm), which are fluorescent proteins (FPs) from the lancelet Branchiostoma lanceolatum (Chordata), were determined together with the structure of a red variant laRFP-ΔS83 (deletion of Ser83) with improved folding. Lancelet FPs are evolutionarily distant and share only ∼20% sequence identity with cnidarian FPs, which have been extensively characterized and widely used as genetically encoded probes. The structure of red-emitting laRFP revealed three exceptional features that have not been observed in wild-type fluorescent proteins from Cnidaria reported to date: (i) an unusual chromophore-forming sequence Gly58-Tyr59-Gly60, (ii) the presence of Gln211 at the position of the conserved catalytic Glu (Glu222 in Aequorea GFP), which proved to be crucial for chromophore formation, and (iii) the absence of modifications typical of known red chromophores and the presence of an extremely unusual covalent bond between the Tyr59 C{sup β} atom and the hydroxyl of the proximal Tyr62. The impact of this covalent bond on the red emission and the large Stokes shift (�

Peroxynitrite mediates muscle insulin resistance in mice via nitration of IRβ/IRS-1 and Akt

International Nuclear Information System (INIS)

Zhou Jun; Huang Kaixun

2009-01-01

Accumulating evidence suggests that peroxynitrite (ONOO - ) is involved in the pathogenesis of insulin resistance. In the current study, we investigated whether insulin resistance in vivo could be mediated by nitration of proteins involved in the early steps of the insulin signal transduction pathway. Exogenous peroxynitrite donated by 3-morpholinosydnonimine hydrochloride (SIN-1) induced in vivo nitration of the insulin receptor β subunit (IRβ), insulin receptor substrate (IRS)-1, and protein kinase B/Akt (Akt) in skeletal muscle of mice and dramatically reduced whole-body insulin sensitivity and muscle insulin signaling. Moreover, in high-fat diet (HFD)-fed insulin-resistant mice, we observed enhanced nitration of IRβ and IRS-1 in skeletal muscle, in parallel with impaired whole-body insulin sensitivity and muscle insulin signaling. Reversal of nitration of these proteins by treatment with the peroxynitrite decomposition catalyst FeTPPS yielded an improvement in whole-body insulin sensitivity and muscle insulin signaling in HFD-fed mice. Taken together, these findings provide new mechanistic insights for the involvement of peroxynitrite in the development of insulin resistance and suggest that nitration of proteins involved in the early steps of insulin signal transduction is a novel molecular mechanism of HFD-induced muscle insulin resistance.

Two phloem nitrate transporters, NRT1.11 and NRT1.12, are important for redistributing xylem-borne nitrate to enhance plant growth.

Science.gov (United States)

Hsu, Po-Kai; Tsay, Yi-Fang

2013-10-01

This study of the Arabidopsis (Arabidopsis thaliana) nitrate transporters NRT1.11 and NRT1.12 reveals how the interplay between xylem and phloem transport of nitrate ensures optimal nitrate distribution in leaves for plant growth. Functional analysis in Xenopus laevis oocytes showed that both NRT1.11 and NRT1.12 are low-affinity nitrate transporters. Quantitative reverse transcription-polymerase chain reaction and immunoblot analysis showed higher expression of these two genes in larger expanded leaves. Green fluorescent protein and β-glucuronidase reporter analyses indicated that NRT1.11 and NRT1.12 are plasma membrane transporters expressed in the companion cells of the major vein. In nrt1.11 nrt1.12 double mutants, more root-fed (15)NO3(-) was translocated to mature and larger expanded leaves but less to the youngest tissues, suggesting that NRT1.11 and NRT1.12 are required for transferring root-derived nitrate into phloem in the major veins of mature and larger expanded leaves for redistributing to the youngest tissues. Distinct from the wild type, nrt1.11 nrt1.12 double mutants show no increase of plant growth at high nitrate supply. These data suggested that NRT1.11 and NRT1.12 are involved in xylem-to-phloem transfer for redistributing nitrate into developing leaves, and such nitrate redistribution is a critical step for optimal plant growth enhanced by increasing external nitrate.

Contribution of the Tyr-1 in Plantaricin149a to Disrupt Phospholipid Model Membranes

Directory of Open Access Journals (Sweden)

Georgina Tonarelli

2013-06-01

Full Text Available Plantaricin149a (Pln149a is a cationic antimicrobial peptide, which was suggested to cause membrane destabilization via the carpet mechanism. The mode of action proposed to this antimicrobial peptide describes the induction of an amphipathic α-helix from Ala7 to Lys20, while the N-terminus residues remain in a coil conformation after binding. To better investigate this assumption, the purpose of this study was to determine the contributions of the Tyr1 in Pln149a in the binding to model membranes to promote its destabilization. The Tyr to Ser substitution increased the dissociation constant (KD of the antimicrobial peptide from the liposomes (approximately three-fold higher, and decreased the enthalpy of binding to anionic vesicles from −17.2 kcal/mol to −10.2 kcal/mol. The peptide adsorption/incorporation into the negatively charged lipid vesicles was less effective with the Tyr1 substitution and peptide Pln149a perturbed the liposome integrity more than the analog, Pln149S. Taken together, the peptide-lipid interactions that govern the Pln149a antimicrobial activity are found not only in the amphipathic helix, but also in the N-terminus residues, which take part in enthalpic contributions due to the allocation at a lipid-aqueous interface.

The systems lanthanum (cerium, samarium) nitrate-tetramethyl-ammonium nitrate-water

International Nuclear Information System (INIS)

Zhuravlev, E.F.; Khisaeva, D.A.; Semenova, Eh.B.

1984-01-01

The method of cross sections at 25 and 50 deg C has been applied to study solubility in the systems lanthanum nitrate-tetramethyl ammonium nitrate-water (1), cesium (3) nitrate-tetramethyl ammonium nitrate-water (2) and samarium nitrate-tetramethyl ammonium nitrate-water (3). Crystallization fields of congruently dissolving compounds with 1:3 ratio of salt components (in system 1) and 1:2 ratio (in systems 2 and 3) are found in the systems. New solid phases are separated preparatively and subjected to chemical, differential thermal and IR spectroscopic analyses. Compositions of formed compounds are compared with the compositions known for nitrates of other representatives of light lanthanides

The ytterbium nitrate-quinoline (piperidine) nitrate-water system

International Nuclear Information System (INIS)

Khisaeva, D.A.; Boeva, M.K.; Zhuravlev, E.F.

1985-01-01

Using the method of cross sections the solubility of solid phases in the ytterbium nitrate-quinoline nitrate - water (1) and ytterbium nitrate-piperidine nitrate-water (2) systems is studied at 25 and 50 deg C. It is established, that in system 1 congruently melting compound of the composition Yb(NO 3 ) 3 x2C 9 H 7 NxHNO 3 x3H 2 O is formed. The new solid phase has been isolated as a preparation and subjected to chemical X-ray diffraction, differential thermal and IR spectroscopic analyses. Isotherms of system 2 in the studied range of concentrations and temperatures consist of two branches, corresponding to crystallization of tetruaqueous ytterbi um nitrate and nitric acid piperidine

Assimilation of ammonium and nitrate nitrogen by bean plants

International Nuclear Information System (INIS)

Volk, R.J.; Chaillou, S.; Morot-Gaudry, J.F.; Mariotti, A.

1989-01-01

Enhanced growth is often observed in plants growing on combined ammonium and nitrate nutrition. The physiological basis for such enhancement was examined by exposing non-nodulated bean (Phaseolus vulgaris L.) plants to 15 N-labeled, 1.0 mM N solutions containing 0, 33, 67 or 100% of the N as ammonium, the balance being nitrate. Maximal total N uptake and biomass production were attained by plants receiving 33% ammonium. A higher proportion of incoming ammonium than nitrate was incorporated into root protein. This was accompanied by increased partitioning of plant biomass to roots. It was concluded that as a consequence of greater N metabolism in the root under mixed ammonium and nitrate nutrition, the root became a more active sink for photosynthate. Concurrently, the augmented supply of N to the shoot enhanced net photosynthesis as reflected in increased plant biomass

Lanthanum (samarium) nitrate-4-aminoantipyrine nitrate-water systems

International Nuclear Information System (INIS)

Starikova, L.I.; Zhuravlev, E.F.

1985-01-01

Using the isothermal method of cross-sections at 50 deg C systems lanthanum nitrate-4-aminoantipyrine nitrate-water (1), samarium nitrate-4-aminoantipyrine nitrate-water (2), are studied. Isotherms of system 1 consist of two crystallization branches of initial salt components. In system 2 formation of congruently soluble compounds of the composition Sm(No) 3 ) 3 xC 11 H 13 ON 3 xHNO 3 is established. Analytical, X-ray phase and thermogravimetric analysis of the isolated binary salt are carried out

Selective mono-radioiodination and characterization of a cell-penetrating peptide. L-Tyr-maurocalcine

Energy Technology Data Exchange (ETDEWEB)

Ahmadi, Mitra; Bacot, Sandrine; Perret, Pascale; Riou, Laurent; Ghezzi, Catherine [Universite Joseph Fourier, Grenoble (France); INSERM U1039, Grenoble (France). Radiopharmaceutiques Biocliniques; Poillot, Cathy; Cestele, Sandrine [INSERM U836, Grenoble (France). Grenoble Inst. of Neuroscience; Universite Joseph Fourier, Grenoble (France); Desruet, Marie-Dominique [INSERM U1039, Grenoble (France). Radiopharmaceutiques Biocliniques; Couvet, Morgane; Bourgoin, Sandrine; Seve, Michel [CRI-INSERM U823, Grenoble (France). Inst. of Albert Bonniot; Universite Joseph Fourier, Grenoble (France); Waard, Michel de [INSERM U836, Grenoble (France). Grenoble Inst. of Neuroscience; Universite Joseph Fourier, Grenoble (France); Smartox Biotechnologies, Grenoble (France)

2014-07-01

Mono-and poly-iodinated peptides form frequently during radioiodination procedures. However, the formation of a single species in its mono-iodinated form is essential for quantitative studies such as determination of tissue concentration or image quantification. Therefore, the aim of the present study was to define the optimal experimental conditions in order to exclusively obtain the mono-iodinated form of L-maurocalcine (L-MCa). L-MCa is an animal venom toxin which was shown to act as a cell-penetrating peptide. In order to apply the current direct radioiodination technique using oxidative agents including chloramine T, Iodo-Gen {sup registered} or lactoperoxidase, an analogue of this peptide containing a tyrosine residue (Tyr-L-MCa) was synthesized and was shown to fold/oxidize properly. The enzymatic approach using lactoperoxidase/H{sub 2}O{sub 2} was found to be the best method for radioiodination of Tyr-L-MCa. MALDI-TOF mass spectrometry analyses were then used for identification of the chromatographic eluting components of the reaction mixtures. We observed that the production of different radioiodinated species depended upon the reaction conditions. Our results successfully described the experimental conditions of peptide radioiodination allowing the exclusive production of the mono-iodinated form with high radiochemical purity and without the need for a purification step. Mono-radioiodination of L-Tyr-MCa will be crucial for future quantitative studies, investigating the mechanism of cell penetration and in vivo biodistribution.

Synthesis of the tritium labelled β-casomorphine analogues 3H-Phe-Pro-Gly-OH and 3H2-Tyr-Pro-3H-Phe-pyrrolidide

International Nuclear Information System (INIS)

Oehlke, J.; Niedrich, H.; Born, I.; Neubert, K.; Mittag, E.

1991-01-01

The precursor peptides H-Phe(I)-Pro-Gly-OH (III) and H-Tyr(I 2 )-Pro-Phe(I)-pyrrolidide (VIII) were synthesized by stepwise elongation from the C-terminal end and by coupling of Boc-Tyr(I 2 )-Pro-OH with H-Phe(I)-pyrrolidide and following deprotection of the Boc-residue respectively. Catalytic dehalotritiation yielded tritated peptides with specific radioactivities of 450 and 1500 GBq/mmol respectively. Cleavage of 3 H 2 -Tyr-Pro- 3 H-Phe-pyrrolidide by dipeptidylpeptidase IV resulted in fragments with specific radioactivities of 950 ( 3 H 2 -Tyr-Pro) and 590 GBq/ mmol ( 3 H-Phe-pyrrolidide). (author)

Neodymium nitrate-tetraethylammonium nitrate-water system

International Nuclear Information System (INIS)

Khisaeva, D.A.; Boeva, M.K.

1987-01-01

Method of isothermal cross sections at 25 and 50 deg C is used to study solid phase solubility in the neodymium nitrate-tetraethylammonium nitrate-water system. Crystallization fields of congruently soluble compounds, the salt component ratio being 1:1:4H 2 O and 1:3:2H 2 O are detected. New solid phases are preparatively obtained and subjected to chemical, differential thermal, IR spectroscopic and X-ray diffraction analyses. The obtained compounds are acido-complexes in which nitrate groups enter into the first coordination sphere

Analysis of 125I-[Tyr3] octreotide receptors of NCI-H466 cell line

International Nuclear Information System (INIS)

Sun Junjie; Fan Wo; Xu Yujie; Zhang Youjiu; Zhu Ran

2002-01-01

Objective: To study the affinity of small cell lung carcinoma to [Tyr 3 ] octreotide (TOC). Methods: Taking 125 I-[Tyr 3 ] octreotide (labeled by chloramine-T method), as the ligand, small cell lung carcinoma NCI-H466 cell line was inspected for the receptor-binding points and affinity constant. Results: The radio-chemical purity of 125 I-TOC purified through sephadex G-10 was higher than 95%. Receptor analysis study showed that the expression of somatostatin receptors on NCI-H446 cells was numerous (Bmax = 1.17 x 10 5 /cell) with strong affinity to 125 I-TOC (Kd = 0.56 nM). Conclusion: Labeled TOC could be used for small cell lung carcinoma receptor imaging and radio-pharmaceutical therapy

Biosensors Used for Quantification of Nitrates in Plants

Directory of Open Access Journals (Sweden)

Romero-Galindo Raul

2016-01-01

Full Text Available Nitrogen is essential for the plant because it is used for the production of chlorophyll, proteins, nucleic acids, amino acids, and other cellular compounds; nitrogen is available in two forms: ammonium and nitrate. Several tools have been used to quantify nitrates in plants such as the Kjeldahl method and Dumas combustion digestion; however, they are destructive and long time-consuming methods. To solve these disadvantages, methods such as selective electrodes, optical sensors, reflectometers, and images based sensors have been developed; nonetheless, all these techniques show interference when carrying out measurements. Currently, biosensors based on genetic constructions, based on the response of promoter gene fused to Gene Fluorescent Protein (GFP, are gaining popularity, because they improve the accuracy of measurements of nitrate by avoiding the interference of carriers ion, high salt conditions, and other factors. The present review shows the different methods to quantify the nitrogen in plants; later, a biosensors perspective is presented, mainly focused on biosensors based on organism genetically modified. The review presents a list of promoter and reporter genes that could be used to develop different kind of sensors, and a perspective of sensors to measure quantitatively the nitrogen is presented.

Yellow fluorescent protein phiYFPv (Phialidium): structure and structure-based mutagenesis

Energy Technology Data Exchange (ETDEWEB)

Pletneva, Nadya V.; Pletnev, Vladimir Z., E-mail: vzpletnev@gmail.com; Souslova, Ekaterina; Chudakov, Dmitry M. [Russian Academy of Sciences, Moscow (Russian Federation); Lukyanov, Sergey [Russian Academy of Sciences, Moscow (Russian Federation); Nizhny Novgorod State Medical Academy, Nizhny Novgorod (Russian Federation); Martynov, Vladimir I.; Arhipova, Svetlena; Artemyev, Igor [Russian Academy of Sciences, Moscow (Russian Federation); Wlodawer, Alexander [National Cancer Institute, Frederick, MD 21702 (United States); Dauter, Zbigniew [National Cancer Institute, Argonne, IL 60439 (United States); Pletnev, Sergei [National Cancer Institute, Argonne, IL 60439 (United States); SAIC-Frederick, 9700 South Cass Avenue, Argonne, IL 60439 (United States); Russian Academy of Sciences, Moscow (Russian Federation)

2013-06-01

The yellow fluorescent protein phiYFPv with improved folding has been developed from the spectrally identical wild-type phiYFP found in the marine jellyfish Phialidium. The yellow fluorescent protein phiYFPv (λ{sub em}{sup max} ≃ 537 nm) with improved folding has been developed from the spectrally identical wild-type phiYFP found in the marine jellyfish Phialidium. The latter fluorescent protein is one of only two known cases of naturally occurring proteins that exhibit emission spectra in the yellow–orange range (535–555 nm). Here, the crystal structure of phiYFPv has been determined at 2.05 Å resolution. The ‘yellow’ chromophore formed from the sequence triad Thr65-Tyr66-Gly67 adopts the bicyclic structure typical of fluorophores emitting in the green spectral range. It was demonstrated that perfect antiparallel π-stacking of chromophore Tyr66 and the proximal Tyr203, as well as Val205, facing the chromophore phenolic ring are chiefly responsible for the observed yellow emission of phiYFPv at 537 nm. Structure-based site-directed mutagenesis has been used to identify the key functional residues in the chromophore environment. The obtained results have been utilized to improve the properties of phiYFPv and its homologous monomeric biomarker tagYFP.

The tetrapeptide Arg-Leu-Tyr-Glu inhibits VEGF-induced angiogenesis

International Nuclear Information System (INIS)

Baek, Yi-Yong; Lee, Dong-Keon; So, Ju-Hoon; Kim, Cheol-Hee; Jeoung, Dooil; Lee, Hansoo; Choe, Jongseon; Won, Moo-Ho; Ha, Kwon-Soo; Kwon, Young-Guen; Kim, Young-Myeong

2015-01-01

Kringle 5, derived from plasminogen, is highly capable of inhibiting angiogenesis. Here, we have designed and synthesized 10 tetrapeptides, based on the amino acid properties of the core tetrapeptide Lys-Leu-Tyr-Asp (KLYD) originating from anti-angiogenic kringle 5 of human plasminogen. Of these, Arg-Leu-Tyr-Glu (RLYE) effectively inhibited vascular endothelial growth factor (VEGF)-induced endothelial cell proliferation, migration and tube formation, with an IC 50 of 0.06–0.08 nM, which was about ten-fold lower than that of the control peptide KLYD (0.79 nM), as well as suppressed developmental angiogenesis in a zebrafish model. Furthermore, this peptide effectively inhibited the cellular events that precede angiogenesis, such as ERK and eNOS phosphorylation and nitric oxide production, in endothelial cells stimulated with VEGF. Collectively, these data demonstrate that RLYE is a potent anti-angiogenic peptide that targets the VEGF signaling pathway. - Highlights: • The tetrapeptide RLYE inhibited VEGF-induced angiogenesis in vitro. • RLYE also suppressed neovascularization in a zebrafish model. • Its effect was correlated with inhibition of VEGF-induced ERK and eNOS activation. • RLYE may be used as a therapeutic drug for angiogenesis-related diseases

The tetrapeptide Arg-Leu-Tyr-Glu inhibits VEGF-induced angiogenesis

Energy Technology Data Exchange (ETDEWEB)

Baek, Yi-Yong; Lee, Dong-Keon [Department of Molecular and Cellular Biochemistry, School of Medicine, Kangwon National University, Chuncheon, Gangwon-do, 200-702 (Korea, Republic of); So, Ju-Hoon; Kim, Cheol-Hee [Department of Biology, Chungnam National University, Daejeon, 305-764 (Korea, Republic of); Jeoung, Dooil [Department of Biochemistry, College of Natural Sciences, Kangwon National University, Chuncheon, Gangwon-do, 200-702 (Korea, Republic of); Lee, Hansoo [Department of Life Sciences, College of Natural Sciences, Kangwon National University, Chuncheon, Gangwon-do, 200-702 (Korea, Republic of); Choe, Jongseon [Department of Immunology, School of Medicine, Kangwon National University, Chuncheon, Gangwon-do, 200-702 (Korea, Republic of); Won, Moo-Ho [Department of Neurobiology, School of Medicine, Kangwon National University, Chuncheon, Gangwon-do, 200-702 (Korea, Republic of); Ha, Kwon-Soo [Department of Molecular and Cellular Biochemistry, School of Medicine, Kangwon National University, Chuncheon, Gangwon-do, 200-702 (Korea, Republic of); Kwon, Young-Guen [Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul, 120-752 (Korea, Republic of); Kim, Young-Myeong, E-mail: ymkim@kangwon.ac.kr [Department of Molecular and Cellular Biochemistry, School of Medicine, Kangwon National University, Chuncheon, Gangwon-do, 200-702 (Korea, Republic of)

2015-08-07

Kringle 5, derived from plasminogen, is highly capable of inhibiting angiogenesis. Here, we have designed and synthesized 10 tetrapeptides, based on the amino acid properties of the core tetrapeptide Lys-Leu-Tyr-Asp (KLYD) originating from anti-angiogenic kringle 5 of human plasminogen. Of these, Arg-Leu-Tyr-Glu (RLYE) effectively inhibited vascular endothelial growth factor (VEGF)-induced endothelial cell proliferation, migration and tube formation, with an IC{sub 50} of 0.06–0.08 nM, which was about ten-fold lower than that of the control peptide KLYD (0.79 nM), as well as suppressed developmental angiogenesis in a zebrafish model. Furthermore, this peptide effectively inhibited the cellular events that precede angiogenesis, such as ERK and eNOS phosphorylation and nitric oxide production, in endothelial cells stimulated with VEGF. Collectively, these data demonstrate that RLYE is a potent anti-angiogenic peptide that targets the VEGF signaling pathway. - Highlights: • The tetrapeptide RLYE inhibited VEGF-induced angiogenesis in vitro. • RLYE also suppressed neovascularization in a zebrafish model. • Its effect was correlated with inhibition of VEGF-induced ERK and eNOS activation. • RLYE may be used as a therapeutic drug for angiogenesis-related diseases.

Quantitative proteome and phosphoproteome analyses of Streptomyces coelicolor reveal proteins and phosphoproteins modulating differentiation and secondary metabolism

DEFF Research Database (Denmark)

Rioseras, Beatriz; Sliaha, Pavel V; Gorshkov, Vladimir

2018-01-01

identified and quantified 3461 proteins corresponding to 44.3% of the S. coelicolor proteome across three developmental stages: vegetative hypha (MI); secondary metabolite producing hyphae (MII); and sporulating hyphae. A total of 1350 proteins exhibited more than 2-fold expression changes during....../Thr/Tyr kinases, making this genus an outstanding model for the study of bacterial protein phosphorylation events. We used mass spectrometry based quantitative proteomics and phosphoproteomics to characterize bacterial differentiation and activation of secondary metabolism of Streptomyces coelicolor. We...... the bacterial differentiation process. These proteins include 136 regulators (transcriptional regulators, transducers, Ser/Thr/Tyr kinases, signalling proteins), as well as 542 putative proteins with no clear homology to known proteins which are likely to play a role in differentiation and secondary metabolism...

Nitrate in drinking water and risk of colorectal cancer in Yogyakarta, Indonesia.

Science.gov (United States)

Fathmawati; Fachiroh, Jajah; Gravitiani, Evi; Sarto; Husodo, Adi Heru

2017-01-01

Nitrate concentration in well water in Yogyakarta, Indonesia, and its surroundings tended to increase rapidly from time to time, and it may be associated with an elevated risk for several types of cancer. The purpose of this study was to examine the association between nitrate in drinking water and colorectal cancer (CRC) risk occurrence. A case-control study was conducted in Yogyakarta Special Province. Pathologically confirmed 75 CRC patients and 75 controls were consulted and their individual well water was sampled and examined for nitrate concentrations. Logistic regression analysis was conducted to establish the association between nitrate and CRC risk development. There was a significant correlation between nitrate in drinking water and CRC occurrence, and this value was relatively stable after being adjusted for protein intake, smoking history, age, and family history of cancer. These findings demonstrated that the risk of CRC development was fourfold among those with >10 years of nitrate exposure from well water compared with those with ≤10 years of nitrate exposure. Consequently, a significant association between nitrate in drinking water and occurrence of CRC in Yogyakarta was established.

The cisproline(i - 1)-aromatic(i) interaction: Folding of the Ala-cisPro-Tyr peptide characterized by NMR and theoretical approaches

Energy Technology Data Exchange (ETDEWEB)

Nardi, Frederico; Kemmink, Johan; Sattler, Michael; Wade, Rebecca C. [European Molecular Biology Laboratory (Germany)

2000-05-15

Cisproline(i-1)-aromatic(i) interactions have been detected in several short peptides in aqueous solution by analysis of anomalous chemical shifts measured by {sup 1}H-NMR spectroscopy. This formation of local structure is of importance for protein folding and binding properties. To obtain an atomic-detail characterisation of the cisproline(i-1)-aromatic(i) interaction in terms of structure, energetics and dynamics, we studied the minimal peptide unit, blocked Ala-cisPro-Tyr, using computational and experimental techniques. Structural database analyses and a systematic search revealed two groups of conformations displaying a cisproline(i-1)-aromatic(i) interaction. These conformations were taken as seeds for molecular dynamics simulations in explicit solvent at 278 K. During a total of 33.6 ns of simulation, all the 'folded' conformations and some 'unfolded' states were sampled. {sup 1}H- and {sup 13}C-chemical shifts and {sup 3}J-coupling constants were measured for the Ala-Pro-Tyr peptide. Excellent agreement was found between all the measured and computed NMR properties, showing the good quality of the force field. We find that under the experimental and simulation conditions, the Ala-cisPro-Tyr peptide is folded 90% of the time and displays two types of folded conformation which we denote 'a' and 'b'. The type a conformations are twice as populated as the type b conformations. The former have the tyrosine ring interacting with the alanine {alpha} proton and are enthalpically stabilised. The latter have the aromatic ring interacting with the proline side chain and are entropically stabilised. The combined and complementary use of computational and experimental techniques permitted derivation of a detailed scenario of the 'folding' of this peptide.

Probing hydrogen bonding interactions and proton transfer in proteins

Science.gov (United States)

Nie, Beining

Scope and method of study. Hydrogen bonding is a fundamental element in protein structure and function. Breaking a single hydrogen bond may impair the stability of a protein. It is therefore important to probe dynamic changes in hydrogen bonding interactions during protein folding and function. Time-resolved Fourier transform infrared spectroscopy is highly sensitive to hydrogen bonding interactions. However, it lacks quantitative correlation between the vibrational frequencies and the number, type, and strength of hydrogen bonding interactions of ionizable and polar residues. We employ quantum physics theory based ab initio calculations to study the effects of hydrogen bonding interactions on vibrational frequencies of Asp, Glu, and Tyr residues and to develop vibrational spectral markers for probing hydrogen bonding interactions using infrared spectroscopy. In addition, proton transfer process plays a crucial role in a wide range of energy transduction, signal transduction, and enzymatic reactions. We study the structural basis for proton transfer using photoactive yellow protein as an excellent model system. Molecular dynamics simulation is employed to investigate the structures of early intermediate states. Quantum theory based ab initio calculations are used to study the impact of hydrogen bond interactions on proton affinity and proton transfer. Findings and conclusions. Our extensive density function theory based calculations provide rich structural, spectral, and energetic information on hydrogen bonding properties of protonated side chain groups of Asp/Glu and Tyr. We developed vibrational spectral markers and 2D FTIR spectroscopy for structural characterization on the number and the type of hydrogen bonding interactions of the COOH group of Asp/Glu and neutral phenolic group of Tyr. These developments greatly enhance the power of time-resolved FTIR spectroscopy as a major experimental tool for structural characterization of functionally important

The outer membrane protein Omp35 affects the reduction of Fe(III, nitrate, and fumarate by Shewanella oneidensis MR-1

Directory of Open Access Journals (Sweden)

Myers Charles R

2004-06-01

Full Text Available Abstract Background Shewanella oneidensis MR-1 uses several electron acceptors to support anaerobic respiration including insoluble species such as iron(III and manganese(IV oxides, and soluble species such as nitrate, fumarate, dimethylsulfoxide and many others. MR-1 has complex branched electron transport chains that include components in the cytoplasmic membrane, periplasm, and outer membrane (OM. Previous studies have implicated a role for anaerobically upregulated OM electron transport components in the use of insoluble electron acceptors, and have suggested that other OM components may also contribute to insoluble electron acceptor use. In this study, the role for an anaerobically upregulated 35-kDa OM protein (Omp35 in the use of anaerobic electron acceptors was explored. Results Omp35 was purified from the OM of anaerobically grown cells, the gene encoding Omp35 was identified, and an omp35 null mutant (OMP35-1 was isolated and characterized. Although OMP35-1 grew on all electron acceptors tested, a significant lag was seen when grown on fumarate, nitrate, and Fe(III. Complementation studies confirmed that the phenotype of OMP35-1 was due to the loss of Omp35. Despite its requirement for wild-type rates of electron acceptor use, analysis of Omp35 protein and predicted sequence did not identify any electron transport moieties or predicted motifs. OMP35-1 had normal levels and distribution of known electron transport components including quinones, cytochromes, and fumarate reductase. Omp35 is related to putative porins from MR-1 and S. frigidimarina as well as to the PorA porin from Neisseria meningitidis. Subcellular fraction analysis confirmed that Omp35 is an OM protein. The seven-fold anaerobic upregulation of Omp35 is mediated post-transcriptionally. Conclusion Omp35 is a putative porin in the OM of MR-1 that is markedly upregulated anaerobically by a post-transcriptional mechanism. Omp35 is required for normal rates of growth on Fe

The stress caused by nitrite with titanium dioxide nanoparticles under UVA irradiation in human keratinocyte cell

International Nuclear Information System (INIS)

Tu, Min; Huang, Yi; Li, Hai-Ling; Gao, Zhong-Hong

2012-01-01

Highlights: ► Nitrite increased photo-toxicity of nano-TiO 2 on human keratinocyte cells in a dose-dependant manner. ► Morphological study suggested the cell death may be mediated by apoptosis inducing factor. ► Protein nitration was generated in the cells, and the most abundant nitrated protein was identified as cystatin-A. ► Tyr35 was the most likely site to be nitrated in cystatin-A. -- Abstract: Our previous work found that in the presence of nitrite, titanium dioxide nanoparticles can cause protein tyrosine nitration under UVA irradiation in vivo. In this paper, the human keratinocyte cells was used as a skin cell model to further study the photo-toxicity of titanium dioxide nanoparticles when nitrite was present. The results showed that nitrite increased the photo-toxicity of titanium dioxide in a dose-dependant manner, and generated protein tyrosine nitration in keratinocyte cells. Morphological study of keratinocyte cells suggested a specific apoptosis mediated by apoptosis inducing factor. It was also found the main target nitrated in cells was cystatin-A, which expressed abundantly in cytoplasm and functioned as a cysteine protease inhibitor. The stress induced by titanium dioxide with nitrite under UVA irradiation in human keratinocyte cells appeared to trigger the apoptosis inducing factor mediated cell death and lose the inhibition of active caspase by cystatin-A. We conclude that nitrite can bring new damage and stress to human keratinocyte cells with titanium dioxide nanoparticles under UVA irradiation.

Frequency of the GPR7 Tyr135Phe allelic variant in lean and obese subjects.

Science.gov (United States)

Pelosini, C; Maffei, M; Ceccarini, G; Marchi, M; Marsili, A; Galli, G; Scartabelli, G; Tamberi, A; Latrofa, F; Fierabracci, P; Vitti, P; Pinchera, A; Santini, F

2013-10-01

GPR7, the endogenous coupled receptor for neuropeptide B and neuropeptide W, is expressed in several regions of the central nervous system, which are involved in the regulation of feeding behavior. GPR7 affects the regulation of energy balance through a mechanism independent of leptin and melanocortin pathways. Aim of this study was to investigate whether GPR7 gene mutations can be detected in human subjects and, in that event, if they are differently distributed among lean and obese subjects. The coding region of GPR7 were sequenced in 150 obese patients and 100 normal-weight unrelated controls. Functional studies of the allelic variants were performed. One genetic GPR7 variant was found (Tyr135Phe - rs33977775) in obese subjects (13.3%) and lean control (25%). Functional studies did not reveal significant differences between the wild type and the Tyr135Phe allelic variants in their NPW-mediated capacity to inhibit forskolin-induced cAMP production. Screening of GPR7 gene mutations among lean and obese subjects revealed a Tyr135Phe allelic variant that was fairly common in the study population. As indicated by in vitro and in silico studies, this variant is unlikely to cause a functional derangement of the receptor.

Adaptor protein GRB2 promotes Src tyrosine kinase activation and podosomal organization by protein-tyrosine phosphatase ϵ in osteoclasts.

Science.gov (United States)

Levy-Apter, Einat; Finkelshtein, Eynat; Vemulapalli, Vidyasiri; Li, Shawn S-C; Bedford, Mark T; Elson, Ari

2014-12-26

The non-receptor isoform of protein-tyrosine phosphatase ϵ (cyt-PTPe) supports adhesion of bone-resorbing osteoclasts by activating Src downstream of integrins. Loss of cyt-PTPe reduces Src activity in osteoclasts, reduces resorption of mineralized matrix both in vivo and in cell culture, and induces mild osteopetrosis in young female PTPe KO mice. Activation of Src by cyt-PTPe is dependent upon this phosphatase undergoing phosphorylation at its C-terminal Tyr-638 by partially active Src. To understand how cyt-PTPe activates Src, we screened 73 Src homology 2 (SH2) domains for binding to Tyr(P)-638 of cyt-PTPe. The SH2 domain of GRB2 bound Tyr(P)-638 of cyt-PTPe most prominently, whereas the Src SH2 domain did not bind at all, suggesting that GRB2 may link PTPe with downstream molecules. Further studies indicated that GRB2 is required for activation of Src by cyt-PTPe in osteoclast-like cells (OCLs) in culture. Overexpression of GRB2 in OCLs increased activating phosphorylation of Src at Tyr-416 and of cyt-PTPe at Tyr-638; opposite results were obtained when GRB2 expression was reduced by shRNA or by gene inactivation. Phosphorylation of cyt-PTPe at Tyr-683 and its association with GRB2 are integrin-driven processes in OCLs, and cyt-PTPe undergoes autodephosphorylation at Tyr-683, thus limiting Src activation by integrins. Reduced GRB2 expression also reduced the ability of bone marrow precursors to differentiate into OCLs and reduced the fraction of OCLs in which podosomal adhesion structures assume organization typical of active, resorbing cells. We conclude that GRB2 physically links cyt-PTPe with Src and enables cyt-PTPe to activate Src downstream of activated integrins in OCLs. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Dissimilatory nitrate reduction by Aspergillus terreus isolated from the seasonal oxygen minimum zone in the Arabian Sea

DEFF Research Database (Denmark)

Stief, Peter; Fuchs-Ocklenburg, Silvia; Kamp, Anja

2014-01-01

Background: A wealth of microbial eukaryotes is adapted to life in oxygen-deficient marine environments. Evidence is accumulating that some of these eukaryotes survive anoxia by employing dissimilatory nitrate reduction, a strategy that otherwise is widespread in prokaryotes. Here, we report...... the dissimilatory nature of nitrate reduction. Interestingly, An-4 used intracellular nitrate stores (up to 6-8 μmol NO3 - g-1 protein) for dissimilatory nitrate reduction. Conclusions: Our findings expand the short list of microbial eukaryotes that store nitrate intracellularly and carry out dissimilatory nitrate...

Side-chain-anchored N(alpha)-Fmoc-Tyr-OPfp for bidirectional solid-phase synthesis

DEFF Research Database (Denmark)

Olsen, Christian A; Jørgensen, Malene; Hansen, Steen H

2005-01-01

[reaction: see text] A mild resin-immobilization strategy employing a readily prepared trityl bromide resin for anchoring building blocks via a phenol group has been developed. With N(alpha)-Fmoc-Tyr-OPfp as a starter building block, it was possible to prepare asymmetrically substituted hybrids...

Rapid kit-based (68)Ga-labelling and PET imaging with THP-Tyr(3)-octreotate: a preliminary comparison with DOTA-Tyr(3)-octreotate.

Science.gov (United States)

Ma, Michelle T; Cullinane, Carleen; Waldeck, Kelly; Roselt, Peter; Hicks, Rodney J; Blower, Philip J

2015-12-01

Ge/(68)Ga generators provide an inexpensive source of a PET isotope to hospitals without cyclotron facilities. The development of new (68)Ga-based molecular imaging agents and subsequent clinical translation would be greatly facilitated by simplification of radiochemical syntheses. We report the properties of a tris(hydroxypyridinone) conjugate of the SSTR2-targeted peptide, Tyr(3)-octreotate (TATE), and compare the (68)Ga-labelling and biodistribution of [(68)Ga(THP-TATE)] with the clinical radiopharmaceutical [(68)Ga(DOTATATE)]. A tris(hydroxypyridinone) with a pendant isothiocyanate group was conjugated to the primary amine terminus of H2N-PEG2-Lys(iv-Dde)(5)-TATE, and the resulting conjugate was deprotected to provide THP-TATE. THP-TATE was radiolabelled with (68)Ga(3+) from a (68)Ge/(68)Ga generator. In vitro uptake was assessed in SSTR2-positive 427-7 cells and SSTR2-negative 427 (parental) cells. Biodistribution of [(68)Ga(THP-TATE)] was compared with that of [(68)Ga(DOTATATE)] in Balb/c nude mice bearing SSTR2-positive AR42J tumours. PET scans were obtained 1 h post-injection, after which animals were euthanised and tissues/organs harvested and counted. [(68)Ga(THP-TATE)] was radiolabelled and formulated rapidly in negative cells, and receptor binding and internalisation were specific. Animals administered [(68)Ga(THP-TATE)] demonstrated comparable SSTR2-positive tumour activity (11.5 ± 0.6 %ID g(-1)) compared to animals administered [(68)Ga(DOTATATE)] (14.4 ± 0.8 %ID g(-1)). Co-administration of unconjugated Tyr(3)-octreotate effectively blocked tumour accumulation of [(68)Ga(THP-TATE)] (2.7 ± 0.6 %ID g(-1)). Blood clearance of [(68)Ga(THP-TATE)] was rapid and excretion was predominantly renal, although compared to [(68)Ga(DOTATATE)], [(68)Ga(THP-TATE)] exhibited comparatively longer kidney retention. Radiochemical synthesis of [(68)Ga(THP-TATE)] is significantly faster, proceeds under milder conditions, and requires less manipulation

Structure and Function of Human Tyrosinase and Tyrosinase-Related Proteins

NARCIS (Netherlands)

Lai, Xuelei; Wichers, Harry J.; Soler-Lopez, Montserrat; Dijkstra, Bauke W.

2018-01-01

Melanin is the main pigment responsible for the color of human skin, hair and eye. Its biosynthesis requires three melanogenic enzymes, tyrosinase (TYR), and the tyrosinase-related proteins TYRP1 and TYRP2. The difficulty of isolating pure and homogeneous proteins from endogenous sources has

Simultaneous effect of nitrate (NO3- concentration, carbon dioxide (CO2 supply and nitrogen limitation on biomass, lipids, carbohydrates and proteins accumulation in Nannochloropsis oculata

Directory of Open Access Journals (Sweden)

Aarón Millán-Oropeza

2015-03-01

Full Text Available Biodiesel from microalgae is a promising technology. Nutrient limitation and the addition of CO2 are two strategies to increase lipid content in microalgae. There are two different types of nitrogen limitation, progressive and abrupt limitation. In this work, the simultaneous effect of initial nitrate concentration, addition of CO2, and nitrogen limitation on biomass, lipid, protein and carbohydrates accumulation were analyzed. An experimental design was established in which initial nitrogen concentration, culture time and CO2 aeration as independent numerical variables with three levels were considered. Nitrogen limitation was taken into account as a categorical independent variable. For the experimental design, all the experiments were performed with progressive nitrogen limitation. The dependent response variables were biomass, lipid production, carbohydrates and proteins. Subsequently, comparison of both types of limitation i.e. progressive and abrupt limitation, was performed. Nitrogen limitation in a progressive mode exerted a greater effect on lipid accumulation. Culture time, nitrogen limitation and the interaction of initial nitrate concentration with nitrogen limitation had higher influences on lipids and biomass production. The highest lipid production and productivity were at 582 mgL-1 (49.7 % lipid, dry weight basis and 41.5 mgL-1d-1, respectively; under the following conditions: 250 mgL-1 of initial nitrate concentration, CO2 supply of 4% (v/v, 12 d of culturing and 2 d in state of nitrogen starvation induced by progressive limitation. This work presents a novel way to perform simultaneous analysis of the effect of the initial concentration of nitrate, nitrogen limitation, and CO2 supply on growth and lipid production of Nannochloropsis oculata, with the aim to produce potential biofuels feedstock.

Protein analysis by 31p NMR spectroscopy in ionic liquid: quantitative determination of enzymatically created cross-links.

Science.gov (United States)

Monogioudi, Evanthia; Permi, Perttu; Filpponen, Ilari; Lienemann, Michael; Li, Bin; Argyropoulos, Dimitris; Buchert, Johanna; Mattinen, Maija-Liisa

2011-02-23

Cross-linking of β-casein by Trichoderma reesei tyrosinase (TrTyr) and Streptoverticillium mobaraense transglutaminase (Tgase) was analyzed by (31)P nuclear magnetic resonance (NMR) spectroscopy in ionic liquid (IL). According to (31)P NMR, 91% of the tyrosine side chains were cross-linked by TrTyr at high dosages. When Tgase was used, no changes were observed because a different cross-linking mechanism was operational. However, this verified the success of the phosphitylation of phenolics within the protein matrix in the IL. Atomic force microscopy (AFM) in solid state showed that disk-shaped nanoparticles were formed in the reactions with average diameters of 80 and 20 nm for TrTyr and Tgase, respectively. These data further advance the current understanding of the action of tyrosinases on proteins on molecular and chemical bond levels. Quantitative (31)P NMR in IL was shown to be a simple and efficient method for the study of protein modification.

Alkaline transition of pseudoazurin Met16X mutant proteins: protein stability influenced by the substitution of Met16 in the second sphere coordination.

Science.gov (United States)

Abdelhamid, Rehab F; Obara, Yuji; Kohzuma, Takamitsu

2008-01-01

Several blue copper proteins are known to change the active site structure at alkaline pH (alkaline transition). Spectroscopic studies of Met16Phe, Met16Tyr, Met16Trp, and Met16Val pseudoazurin variants were performed to investigate the second sphere role through alkaline transition. The visible electronic absorption and resonance Raman spectra of Met16Phe, Met16Tyr, and Met16Trp variants showed the increasing of axial component at pH approximately 11 like wild-type PAz. The visible electronic absorption and far-UV CD spectra of Met16Val demonstrated that the destabilization of the protein structure was triggered at pH>11. Resonance Raman (RR) spectra of PAz showed that the intensity-weighted averaged Cu-S(Cys) stretching frequency was shifted to higher frequency region at pH approximately 11. The higher frequency shift of Cu-S(Cys) bond is implied the stronger Cu-S(Cys) bond at alkaline transition pH approximately 11. The visible electronic absorption and far-UV CD spectra of Met16X PAz revealed that the Met16Val variant is denatured at pH>11, but Met16Phe, Met16Tyr, and Met16Trp mutant proteins are not denatured even at pH>11. These observations suggest that Met16 is important to maintain the protein structure through the possible weak interaction between methionine -SCH3 part and coordinated histidine imidazole moiety. The introduction of pi-pi interaction in the second coordination sphere may be contributed to the enhancement of protein structure stability.

Reexamining the risks of drinking-water nitrates on public health.

Science.gov (United States)

Richard, Alyce M; Diaz, James H; Kaye, Alan David

2014-01-01

Nitrates in drinking water are generally considered the sole source of nitrite poisoning with methemoglobinemia in infantile methomoglobinemia (IM). However, IM, which occurs during the first 4 months of life, is actually a constellation of cyanosis and hypoxia associated with methemoglobinemia that can result from several other causes. This review reexamines the role of nitrate levels in drinking water as a cause of IM and identifies other sources of nitrates that can affect public health and cause chronic diseases. Causes of IM include nitrites in foods, environmental chemical exposures, commonly prescribed pharmaceuticals, and the endogenous generation of oxides of nitrogen. Infants with congenital enzyme deficiencies in glucose-6-phosphate dehydrogenase and methemoglobin reductase are at greater risk of nitrite-induced methemoglobinemia from nitrates in water and food and from exposures to hemoglobin oxidizers. Early epidemiological studies demonstrated significant associations between high groundwater nitrate levels and elevated methemoglobin levels in infants fed drinking water-diluted formulas. However, more recent epidemiological investigations suggest other sources of nitrogenous substance exposures in infants, including protein-based formulas and foods and the production of nitrate precursors (nitric acid) by bacterial action in the infant gut in response to inflammation and infection.

The systems cerium(3) (samarium) nitrate-quinoline nitrate-water

International Nuclear Information System (INIS)

Khisaeva, D.A.; Zhuravlev, E.F.; Semenova, Eh.B.

1982-01-01

Using the method of cross sections at 25 and 50 deg C the solubility in the systems cerium (3) nitrate-quinoline nitrate-water and samarium nitrate-quinoline nitrate-water has been studied. It is established that in the systems during chemical interaction of components congruently melting compounds of the composition: Ce(NO 3 ) 2 x2[C 9 H 7 NxHNO 3 ]x6H 2 O and Sm(NO 3 ) 3 x2[C 9 H 7 NxHNO 3 ]x2H 2 O are formed. New solid phases are separated preparatively and are subjected to chemical, differential thermal and IR spectroscopic analyses. The investigation results are compared with similar ones for nitrates of other representatives of lanthanide group

Nitrate storage and dissimilatory nitrate reduction by eukaryotic microbes

DEFF Research Database (Denmark)

Kamp, Anja; Høgslund, Signe; Risgaard-Petersen, Nils

2015-01-01

The microbial nitrogen cycle is one of the most complex and environmentally important element cycles on Earth and has long been thought to be mediated exclusively by prokaryotic microbes. Rather recently, it was discovered that certain eukaryotic microbes are able to store nitrate intracellularly......, suggesting that eukaryotes may rival prokaryotes in terms of dissimilatory nitrate reduction. Finally, this review article sketches some evolutionary perspectives of eukaryotic nitrate metabolism and identifies open questions that need to be addressed in future investigations....... and use it for dissimilatory nitrate reduction in the absence of oxygen. The paradigm shift that this entailed is ecologically significant because the eukaryotes in question comprise global players like diatoms, foraminifers, and fungi. This review article provides an unprecedented overview of nitrate...

Phosphoproteome of the cyanobacterium Synechocystis sp. PCC 6803 and its dynamics during nitrogen starvation.

Directory of Open Access Journals (Sweden)

Philipp eSpät

2015-03-01

Full Text Available Cyanobacteria have shaped the earth’s biosphere as the first oxygenic photoautotrophs and still play an important role in many ecosystems. The ability to adapt to changing environmental conditions is an essential characteristic in order to ensure survival. To this end, numerous studies have shown that bacteria use protein post-translational modifications such as Ser/Thr/Tyr phosphorylation in cell signalling, adaptation and regulation. Nevertheless, our knowledge of cyanobacterial phosphoproteomes and their dynamic response to environmental stimuli is relatively limited. In this study, we applied gel-free methods and high accuracy mass spectrometry towards the unbiased detection of Ser/Thr/Tyr phosphorylation events in the model cyanobacterium Synechocystis sp. PCC 6803. We could identify over 300 phosphorylation events in cultures grown on nitrate as exclusive nitrogen source. Chemical dimethylation labelling was applied to investigate proteome and phosphoproteome dynamics during nitrogen starvation. Our dataset describes the most comprehensive (phosphoproteome of Synechocystis to date, identifying 2,382 proteins and 183 phosphorylation events and quantifying 2,111 proteins and 148 phosphorylation events during nitrogen starvation. Global protein phosphorylation levels were increased in response to nitrogen depletion after 24 hours. Among the proteins with increased phosphorylation, the PII signalling protein showed the highest fold-change, serving as positive control. Other proteins with increased phosphorylation levels comprised functions in photosynthesis and in carbon and nitrogen metabolism. This study reveals dynamics of Synechocystis phosphoproteome in response to environmental stimuli and suggests an important role of protein Ser/Thr/Tyr phosphorylation in fundamental mechanisms of homeostatic control in cyanobacteria.

Transforming Growth Factor-β and Nitrates in Epithelial Ovarian Cancer

Science.gov (United States)

Khalifa, Ali; Kassim, Samar K.; Ahmed, Maha I.; Fayed, Salah T.

1999-01-01

The role of transforming growth factor-β (TGF-β) and nitric oxide (NO) in ovarian neoplasia is still not clear. We studied the expression of TGF-β by enzyme immunoassay, and nitrates (as a stable end product of NO) in 127 ovarian tissues (36 normal, 37 benign, and 54 malignant). Ploidy status and synthetic phase fraction (SPF) were also assessed by flow cytometry. Mean ranks of TGF-β, nitrate, and SPF were significant among different groups (X2 = 12.01, P = 0.0025, X2 = 67.42, P = 0.000, X2 = 9.06, P = 0.011 respectively). Nitrate mean ranks were significant among different FIGO stages of the disease (X2 = 17.6, P = 0.000). A significant correlation was shown between TGF-â, and nitrate levels in all tissues (r = 0.24, P = 0.01), as well as in malignant tissues (r = 0.3, P = 0.026). Cutoff values were determined for both TGF-β (290 pg/mg protein), and nitrates (310 nmole/mg non protein nitrogenous substances). At these cut-offs, nitrates showed a sensitivity of 93% and 84% specificity for malignant versus normal cases, while TGF-β had 76% sensitivity, and 82.4% specificity for poor versus good outcome. Patients with epithelial ovarian cancer were followed up for a total of 40 months. Survival analysis showed that patients with TGF-β above the cut-off had worse prognosis (X2 = 12.69, P = 0.004). The present results suggest that malignant transformation of ovarian tissues is associated with increased TGF-β and NO production. NO level is related to the development and progression of epithelial ovarian cancer, while high levels of TGF-β could be of prognostic significance. PMID:10689548

Transforming Growth Factor-β and Nitrates in Epithelial Ovarian Cancer

Directory of Open Access Journals (Sweden)

Ali Khalifa

1999-01-01

Full Text Available The role of transforming growth factor-β (TGF-β and nitric oxide (NO in ovarian neoplasia is still not clear. We studied the expression of TGF-β by enzyme immunoassay, and nitrates (as a stable end product of NO in 127 ovarian tissues (36 normal, 37 benign, and 54 malignant. Ploidy status and synthetic phase fraction (SPF were also assessed by flow cytometry. Mean ranks of TGF-β, nitrate, and SPF were significant among different groups (X2 = 12.01, P = 0.0025, X2 = 67.42, P = 0.000, X2 = 9.06, P = 0.011 respectively. Nitrate mean ranks were significant among different FIGO stages of the disease (X2 = 17.6, P = 0.000. A significant correlation was shown between TGF-â, and nitrate levels in all tissues (r = 0.24, P = 0.01, as well as in malignant tissues (r = 0.3, P = 0.026. Cutoff values were determined for both TGF-β (290 pg/mg protein, and nitrates (310 nmole/mg non protein nitrogenous substances. At these cut-offs, nitrates showed a sensitivity of 93% and 84% specificity for malignant versus normal cases, while TGF-β had 76% sensitivity, and 82.4% specificity for poor versus good outcome. Patients with epithelial ovarian cancer were followed up for a total of 40 months. Survival analysis showed that patients with TGF-β above the cut-off had worse prognosis (X2 = 12.69, P = 0.004. The present results suggest that malignant transformation of ovarian tissues is associated with increased TGF-β and NO production. NO level is related to the development and progression of epithelial ovarian cancer, while high levels of TGF-β could be of prognostic significance.

Transforming growth factor-beta and nitrates in epithelial ovarian cancer.

Science.gov (United States)

Khalifa, A; Kassim, S K; Ahmed, M I; Fayed, S T

1999-12-01

The role of transforming growth factor-beta (TGF-beta) and nitric oxide (NO) in ovarian neoplasia is still not clear. We studied the expression of TGF-beta by enzyme immunoassay, and nitrates (as a stable end product of NO) in 127 ovarian tissues (36 normal, 37 benign, and 54 malignant). Ploidy status and synthetic phase fraction (SPF) were also assessed by flow cytometry. Mean ranks of TGF-beta, nitrate, and SPF were significant among different groups (X2 = 12.01, P = 0.0025, X2 = 67.42, P = 0.000, X2 = 9.06, P = 0.011 respectively). Nitrate mean ranks were significant among different FIGO stages of the disease (X2 = 17.6, P = 0.000). A significant correlation was shown between TGF-beta, and nitrate levels in all tissues (r = 0.24, P = 0.01), as well as in malignant tissues (r = 0.3, P = 0.026). Cutoff values were determined for both TGF-beta (290 pg/mg protein), and nitrates (310 nmole/mg non protein nitrogenous substances). At these cut-offs, nitrates showed a sensitivity of 93% and 84% specificity for malignant versus normal cases, while TGF-beta had 76% sensitivity, and 82.4% specificity for poor versus good outcome. Patients with epithelial ovarian cancer were followed up for a total of 40 months. Survival analysis showed that patients with TGF-beta above the cut-off had worse prognosis (X2 = 12.69, P = 0.004). The present results suggest that malignant transformation of ovarian tissues is associated with increased TGF-beta and NO production. NO level is related to the development and progression of epithelial ovarian cancer, while high levels of TGF-beta could be of prognostic significance.

Mechanical wounding induces a nitrosative stress by down-regulation of GSNO reductase and an increase in S-nitrosothiols in sunflower (Helianthus annuus) seedlings

Science.gov (United States)

Chaki, Mounira; Valderrama, Raquel; Fernández-Ocaña, Ana M.; Carreras, Alfonso; Gómez-Rodríguez, Maria. V.; Pedrajas, José R.; Begara-Morales, Juan C.; Sánchez-Calvo, Beatriz; Luque, Francisco; Leterrier, Marina; Corpas, Francisco J.; Barroso, Juan B.

2011-01-01

Nitric oxide (NO) and related molecules such as peroxynitrite, S-nitrosoglutathione (GSNO), and nitrotyrosine, among others, are involved in physiological processes as well in the mechanisms of response to stress conditions. In sunflower seedlings exposed to five different adverse environmental conditions (low temperature, mechanical wounding, high light intensity, continuous light, and continuous darkness), key components of the metabolism of reactive nitrogen species (RNS) and reactive oxygen species (ROS), including the enzyme activities L-arginine-dependent nitric oxide synthase (NOS), S-nitrosogluthathione reductase (GSNOR), nitrate reductase (NR), catalase, and superoxide dismutase, the content of lipid hydroperoxide, hydrogen peroxide, S-nitrosothiols (SNOs), the cellular level of NO, GSNO, and GSNOR, and protein tyrosine nitration [nitrotyrosine (NO2-Tyr)] were analysed. Among the stress conditions studied, mechanical wounding was the only one that caused a down-regulation of NOS and GSNOR activities, which in turn provoked an accumulation of SNOs. The analyses of the cellular content of NO, GSNO, GSNOR, and NO2-Tyr by confocal laser scanning microscopy confirmed these biochemical data. Therefore, it is proposed that mechanical wounding triggers the accumulation of SNOs, specifically GSNO, due to a down-regulation of GSNOR activity, while NO2-Tyr increases. Consequently a process of nitrosative stress is induced in sunflower seedlings and SNOs constitute a new wound signal in plants. PMID:21172815

Plasma nitrate and nitrite are increased by a high nitrate supplement, but not by high nitrate foods in older adults

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Miller, Gary D.; Marsh, Anthony P.; Dove, Robin W.; Beavers, Daniel; Presley, Tennille; Helms, Christine; Bechtold, Erika; King, S. Bruce; Kim-Shapiro, Daniel

2012-01-01

Little is known about the effect of dietary nitrate on the nitrate/nitrite/NO (nitric oxide) cycle in older adults. We examined the effect of a 3-day control diet vs. high nitrate diet, with and without a high nitrate supplement (beetroot juice), on plasma nitrate and nitrite kinetics, and blood pressure using a randomized four period cross-over controlled design. We hypothesized that the high nitrate diet would show higher levels of plasma nitrate/nitrite and blood pressure compared to the control diet, which would be potentiated by the supplement. Participants were eight normotensive older men and women (5 female, 3 male, 72.5±4.7 yrs) with no overt disease or medications that affect NO metabolism. Plasma nitrate and nitrite levels and blood pressure were measured prior to and hourly for 3 hours after each meal. The mean daily changes in plasma nitrate and nitrite were significantly different from baseline for both control diet+supplement (pnitrate and nitrite, respectively) and high nitrate diet+supplement (p=0.001 and 0.002), but not for control diet (p=0.713 and 0.741) or high nitrate diet (p=0.852 and 0.500). Blood pressure decreased from the morning baseline measure to the three 2 hr post-meal follow-up time-points for all treatments, but there was no main effect for treatment. In healthy older adults, a high nitrate supplement consumed at breakfast elevated plasma nitrate and nitrite levels throughout the day. This observation may have practical utility for the timing of intake of a nitrate supplement with physical activity for older adults with vascular dysfunction. PMID:22464802

Amifostine protects rat kidneys during peptide receptor radionuclide therapy with [{sup 177}Lu-DOTA{sup 0},Tyr{sup 3}]octreotate

Energy Technology Data Exchange (ETDEWEB)

Rolleman, Edgar J.; Forrer, Flavio; Bernard, Bert; Bijster, Magda; Valkema, Roelf; Krenning, Eric P.; Jong, Marion de [Erasmus MC, Department of Nuclear Medicine, Rotterdam (Netherlands); Vermeij, Marcel [Erasmus MC, Department of Pathology, Rotterdam (Netherlands)

2007-05-15

In peptide receptor radionuclide therapy (PRRT) using radiolabelled somatostatin analogues, the kidneys are the major dose-limiting organs, because of tubular reabsorption and retention of radioactivity. Preventing renal uptake or toxicity will allow for higher tumour radiation doses. We tested the cytoprotective drug amifostine, which selectively protects healthy tissue during chemo- and radiotherapy, for its renoprotective capacities after PRRT with high-dose [{sup 177}Lu-DOTA{sup 0},Tyr{sup 3}]octreotate. Male Lewis rats were injected with 278 or 555 MBq [{sup 177}Lu-DOTA{sup 0},Tyr{sup 3}]octreotate to create renal damage and were followed up for 130 days. For renoprotection, rats received either amifostine or co-injection with lysine. Kidneys, blood and urine were collected for toxicity measurements. At 130 days after PRRT, a single-photon emission computed tomography (SPECT) scan was performed to quantify tubular uptake of {sup 99m}Tc-dimercaptosuccinic acid (DMSA), a measure of tubular function. Treatment with 555 MBq [{sup 177}Lu-DOTA{sup 0},Tyr{sup 3}]octreotate resulted in body weight loss, elevated creatinine and proteinuria. Amifostine and lysine treatment significantly prevented this rise in creatinine and the level of proteinuria, but did not improve the histological damage. In contrast, after 278 MBq [{sup 177}Lu-DOTA{sup 0},Tyr{sup 3}]octreotate, creatinine values were slightly, but not significantly, elevated compared with the control rats. Proteinuria and histological damage were different from controls and were significantly improved by amifostine treatment. Quantification of {sup 99m}Tc-DMSA SPECT scintigrams at 130 days after [{sup 177}Lu-DOTA{sup 0},Tyr{sup 3}]octreotate therapy correlated well with 1/creatinine (r {sup 2} = 0.772, p < 0.001). Amifostine and lysine effectively decreased functional renal damage caused by high-dose [{sup 177}Lu-DOTA{sup 0},Tyr{sup 3}]octreotate. Besides lysine, amifostine might be used in clinical PRRT as well

Tumour uptake of the radiolabelled somatostatin analogue [DOTA0,TYR3]octreotide is dependent on the peptide amount

International Nuclear Information System (INIS)

Jong, M. de; Breeman, W.A.P.; Bernard, B.F.; Gameren, A. van; Bruin, E. de; Bakker, W.H.; Van der Pluijm, M.E.; Krenning, E.P.; Visser, T.J.; Maecke, H.R.

1999-01-01

Radiolabelled tumour receptor-binding peptides can be used for in vivo scintigraphic imaging. Recently, the somatostatin analogue [Tyr 3 ]octreotide (d-Phe-c(Cys-Tyr-d-Trp-Lys-Thr-Cys)-Thr(ol)) was derivatized with the chelator DOTA (tetra-azacyclododecane-tetra-acetic acid), enabling stable radiolabelling with both the high-energy beta particle-emitter yttrium-90 and the Auger electron-emitter indium-111. The thus produced radiolabelled compounds are promising for peptide receptor radionuclide therapy. Our previous in vitro and in vivo (rat) experiments with these radiolabelled compounds showed favourable binding and biodistribution characteristics with high uptake and retention in the target organs. We also demonstrated receptor-specific, time- and temperature-dependent internalization of radiolabelled [DOTA 0 ,Tyr 3 ]octreotide in somatostatin receptor subtype 2 (sst 2 )-positive rat pancreatic tumour cell lines. In this study we have investigated the effects of differences in the amount of injected peptide on tissue distribution of 111 In-labelled [DOTA 0 ,Tyr 3 ]octreotide in normal, i.e. non-tumour-bearing, and CA20948 tumour-bearing rats. This was done in order to find the amount of peptide at which the highest uptake in target tissues is achieved, and thereby to increase the potential of radionuclide therapy while simultaneously ensuring the lowest possible radiotoxicity in normal organs. Uptake of radiolabelled [DOTA 0 ,Tyr 3 ]octreotide in sst 2 -positive organs showed different bell-shaped functions of the amount of injected peptide, being highest at 0.05 (adrenals), 0.05-0.1 (pituitary and stomach) and 0.25 (pancreas) μg. Uptake in the tumour was highest at 0.5 μg injected peptide. The highest uptake was found at peptide amounts that were lower than those reported for [ 111 In-DTPA 0 ]octreotide (d-Phe-c(Cys-Phe-d-Trp-Lys-Thr-Cys)-Thr(ol), DTPA = diethylene-triamine-penta-acetic acid), consistent with the higher receptor affinity of the first compound

p56Lck and p59Fyn Regulate CD28 Binding to Phosphatidylinositol 3-Kinase, Growth Factor Receptor-Bound Protein GRB-2, and T Cell-Specific Protein-Tyrosine Kinase ITK: Implications for T-Cell Costimulation

Science.gov (United States)

Raab, Monika; Cai, Yun-Cai; Bunnell, Stephen C.; Heyeck, Stephanie D.; Berg, Leslie J.; Rudd, Christopher E.

1995-09-01

T-cell activation requires cooperative signals generated by the T-cell antigen receptor ξ-chain complex (TCRξ-CD3) and the costimulatory antigen CD28. CD28 interacts with three intracellular proteins-phosphatidylinositol 3-kinase (PI 3-kinase), T cell-specific protein-tyrosine kinase ITK (formerly TSK or EMT), and the complex between growth factor receptor-bound protein 2 and son of sevenless guanine nucleotide exchange protein (GRB-2-SOS). PI 3-kinase and GRB-2 bind to the CD28 phosphotyrosine-based Tyr-Met-Asn-Met motif by means of intrinsic Src-homology 2 (SH2) domains. The requirement for tyrosine phosphorylation of the Tyr-Met-Asn-Met motif for SH2 domain binding implicates an intervening protein-tyrosine kinase in the recruitment of PI 3-kinase and GRB-2 by CD28. Candidate kinases include p56Lck, p59Fyn, ξ-chain-associated 70-kDa protein (ZAP-70), and ITK. In this study, we demonstrate in coexpression studies that p56Lck and p59Fyn phosphorylate CD28 primarily at Tyr-191 of the Tyr-Met-Asn-Met motif, inducing a 3- to 8-fold increase in p85 (subunit of PI 3-kinase) and GRB-2 SH2 binding to CD28. Phosphatase digestion of CD28 eliminated binding. In contrast to Src kinases, ZAP-70 and ITK failed to induce these events. Further, ITK binding to CD28 was dependent on the presence of p56Lck and is thus likely to act downstream of p56Lck/p59Fyn in a signaling cascade. p56Lck is therefore likely to be a central switch in T-cell activation, with the dual function of regulating CD28-mediated costimulation as well as TCR-CD3-CD4 signaling.

Understanding Strategy of Nitrate and Urea Assimilation in a Chinese Strain of Aureococcus anophagefferens through RNA-Seq Analysis

Science.gov (United States)

Dong, Hong-Po; Huang, Kai-Xuan; Wang, Hua-Long; Lu, Song-Hui; Cen, Jing-Yi; Dong, Yue-Lei

2014-01-01

Aureococcus anophagefferens is a harmful alga that dominates plankton communities during brown tides in North America, Africa, and Asia. Here, RNA-seq technology was used to profile the transcriptome of a Chinese strain of A. anophagefferens that was grown on urea, nitrate, and a mixture of urea and nitrate, and that was under N-replete, limited and recovery conditions to understand the molecular mechanisms that underlie nitrate and urea utilization. The number of differentially expressed genes between urea-grown and mixture N-grown cells were much less than those between urea-grown and nitrate-grown cells. Compared with nitrate-grown cells, mixture N-grown cells contained much lower levels of transcripts encoding proteins that are involved in nitrate transport and assimilation. Together with profiles of nutrient changes in media, these results suggest that A. anophagefferens primarily feeds on urea instead of nitrate when urea and nitrate co-exist. Furthermore, we noted that transcripts upregulated by nitrate and N-limitation included those encoding proteins involved in amino acid and nucleotide transport, degradation of amides and cyanates, and nitrate assimilation pathway. The data suggest that A. anophagefferens possesses an ability to utilize a variety of dissolved organic nitrogen. Moreover, transcripts for synthesis of proteins, glutamate-derived amino acids, spermines and sterols were upregulated by urea. Transcripts encoding key enzymes that are involved in the ornithine-urea and TCA cycles were differentially regulated by urea and nitrogen concentration, which suggests that the OUC may be linked to the TCA cycle and involved in reallocation of intracellular carbon and nitrogen. These genes regulated by urea may be crucial for the rapid proliferation of A. anophagefferens when urea is provided as the N source. PMID:25338000

Nitrates of rare earths

International Nuclear Information System (INIS)

Komissarova, L.N.; Pushkina, L.Ya.

1984-01-01

The systematization of experimental data with account of the last achievements in the field of studying the RE nitrate properties is realized. The methods of production, solubility in aqueous solutions structure, thermodynamic characteristics and thermal stability of nitrate hydrates, RE anhydrous and basic nitrates are considered. The data on RE nirtrate complexing in aqueous solutions are given. Binary nitrates, nitrate solvates and RE nitrate adducts with organic compounds are described. The use of RE nitrates in the course of RE production, in the processes of separation and fine cleaning of RE preparations is considered

Challenges with nitrate therapy and nitrate tolerance: prevalence, prevention, and clinical relevance.

Science.gov (United States)

Thadani, Udho

2014-08-01

Nitrate therapy has been an effective treatment for ischemic heart disease for over 100 years. The anti-ischemic and exercise-promoting benefits of sublingually administered nitrates are well established. Nitroglycerin is indicated for the relief of an established attack of angina and for prophylactic use, but its effects are short lived. In an effort to increase the duration of beneficial effects, long-acting orally administered and topical applications of nitrates have been developed; however, following their continued or frequent daily use, patients soon develop tolerance to these long-acting nitrate preparations. Once tolerance develops, patients begin losing the protective effects of the long-acting nitrate therapy. By providing a nitrate-free interval, or declining nitrate levels at night, one can overcome or reduce the development of tolerance, but cannot provide 24-h anti-anginal and anti-ischemic protection. In addition, patients may be vulnerable to occurrence of rebound angina and myocardial ischemia during periods of absent nitrate levels at night and early hours of the morning, and worsening of exercise capacity prior to the morning dose of the medication. This has been a concern with nitroglycerin patches but not with oral formulations of isosorbide-5 mononitrates, and has not been adequately studied with isosorbide dinitrate. This paper describes problems associated with nitrate tolerance, reviews mechanisms by which nitrate tolerance and loss of efficacy develop, and presents strategies to avoid nitrate tolerance and maintain efficacy when using long-acting nitrate formulations.

Data on nitrate and nitrate of Taham dam in Zanjan (Iran

Directory of Open Access Journals (Sweden)

Mohammadreza Massoudinejad

2018-04-01

Full Text Available In recent years, contamination of water resources, with pollutants such as nitrate and nitrite, has significantly increased. These compounds can have harmful effects on human health, especially children such as methemoglobinemia. The main objective of this study was to measure the concentration of nitrate and nitrite and its health-risk assessment in the rivers entering Taham dam in Zanjan. USEPA Method was used to assess the health-risk of nitrate and nitrite. According to the obtained results, the concentration of nitrate and nitrite was in the range of 0.51–14.93 mg/l and 0.001–0.061 mg/l, respectively. According to the results, the mean of the CDI for nitrate and nitrite was 9.52*10−2 and 3.63*10−4 mg/kg/day, respectively. Furthermore, the mean HI for nitrate and nitrite was 5.97*10−2 and 3.63*10−3, respectively. The concentration of nitrate and nitrite in rivers was lower than the WHO and Iran guidelines. Based on the results, the HI value in all samples was less than 1 which indicating the non-carcinogenic effects of nitrate and nitrite in these rivers. Keywords: Nitrate, Nitrite, Water quality, Dam

Identifying three-dimensional structures of autophosphorylation complexes in crystals of protein kinases

Science.gov (United States)

Xu, Qifang; Malecka, Kimberly L.; Fink, Lauren; Jordan, E. Joseph; Duffy, Erin; Kolander, Samuel; Peterson, Jeffrey; Dunbrack, Roland L.

2016-01-01

Protein kinase autophosphorylation is a common regulatory mechanism in cell signaling pathways. Crystal structures of several homomeric protein kinase complexes have a serine, threonine, or tyrosine autophosphorylation site of one kinase monomer located in the active site of another monomer, a structural complex that we call an “autophosphorylation complex.” We developed and applied a structural bioinformatics method to identify all such autophosphorylation kinase complexes in X-ray crystallographic structures in the Protein Data Bank (PDB). We identified 15 autophosphorylation complexes in the PDB, of which 5 complexes had not previously been described in the publications describing the crystal structures. These 5 consist of tyrosine residues in the N-terminal juxtamembrane regions of colony stimulating factor 1 receptor (CSF1R, Tyr561) and EPH receptor A2 (EPHA2, Tyr594), tyrosine residues in the activation loops of the SRC kinase family member LCK (Tyr394) and insulin-like growth factor 1 receptor (IGF1R, Tyr1166), and a serine in a nuclear localization signal region of CDC-like kinase 2 (CLK2, Ser142). Mutations in the complex interface may alter autophosphorylation activity and contribute to disease; therefore we mutated residues in the autophosphorylation complex interface of LCK and found that two mutations impaired autophosphorylation (T445V and N446A) and mutation of Pro447 to Ala, Gly, or Leu increased autophosphorylation. The identified autophosphorylation sites are conserved in many kinases, suggesting that, by homology, these complexes may provide insight into autophosphorylation complex interfaces of kinases that are relevant drug targets. PMID:26628682

Signaling by Kit protein-tyrosine kinase--the stem cell factor receptor.

Science.gov (United States)

Roskoski, Robert

2005-11-11

Signaling by stem cell factor and Kit, its receptor, plays important roles in gametogenesis, hematopoiesis, mast cell development and function, and melanogenesis. Moreover, human and mouse embryonic stem cells express Kit transcripts. Stem cell factor exists as both a soluble and a membrane-bound glycoprotein while Kit is a receptor protein-tyrosine kinase. The complete absence of stem cell factor or Kit is lethal. Deficiencies of either produce defects in red and white blood cell production, hypopigmentation, and sterility. Gain-of-function mutations of Kit are associated with several human neoplasms including acute myelogenous leukemia, gastrointestinal stromal tumors, and mastocytomas. Kit consists of an extracellular domain, a transmembrane segment, a juxtamembrane segment, and a protein kinase domain that contains an insert of about 80 amino acid residues. Binding of stem cell factor to Kit results in receptor dimerization and activation of protein kinase activity. The activated receptor becomes autophosphorylated at tyrosine residues that serve as docking sites for signal transduction molecules containing SH2 domains. The adaptor protein APS, Src family kinases, and Shp2 tyrosyl phosphatase bind to phosphotyrosine 568. Shp1 tyrosyl phosphatase and the adaptor protein Shc bind to phosphotyrosine 570. C-terminal Src kinase homologous kinase and the adaptor Shc bind to both phosphotyrosines 568 and 570. These residues occur in the juxtamembrane segment of Kit. Three residues in the kinase insert domain are phosphorylated and attract the adaptor protein Grb2 (Tyr703), phosphatidylinositol 3-kinase (Tyr721), and phospholipase Cgamma (Tyr730). Phosphotyrosine 900 in the distal kinase domain binds phosphatidylinositol 3-kinase which in turn binds the adaptor protein Crk. Phosphotyrosine 936, also in the distal kinase domain, binds the adaptor proteins APS, Grb2, and Grb7. Kit has the potential to participate in multiple signal transduction pathways as a result of

Leghemoglobin-derived radicals. Evidence for multiple protein-derived radicals and the initiation of peribacteroid membrane damage

DEFF Research Database (Denmark)

Moreau, S; Davies, Michael Jonathan; Mathieu, C

1996-01-01

, with the consequent generation of lipid-derived radicals. The formation of such radicals may result in the depletion of membrane antioxidants and the initiation of lipid peroxidation. This transfer of damage from the heme center via the protein surface to neighboring membranes may be of considerable biological......-derived phenoxyl radical present at Tyr-133 in the soybean protein and Tyr-138 in the lupin protein. To obtain further information on these protein radicals and their potential interaction with the physiologically important peribacteroid membrane (which surrounds the microsymbiont in vivo), EPR spin trapping......); these radicals may be side chain- or alpha-carbon-derived, their exact sites have not been determined. Some of these radicals are on the protein surface and may be key intermediates in the formation of protein dimers. These radicals have been shown to be capable of reacting with peribacteroid membrane fractions...

Insight into the intermolecular recognition mechanism between Keap1 and IKKβ combining homology modelling, protein-protein docking, molecular dynamics simulations and virtual alanine mutation.

Directory of Open Access Journals (Sweden)

Zheng-Yu Jiang

Full Text Available Degradation of certain proteins through the ubiquitin-proteasome pathway is a common strategy taken by the key modulators responsible for stress responses. Kelch-like ECH-associated protein-1(Keap1, a substrate adaptor component of the Cullin3 (Cul3-based ubiquitin E3 ligase complex, mediates the ubiquitination of two key modulators, NF-E2-related factor 2 (Nrf2 and IκB kinase β (IKKβ, which are involved in the redox control of gene transcription. However, compared to the Keap1-Nrf2 protein-protein interaction (PPI, the intermolecular recognition mechanism of Keap1 and IKKβ has been poorly investigated. In order to explore the binding pattern between Keap1 and IKKβ, the PPI model of Keap1 and IKKβ was investigated. The structure of human IKKβ was constructed by means of the homology modeling method and using reported crystal structure of Xenopus laevis IKKβ as the template. A protein-protein docking method was applied to develop the Keap1-IKKβ complex model. After the refinement and visual analysis of docked proteins, the chosen pose was further optimized through molecular dynamics simulations. The resulting structure was utilized to conduct the virtual alanine mutation for the exploration of hot-spots significant for the intermolecular interaction. Overall, our results provided structural insights into the PPI model of Keap1-IKKβ and suggest that the substrate specificity of Keap1 depend on the interaction with the key tyrosines, namely Tyr525, Tyr574 and Tyr334. The study presented in the current project may be useful to design molecules that selectively modulate Keap1. The selective recognition mechanism of Keap1 with IKKβ or Nrf2 will be helpful to further know the crosstalk between NF-κB and Nrf2 signaling.

Do nitrates differ?

Science.gov (United States)

Fung, H.-L.

1992-01-01

1 The organic nitrates all share a common biochemical and physiological mechanism of action. 2 The organic nitrates differ substantially in their pharmacologic potency and pharmacokinetics. In vitro potency differences appear larger than the corresponding in vivo activities. 3 The duration of action of organic nitrates, after a single immediate-release dose, is governed by the pharmacokinetics of the drug. However, the duration of action of available sustained-release preparations, whatever the nitrate or formulation, is limited to about 12 h, due to the development of pharmacologic tolerance. 4 Nitrates do not appear to differ in their production of undesirable effects. PMID:1633079

Involvement of the Cys-Tyr cofactor on iron binding in the active site of human cysteine dioxygenase.

Science.gov (United States)

Arjune, Sita; Schwarz, Guenter; Belaidi, Abdel A

2015-01-01

Sulfur metabolism has gained increasing medical interest over the last years. In particular, cysteine dioxygenase (CDO) has been recognized as a potential marker in oncology due to its altered gene expression in various cancer types. Human CDO is a non-heme iron-dependent enzyme, which catalyzes the irreversible oxidation of cysteine to cysteine sulfinic acid, which is further metabolized to taurine or pyruvate and sulfate. Several studies have reported a unique post-translational modification of human CDO consisting of a cross-link between cysteine 93 and tyrosine 157 (Cys-Tyr), which increases catalytic efficiency in a substrate-dependent manner. However, the reaction mechanism by which the Cys-Tyr cofactor increases catalytic efficiency remains unclear. In this study, steady-state kinetics were determined for wild type CDO and two different variants being either impaired or saturated with the Cys-Tyr cofactor. Cofactor formation in CDO resulted in an approximately fivefold increase in k cat and tenfold increase in k cat/K m over the cofactor-free CDO variant. Furthermore, iron titration experiments revealed an 18-fold decrease in K d of iron upon cross-link formation. This finding suggests a structural role of the Cys-Tyr cofactor in coordinating the ferrous iron in the active site of CDO in accordance with the previously postulated reaction mechanism of human CDO. Finally, we identified product-based inhibition and α-ketoglutarate and glutarate as CDO inhibitors using a simplified well plate-based activity assay. This assay can be used for high-throughput identification of additional inhibitors, which may contribute to understand the functional importance of CDO in sulfur amino acid metabolism and related diseases.

The intensity of tyrosine nitration is associated with selenite and selenate toxicity in Brassica juncea L.

Science.gov (United States)

Molnár, Árpád; Feigl, Gábor; Trifán, Vanda; Ördög, Attila; Szőllősi, Réka; Erdei, László; Kolbert, Zsuzsanna

2018-01-01

Selenium phytotoxicity involves processes like reactive nitrogen species overproduction and nitrosative protein modifications. This study evaluates the toxicity of two selenium forms (selenite and selenate at 0µM, 20µM, 50µM and 100µM concentrations) and its correlation with protein tyrosine nitration in the organs of hydroponically grown Indian mustard (Brassica juncea L.). Selenate treatment resulted in large selenium accumulation in both Brassica organs, while selenite showed slight root-to-shoot translocation resulting in a much lower selenium accumulation in the shoot. Shoot and root growth inhibition and cell viability loss revealed that Brassica tolerates selenate better than selenite. Results also show that relative high amounts of selenium are able to accumulate in Brassica leaves without obvious visible symptoms such as chlorosis or necrosis. The more severe phytotoxicity of selenite was accompanied by more intense protein tyrosine nitration as well as alterations in nitration pattern suggesting a correlation between the degree of Se forms-induced toxicities and nitroproteome size, composition in Brassica organs. These results imply the possibility of considering protein tyrosine nitration as novel biomarker of selenium phytotoxicity, which could help the evaluation of asymptomatic selenium stress of plants. Copyright © 2017 Elsevier Inc. All rights reserved.

Nitrate glass

International Nuclear Information System (INIS)

Kirilenko, I.A.; Vinogradov, E.E.

1977-01-01

Experimental evidence on behaviour of nitrate glasses is reviewed in terms of relationships between the presence of water in vitrescent nitrate systems and the properties of the systems. The glasses considered belong to systems of Mg(NO 3 ) 2 - Nd(NO 3 ) 3 ; Hg(NO 3 ) 2 -Nd(NO 3 ) 3 ; NaNO 3 -Mg(NO 3 ) 2 -Nd(NO 3 ) 3 ; M-Zn(NO 3 ) 3 , where M is a mixture of 20% mass NaNO 3 and 80% mass Mg(NO 3 ) 2 , and Zn is a rare earth ion. Nitrate glass is shown to be a product of dehydration. Vitrification may be regarded as a resusl of formation of molecular complexes in the chain due to hydrogen bonds of two types, i.e. water-water, or water-nicrate group. Chain formation, along with low melting points of the nitrates, hinder crystallization of nitrate melts. Provided there is enough water, this results in vitrification

Preparation of acid deficient solutions of uranyl nitrate and thorium nitrate by steam denitration

International Nuclear Information System (INIS)

Yamagishi, Shigeru; Takahashi, Yoshihisa

1996-01-01

Acid deficient heavy metal (HM) nitrate solutions are often required in the internal gelation processes for nuclear fuel fabrication. The stoichiometric HM-nitrate solutions are needed in a sol-gel process for fuel fabrication. A method for preparing such nitrate solutions with a controlled molar ratio of nitrate/metal by denitration of acid-excess nitrate solutions was developed. The denitration was conducted by bubbling a nitrate solution with a mixture of steam+Ar. It was found that steam was more effective for the denitration than Ar. The acid deficient uranyl nitrate solution with nitrate/U=1.55 was yielded by steam bubbling, while not by only Ar bubbling. As for thorium nitrate, acid deficient solutions of nitrate/Th≥3.1 were obtained by steam bubbling. (author)

Dissimilatory nitrate reduction by Aspergillus terreus isolated from the seasonal oxygen minimum zone in the Arabian Sea.

Science.gov (United States)

Stief, Peter; Fuchs-Ocklenburg, Silvia; Kamp, Anja; Manohar, Cathrine-Sumathi; Houbraken, Jos; Boekhout, Teun; de Beer, Dirk; Stoeck, Thorsten

2014-02-11

A wealth of microbial eukaryotes is adapted to life in oxygen-deficient marine environments. Evidence is accumulating that some of these eukaryotes survive anoxia by employing dissimilatory nitrate reduction, a strategy that otherwise is widespread in prokaryotes. Here, we report on the anaerobic nitrate metabolism of the fungus Aspergillus terreus (isolate An-4) that was obtained from sediment in the seasonal oxygen minimum zone in the Arabian Sea, a globally important site of oceanic nitrogen loss and nitrous oxide emission. Axenic incubations of An-4 in the presence and absence of oxygen and nitrate revealed that this fungal isolate is capable of dissimilatory nitrate reduction to ammonium under anoxic conditions. A ¹⁵N-labeling experiment proved that An-4 produced and excreted ammonium through nitrate reduction at a rate of up to 175 nmol ¹⁵NH₄⁺ g⁻¹ protein h⁻¹. The products of dissimilatory nitrate reduction were ammonium (83%), nitrous oxide (15.5%), and nitrite (1.5%), while dinitrogen production was not observed. The process led to substantial cellular ATP production and biomass growth and also occurred when ammonium was added to suppress nitrate assimilation, stressing the dissimilatory nature of nitrate reduction. Interestingly, An-4 used intracellular nitrate stores (up to 6-8 μmol NO₃⁻ g⁻¹ protein) for dissimilatory nitrate reduction. Our findings expand the short list of microbial eukaryotes that store nitrate intracellularly and carry out dissimilatory nitrate reduction when oxygen is absent. In the currently spreading oxygen-deficient zones in the ocean, an as yet unexplored diversity of fungi may recycle nitrate to ammonium and nitrite, the substrates of the major nitrogen loss process anaerobic ammonium oxidation, and the potent greenhouse gas nitrous oxide.

Evidence for in vivo phosphorylation of the Grb2 SH2-domain binding site on focal adhesion kinase by Src-family protein-tyrosine kinases.

Science.gov (United States)

Schlaepfer, D D; Hunter, T

1996-10-01

Focal adhesion kinase (FAK) is a nonreceptor protein-tyrosine kinase (PTK) that associates with integrin receptors and participates in extracellular matrix-mediated signal transduction events. We showed previously that the c-Src nonreceptor PTK and the Grb2 SH2/SH3 adaptor protein bound directly to FAK after fibronectin stimulation (D. D. Schlaepfer, S.K. Hanks, T. Hunter, and P. van der Geer, Nature [London] 372:786-791, 1994). Here, we present evidence that c-Src association with FAK is required for Grb2 binding to FAK. Using a tryptic phosphopeptide mapping approach, the in vivo phosphorylation of the Grb2 binding site on FAK (Tyr-925) was detected after fibronectin stimulation of NIH 3T3 cells and was constitutively phosphorylated in v-Src-transformed NIH 3T3 cells. In vitro, c-Src phosphorylated FAK Tyr-925 in a glutathione S-transferase-FAK C-terminal domain fusion protein, whereas FAK did not. Using epitope-tagged FAK constructs, transiently expressed in human 293 cells, we determined the effect of site-directed mutations on c-Src and Grb2 binding to FAK. Mutation of FAK Tyr-925 disrupted Grb2 binding, whereas mutation of the c-Src binding site on FAK (Tyr-397) disrupted both c-Src and Grb2 binding to FAK in vivo. These results support a model whereby Src-family PTKs are recruited to FAK and focal adhesions following integrin-induced autophosphorylation and exposure of FAK Tyr-397. Src-family binding and phosphorylation of FAK at Tyr-925 creates a Grb2 SH2-domain binding site and provides a link to the activation of the Ras signal transduction pathway. In Src-transformed cells, this pathway may be constitutively activated as a result of FAK Tyr-925 phosphorylation in the absence of integrin stimulation.

The G2/M DNA damage checkpoint inhibits mitosis through Tyr15 phosphorylation of p34cdc2 in Aspergillus nidulans.

Science.gov (United States)

Ye, X S; Fincher, R R; Tang, A; Osmani, S A

1997-01-02

It is possible to cause G2 arrest in Aspergillus nidulans by inactivating either p34cdc2 or NIMA. We therefore investigated the negative control of these two mitosis-promoting kinases after DNA damage. DNA damage caused rapid Tyr15 phosphorylation of p34cdc2 and transient cell cycle arrest but had little effect on the activity of NIMA. Dividing cells deficient in Tyr15 phosphorylation of p34cdc2 were sensitive to both MMS and UV irradiation and entered lethal premature mitosis with damaged DNA. However, non-dividing quiescent conidiospores of the Tyr15 mutant strain were not sensitive to DNA damage. The UV and MMS sensitivity of cells unable to tyrosine phosphorylate p34cdc2 is therefore caused by defects in DNA damage checkpoint regulation over mitosis. Both the nimA5 and nimT23 temperature-sensitive mutations cause an arrest in G2 at 42 degrees C. Addition of MMS to nimT23 G2-arrested cells caused a marked delay in their entry into mitosis upon downshift to 32 degrees C and this delay was correlated with a long delay in the dephosphorylation and activation of p34cdc2. Addition of MMS to nimA5 G2-arrested cells caused inactivation of the H1 kinase activity of p34cdc2 due to an increase in its Tyr15 phosphorylation level and delayed entry into mitosis upon return to 32 degrees C. However, if Tyr15 phosphorylation of p34cdc2 was prevented then its H1 kinase activity was not inactivated upon MMS addition to nimA5 G2-arrested cells and they rapidly progressed into a lethal mitosis upon release to 32 degrees C. Thus, Tyr15 phosphorylation of p34cdc2 in G2 arrests initiation of mitosis after DNA damage in A. nidulans.

Tumoral fibrosis effect on the radiation absorbed dose of {sup 177}Lu-Tyr{sup 3}-octreotate-gold nanoparticles and {sup 177}Lu-Tyr{sup 3}-octreotate radiopharmaceuticals

Energy Technology Data Exchange (ETDEWEB)

Zambrano R, O. D.

2015-07-01

In this work was comparatively evaluated the effect of tumoral fibrosis in the radiation absorbed dose of the radiopharmaceutical {sup 177}Lu-Tyr{sup 3}-octreotate with and without gold nanoparticles. For this, was used an experimental array of tumoral fibrosis and computer models based on Monte Carlo calculations to simulate tumoral micro environments without fibrosis and with fibrosis. The computer simulation code Penelope (Penetration Energy Loss of Positron and Electrons) and MCNP (Monte Carlo N-particle Transport Code System) which are based on the Monte Carlo methodology were used to create the computer models for the simulation of the transport of particles (emitted by {sup 177}Lu) in the micro environments (without fibrosis and with fibrosis) with the purpose of calculating the radiation absorbed dose in the interstitial space and in the nucleus of cancer cells. The first computational model consisted of multiple concentric spheres (as onion shells) with the radioactive source homogeneously distributed in the shell between 5 and 10 μm in diameter which represents the internalization of the radioactive source into the cell cytoplasm as it occurs in target specific radiotherapy. The concentric spheres were useful to calculate the radiation absorbed dose in depth in the models without fibrosis and with fibrosis. Furthermore, there were constructed other computer models using two different codes that simulate the transport of radiation (Penelope and MCNP). These models consist of seven spheres that represent cancer cells (HeLa cells) of 10 μm in diameter and each one of them contain another smaller sphere in the center that represents the cell nucleus. A comparison was done of the radiation absorbed dose in the nucleus of the cells, calculated with both codes, Penelope and MCNP. The radioactive source ({sup 177}Lu) used for the simulations was given to the codes by means of a convoluted spectrum of the most important beta particles (high percentage emission

Are tyrosine residues involved in the photoconversion of the water-soluble chlorophyll-binding protein of Chenopodium album?

Science.gov (United States)

Takahashi, S; Seki, Y; Uchida, A; Nakayama, K; Satoh, H

2015-05-01

Non-photosynthetic and hydrophilic chlorophyll (Chl) proteins, called water-soluble Chl-binding proteins (WSCPs), are distributed in various species of Chenopodiaceae, Amaranthaceae, Polygonaceae and Brassicaceae. Based on their photoconvertibility, WSCPs are categorised into two classes: Class I (photoconvertible) and Class II (non-photoconvertible). Chenopodium album WSCP (CaWSCP; Class I) is able to convert the chlorin skeleton of Chl a into a bacteriochlorin-like skeleton under light in the presence of molecular oxygen. Potassium iodide (KI) is a strong inhibitor of the photoconversion. Because KI attacks tyrosine residues in proteins, tyrosine residues in CaWSCP are considered to be important amino acid residues for the photoconversion. Recently, we identified the gene encoding CaWSCP and found that the mature region of CaWSCP contained four tyrosine residues: Tyr13, Tyr14, Tyr87 and Tyr134. To gain insight into the effect of the tyrosine residues on the photoconversion, we constructed 15 mutant proteins (Y13A, Y14A, Y87A, Y134A, Y13-14A, Y13-87A, Y13-134A, Y14-87A, Y14-134A, Y87-134A, Y13-14-87A, Y13-14-134A, Y13-87-134A, Y14-87-134A and Y13-14-87-134A) using site-directed mutagenesis. Amazingly, all the mutant proteins retained not only chlorophyll-binding activity, but also photoconvertibility. Furthermore, we found that KI strongly inhibited the photoconversion of Y13-14-87-134A. These findings indicated that the four tyrosine residues are not essential for the photoconversion. © 2014 German Botanical Society and The Royal Botanical Society of the Netherlands.

Efficient syntheses of climate relevant isoprene nitrates and (1R,5S)-(-)-myrtenol nitrate.

Science.gov (United States)

Bew, Sean P; Hiatt-Gipson, Glyn D; Mills, Graham P; Reeves, Claire E

2016-01-01

Here we report the chemoselective synthesis of several important, climate relevant isoprene nitrates using silver nitrate to mediate a 'halide for nitrate' substitution. Employing readily available starting materials, reagents and Horner-Wadsworth-Emmons chemistry the synthesis of easily separable, synthetically versatile 'key building blocks' (E)- and (Z)-3-methyl-4-chlorobut-2-en-1-ol as well as (E)- and (Z)-1-((2-methyl-4-bromobut-2-enyloxy)methyl)-4-methoxybenzene has been achieved using cheap, 'off the shelf' materials. Exploiting their reactivity we have studied their ability to undergo an 'allylic halide for allylic nitrate' substitution reaction which we demonstrate generates (E)- and (Z)-3-methyl-4-hydroxybut-2-enyl nitrate, and (E)- and (Z)-2-methyl-4-hydroxybut-2-enyl nitrates ('isoprene nitrates') in 66-80% overall yields. Using NOESY experiments the elucidation of the carbon-carbon double bond configuration within the purified isoprene nitrates has been established. Further exemplifying our 'halide for nitrate' substitution chemistry we outline the straightforward transformation of (1R,2S)-(-)-myrtenol bromide into the previously unknown monoterpene nitrate (1R,2S)-(-)-myrtenol nitrate.

Pre-therapeutic dosimetry and biodistribution of 86Y-DOTA-Phe1-Tyr3-octreotide versus 111In-pentetreotide in patients with advanced neuroendocrine tumours

International Nuclear Information System (INIS)

Helisch, Andreas; Foerster, Gregor J.; Reber, Helmut; Buchholz, Hans-Georg; Bartenstein, Peter; Arnold, Rudolf; Goeke, Burkhard; Weber, Matthias M.; Wiedenmann, Bertram; Pauwels, Stanislas; Haus, Ulrike; Bouterfa, Hakim

2004-01-01

For the internal radiotherapy of neuroendocrine tumours, the somatostatin analogue DOTATOC labelled with 90 Y is frequently used [ 90 Y-DOTA-Phe 1 -Tyr 3 -octreotide (SMT487-OctreoTher)]. Radiation exposure to the kidneys is critical in this therapy as it may result in renal failure. The aim of this study was to compare cumulative organ and tumour doses based upon dosimetric data acquired with the chemically identical 86 Y-DOTA-Phe 1 -Tyr 3 -octreotide (considered as the gold standard) and the commercially available 111 In-pentetreotide. The cumulative organ and tumour doses for the therapeutic administration of 13.32 GBq 90 Y-DOTA-Phe 1 -Tyr 3 -octreotide (three cycles, each of 4.44 GBq) were estimated based on the MIRD concept (MIRDOSE 3.1 and IMEDOSE). Patients with a cumulative kidney dose exceeding 27 Gy had to be excluded from subsequent therapy with 90 Y-DOTA-Phe 1 -Tyr 3 -octreotide, in accordance with the directives of the German radiation protection authorities. The range of doses (mGy/MBq 90 Y-DOTA-Phe 1 -Tyr 3 -octreotide) for kidneys, spleen, liver and tumour masses was 0.6-2.8, 1.5-4.2, 0.3-1.3 and 2.1-29.5 ( 86 Y-DOTA-Phe 1 -Tyr 3 -octreotide), respectively, versus 1.3-3.0, 1.8-4.4, 0.2-0.8 and 1.4-19.7 ( 111 In-pentetreotide), with wide inter-subject variability. Despite renal protection with amino acid infusions, estimated cumulative kidney doses in two patients exceeded 27 Gy. Compared with 86 Y-DOTA-Phe 1 -Tyr 3 -octreotide, dosimetry with 111 In-pentetreotide overestimated doses to kidneys and spleen, whereas the radiation dose to the tumour-free liver was underestimated. However, both dosimetric approaches detected the two patients with an exceptionally high radiation burden to the kidneys that carried a potential risk of renal failure following radionuclide therapy. (orig.)

Pre-therapeutic dosimetry and biodistribution of 86Y-DOTA-Phe1-Tyr3-octreotide versus 111In-pentetreotide in patients with advanced neuroendocrine tumours.

Science.gov (United States)

Helisch, Andreas; Förster, Gregor J; Reber, Helmut; Buchholz, Hans-Georg; Arnold, Rudolf; Göke, Burkhard; Weber, Matthias M; Wiedenmann, Bertram; Pauwels, Stanislas; Haus, Ulrike; Bouterfa, Hakim; Bartenstein, Peter

2004-10-01

For the internal radiotherapy of neuroendocrine tumours, the somatostatin analogue DOTATOC labelled with 90Y is frequently used [90Y-DOTA-Phe1-Tyr3)-octreotide (SMT487-OctreoTher)]. Radiation exposure to the kidneys is critical in this therapy as it may result in renal failure. The aim of this study was to compare cumulative organ and tumour doses based upon dosimetric data acquired with the chemically identical 86Y-DOTA-Phe1-Tyr3-octreotide (considered as the gold standard) and the commercially available 111In-pentetreotide. The cumulative organ and tumour doses for the therapeutic administration of 13.32 GBq 90Y-DOTA-Phe1-Tyr3-octreotide (three cycles, each of 4.44 GBq) were estimated based on the MIRD concept (MIRDOSE 3.1 and IMEDOSE). Patients with a cumulative kidney dose exceeding 27 Gy had to be excluded from subsequent therapy with 90Y-DOTA-Phe1-Tyr3-octreotide, in accordance with the directives of the German radiation protection authorities. The range of doses (mGy/MBq 90Y-DOTA-Phe1-Tyr3-octreotide) for kidneys, spleen, liver and tumour masses was 0.6-2.8, 1.5-4.2, 0.3-1.3 and 2.1-29.5 (86Y-DOTA-Phe1-Tyr3-octreotide), respectively, versus 1.3-3.0, 1.8-4.4, 0.2-0.8 and 1.4-19.7 (111In-pentetreotide), with wide inter-subject variability. Despite renal protection with amino acid infusions, estimated cumulative kidney doses in two patients exceeded 27 Gy. Compared with 86Y-DOTA-Phe1-Tyr3-octreotide, dosimetry with 111In-pentetreotide overestimated doses to kidneys and spleen, whereas the radiation dose to the tumour-free liver was underestimated. However, both dosimetric approaches detected the two patients with an exceptionally high radiation burden to the kidneys that carried a potential risk of renal failure following radionuclide therapy.

Solubility isotherms in ternary systems of samarium nitrate, water and nitrates of amidopyrine, benzotriazole

International Nuclear Information System (INIS)

Starikova, L.I.

1991-01-01

Solubility in the system of samarium nitrate-amidopyrine nitrate-water at 25 and 50 deg C was studied. Solubility isotherms consist of three branches, corresponding to crystallization of samarium nitrate tetrahydrate, amidopyrine nitrate and congruently soluble compounds of Sm(NO 3 ) 3 · 2C 13 H 17 ON 3 ·HNO 3 composition. Its thermal behaviour was studied. The system of samarium nitrate-benzotriazole nitrate-water is referred to eutonic type

Volatile compounds and some physico-chemical properties of pastırma produced with different nitrate levels

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Ahmet Akköse

2017-08-01

Full Text Available Objective The aim of the study was to evaluate the effects of different nitrate levels (150, 300, 450, and 600 ppm KNO3 on the volatile compounds and some other properties of pastırma. Methods Pastırma samples were produced under the controlled condition and analyses of volatile compounds, and thiobarbituric acid reactive substances (TBARS as an indicator of lipid oxidation, non-protein nitrogenous matter content as an indicator of proteolysis, color and residual nitrite were carried out on the final product. The profile of volatile compounds of pastırma samples was analyzed by gas chromatography/mass spectrometry using a solid phase microextraction. Results Nitrate level had a significant effect on pH value (p<0.05 and a very significant effect on TBARS value (p<0.01. No significant differences were determined in terms of aw value, non-protein nitrogenous substance content, color and residual nitrite between pastırma groups produced by using different nitrate levels. Nitrate level had a significant (p<0.05 or a very significant (p<0.01 effect on some volatile compounds. It was determined that the amounts and counts of volatile compounds were lower in the 450 and especially 600 ppm nitrate levels than 150 and 300 ppm nitrate levels (p<0.05. While the use of 600 ppm nitrate did not cause an increase in residual nitrite levels, the use of 150 ppm nitrate did not negatively affect the color of pastırma. However, the levels of volatile compounds decreased with an increasing level of nitrate. Conclusion The use of 600 ppm nitrate is not a risk in terms of residual nitrite in pastırma produced under controlled condition, however, this level is not suitable due to decrease in the amount of volatile compounds.

Nitration of naphthalene and remarks on the mechanism of electrophilic aromatic nitration*

Science.gov (United States)

Olah, George A.; Narang, Subhash C.; Olah, Judith A.

1981-01-01

Naphthalene was nitrated with a variety of nitrating agents. Comparison of data with Perrin's electrochemical nitration [Perrin, C. L. (1977) J. Am. Chem. Soc. 99, 5516-5518] shows that nitration of naphthalene gives an α-nitronaphthalene to β-nitronaphthalene ratio that varies between 9 and 29 and is thus not constant. Perrin's data, therefore, are considered to be inconclusive evidence for the proposed one-electron transfer mechanism for the nitration of naphthalene and other reactive aromatics. Moodie and Schoefield [Hoggett, J. G., Moodie, R. B., Penton, J. R. & Schoefield, K. (1971) Nitration and Aromatic Reactivity (Cambridge Univ. Press, London)], as well as Perrin, independently concluded that, in the general scheme of nitration of reactive aromatics, there is the necessity to introduce into the classical Ingold mechanism an additional step involving a distinct intermediate preceding the formation of the Wheland intermediate (σ complexes). This view coincides with our two-step mechanistic picture [Kuhn, S. J. & Olah, G. A. (1961) J. Am. Chem. Soc. 83, 4564-4571] of the nitronium salt nitration of aromatic hydrocarbons (including benzene and toluene), in which low substrate selectivity but high positional selectivity was found, indicating the independence of substrate from positional selectivity. PMID:16593026

Nitration of naphthalene and remarks on the mechanism of electrophilic aromatic nitration.

Science.gov (United States)

Olah, G A; Narang, S C; Olah, J A

1981-06-01

Naphthalene was nitrated with a variety of nitrating agents. Comparison of data with Perrin's electrochemical nitration [Perrin, C. L. (1977) J. Am. Chem. Soc. 99, 5516-5518] shows that nitration of naphthalene gives an alpha-nitronaphthalene to beta-nitronaphthalene ratio that varies between 9 and 29 and is thus not constant. Perrin's data, therefore, are considered to be inconclusive evidence for the proposed one-electron transfer mechanism for the nitration of naphthalene and other reactive aromatics. Moodie and Schoefield [Hoggett, J. G., Moodie, R. B., Penton, J. R. & Schoefield, K. (1971) Nitration and Aromatic Reactivity (Cambridge Univ. Press, London)], as well as Perrin, independently concluded that, in the general scheme of nitration of reactive aromatics, there is the necessity to introduce into the classical Ingold mechanism an additional step involving a distinct intermediate preceding the formation of the Wheland intermediate (sigma complexes). This view coincides with our two-step mechanistic picture [Kuhn, S. J. & Olah, G. A. (1961) J. Am. Chem. Soc. 83, 4564-4571] of the nitronium salt nitration of aromatic hydrocarbons (including benzene and toluene), in which low substrate selectivity but high positional selectivity was found, indicating the independence of substrate from positional selectivity.

The NtrY-NtrX two-component system is involved in controlling nitrate assimilation in Herbaspirillum seropedicae strain SmR1.

Science.gov (United States)

Bonato, Paloma; Alves, Lysangela R; Osaki, Juliana H; Rigo, Liu U; Pedrosa, Fabio O; Souza, Emanuel M; Zhang, Nan; Schumacher, Jörg; Buck, Martin; Wassem, Roseli; Chubatsu, Leda S

2016-11-01

Herbaspirillum seropedicae is a diazotrophic β-Proteobacterium found endophytically associated with gramineae (Poaceae or graminaceous plants) such as rice, sorghum and sugar cane. In this work we show that nitrate-dependent growth in this organism is regulated by the master nitrogen regulatory two-component system NtrB-NtrC, and by NtrY-NtrX, which functions to specifically regulate nitrate metabolism. NtrY is a histidine kinase sensor protein predicted to be associated with the membrane and NtrX is the response regulator partner. The ntrYntrX genes are widely distributed in Proteobacteria. In α-Proteobacteria they are frequently located downstream from ntrBC, whereas in β-Proteobacteria these genes are located downstream from genes encoding an RNA methyltransferase and a proline-rich protein with unknown function. The NtrX protein of α-Proteobacteria has an AAA+ domain, absent in those from β-Proteobacteria. An ntrY mutant of H. seropedicae showed the wild-type nitrogen fixation phenotype, but the nitrate-dependent growth was abolished. Gene fusion assays indicated that NtrY is involved in the expression of genes coding for the assimilatory nitrate reductase as well as the nitrate-responsive two-component system NarX-NarL (narK and narX promoters, respectively). The purified NtrX protein was capable of binding the narK and narX promoters, and the binding site at the narX promoter for the NtrX protein was determined by DNA footprinting. In silico analyses revealed similar sequences in other promoter regions of H. seropedicae that are related to nitrate assimilation, supporting the role of the NtrY-NtrX system in regulating nitrate metabolism in H. seropedicae. © 2016 Federation of European Biochemical Societies.

Vacuolar respiration of nitrate coupled to energy conservation in filamentous Beggiatoaceae.

Science.gov (United States)

Beutler, Martin; Milucka, Jana; Hinck, Susanne; Schreiber, Frank; Brock, Jörg; Mussmann, Marc; Schulz-Vogt, Heide N; de Beer, Dirk

2012-11-01

We show that the nitrate storing vacuole of the sulfide-oxidizing bacterium Candidatus Allobeggiatoa halophila has an electron transport chain (ETC), which generates a proton motive force (PMF) used for cellular energy conservation. Immunostaining by antibodies showed that cytochrome c oxidase, an ETC protein and a vacuolar ATPase are present in the vacuolar membrane and cytochrome c in the vacuolar lumen. The effect of different inhibitors on the vacuolar pH was studied by pH imaging. Inhibition of vacuolar ATPases and pyrophosphatases resulted in a pH decrease in the vacuole, showing that the proton gradient over the vacuolar membrane is used for ATP and pyrophosphate generation. Blockage of the ETC decreased the vacuolar PMF, indicating that the proton gradient is build up by an ETC. Furthermore, addition of nitrate resulted in an increase of the vacuolar PMF. Inhibition of nitrate reduction, led to a decreased PMF. Nitric oxide was detected in vacuoles of cells exposed to nitrate showing that nitrite, the product of nitrate reduction, is reduced inside the vacuole. These findings show consistently that nitrate respiration contributes to the high proton concentration within the vacuole and the PMF over the vacuolar membrane is actively used for energy conservation. © 2012 Society for Applied Microbiology and Blackwell Publishing Ltd.

MITIGASI PELINDIAN NITRAT PADA TANAH INCEPTISOL MELALUI PEMANFAATAN BAHAN NITRAT INHIBITOR ALAMI

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Joko Pramono

2012-05-01

Full Text Available Mitigation of Nitrate Leaching in Inceptisol Soil Through the Use of Natural Nitrate Inhibitor ABSTRAK Pelindian NO3- merupakan salah satu mekanisme kehilangan N dalam aktivitas pertanian, yang dapat berdampak terhadap pencemaran lingkungan. Tujuan dari penelitian adalah untuk mengetahui penggunaan bahan alami sebagai nitrat inhibitor terhadap pelindian nitrat pada tanah Inceptisol. Pada penelitian ini diuji tiga jenis bahan nitrat inhibitor (NI alami yang berasal dari; serbuk biji Mimba (SBM, serbuk kulit kayu bakau (SKKB, dan serbuk daun kopi (SDK,yang dikombinasikan dengan tiga taraf dosis NI, yaitu: 20 %, 30 % dan 40 % dari urea yang diberikan, dan ditambah satu perlakuan kontrol tanpa NI. Bahan nitrat inhibitor diberikan bersama urea pada permukaan tanah dalam pot percobaan yang telah dibasahi dengan air suling. Hasil penelitian menunjukkan bahwa bahan NI yang berbeda memberikan respon terhadap penghambatan nitrifi kasi yang berbeda. Bahan NI yang berasal dari serbuk biji mimba memberikan tingkat penghambatan tertinggi sebesar (25,6 %, serbuk kulit kayu bakau sebesar (19,1 %, dan serbuk daun kopi sebesar 11,8 %. Bahan NI alami mampu menghambat nitrifi kasi melalui penghambatan pertumbuhan bakteri nitrifi kasi (pengoksida ammonium yang bersifat sementara pada kisaran 7-14 hari setelah aplikasi. Perlakuan berbagai bahan dan dosis NI mampu menekan pelindian nitrat rata-rata pada kisaran antara 56,6 sampai 62,8 % dan berbeda sangat nyata terhadap perlakuan kontrol tanpa NI. Bahan NI yang mampu menurunkan rata-rata pelindian nitrat pada pengamatan 14 hari setelah aplikasi tertinggi adalah SBM sebesar 74,15 %. Dosis optimal dua bahan NI terpilih yang menunjukkan kinerja penghambatan nitrifi kasi terbaik (SBM dan SKKB pada 7 hsa, masing-masing 18,30 % (R2 = 0,694 dan 21,67 % (R2=0.691 dari dosis urea yang diberikan. Kata kunci: Nitrifi kasi, nitrat inhibitor, pelindian nitrat ABSTRACT NO3 - leaching is one mechanism of N reduction in agricultural

Nitrate pollution of groundwater

International Nuclear Information System (INIS)

Heaton, T.H.E.

1986-01-01

Concern about the possible health risks associated with the consumption of nitrate has led many countries, including South Africa, to propose that 10mg of nitrogen (as nitrate or nitrite) per liter should be the maximum allowable limit for domestic water supplies. Groundwater in certain parts of South Africa and Namibia contains nitrate in concentrations which exceed this limit. The CSIR's Natural Isotope Division has been studying the nitrogen isotope composition of the nitrate as an aid to investigation into the sources of this nitrate contamination

Impact of Sulfide on Nitrate Conversion in Eutrophic Nitrate-Rich Marine Sludge

DEFF Research Database (Denmark)

Schwermer, Carsten U.; Krieger, Bärbel; Lavik, Gaute

2006-01-01

IMPACT OF SULFIDE ON NITRATE CONVERSION IN EUTROPHIC NITRATE-RICH MARINE SLUDGE C.U. Schwermer 1, B.U. Krieger 2, G. Lavik 1, A. Schramm 3, J. van Rijn 4, D. de Beer 1, D. Minz 5, E. Cytryn 4, M. Kuypers 1, A. Gieseke 1 1 Max Planck Institute for Marine Microbiology, Bremen, Germany; 2 Dept...... nitrate conversion from denitrification to dissimilatory nitrate-reduction to ammonium (DNRA). In situ microsensor profiling in stagnant sludge revealed the typical stratification of nitrate reduction on top of sulfate reduction. Increasing the bulk nitrate concentration lead to a downward shift....... Our results show that the presence of sulfide generally decreased growth rates but increased N2O production. We conclude that sulfide plays a key role in causing incomplete denitrification, presumably by inhibiting the N2O reductase, and enhancing DNRA compared to denitrification.  ...

Proton NMR studies of Cucurbita maxima trypsin inhibitors: evidence for pH-dependent conformational change and His25-Tyr27 interaction.

Science.gov (United States)

Krishnamoorthi, R; Lin, C L; Gong, Y X; VanderVelde, D; Hahn, K

1992-01-28

A pH-dependent His25-Tyr27 interaction was demonstrated in the case of Cucurbita maxima trypsin inhibitors (CMTI-I and CMTI-III) by means of nuclear magnetic resonance (NMR) spectroscopy. pH titration, line widths, peak shapes, deuterium exchange kinetics, and two-dimensional nuclear Overhauser effect spectroscopy (NOESY) were employed to characterize a conformational change involving Tyr27, which was shown to be triggered by deprotonation of His25 around pH 6. A hydrogen bond is proposed to be formed between N epsilon of His25 and OH of Tyr27, as a distance between the atoms, His25 N epsilon and Tyr27 OH, of 3.02 A is consistent with a model built with NOE-derived distance constraints. Both the X-ray [Bode, W., Greyling, J.H., Huber, R., Otlewski, J., & Wilusz, T. (1989) FEBS Lett. 242, 282-292] and NMR [Holak, T.A., Gondol, D., Otlewski, J., & Wilusz, T. (1989) J. Mol. Biol. 210, 635-648] structures of CMTI-I at low pH (4.7-5.3) rule out such an interaction between the two aromatic rings, as the ring planes are oriented about 10 A away from each other. The presently characterized relative orientations of His25 and Tyr27 are of functional significance, as these residues make contact with the enzyme in the enzyme-inhibitor complex. Furthermore, trypsin assay and inhibitor-binding studies showed that conformations of trypsin and the squash inhibitor were functionally relevant only in the pH range 6-8. The pKa of His25 was determined and found to be influenced by Glu9/Lys substitution and by the hydrolysis of the reactive-site peptide bond between Arg5 and Ile6.(ABSTRACT TRUNCATED AT 250 WORDS)

SH2 Domains Serve as Lipid-Binding Modules for pTyr-Signaling Proteins.

Science.gov (United States)

Park, Mi-Jeong; Sheng, Ren; Silkov, Antonina; Jung, Da-Jung; Wang, Zhi-Gang; Xin, Yao; Kim, Hyunjin; Thiagarajan-Rosenkranz, Pallavi; Song, Seohyeon; Yoon, Youngdae; Nam, Wonhee; Kim, Ilshin; Kim, Eui; Lee, Dong-Gyu; Chen, Yong; Singaram, Indira; Wang, Li; Jang, Myoung Ho; Hwang, Cheol-Sang; Honig, Barry; Ryu, Sungho; Lorieau, Justin; Kim, You-Me; Cho, Wonhwa

2016-04-07

The Src-homology 2 (SH2) domain is a protein interaction domain that directs myriad phosphotyrosine (pY)-signaling pathways. Genome-wide screening of human SH2 domains reveals that ∼90% of SH2 domains bind plasma membrane lipids and many have high phosphoinositide specificity. They bind lipids using surface cationic patches separate from pY-binding pockets, thus binding lipids and the pY motif independently. The patches form grooves for specific lipid headgroup recognition or flat surfaces for non-specific membrane binding and both types of interaction are important for cellular function and regulation of SH2 domain-containing proteins. Cellular studies with ZAP70 showed that multiple lipids bind its C-terminal SH2 domain in a spatiotemporally specific manner and thereby exert exquisite spatiotemporal control over its protein binding and signaling activities in T cells. Collectively, this study reveals how lipids control SH2 domain-mediated cellular protein-protein interaction networks and suggest a new strategy for therapeutic modulation of pY-signaling pathways. Copyright © 2016 Elsevier Inc. All rights reserved.

Ammonium nitrate explosion hazards

Directory of Open Access Journals (Sweden)

Negovanović Milanka

2015-01-01

Full Text Available Ammonium nitrate (AN primarily is used as a fertilizer but it is also very important compound in the production of industrial explosives. The application of ammonium nitrate in the production of industrial explosives was related with the early era of Nobel dynamite and widely increased with the appearance of blasting agents such as ANFO and Slurry, in the middle of the last Century. Throughout the world millions of tons of ammonium nitrate are produced annually and handled without incident. Although ammonium nitrate generally is used safely, accidental explosions involving AN have high impact resulting in loss of lives and destruction of property. The paper presents the basic properties of ammonium nitrate as well as hazards in handling of ammonium nitrate in order to prevent accidents. Several accidents with explosions of ammonium nitrate resulted in catastrophic consequences are listed in the paper as examples of non-compliance with prescribed procedures.

Ternary systems, consist of erbium nitrates, water and nitrates of pyridines, quinolines

International Nuclear Information System (INIS)

Starikova, L.I.; Zhuravlev, E.F.; Khalfina, L.R.

1979-01-01

At 25 and 50 deg C investigated is solubility of solid phases in ternary water salt systems: erbium nitrate-pyridine nitrate-water; erbium nitrate-quinoline nitrate-water. Formation of congruently soluble compounds of the Er(NO 3 ) 3 x2C 5 H 5 NxHNO 3 , Er(NO 3 ) 3 x2C 9 H 7 NxHNO 3 x4H 2 O composition is established. X-ray phase and thermogravimetric analyses have been carried out

Density and electrical conductivity of molten salts. Comparative study of binary mixtures of alkali nitrates with silver nitrate and with thallium nitrate

International Nuclear Information System (INIS)

Brillant, S.

1968-01-01

The choice of methods and the number of measurements made enable us to give results on the density and electrical conductivity of molten binary mixtures, alkali nitrate and silver nitrate, and alkali nitrate and thallium nitrate, in the form of equations. The deviations from linearity of the volume and the molar conductivity are determined by calculating the corresponding excess values whose variations are analyzed as a function of the Tobolsky parameter. The absence of any relationship in the sign of the entropy and the excess volume is justified. It is shown that the silver and thallium nitrates, in contrast to the thermodynamic properties, behave as the alkali nitrates in so far as the excess conductivity is concerned. This result is confirmed by the study of changes in the activation enthalpy for the partial molar conductivity; this study also shows the particular behaviour of lithium nitrate. (author) [fr

Neoadjuvant Treatment of Nonfunctioning Pancreatic Neuroendocrine Tumors with [177Lu-DOTA0,Tyr3]Octreotate

NARCIS (Netherlands)

van Vliet, Esther I.; van Eijck, Casper H.; de Krijger, Ronald R.; Nieveen van Dijkum, Elisabeth J.; Teunissen, Jaap J.; Kam, Boen L.; de Herder, Wouter W.; Feelders, Richard A.; Bonsing, Bert A.; Brabander, Tessa; Krenning, Eric P.; Kwekkeboom, Dik J.

2015-01-01

Pancreatic neuroendocrine tumors (NETs) are rare neoplasms for which surgery has almost the only potential for cure. When surgery is not possible because of tumor size and vascular involvement, neoadjuvant treatment with [(177)Lu-DOTA(0),Tyr(3)]octreotate ((177)Lu-octreotate) may be an option. We

49 CFR 176.410 - Division 1.5 materials, ammonium nitrate and ammonium nitrate mixtures.

Science.gov (United States)

2010-10-01

... 49 Transportation 2 2010-10-01 2010-10-01 false Division 1.5 materials, ammonium nitrate and ammonium nitrate mixtures. 176.410 Section 176.410 Transportation Other Regulations Relating to... nitrate and ammonium nitrate mixtures. (a) This section prescribes requirements to be observed with...

Organic Nitrate Therapy, Nitrate Tolerance, and Nitrate-Induced Endothelial Dysfunction: Emphasis on Redox Biology and Oxidative Stress.

Science.gov (United States)

Daiber, Andreas; Münzel, Thomas

2015-10-10

Organic nitrates, such as nitroglycerin (GTN), isosorbide-5-mononitrate and isosorbide dinitrate, and pentaerithrityl tetranitrate (PETN), when given acutely, have potent vasodilator effects improving symptoms in patients with acute and chronic congestive heart failure, stable coronary artery disease, acute coronary syndromes, or arterial hypertension. The mechanisms underlying vasodilation include the release of •NO or a related compound in response to intracellular bioactivation (for GTN, the mitochondrial aldehyde dehydrogenase [ALDH-2]) and activation of the enzyme, soluble guanylyl cyclase. Increasing cyclic guanosine-3',-5'-monophosphate (cGMP) levels lead to an activation of the cGMP-dependent kinase I, thereby causing the relaxation of the vascular smooth muscle by decreasing intracellular calcium concentrations. The hemodynamic and anti-ischemic effects of organic nitrates are rapidly lost upon long-term (low-dose) administration due to the rapid development of tolerance and endothelial dysfunction, which is in most cases linked to increased intracellular oxidative stress. Enzymatic sources of reactive oxygen species under nitrate therapy include mitochondria, NADPH oxidases, and an uncoupled •NO synthase. Acute high-dose challenges with organic nitrates cause a similar loss of potency (tachyphylaxis), but with distinct pathomechanism. The differences among organic nitrates are highlighted regarding their potency to induce oxidative stress and subsequent tolerance and endothelial dysfunction. We also address pleiotropic effects of organic nitrates, for example, their capacity to stimulate antioxidant pathways like those demonstrated for PETN, all of which may prevent adverse effects in response to long-term therapy. Based on these considerations, we will discuss and present some preclinical data on how the nitrate of the future should be designed.

Organic Nitrate Therapy, Nitrate Tolerance, and Nitrate-Induced Endothelial Dysfunction: Emphasis on Redox Biology and Oxidative Stress

Science.gov (United States)

2015-01-01

Abstract Organic nitrates, such as nitroglycerin (GTN), isosorbide-5-mononitrate and isosorbide dinitrate, and pentaerithrityl tetranitrate (PETN), when given acutely, have potent vasodilator effects improving symptoms in patients with acute and chronic congestive heart failure, stable coronary artery disease, acute coronary syndromes, or arterial hypertension. The mechanisms underlying vasodilation include the release of •NO or a related compound in response to intracellular bioactivation (for GTN, the mitochondrial aldehyde dehydrogenase [ALDH-2]) and activation of the enzyme, soluble guanylyl cyclase. Increasing cyclic guanosine-3′,-5′-monophosphate (cGMP) levels lead to an activation of the cGMP-dependent kinase I, thereby causing the relaxation of the vascular smooth muscle by decreasing intracellular calcium concentrations. The hemodynamic and anti-ischemic effects of organic nitrates are rapidly lost upon long-term (low-dose) administration due to the rapid development of tolerance and endothelial dysfunction, which is in most cases linked to increased intracellular oxidative stress. Enzymatic sources of reactive oxygen species under nitrate therapy include mitochondria, NADPH oxidases, and an uncoupled •NO synthase. Acute high-dose challenges with organic nitrates cause a similar loss of potency (tachyphylaxis), but with distinct pathomechanism. The differences among organic nitrates are highlighted regarding their potency to induce oxidative stress and subsequent tolerance and endothelial dysfunction. We also address pleiotropic effects of organic nitrates, for example, their capacity to stimulate antioxidant pathways like those demonstrated for PETN, all of which may prevent adverse effects in response to long-term therapy. Based on these considerations, we will discuss and present some preclinical data on how the nitrate of the future should be designed. Antioxid. Redox Signal. 23, 899–942. PMID:26261901

Efficiency of nitrate uptake in spinach : impact of external nitrate concentration and relative growth rate on nitrate influx and efflux

NARCIS (Netherlands)

Ter Steege, MW; Stulen, [No Value; Wiersema, PK; Posthumus, F; Vaalburg, W

1999-01-01

Regulation of nitrate influx and efflux in spinach (Spinacia oleracea L., cv. Subito), was studied in short-term label experiments with N-13- and N-15-nitrate. Nitrate fluxes were examined in relation to the N demand for growth, defined as relative growth rate (RGR) times plant N concentration.

Solvent selection and optimization of α-chymotrypsin-catalyzed synthesis of N-Ac-Phe-Tyr-NH2 using mixture design and response surface methodology.

Science.gov (United States)

Hu, Shih-Hao; Kuo, Chia-Hung; Chang, Chieh-Ming J; Liu, Yung-Chuan; Chiang, Wen-Dee; Shieh, Chwen-Jen

2012-01-01

A peptide, N-Ac-Phe-Tyr-NH(2) , with angiotensin I-converting enzyme (ACE) inhibitor activity was synthesized by an α-chymotrypsin-catalyzed condensation reaction of N-acetyl phenylalanine ethyl ester (N-Ac-Phe-OEt) and tyrosinamide (Tyr-NH(2) ). Three kinds of solvents: a Tris-HCl buffer (80 mM, pH 9.0), dimethylsulfoxide (DMSO), and acetonitrile were employed in this study. The optimum reaction solvent component was determined by simplex centroid mixture design. The synthesis efficiency was enhanced in an organic-aqueous solvent (Tris-HCl buffer: DMSO: acetonitrile = 2:1:1) in which 73.55% of the yield of N-Ac-Phe-Tyr-NH(2) could be achieved. Furthermore, the effect of reaction parameters on the yield was evaluated by response surface methodology (RSM) using a central composite rotatable design (CCRD). Based on a ridge max analysis, the optimum condition for this peptide synthesis included a reaction time of 7.4 min, a reaction temperature of 28.1°C, an enzyme activity of 98.9 U, and a substrate molar ratio (Phe:Tyr) of 1:2.8. The predicted and the actual (experimental) yields were 87.6 and 85.5%, respectively. The experimental design and RSM performed well in the optimization of synthesis of N-Ac-Phe-Tyr-NH(2) , so it is expected to be an effective method for obtaining a good yield of enzymatic peptide. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2012. Copyright © 2012 American Institute of Chemical Engineers (AIChE).

Sodium nitrate-cerium nitrate-water ternary system at 25 deg C

International Nuclear Information System (INIS)

Fedorenko, T.P.; Onishchenko, M.K.

1978-01-01

Solubility isotherm of sodium nitrate-cerium nitrate-water system at 25 deg C consists of three crystallization branches of initial salts and double compound of the composition 2NaNO 3 xCe(NO 3 ) 3 x2H 2 O. Sodium nitrate introduced in the solution strengthens complexing. Physico-chemical characteristics are in a good agreement with solubility curve

A new class of organic nitrates: investigations on bioactivation, tolerance and cross-tolerance phenomena.

Science.gov (United States)

Schuhmacher, S; Schulz, E; Oelze, M; König, A; Roegler, C; Lange, K; Sydow, L; Kawamoto, T; Wenzel, P; Münzel, T; Lehmann, J; Daiber, A

2009-09-01

The chronic use of organic nitrates is limited by serious side effects including oxidative stress, nitrate tolerance and/or endothelial dysfunction. The side effects and potency of nitroglycerine depend on mitochondrial aldehyde dehydrogenase (ALDH-2). We sought to determine whether this concept can be extended to a new class of organic nitrates with amino moieties (aminoalkyl nitrates). Vasodilator potency of the organic nitrates, in vitro tolerance and in vivo tolerance (after continuous infusion for 3 days) were assessed in wild-type and ALDH-2 knockout mice by isometric tension studies. Mitochondrial oxidative stress was analysed by L-012-dependent chemiluminescence and protein tyrosine nitration. Aminoethyl nitrate (AEN) showed an almost similar potency to glyceryl trinitrate (GTN), even though it is only a mononitrate. AEN-dependent vasodilatation was mediated by cGMP and nitric oxide. In contrast to triethanolamine trinitrate (TEAN) and GTN, AEN bioactivation did not depend on ALDH-2 and caused no in vitro tolerance. In vivo treatment with TEAN and GTN, but not with AEN, induced cross-tolerance to acetylcholine (ACh)-dependent and GTN-dependent relaxation. Although all nitrates tested induced tolerance to themselves, only TEAN and GTN significantly increased mitochondrial oxidative stress in vitro and in vivo. The present results demonstrate that not all high potency nitrates are bioactivated by ALDH-2 and that high potency of a given nitrate is not necessarily associated with induction of oxidative stress or nitrate tolerance. Obviously, there are distinct pathways for bioactivation of organic nitrates, which for AEN may involve xanthine oxidoreductase rather than P450 enzymes.

Formation, Evaporation, and Hydrolysis of Organic Nitrates from Nitrate Radical Oxidation of Monoterpenes

Science.gov (United States)

Ng, N. L.; Takeuchi, M.; Eris, G.; Berkemeier, T.; Boyd, C.; Nah, T.; Xu, L.

2017-12-01

Organic nitrates play an important role in the cycling of NOx and secondary organic aerosol (SOA) formation, yet their formation mechanisms and fates remain highly uncertain. The interactions of biogenic VOCs with NO3 radicals represent a direct way for positively linking anthropogenic and biogenic emissions. Results from ambient studies suggest that organic nitrates have a relatively short lifetime, though corresponding laboratory data are limited. SOA and organic nitrates produced at night may evaporate the following morning due to increasing temperatures or dilution of semi-volatile compounds. Once formed, organic nitrates can also undergo hydrolysis in the presence of particle water. In this work, we investigate the formation, evaporation, and hydrolysis of organic nitrates generated from the nitrate radical oxidation of a-pinene, b-pinene, and limonene. Experiments are conducted in the Georgia Environmental Chamber facility (GTEC) under dry and humid conditions and different temperatures. Experiments are also designed to probe different peroxy radical pathways (RO2+HO2 vs RO2+NO3). Speciated gas-phase and particle-phase organic nitrates are continuously monitored by a Filter Inlet for Gases and AEROsols High Resolution Time-of-Flight Chemical Ionization Mass Spectrometer (FIGAERO-HR-ToF-CIMS). Bulk aerosol composition is measured by a High Resolution Time-of-Flight Aerosol Mass Spectrometer (HR-ToF-AMS). A large suite of highly oxygenated gas- and particle-phase organic nitrates are formed rapidly. We find a resistance to aerosol evaporation when it is heated. The extent of organic nitrate hydrolysis in the humid experiments is evaluated. The dynamics of the speciated organic nitrates over the course of the experiments will also be discussed. Results from this chamber study provide fundamental data for understanding the dynamics of organic nitrate aerosols over its atmospheric lifetime.

The influence of Glyceria maxima and nitrate input on the composition and nitrate metabolism of the dissimilatory nitrate-reducing bacterial community

NARCIS (Netherlands)

Nijburg, J.W.; Laanbroek, H.J.

1997-01-01

The influence of nitrate addition and the presence of Glyceria maxima (reed sweetgrass) on the composition and nitrate metabolism of the dissimilatory nitrate-reducing bacterial community was investigated. Anoxic freshwater sediment was incubated in pots with or without G. maxima and with or

Mutation of androgen receptor N-terminal phosphorylation site Tyr-267 leads to inhibition of nuclear translocation and DNA binding.

Directory of Open Access Journals (Sweden)

Mehmet Karaca

Full Text Available Reactivation of androgen receptor (AR may drive recurrent prostate cancer in castrate patients. Ack1 tyrosine kinase is overexpressed in prostate cancer and promotes castrate resistant xenograft tumor growth and enhances androgen target gene expression and AR recruitment to enhancers. Ack1 phosphorylates AR at Tyr-267 and possibly Tyr-363, both in the N-terminal transactivation domain. In this study, the role of these phosphorylation sites was investigated by characterizing the phosphorylation site mutants in the context of full length and truncated AR lacking the ligand-binding domain. Y267F and Y363F mutants showed decreased transactivation of reporters. Expression of wild type full length and truncated AR in LNCaP cells increased cell proliferation in androgen-depleted conditions and increased colony formation. However, the Y267F mutant of full length and truncated AR was defective in stimulating cell proliferation. The Y363F mutant was less severely affected than the Y267F mutant. The full length AR Y267F mutant was defective in nuclear translocation induced by androgen or Ack1 kinase. The truncated AR was constitutively localized to the nucleus. Chromatin immunoprecipitation analysis showed that it was recruited to the target enhancers without androgen. The truncated Y267F AR mutant did not exhibit constitutive nuclear localization and androgen enhancer binding activity. These results support the concept that phosphorylation of Tyr-267, and to a lesser extent Tyr-363, is required for AR nuclear translocation and recruitment and DNA binding and provide a rationale for development of novel approaches to inhibit AR activity.

Constitutive expression of nitrate reductase allows normal growth and development of Nicotiana plumbaginifolia plants.

Science.gov (United States)

Vincentz, M; Caboche, M

1991-01-01

A nitrate reductase (NR) deficient mutant of Nicotiana plumbaginifolia totally impaired in the production of NR transcript and protein was restored for NR activity by transformation with a chimaeric NR gene. This gene was composed of a full-length tobacco NR cDNA fused to the CaMV 35S promoter and to termination signals from the tobacco NR gene. The transgenic plants we obtained were viable and fertile and expressed from one-fifth to three times the wild-type NR activity in their leaves. The analysis of chimeric NR gene expression in these plants showed, by comparison with wild-type plants, that the regulation of NR gene expression by light, nitrate and circadian rhythm takes place at the transcriptional level. However, unlike nitrate, light was required for the accumulation of NR protein in transgenic plants, suggesting that NR expression is also controlled at the translational and/or post-translational level. Images PMID:2022181

Cold adaptation of the mononuclear molybdoenzyme periplasmic nitrate reductase from the Antarctic bacterium Shewanella gelidimarina

Energy Technology Data Exchange (ETDEWEB)

Simpson, Philippa J.L. [School of Chemistry, University of Sydney, New South Wales 2006 (Australia); Codd, Rachel, E-mail: rachel.codd@sydney.edu.au [School of Chemistry, University of Sydney, New South Wales 2006 (Australia); School of Medical Sciences (Pharmacology) and Bosch Institute, University of New South Wales, New South Wales 2006 (Australia)

2011-11-04

Highlights: Black-Right-Pointing-Pointer Cold-adapted phenotype of NapA from the Antarctic bacterium Shewanella gelidimarina. Black-Right-Pointing-Pointer Protein homology model of NapA from S. gelidimarina and mesophilic homologue. Black-Right-Pointing-Pointer Six amino acid residues identified as lead candidates governing NapA cold adaptation. Black-Right-Pointing-Pointer Molecular-level understanding of designing cool-temperature in situ oxyanion sensors. -- Abstract: The reduction of nitrate to nitrite is catalysed in bacteria by periplasmic nitrate reductase (Nap) which describes a system of variable protein subunits encoded by the nap operon. Nitrate reduction occurs in the NapA subunit, which contains a bis-molybdopterin guanine dinucleotide (Mo-MGD) cofactor and one [4Fe-4S] iron-sulfur cluster. The activity of periplasmic nitrate reductase (Nap) isolated as native protein from the cold-adapted (psychrophilic) Antarctic bacterium Shewanella gelidimarina (Nap{sub Sgel}) and middle-temperature adapted (mesophilic) Shewanella putrefaciens (Nap{sub Sput}) was examined at varied temperature. Irreversible deactivation of Nap{sub Sgel} and Nap{sub Sput} occurred at 54.5 and 65 Degree-Sign C, respectively. When Nap{sub Sgel} was preincubated at 21-70 Degree-Sign C for 30 min, the room-temperature nitrate reductase activity was maximal and invariant between 21 and 54 Degree-Sign C, which suggested that Nap{sub Sgel} was poised for optimal catalysis at modest temperatures and, unlike Nap{sub Sput}, did not benefit from thermally-induced refolding. At 20 Degree-Sign C, Nap{sub Sgel} reduced selenate at 16% of the rate of nitrate reduction. Nap{sub Sput} did not reduce selenate. Sequence alignment showed 46 amino acid residue substitutions in Nap{sub Sgel} that were conserved in NapA from mesophilic Shewanella, Rhodobacter and Escherichia species and could be associated with the Nap{sub Sgel} cold-adapted phenotype. Protein homology modeling of Nap{sub Sgel} using a

Cold adaptation of the mononuclear molybdoenzyme periplasmic nitrate reductase from the Antarctic bacterium Shewanella gelidimarina

International Nuclear Information System (INIS)

Simpson, Philippa J.L.; Codd, Rachel

2011-01-01

Highlights: ► Cold-adapted phenotype of NapA from the Antarctic bacterium Shewanella gelidimarina. ► Protein homology model of NapA from S. gelidimarina and mesophilic homologue. ► Six amino acid residues identified as lead candidates governing NapA cold adaptation. ► Molecular-level understanding of designing cool-temperature in situ oxyanion sensors. -- Abstract: The reduction of nitrate to nitrite is catalysed in bacteria by periplasmic nitrate reductase (Nap) which describes a system of variable protein subunits encoded by the nap operon. Nitrate reduction occurs in the NapA subunit, which contains a bis-molybdopterin guanine dinucleotide (Mo–MGD) cofactor and one [4Fe–4S] iron–sulfur cluster. The activity of periplasmic nitrate reductase (Nap) isolated as native protein from the cold-adapted (psychrophilic) Antarctic bacterium Shewanella gelidimarina (Nap Sgel ) and middle-temperature adapted (mesophilic) Shewanella putrefaciens (Nap Sput ) was examined at varied temperature. Irreversible deactivation of Nap Sgel and Nap Sput occurred at 54.5 and 65 °C, respectively. When Nap Sgel was preincubated at 21–70 °C for 30 min, the room-temperature nitrate reductase activity was maximal and invariant between 21 and 54 °C, which suggested that Nap Sgel was poised for optimal catalysis at modest temperatures and, unlike Nap Sput , did not benefit from thermally-induced refolding. At 20 °C, Nap Sgel reduced selenate at 16% of the rate of nitrate reduction. Nap Sput did not reduce selenate. Sequence alignment showed 46 amino acid residue substitutions in Nap Sgel that were conserved in NapA from mesophilic Shewanella, Rhodobacter and Escherichia species and could be associated with the Nap Sgel cold-adapted phenotype. Protein homology modeling of Nap Sgel using a mesophilic template with 66% amino acid identity showed the majority of substitutions occurred at the protein surface distal to the Mo–MGD cofactor. Two mesophilic ↔ psychrophilic

The influence of Glyceria maxima and nitrate input on the composition and nitrate metabolism of the dissimilatory nitrate-reducing bacterial community

NARCIS (Netherlands)

Nijburg, J.W.; Laanbroek, H.J.

1997-01-01

The influence of nitrate addition and the presence of Glyceria maxima (reed sweetgrass) on the composition and nitrate metabolism of the dissimilatory nitrate-reducing bacterial community was investigated. Anoxic freshwater sediment was incubated in pots with or without G. maxima and with or without

49 CFR 176.415 - Permit requirements for Division 1.5, ammonium nitrates, and certain ammonium nitrate fertilizers.

Science.gov (United States)

2010-10-01

... nitrates, and certain ammonium nitrate fertilizers. 176.415 Section 176.415 Transportation Other... requirements for Division 1.5, ammonium nitrates, and certain ammonium nitrate fertilizers. (a) Except as... Captain of the Port (COTP). (1) Ammonium nitrate UN1942, ammonium nitrate fertilizers containing more than...

Respiration of Nitrate and Nitrite.

Science.gov (United States)

Cole, Jeffrey A; Richardson, David J

2008-09-01

Nitrate reduction to ammonia via nitrite occurs widely as an anabolic process through which bacteria, archaea, and plants can assimilate nitrate into cellular biomass. Escherichia coli and related enteric bacteria can couple the eight-electron reduction of nitrate to ammonium to growth by coupling the nitrate and nitrite reductases involved to energy-conserving respiratory electron transport systems. In global terms, the respiratory reduction of nitrate to ammonium dominates nitrate and nitrite reduction in many electron-rich environments such as anoxic marine sediments and sulfide-rich thermal vents, the human gastrointestinal tract, and the bodies of warm-blooded animals. This review reviews the regulation and enzymology of this process in E. coli and, where relevant detail is available, also in Salmonella and draws comparisons with and implications for the process in other bacteria where it is pertinent to do so. Fatty acids may be present in high levels in many of the natural environments of E. coli and Salmonella in which oxygen is limited but nitrate is available to support respiration. In E. coli, nitrate reduction in the periplasm involves the products of two seven-gene operons, napFDAGHBC, encoding the periplasmic nitrate reductase, and nrfABCDEFG, encoding the periplasmic nitrite reductase. No bacterium has yet been shown to couple a periplasmic nitrate reductase solely to the cytoplasmic nitrite reductase NirB. The cytoplasmic pathway for nitrate reduction to ammonia is restricted almost exclusively to a few groups of facultative anaerobic bacteria that encounter high concentrations of environmental nitrate.

Interaction in triple systems of neodymium nitrate, water and nitrates of trimethylammonium and tetramethylammonium

International Nuclear Information System (INIS)

Boeva, M.K.; Zhuravlev, E.F.

1977-01-01

At 20 and 40 deg C the mutual solubility is studied in systems neodymium nitrate-water-trimethylamine nitrate and neodymium nitrate-water-tetramethylammonium nitrate. It has been established that the above systems belong to those with chemical interaction of the components. The compounds have been isolated preparatively, their composition has been confirmed analytically, and their thermal behaviour studied

Nitrate accumulation in spinach

NARCIS (Netherlands)

Steingröver, Eveliene Geertruda

1986-01-01

Leafy vegetables, like spinach, may contain high concentrations of nitrate. In the Netherlands, about 75% of mean daily intake of nitrate orginates from the consumption of vegatables. Hazards to human health are associated with the reduction of nitrate to nitrite. Acute nitrite poisoning causes

Impact of elevated nitrate on sulfate-reducing bacteria: A comparative study of Desulfovibrio vulgaris

Energy Technology Data Exchange (ETDEWEB)

He, Q.; He, Z.; Joyner, D.C.; Joachimiak, M.; Price, M.N.; Yang, Z.K.; Yen, H.-C. B.; Hemme, C. L.; Chen, W.; Fields, M.; Stahl, D. A.; Keasling, J. D.; Keller, M.; Arkin, A. P.; Hazen, T. C.; Wall, J. D.; Zhou, J.

2010-07-15

Sulfate-reducing bacteria have been extensively studied for their potential in heavy-metal bioremediation. However, the occurrence of elevated nitrate in contaminated environments has been shown to inhibit sulfate reduction activity. Although the inhibition has been suggested to result from the competition with nitrate-reducing bacteria, the possibility of direct inhibition of sulfate reducers by elevated nitrate needs to be explored. Using Desulfovibrio vulgaris as a model sulfate-reducing bacterium, functional genomics analysis reveals that osmotic stress contributed to growth inhibition by nitrate as shown by the upregulation of the glycine/betaine transporter genes and the relief of nitrate inhibition by osmoprotectants. The observation that significant growth inhibition was effected by 70 mM NaNO{sub 3} but not by 70 mM NaCl suggests the presence of inhibitory mechanisms in addition to osmotic stress. The differential expression of genes characteristic of nitrite stress responses, such as the hybrid cluster protein gene, under nitrate stress condition further indicates that nitrate stress response by D. vulgaris was linked to components of both osmotic and nitrite stress responses. The involvement of the oxidative stress response pathway, however, might be the result of a more general stress response. Given the low similarities between the response profiles to nitrate and other stresses, less-defined stress response pathways could also be important in nitrate stress, which might involve the shift in energy metabolism. The involvement of nitrite stress response upon exposure to nitrate may provide detoxification mechanisms for nitrite, which is inhibitory to sulfate-reducing bacteria, produced by microbial nitrate reduction as a metabolic intermediate and may enhance the survival of sulfate-reducing bacteria in environments with elevated nitrate level.

The G2/M DNA damage checkpoint inhibits mitosis through Tyr15 phosphorylation of p34cdc2 in Aspergillus nidulans.

OpenAIRE

Ye, X S; Fincher, R R; Tang, A; Osmani, S A

1997-01-01

It is possible to cause G2 arrest in Aspergillus nidulans by inactivating either p34cdc2 or NIMA. We therefore investigated the negative control of these two mitosis-promoting kinases after DNA damage. DNA damage caused rapid Tyr15 phosphorylation of p34cdc2 and transient cell cycle arrest but had little effect on the activity of NIMA. Dividing cells deficient in Tyr15 phosphorylation of p34cdc2 were sensitive to both MMS and UV irradiation and entered lethal premature mitosis with damaged DN...

The nitrate transporter (NRT gene family in poplar.

Directory of Open Access Journals (Sweden)

Hua Bai

Full Text Available Nitrate is an important nutrient required for plant growth. It also acts as a signal regulating plant development. Nitrate is actively taken up and transported by nitrate transporters (NRT, which form a large family with many members and distinct functions. In contrast to Arabidopsis and rice there is little information about the NRT family in woody plants such as Populus. In this study, a comprehensive analysis of the Populus NRT family was performed. Sixty-eight PtNRT1/PTR, 6 PtNRT2, and 5 PtNRT3 genes were identified in the P. trichocarpa genome. Phylogenetic analysis confirmed that the genes of the NRT family are divided into three clades: NRT1/PTR with four subclades, NRT2, and NRT3. Topological analysis indicated that all members of PtNRT1/PTR and PtNRT2 have 8 to 12 trans-membrane domains, whereas the PtNRT3 proteins have no or up to two trans-membrane domains. Four PtNRT3 members were predicted as secreted proteins. Microarray analyses revealed tissue-specific expression patterns of PtNRT genes with distinct clusters of NRTs for roots, for the elongation zone of the apical stem segment and the developing xylem and a further cluster for leaves, bark and wood. A comparison of different poplar species (P. trichocarpa, P. tremula, P. euphratica, P. fremontii x P. angustifolia, and P. x canescens showed that the tissue-specific patterns of the NRT genes varied to some extent with species. Bioinformatic analysis of putative cis-regulatory elements in the promoter regions of PtNRT family retrieved motifs suggesting the regulation of the NRT genes by N metabolism, by energy and carbon metabolism, and by phytohormones and stress. Multivariate analysis suggested that the combination and abundance of motifs in distinct promoters may lead to tissue-specificity. Our genome wide analysis of the PtNRT genes provides a valuable basis for functional analysis towards understanding the role of nitrate transporters for tree growth.

Transglutaminase-catalyzed amination of pea protein peptides using the biogenic amines histamine and tyramine.

Science.gov (United States)

Lu, Xinyao; Hrynets, Yuliya; Betti, Mirko

2017-06-01

Biogenic amines (BAs) are produced by the enzymatic decarboxylation of amino acids, and are well-known for their toxicity to humans. This study describes a new method using microbial transglutaminase (MTGase) to covalently link BAs such as histamine (HIS) and tyramine (TYR) to the glutamine residues of alcalase-hydrolyzed pea protein (PPH). The incubation of PPH and HIS and TYR in the presence of MTGase at 37 °C led to the formation of conjugates, as determined by liquid chromatography, after derivatization with dansyl chloride. Seventy-six % of HIS and 65% of TYR were covalently incorporated to PPH by MTGase. The incubation of PPH and TYR in the presence of MTGase exhibited a 52% DPPH radical scavenging activity at 10 mg mL -1 . Conjugation via MTGase improved the antioxidant status by reducing lipid peroxidation. This study emphasizes that the application of MTGase can effectively reduce histamine and tyramine content while simultaneously enhancing antioxidative capacity of PPH. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

Dancing with Hormones: A Current Perspective of Nitrate Signaling and Regulation in Arabidopsis

Directory of Open Access Journals (Sweden)

Peizhu Guan

2017-09-01

Full Text Available In nature and agriculture, nitrate availability is a main environmental cue for plant growth, development and stress responses. Nitrate signaling and regulation are hence at the center of communications between plant intrinsic programs and the environment. It is also well known that endogenous phytohormones play numerous critical roles in integrating extrinsic cues and intrinsic responses, regulating and refining almost all aspects of plant growth, development and stress responses. Therefore, interaction between nitrate and phytohormones, such as auxins, cytokinins, abscisic acid, gibberellins, and ethylene, is prevalent. The growing evidence indicates that biosynthesis, de-conjugation, transport, and signaling of hormones are partly controlled by nitrate signaling. Recent advances with nitrate signaling and transcriptional regulation in Arabidopsis give rise to new paradigms. Given the comprehensive nitrate transport, sensing, signaling and regulations at the level of the cell and organism, nitrate itself is a local and long-distance signal molecule, conveying N status at the whole-plant level. A direct molecular link between nitrate signaling and cell cycle progression was revealed with TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR1-20 (TCP20 – NIN-LIKE PROTEIN 6/7 (NLP6/7 regulatory nexus. NLPs are key regulators of nitrogen responses in plants. TCPs function as the main regulators of plant morphology and architecture, with the emerging role as integrators of plant developmental responses to the environment. By analogy with auxin being proposed as a plant morphogen, nitrate may be an environmental morphogen. The morphogen-gradient-dependent and cell-autonomous mechanisms of nitrate signaling and regulation are an integral part of cell growth and cell identification. This is especially true in root meristem growth that is regulated by intertwined nitrate, phytohormones, and glucose-TOR signaling pathways. Furthermore, the nitrate

The conservation pattern of short linear motifs is highly correlated with the function of interacting protein domains

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Wang Yiguo

2008-10-01

Full Text Available Abstract Background Many well-represented domains recognize primary sequences usually less than 10 amino acids in length, called Short Linear Motifs (SLiMs. Accurate prediction of SLiMs has been difficult because they are short (often Results Our combined approach revealed that SLiMs are highly conserved in proteins from functional classes that are known to interact with a specific domain, but that they are not conserved in most other protein groups. We found that SLiMs recognized by SH2 domains were highly conserved in receptor kinases/phosphatases, adaptor molecules, and tyrosine kinases/phosphatases, that SLiMs recognized by SH3 domains were highly conserved in cytoskeletal and cytoskeletal-associated proteins, that SLiMs recognized by PDZ domains were highly conserved in membrane proteins such as channels and receptors, and that SLiMs recognized by S/T kinase domains were highly conserved in adaptor molecules, S/T kinases/phosphatases, and proteins involved in transcription or cell cycle control. We studied Tyr-SLiMs recognized by SH2 domains in more detail, and found that SH2-recognized Tyr-SLiMs on the cytoplasmic side of membrane proteins are more highly conserved than those on the extra-cellular side. Also, we found that SH2-recognized Tyr-SLiMs that are associated with SH3 motifs and a tyrosine kinase phosphorylation motif are more highly conserved. Conclusion The interactome of protein domains is reflected by the evolutionary conservation of SLiMs recognized by these domains. Combining scoring matrixes derived from peptide libraries and conservation analysis, we would be able to find those protein groups that are more likely to interact with specific domains.

The Cytokinin Requirement for Cell Division in Cultured Nicotiana plumbaginifolia Cells Can Be Satisfied by Yeast Cdc25 Protein Tyrosine Phosphatase. Implications for Mechanisms of Cytokinin Response and Plant Development

Science.gov (United States)

Zhang, Kerong; Diederich, Ludger; John, Peter C.L.

2005-01-01

Cultured cells of Nicotiana plumbaginifolia, when deprived of exogenous cytokinin, arrest in G2 phase prior to mitosis and then contain cyclin-dependent protein kinase (CDK) that is inactive because phosphorylated on tyrosine (Tyr). The action of cytokinin in stimulating the activation of CDK by removal of inhibitory phosphorylation from Tyr is not a secondary downstream consequence of other hormone actions but is the key primary effect of the hormone in its stimulation of cell proliferation, since cytokinin could be replaced by expression of cdc25, which encodes the main Cdc2 (CDK)-Tyr dephosphorylating enzyme of yeast (Saccharomyces cerevisiae). The cdc25 gene, under control of a steroid-inducible promoter, induced a rise in cdc25 mRNA, accumulation of p67Cdc25 protein, and increase in Cdc25 phosphatase activity that was measured in vitro with Tyr-phosphorylated Cdc2 as substrate. Cdc25 phosphatase activity peaked during mitotic prophase at the time CDK activation was most rapid. Mitosis that was induced by cytokinin also involved increase in endogenous plant CDK Tyr phosphatase activity during prophase, therefore indicating that this is a normal part of plant mitosis. These results suggest a biochemical mechanism for several previously described transgene phenotypes in whole plants and suggest that a primary signal from cytokinin leading to progression through mitosis is the activation of CDK by dephosphorylation of Tyr. PMID:15618425

The cytokinin requirement for cell division in cultured Nicotiana plumbaginifolia cells can be satisfied by yeast Cdc25 protein tyrosine phosphatase: implications for mechanisms of cytokinin response and plant development.

Science.gov (United States)

Zhang, Kerong; Diederich, Ludger; John, Peter C L

2005-01-01

Cultured cells of Nicotiana plumbaginifolia, when deprived of exogenous cytokinin, arrest in G2 phase prior to mitosis and then contain cyclin-dependent protein kinase (CDK) that is inactive because phosphorylated on tyrosine (Tyr). The action of cytokinin in stimulating the activation of CDK by removal of inhibitory phosphorylation from Tyr is not a secondary downstream consequence of other hormone actions but is the key primary effect of the hormone in its stimulation of cell proliferation, since cytokinin could be replaced by expression of cdc25, which encodes the main Cdc2 (CDK)-Tyr dephosphorylating enzyme of yeast (Saccharomyces cerevisiae). The cdc25 gene, under control of a steroid-inducible promoter, induced a rise in cdc25 mRNA, accumulation of p67(Cdc25) protein, and increase in Cdc25 phosphatase activity that was measured in vitro with Tyr-phosphorylated Cdc2 as substrate. Cdc25 phosphatase activity peaked during mitotic prophase at the time CDK activation was most rapid. Mitosis that was induced by cytokinin also involved increase in endogenous plant CDK Tyr phosphatase activity during prophase, therefore indicating that this is a normal part of plant mitosis. These results suggest a biochemical mechanism for several previously described transgene phenotypes in whole plants and suggest that a primary signal from cytokinin leading to progression through mitosis is the activation of CDK by dephosphorylation of Tyr.

Feeding nitrate and docosahexaenoic acid affects enteric methane production and milk fatty acid composition in lactating dairy cows.

Science.gov (United States)

Klop, G; Hatew, B; Bannink, A; Dijkstra, J

2016-02-01

An experiment was conducted to study potential interaction between the effects of feeding nitrate and docosahexaenoic acid (DHA; C22:6 n-3) on enteric CH4 production and performance of lactating dairy cows. Twenty-eight lactating Holstein dairy cows were grouped into 7 blocks of 4 cows. Within blocks, cows were randomly assigned to 1 of 4 treatments: control (CON; urea as alternative nonprotein N source to nitrate), NO3 [21 g of nitrate/kg of dry matter (DM)], DHA (3 g of DHA/kg of DM and urea as alternative nonprotein N source to nitrate), or NO3 + DHA (21 g of nitrate/kg of DM and 3 g of DHA/kg of DM, respectively). Cows were fed a total mixed ration consisting of 21% grass silage, 49% corn silage, and 30% concentrates on a DM basis. Feed additives were included in the concentrates. Cows assigned to a treatment including nitrate were gradually adapted to the treatment dose of nitrate over a period of 21 d during which no DHA was fed. The experimental period lasted 17 d, and CH4 production was measured during the last 5d in climate respiration chambers. Cows produced on average 363, 263, 369, and 298 g of CH4/d on CON, NO3, DHA, and NO3 + DHA treatments, respectively, and a tendency for a nitrate × DHA interaction effect was found where the CH4-mitigating effect of nitrate decreased when combined with DHA. This tendency was not obtained for CH4 production relative to dry matter intake (DMI) or to fat- and protein corrected milk (FPCM). The NO3 treatment decreased CH4 production irrespective of the unit in which it was expressed, whereas DHA did not affect CH4 production per kilogram of DMI, but resulted in a higher CH4 production per kilogram of fat- and protein-corrected milk (FPCM) production. The FPCM production (27.9, 24.7, 24.2, and 23. 8 kg/d for CON, NO3, DHA, and NO3 + DHA, respectively) was lower for DHA-fed cows because of decreased milk fat concentration. The proportion of saturated fatty acids in milk fat was decreased by DHA, and the proportion of

The effect of peroxynitrite decomposition catalyst MnTBAP on aldehyde dehydrogenase-2 nitration by organic nitrates: role in nitrate tolerance.

Science.gov (United States)

Mollace, Vincenzo; Muscoli, Carolina; Dagostino, Concetta; Giancotti, Luigino Antonio; Gliozzi, Micaela; Sacco, Iolanda; Visalli, Valeria; Gratteri, Santo; Palma, Ernesto; Malara, Natalia; Musolino, Vincenzo; Carresi, Cristina; Muscoli, Saverio; Vitale, Cristiana; Salvemini, Daniela; Romeo, Francesco

2014-11-01

Bioconversion of glyceryl trinitrate (GTN) into nitric oxide (NO) by aldehyde dehydrogenase-2 (ALDH-2) is a crucial mechanism which drives vasodilatory and antiplatelet effect of organic nitrates in vitro and in vivo. Oxidative stress generated by overproduction of free radical species, mostly superoxide anions and NO-derived peroxynitrite, has been suggested to play a pivotal role in the development of nitrate tolerance, though the mechanism still remains unclear. Here we studied the free radical-dependent impairment of ALDH-2 in platelets as well as vascular tissues undergoing organic nitrate ester tolerance and potential benefit when using the selective peroxynitrite decomposition catalyst Mn(III) tetrakis (4-Benzoic acid) porphyrin (MnTBAP). Washed human platelets were made tolerant to nitrates via incubation with GTN for 4h. This was expressed by attenuation of platelet aggregation induced by thrombin (40U/mL), an effect accompanied by GTN-related induction of cGMP levels in platelets undergoing thrombin-induced aggregation. Both effects were associated to attenuated GTN-induced nitrite formation in platelets supernatants and to prominent nitration of ALDH-2, the GTN to NO metabolizing enzyme, suggesting that GTN tolerance was associated to reduced NO formation via impairment of ALDH-2. These effects were all antagonized by co-incubation of platelets with MnTBAP, which restored GTN-induced responses in tolerant platelets. Comparable effect was found under in in vivo settings. Indeed, MnTBAP (10mg/kg, i.p.) significantly restored the hypotensive effect of bolus injection of GTN in rats made tolerants to organic nitrates via chronic administration of isosorbide-5-mononitrate (IS-5-MN), thus confirming the role of peroxynitrite overproduction in the development of tolerance to vascular responses induced by organic nitrates. In conclusion, oxidative stress subsequent to prolonged use of organic nitrates, which occurs via nitration of ALDH-2, represents a key event

Nitrate radical oxidation of γ-terpinene: hydroxy nitrate, total organic nitrate, and secondary organic aerosol yields

Science.gov (United States)

Slade, Jonathan H.; de Perre, Chloé; Lee, Linda; Shepson, Paul B.

2017-07-01

Polyolefinic monoterpenes represent a potentially important but understudied source of organic nitrates (ONs) and secondary organic aerosol (SOA) following oxidation due to their high reactivity and propensity for multi-stage chemistry. Recent modeling work suggests that the oxidation of polyolefinic γ-terpinene can be the dominant source of nighttime ON in a mixed forest environment. However, the ON yields, aerosol partitioning behavior, and SOA yields from γ-terpinene oxidation by the nitrate radical (NO3), an important nighttime oxidant, have not been determined experimentally. In this work, we present a comprehensive experimental investigation of the total (gas + particle) ON, hydroxy nitrate, and SOA yields following γ-terpinene oxidation by NO3. Under dry conditions, the hydroxy nitrate yield = 4(+1/-3) %, total ON yield = 14(+3/-2) %, and SOA yield ≤ 10 % under atmospherically relevant particle mass loadings, similar to those for α-pinene + NO3. Using a chemical box model, we show that the measured concentrations of NO2 and γ-terpinene hydroxy nitrates can be reliably simulated from α-pinene + NO3 chemistry. This suggests that NO3 addition to either of the two internal double bonds of γ-terpinene primarily decomposes forming a relatively volatile keto-aldehyde, reconciling the small SOA yield observed here and for other internal olefinic terpenes. Based on aerosol partitioning analysis and identification of speciated particle-phase ON applying high-resolution liquid chromatography-mass spectrometry, we estimate that a significant fraction of the particle-phase ON has the hydroxy nitrate moiety. This work greatly contributes to our understanding of ON and SOA formation from polyolefin monoterpene oxidation, which could be important in the northern continental US and the Midwest, where polyolefinic monoterpene emissions are greatest.

High temperature interaction studies on equimolar nitrate mixture of uranyl nitrate hexahydrate and gadolinium nitrate hexahydrate

International Nuclear Information System (INIS)

Kalekar, Bhupesh B.; Raje, Naina; Reddy, A.V.R.

2015-01-01

Rare earths including gadolinium form a sizeable fraction of the fission products in the nuclear fission of fissile material in the reactor. These fission products can interact with uranium dioxide fuel and can form various compounds which can alter the thermal behavior of the fuel. The mixed oxide formed due to the high temperature interactions of mixture of uranyl nitrate hexahydrate (UNH) and gadolinium nitrate hexahydrate (GdNH) has been studied using thermal and X- ray diffraction techniques. The equimolar mixture of UNH and GdNH was prepared by mixing the weighed amount of individual nitrates and grinding gently with mortar and pestle. Thermogravimetry (TG) measurements were carried out by separately heating 100 mg of mixture and individual nitrates at heating rate of 10°C min -1 using Netzsch thermal analyzer (Model No.: STA 409 PC Luxx) in high purity nitrogen atmosphere with a flow rate of 120 mL min -1 . The XRD measurement was carried out on a Philips X-ray diffractometer (Model PW1710) using nickel-filtered Cu-Kα radiation

Amperometric nitrate biosensor based on Carbon nanotube/Polypyrrole/Nitrate reductase biofilm electrode

Energy Technology Data Exchange (ETDEWEB)

Can, Faruk; Korkut Ozoner, Seyda; Ergenekon, Pinar; Erhan, Elif, E-mail: e.erhan@gyte.edu.tr

2012-01-01

This study describes the construction and characterization of an amperometric nitrate biosensor based on the Polypyrrole (PPy)/Carbon nanotubes (CNTs) film. Nitrate reductase (NR) was both entrapped into the growing PPy film and chemically immobilized via the carboxyl groups of CNTs to the CNT/PPy film electrode. The optimum amperometric response for nitrate was obtained in 0.1 M phosphate buffer solution (PBS), pH 7.5 including 0.1 M lithium chloride and 7 mM potassium ferricyanide with an applied potential of 0.13 V (vs. Ag/AgCl, 3 M NaCl). Sensitivity was found to be 300 nA/mM in a linear range of 0.44-1.45 mM with a regression coefficient of 0.97. The biosensor response showed a higher linear range in comparison to standard nitrate analysis methods which were tested in this study and NADH based nitrate biosensors. A minimum detectable concentration of 0.17 mM (S/N = 3) with a relative standard deviation (RSD) of 5.4% (n = 7) was obtained for the biosensor. Phenol and glucose inhibit the electrochemical reaction strictly at a concentration of 1 {mu}g/L and 20 mg/L, respectively. The biosensor response retained 70% of its initial response over 10 day usage period when used everyday. - Highlights: Black-Right-Pointing-Pointer K{sub 3}Fe(CN){sub 6} has been used for the first time as mediator for nitrate reductase. Black-Right-Pointing-Pointer Better performance was obtained in comparison to other nitrate biosensor studies operated with various mediators. Black-Right-Pointing-Pointer Analytical parameters were better than standard nitrate analysis methods.

Amperometric nitrate biosensor based on Carbon nanotube/Polypyrrole/Nitrate reductase biofilm electrode

International Nuclear Information System (INIS)

Can, Faruk; Korkut Ozoner, Seyda; Ergenekon, Pinar; Erhan, Elif

2012-01-01

This study describes the construction and characterization of an amperometric nitrate biosensor based on the Polypyrrole (PPy)/Carbon nanotubes (CNTs) film. Nitrate reductase (NR) was both entrapped into the growing PPy film and chemically immobilized via the carboxyl groups of CNTs to the CNT/PPy film electrode. The optimum amperometric response for nitrate was obtained in 0.1 M phosphate buffer solution (PBS), pH 7.5 including 0.1 M lithium chloride and 7 mM potassium ferricyanide with an applied potential of 0.13 V (vs. Ag/AgCl, 3 M NaCl). Sensitivity was found to be 300 nA/mM in a linear range of 0.44–1.45 mM with a regression coefficient of 0.97. The biosensor response showed a higher linear range in comparison to standard nitrate analysis methods which were tested in this study and NADH based nitrate biosensors. A minimum detectable concentration of 0.17 mM (S/N = 3) with a relative standard deviation (RSD) of 5.4% (n = 7) was obtained for the biosensor. Phenol and glucose inhibit the electrochemical reaction strictly at a concentration of 1 μg/L and 20 mg/L, respectively. The biosensor response retained 70% of its initial response over 10 day usage period when used everyday. - Highlights: ► K 3 Fe(CN) 6 has been used for the first time as mediator for nitrate reductase. ► Better performance was obtained in comparison to other nitrate biosensor studies operated with various mediators. ► Analytical parameters were better than standard nitrate analysis methods.

Dietary nitrate increases tetanic [Ca2+]i and contractile force in mouse fast-twitch muscle.

Science.gov (United States)

Hernández, Andrés; Schiffer, Tomas A; Ivarsson, Niklas; Cheng, Arthur J; Bruton, Joseph D; Lundberg, Jon O; Weitzberg, Eddie; Westerblad, Håkan

2012-08-01

Dietary inorganic nitrate has profound effects on health and physiological responses to exercise. Here, we examined if nitrate, in doses readily achievable via a normal diet, could improve Ca(2+) handling and contractile function using fast- and slow-twitch skeletal muscles from C57bl/6 male mice given 1 mm sodium nitrate in water for 7 days. Age matched controls were provided water without added nitrate. In fast-twitch muscle fibres dissected from nitrate treated mice, myoplasmic free [Ca(2+)] was significantly greater than in Control fibres at stimulation frequencies from 20 to 150 Hz, which resulted in a major increase in contractile force at ≤ 50 Hz. At 100 Hz stimulation, the rate of force development was ∼35% faster in the nitrate group. These changes in nitrate treated mice were accompanied by increased expression of the Ca(2+) handling proteins calsequestrin 1 and the dihydropyridine receptor. No changes in force or calsequestrin 1 and dihydropyridine receptor expression were measured in slow-twitch muscles. In conclusion, these results show a striking effect of nitrate supplementation on intracellular Ca(2+) handling in fast-twitch muscle resulting in increased force production. A new mechanism is revealed by which nitrate can exert effects on muscle function with applications to performance and a potential therapeutic role in conditions with muscle weakness.

Genotoxic evaluation of [DOTA,Tyr3]octreotate labeled with 131I and 177Lu in human peripheral lymphocytes in vitro by micronucleus assay

International Nuclear Information System (INIS)

Suzauki, Miriam Fussae; Silva, Marcia Augusta da; Caldeira Filho, Jose de Souza; Colturato, Maria Tereza; Araujo, Elaine Bortoleti de; Bartolini, Paolo; Okazaki, Kayo

2005-01-01

The radiolabeled receptor-binding peptides have being used for cancer diagnosis and therapy. The octreotate, a somatostatin analogue peptide, bound to various tumors expressing sst receptors (thyroid, pancreas, prostrate, melanoma and lymphomas). The amount and the type of receptors for somatostatin influence the tissue uptake. The [DOTA, Tyr 3 ]octreotate has been used because of its high affinity to somatostatin subtype receptors sstr 2 and sstr 5 . The pharmacokinetic study showed that the blood clearance is rapid and only 9% of the intravenous injected activity remains in human blood after one hour. The aim of this study was to evaluate the cytogenetic effect of radiolabeled [DOTA, Tyr 3 ]octreotate in blood cells in vitro, using the cytokinesis-block micronucleus (MN) assay. This technique allows evaluating the mutagenic effects of both endogenous and exogenous agents at chromosome level. Blood samples of healthy donors were collected in heparinized syringes and exposed to different activities of [DOTA, Tyr 3 ]octreotate labeled with with 131 I (n=3) and 177 Lu (n=3), where radioactive concentration ranged from 600 to 5600 kBq/mL, corresponding to an injected activity of 3.1 to 28.9 GBq in a reference man of 70 kg weight. 131 I and 177 Lu are beta- and gamma-emitters. After one-hour exposition to radiopharmaceuticals at 37 deg C, the cells were washed with culture medium for removing the non internalised octreotate and cultivated for 72 hours, according to criteria adopted by the IAEA. The results showed a positive correlation between radioactive concentrations (X) and the frequency of binucleated cells with micronuclei (Y) (P 131 I-DOTA, Tyr 3 ]octreotate was Y = (1.634 ± 0.236) + (0.912 ± 0.137) 10 -3 X and for [ 177 Lu-DOTA, Tyr 3 ]octreotate was Y = (1.715 ± 0.342) + (0.743 ± 0.135) 10 -3 X. The non labeled molecule, [DOTA, Tyr 3 ]octreotate, has no influence in the induction of cytogenetic damage. The micronucleus assay with rat pancreatic tumor cells

Nitrate assimilation pathway (NAP): role of structural (nit) and transporter (ntr1) genes in Fusarium oxysporum f.sp. lycopersici growth and pathogenicity.

Science.gov (United States)

Gomez-Gil, Lucia; Camara Almiron, Jesus; Rodriguez Carrillo, Patricia Lizett; Olivares Medina, Cindy Nayely; Bravo Ruiz, Gustavo; Romo Rodriguez, Pamela; Corrales Escobosa, Alma Rosa; Gutierrez Corona, Felix; Roncero, M Isabel

2018-04-01

The tomato pathogen Fusarium oxysporum f.sp. lycopersici possesses the capability to use nitrate as the only nitrogen source under aerobic and anaerobic conditions and to activate virulence-related functions when cultivated in the presence of nitrate, but not in ammonium. The genome of F. oxysporum f.sp. lycopersici encodes three paralogs nitrate reductase (NR) genes (nit1, nit2 and nit3) and one predicted ortholog of the Aspergillus nidulans high-affinity nitrate/nitrite transporters NtrA and NtrB, named ntr1. We set out to clarify the role of nit1, nit2, nit3 and ntr1 genes in nitrate assimilation and in the virulence of F. oxysporum f.sp. lycopersici. Quantitative RT-PCR analysis revealed that only nit1, nit2 and ntr1 are expressed at significant levels during growth in nitrate as the only nitrogen source. Targeted deletion of nit1 and ntr1, but not of nit2 or nit3, severely impaired growth of F. oxysporum on nitrate as nitrogen source, indicating that Nit1 and Ntr1 proteins are involved in nitrate assimilation by the fungus; biochemical analysis of nit mutants indicated that Nit1 and Nit2 enzymes contribute to about 50 and 30% of the total NR activity, respectively. In addition, a spontaneous chlorate-resistant mutant derived from F. oxysporum 4287, denoted NitFG, was characterized, showing inability to grow in nitrate under aerobic and anaerobic conditions and low levels of NR activity, in spite of its increased transcription levels of nit1 and nit2 genes. Tomato plant infection assays showed that NitFG and ∆ntr1 mutants induced an earlier death in tomato plants, whereas the single mutants ∆nit1, ∆nit2 and ∆nit1∆nit2 double mutant showed a mortality rate similar to the wild-type strain. Taken together, these results indicate that the Nit1 and Ntr1 proteins are important for nitrate assimilation of F. oxysporum f.sp. lycopersici incubated under aerobic and anaerobic conditions and that this metabolic process is not essential for the virulence of

Nitrate reductase and nitrous oxide production by Fusarium oxysporum 11dn1 under aerobic and anaerobic conditions.

Science.gov (United States)

Kurakov, A V; Nosikov, A N; Skrynnikova, E V; L'vov, N P

2000-08-01

The fungus Fusarium oxysporum 11dn1 was found to be able to grow and produce nitrous oxide on nitrate-containing medium in anaerobic conditions. The rate of nitrous oxide formation was three to six orders of magnitude lower than the rates of molecular nitrogen production by common denitrifying bacteria. Acetylene and ammonia did not affect the release of nitrous oxide release. It was shown that under anaerobic conditions fast increase of nitrate reductase activity occurred, caused by the synthesis of enzyme de novo and protein dephosphorylation. Reverse transfer of the mycelium to aerobic conditions led to a decline in nitrate reductase activity and stopped nitrous oxide production. The presence of two nitrate reductases was shown, which differed in molecular mass, location, temperature optima, and activity in nitrate- and ammonium-containing media. Two enzymes represent assimilatory and dissimilatory nitrate reductases, which are active in aerobic and anaerobic conditions, respectively.

Purification of alkali metal nitrates

Science.gov (United States)

Fiorucci, Louis C.; Gregory, Kevin M.

1985-05-14

A process is disclosed for removing heavy metal contaminants from impure alkali metal nitrates containing them. The process comprises mixing the impure nitrates with sufficient water to form a concentrated aqueous solution of the impure nitrates, adjusting the pH of the resulting solution to within the range of between about 2 and about 7, adding sufficient reducing agent to react with heavy metal contaminants within said solution, adjusting the pH of the solution containing reducing agent to effect precipitation of heavy metal impurities and separating the solid impurities from the resulting purified aqueous solution of alkali metal nitrates. The resulting purified solution of alkali metal nitrates may be heated to evaporate water therefrom to produce purified molten alkali metal nitrate suitable for use as a heat transfer medium. If desired, the purified molten form may be granulated and cooled to form discrete solid particles of alkali metal nitrates.

The proportion of nitrate in leaf nitrogen, but not changes in root growth, are associated with decreased grain protein in wheat under elevated [CO2].

Science.gov (United States)

Bahrami, Helale; De Kok, Luit J; Armstrong, Roger; Fitzgerald, Glenn J; Bourgault, Maryse; Henty, Samuel; Tausz, Michael; Tausz-Posch, Sabine

2017-09-01

The atmospheric CO 2 concentration ([CO 2 ]) is increasing and predicted to reach ∼550ppm by 2050. Increasing [CO 2 ] typically stimulates crop growth and yield, but decreases concentrations of nutrients, such as nitrogen ([N]), and therefore protein, in plant tissues and grains. Such changes in grain composition are expected to have negative implications for the nutritional and economic value of grains. This study addresses two mechanisms potentially accountable for the phenomenon of elevated [CO 2 ]-induced decreases in [N]: N uptake per unit length of roots as well as inhibition of the assimilation of nitrate (NO 3 - ) into protein are investigated and related to grain protein. We analysed two wheat cultivars from a similar genetic background but contrasting in agronomic features (Triticum aestivum L. cv. Scout and Yitpi). Plants were field-grown within the Australian Grains Free Air CO 2 Enrichment (AGFACE) facility under two atmospheric [CO 2 ] (ambient, ∼400ppm, and elevated, ∼550ppm) and two water treatments (rain-fed and well-watered). Aboveground dry weight (ADW) and root length (RL, captured by a mini-rhizotron root growth monitoring system), as well as [N] and NO 3 - concentrations ([NO 3 - ]) were monitored throughout the growing season and related to grain protein at harvest. RL generally increased under e[CO 2 ] and varied between water supply and cultivars. The ratio of total aboveground N (TN) taken up per RL was affected by CO 2 treatment only later in the season and there was no significant correlation between TN/RL and grain protein concentration across cultivars and [CO 2 ] treatments. In contrast, a greater percentage of N remained as unassimilated [NO 3 - ] in the tissue of e[CO 2 ] grown crops (expressed as the ratio of NO 3 - to total N) and this was significantly correlated with decreased grain protein. These findings suggest that e[CO 2 ] directly affects the nitrate assimilation capacity of wheat with direct negative implications

Effect of dietary nitrate level on enteric methane production, hydrogen emission, rumen fermentation, and nutrient digestibility in dairy cows.

Science.gov (United States)

Olijhoek, D W; Hellwing, A L F; Brask, M; Weisbjerg, M R; Højberg, O; Larsen, M K; Dijkstra, J; Erlandsen, E J; Lund, P

2016-08-01

Nitrate may lower methane production in ruminants by competing with methanogenesis for available hydrogen in the rumen. This study evaluated the effect of 4 levels of dietary nitrate addition on enteric methane production, hydrogen emission, feed intake, rumen fermentation, nutrient digestibility, microbial protein synthesis, and blood methemoglobin. In a 4×4 Latin square design 4 lactating Danish Holstein dairy cows fitted with rumen, duodenal, and ileal cannulas were assigned to 4 calcium ammonium nitrate addition levels: control, low, medium, and high [0, 5.3, 13.6, and 21.1g of nitrate/kg of dry matter (DM), respectively]. Diets were made isonitrogenous by replacing urea. Cows were fed ad libitum and, after a 6-d period of gradual introduction of nitrate, adapted to the corn-silage-based total mixed ration (forage:concentrate ratio 50:50 on DM basis) for 16d before sampling. Digesta content from duodenum, ileum, and feces, and rumen liquid were collected, after which methane production and hydrogen emissions were measured in respiration chambers. Methane production [L/kg of dry matter intake (DMI)] linearly decreased with increasing nitrate concentrations compared with the control, corresponding to a reduction of 6, 13, and 23% for the low, medium, and high diets, respectively. Methane production was lowered with apparent efficiencies (measured methane reduction relative to potential methane reduction) of 82.3, 71.9, and 79.4% for the low, medium, and high diets, respectively. Addition of nitrate increased hydrogen emissions (L/kg of DMI) quadratically by a factor of 2.5, 3.4, and 3.0 (as L/kg of DMI) for the low, medium, and high diets, respectively, compared with the control. Blood methemoglobin levels and nitrate concentrations in milk and urine increased with increasing nitrate intake, but did not constitute a threat for animal health and human food safety. Microbial crude protein synthesis and efficiency were unaffected. Total volatile fatty acid

Thermochemical nitrate reduction

International Nuclear Information System (INIS)

Cox, J.L.; Lilga, M.A.; Hallen, R.T.

1992-09-01

A series of preliminary experiments was conducted directed at thermochemically converting nitrate to nitrogen and water. Nitrates are a major constituent of the waste stored in the underground tanks on the Hanford Site, and the characteristics and effects of nitrate compounds on stabilization techniques must be considered before permanent disposal operations begin. For the thermochemical reduction experiments, six reducing agents (ammonia, formate, urea, glucose, methane, and hydrogen) were mixed separately with ∼3 wt% NO 3 - solutions in a buffered aqueous solution at high pH (13); ammonia and formate were also mixed at low pH (4). Reactions were conducted in an aqueous solution in a batch reactor at temperatures of 200 degrees C to 350 degrees C and pressures of 600 to 2800 psig. Both gas and liquid samples were analyzed. The specific components analyzed were nitrate, nitrite, nitrous oxide, nitrogen, and ammonia. Results of experimental runs showed the following order of nitrate reduction of the six reducing agents in basic solution: formate > glucose > urea > hydrogen > ammonia ∼ methane. Airnmonia was more effective under acidic conditions than basic conditions. Formate was also effective under acidic conditions. A more thorough, fundamental study appears warranted to provide additional data on the mechanism of nitrate reduction. Furthermore, an expanded data base and engineering feasibility study could be used to evaluate conversion conditions for promising reducing agents in more detail and identify new reducing agents with improved performance characteristics

Effects of [Des-Tyr1]-γ-endorphin and α-endorphin on substantia nigra self-stimulation

NARCIS (Netherlands)

Dorsa, D.M.; Ree, J.M. van; Wied, D. de

The β-lipotropin fragments, [des-Tyr1]-γ-endorphin (DTγE, β-LPH62–77) and α-endorphin (β-LPH61–76) affect self-stimulating behavior associated with electrical stimulation of neurons of the ventral tegmentum area of rats in an opposite way. Subcutaneous administration of DTγE (5 and 25 μg) attenuated

Protein tyrosine phosphatase, PTP1B, expression and activity in rat corneal endothelial cells

Science.gov (United States)

Harris, Deshea L.

2007-01-01

Purpose The current studies were conducted to determine whether the protein tyrosine phosphatase, PTP1B, plays a role in regulating epidermal growth factor receptor (EGFR) Tyr992 phosphorylation and cell cycle entry in rat corneal endothelial cells. Methods Corneas were obtained from male Sprague-Dawley rats. PTP1B mRNA and protein expression were compared in confluent and subconfluent cells by RT-PCR and western blots. Immunocytochemistry was used to determine the subcellular localization of both PTP1B and EGFR following epidermal growth factor (EGF) stimulation. Western blots were used to analyze the time-dependent effect of EGF on phosphorylation of EGFR Tyr992 plus or minus CinnGEL 2Me, an inhibitor of PTP1B activity. The effect of PTP1B inhibition on cell cycle entry was determined by calculating the percent of Ki67-positive cells following EGF treatment. Results PTP1B mRNA expression was similar in confluent and subconfluent cells, but PTP1B protein was expressed at 3 fold higher levels in subconfluent cells. Positive staining for PTP1B was localized in vesicular structures below the plasma membrane. EGFR staining was located at cell-cell borders in untreated endothelium, but was mainly cytoplasmic by 15 min after EGF treatment. In control cultures, phosphorylation of EGFR Tyr992 peaked by 5 min following EGF stimulation and rapidly decreased to basal levels by 30 min. In cultures pretreated with CinnGEL 2Me, Tyr992 phosphorylation peaked 2 min following EGF addition and was consistently sustained at a higher level than controls until 60 min after treatment. By 18 h following EGF treatment, cultures pretreated with CinnGEL 2Me exhibited a 1.7 fold increase in the number of Ki67-positive cells compared with control cultures. Conclusions Comparison of PTP1B mRNA and protein levels indicates that PTP1B expression is regulated mainly at the protein level and is higher in subconfluent cells. PTP1B was located in vesicles below the plasma membrane. The fact that

Waterproofing Materials for Ammonium Nitrate

OpenAIRE

R.S. Damse

2004-01-01

This study explores the possibility of overcoming the problem of hygroscopicity of ammonium nitrate by coating the particles with selected waterproofing materials. Gravimetric analysis ofthe samples of ammonium nitrate coated with eight different waterproofing materials, vis-a-vis, uncoated ammonium nitrate, were conducted at different relative humidity and exposuretime. The results indicate that mineral jelly is the promising waterproofing material for ammonium nitrate among the materials te...

Using Nitrate Isotopes to Distinguish Pathways along which Unprocessed Atmospheric Nitrate is Transported through Forests to Streams

Science.gov (United States)

Sebestyen, S. D.

2013-12-01

Evaluation of natural abundance oxygen and nitrogen isotopes in nitrate has revealed that atmospheric deposition of nitrate to forests sometimes has direct effects on the timing and magnitude of stream nitrate concentrations. Large amounts of unprocessed atmospheric nitrate have sometimes been found in streams during snowmelt and stormflow events. Despite increasing evidence that unprocessed atmospheric nitrate may be transported without biological processing to streams at various times and multiple locations, little has been reported about specific hydrological processes. I synthesized research findings from a number of studies in which nitrate isotopes have been measured over the past decade. Unprocessed nitrate may predominate in surficial soil waters after rainfall and snowmelt events relative to nitrate that originated from nitrification. Although transport to deep groundwater may be important in the most nitrogen saturated catchments, the transport of unprocessed atmospheric nitrate along shallow subsurface flowpaths is likely more important in many moderately N-polluted ecosystems, which predominate in the northeastern USA where most of my study sites are located. The presence of unprocessed atmospheric nitrate in surficial soils was linked to stream nitrate concentrations when large amounts of unprocessed nitrate were occasionally routed along lateral, shallow subsurface flowpaths during stormflow events. During these events, water tables rose to saturate shallow-depth soils. When catchments were drying or dryer, atmospheric nitrate was completely consumed by biological processing as flowpaths shifted from lateral to vertical transport through soils. The source areas of unprocessed atmospheric nitrate were usually limited to soils that were adjacent to streams, with little to no near-surface saturation and transport of unprocessed nitrate from more distal hillslope positions. The occasional large amounts of unprocessed atmospheric nitrate in soil water

Receptor protein tyrosine phosphatase alpha activates Src-family kinases and controls integrin-mediated responses in fibroblasts

DEFF Research Database (Denmark)

Su, J; Muranjan, M; Sap, J

1999-01-01

of tyrosine kinases, the activity of which is tightly controlled by inhibitory phosphorylation of a carboxyterminal tyrosine residue (Tyr527 in chicken c-Src); this phosphorylation induces the kinases to form an inactive conformation. Whereas the identity of such inhibitory Tyr527 kinases has been well...... established, no corresponding phosphatases have been identified that, under physiological conditions, function as positive regulators of c-Src and Fyn in fibroblasts. RESULTS: Receptor protein tyrosine phosphatase alpha (RPTPalpha) was inactivated by homologous recombination. Fibroblasts derived from...... these RPTPalpha-/- mice had impaired tyrosine kinase activity of both c-Src and Fyn, and this was accompanied by a concomitant increase in c-Src Tyr527 phosphorylation. RPTPalpha-/- fibroblasts also showed a reduction in the rate of spreading on fibronectin substrates, a trait that is a phenocopy of the effect...

The acute effect of addition of nitrate on in vitro and in vivo methane emission in dairy cows

DEFF Research Database (Denmark)

Lund, Peter; Dahl, R.; Yang, H. J.

2014-01-01

The effects of a 24-h administration of a ration high in nitrate (20 g/kg DM) on DM intake and enteric gas production in lactating dairy cows as well as the effect of different doses of nitrate on in vitro fermentation were studied. Nitrate reduced in vivo methane (CH4) production by 31%, and CH4...... rumen fermentation in terms of DM degradability, pH, ammonia nitrogen, microbial protein and volatile fatty acid production, but it decreased gas production with longer initial delay time before onset of gas production and lower gas production rate. Nitrate added at 7-20 g/kg ration DM significantly...

Membrane-anchored MucR mediates nitrate-dependent regulation of alginate production in Pseudomonas aeruginosa

KAUST Repository

Wang, Yajie

2015-04-29

Alginates exhibit unique material properties suitable for medical and industrial applications. However, if produced by Pseudomonas aeruginosa, it is an important virulence factor in infection of cystic fibrosis patients. The alginate biosynthesis machinery is activated by c-di-GMP imparted by the inner membrane protein, MucR. Here, it was shown that MucR impairs alginate production in response to nitrate in P. aeruginosa. Subsequent site-specific mutagenesis of MucR revealed that the second MHYT sensor motif (MHYT II, amino acids 121–124) of MucR sensor domain was involved in nitrate sensing. We also showed that both c-di-GMP synthesizing and degrading active sites of MucR were important for alginate production. Although nitrate and deletion of MucR impaired alginate promoter activity and global c-di-GMP levels, alginate yields were not directly correlated with alginate promoter activity or c-di-GMP levels, suggesting that nitrate and MucR modulate alginate production at a post-translational level through a localized pool of c-di-GMP. Nitrate increased pel promoter activity in the mucR mutant while in the same mutant the psl promoter activity was independent of nitrate. Nitrate and deletion of mucR did not impact on swarming motility but impaired attachment to solid surfaces. Nitrate and deletion of mucR promoted the formation of biofilms with increased thickness, cell density, and survival. Overall, this study provided insight into the functional role of MucR with respect to nitrate-mediated regulation of alginate biosynthesis. © 2015 Springer-Verlag Berlin Heidelberg

Criticality parameters for uranyl nitrate or plutonium nitrate systems in tributyl phosphate/kerosine and water

International Nuclear Information System (INIS)

Weber, W.

1985-01-01

This report presents the calculated values of smallest critical masses and volumina and neutron physical parameters for uranyl nitrate (3, 4, 5% U-235) or plutonium nitrate (5% Pu-240), each in a 30 per cent solution of tributyl phosphate (TBP)/kerosine. For the corresponding nitrate-water solutions, newly calculated results are presented together with a revised solution density model. A comparison of the data shows to what extent the criticality of nitrate-TBP/kerosine systems can be assessed on the basis of nitrate-water parameters, revealing that such data can be applied to uranyl nitrate/water systems, taking into account that the smallest critical mass of uranyl nitrate-TBP/kerosine systems, up to a 5 p.c. U-235 enrichment, is by 4.5 p.c. at the most smaller than that of UNH-water solutions. Plutonium nitrate (5% Pu-240) in the TBP/kerosine solution will have a smallest critical mass of up to 7 p.c. smaller, as compared with the water data. The suitability of the computing methods and cross-sections used is verified by recalculating experiments carried out to determine the lowest critical enrichment of uranyl nitrate. The calculated results are well in agreement with experimental data. The lowest critical enrichment is calculated to be 2.10 p.c. in the isotope U-235. (orig.) [de

Nitrate-Rich Vegetables Increase Plasma Nitrate and Nitrite Concentrations and Lower Blood Pressure in Healthy Adults.

Science.gov (United States)

Jonvik, Kristin L; Nyakayiru, Jean; Pinckaers, Philippe Jm; Senden, Joan Mg; van Loon, Luc Jc; Verdijk, Lex B

2016-05-01

Dietary nitrate is receiving increased attention due to its reported ergogenic and cardioprotective properties. The extent to which ingestion of various nitrate-rich vegetables increases postprandial plasma nitrate and nitrite concentrations and lowers blood pressure is currently unknown. We aimed to assess the impact of ingesting different nitrate-rich vegetables on subsequent plasma nitrate and nitrite concentrations and resting blood pressure in healthy normotensive individuals. With the use of a semirandomized crossover design, 11 men and 7 women [mean ± SEM age: 28 ± 1 y; mean ± SEM body mass index (BMI, in kg/m(2)): 23 ± 1; exercise: 1-10 h/wk] ingested 4 different beverages, each containing 800 mg (∼12.9 mmol) nitrate: sodium nitrate (NaNO3), concentrated beetroot juice, a rocket salad beverage, and a spinach beverage. Plasma nitrate and nitrite concentrations and blood pressure were determined before and up to 300 min after beverage ingestion. Data were analyzed using repeated-measures ANOVA. Plasma nitrate and nitrite concentrations increased after ingestion of all 4 beverages (P nitrate concentrations were similar for all treatments (all values presented as means ± SEMs: NaNO3: 583 ± 29 μmol/L; beetroot juice: 597 ± 23 μmol/L; rocket salad beverage: 584 ± 24 μmol/L; spinach beverage: 584 ± 23 μmol/L). Peak plasma nitrite concentrations were different between treatments (NaNO3: 580 ± 58 nmol/L; beetroot juice: 557 ± 57 nmol/L; rocket salad beverage: 643 ± 63 nmol/L; spinach beverage: 980 ± 160 nmol/L; P = 0.016). When compared with baseline, systolic blood pressure declined 150 min after ingestion of beetroot juice (from 118 ± 2 to 113 ± 2 mm Hg; P nitrate-rich beetroot juice, rocket salad beverage, and spinach beverage effectively increases plasma nitrate and nitrite concentrations and lowers blood pressure to a greater extent than sodium nitrate. These findings show that nitrate-rich vegetables can be used as dietary nitrate

Ingested nitrate and nitrite and stomach cancer risk: an updated review.

Science.gov (United States)

Bryan, Nathan S; Alexander, Dominik D; Coughlin, James R; Milkowski, Andrew L; Boffetta, Paolo

2012-10-01

Nitrite and nitrate are naturally occurring molecules in vegetables and also added to cured and processed meats to delay spoilage and pathogenic bacteria growth. Research over the past 15 years has led to a paradigm change in our ideas about health effects of both nitrite and nitrate. Whereas, historically nitrite and nitrate were considered harmful food additives and listed as probable human carcinogens under conditions where endogenous nitrosation could take place, they are now considered by some as indispensible nutrients essential for cardiovascular health by promoting nitric oxide (NO) production. We provide an update to the literature and knowledge base concerning their safety. Most nitrite and nitrate exposure comes from naturally occurring and endogenous sources and part of the cell signaling effects of NO involve nitrosation. Nitrosation must now be considered broadly in terms of both S- and N-nitrosated species, since S-nitrosation is kinetically favored. Protein S-nitrosation is a significant part of the role of NO in cellular signal transduction and is involved in critical aspects of cardiovascular health. A critical review of the animal toxicology literature of nitrite indicates that in the absence of co-administration of a carcinogenic nitrosamine precursor, there is no evidence for carcinogenesis. Newly published prospective epidemiological cohort studies indicate that there is no association between estimated intake of nitrite and nitrate in the diet and stomach cancer. This new and growing body of evidence calls for a reconsideration of nitrite and nitrate safety. Copyright © 2012 Elsevier Ltd. All rights reserved.

Dissimilatory nitrate reduction to nitrate, nitrous oxide, and ammonium by Pseudomonas putrefaciens.

OpenAIRE

Samuelsson, M O

1985-01-01

The influence of redox potential on dissimilatory nitrate reduction to ammonium was investigated on a marine bacterium, Pseudomonas putrefaciens. Nitrate was consumed (3.1 mmol liter-1), and ammonium was produced in cultures with glucose and without sodium thioglycolate. When sodium thioglycolate was added, nitrate was consumed at a lower rate (1.1 mmol liter-1), and no significant amounts of nitrite or ammonium were produced. No growth was detected in glucose media either with or without sod...

Arabidopsis Yak1 protein (AtYak1) is a dual specificity protein kinase

KAUST Repository

Kim, Dongjin; Ntui, Valentine Otang; Zhang, Nianshu; Xiong, Liming

2015-01-01

Yak1 is a member of dual-specificity Tyr phosphorylation-regulated kinases (DYRKs) that are evolutionarily conserved. The downstream targets of Yak1 and their functions are largely unknown. Here, a homologous protein AtYAK1 was identified in Arabidopsis thaliana and the phosphoprotein profiles of the wild type and an atyak1 mutant were compared on two-dimensional gel following Pro-Q Diamond phosphoprotein gel staining. Annexin1, Annexin2 and RBD were phosphorylated at serine/ threonine residues by the AtYak1 kinase. Annexin1, Annexin2 and Annexin4 were also phosphorylated at tyrosine residues. Our study demonstrated that AtYak1 is a dual specificity protein kinase in Arabidopsis that may regulate the phosphorylation status of the annexin family proteins.

Arabidopsis Yak1 protein (AtYak1) is a dual specificity protein kinase

KAUST Repository

Kim, Dongjin

2015-10-09

Yak1 is a member of dual-specificity Tyr phosphorylation-regulated kinases (DYRKs) that are evolutionarily conserved. The downstream targets of Yak1 and their functions are largely unknown. Here, a homologous protein AtYAK1 was identified in Arabidopsis thaliana and the phosphoprotein profiles of the wild type and an atyak1 mutant were compared on two-dimensional gel following Pro-Q Diamond phosphoprotein gel staining. Annexin1, Annexin2 and RBD were phosphorylated at serine/ threonine residues by the AtYak1 kinase. Annexin1, Annexin2 and Annexin4 were also phosphorylated at tyrosine residues. Our study demonstrated that AtYak1 is a dual specificity protein kinase in Arabidopsis that may regulate the phosphorylation status of the annexin family proteins.

Nitrated alpha-synuclein immunity accelerates degeneration of nigral dopaminergic neurons.

Directory of Open Access Journals (Sweden)

Eric J Benner

2008-01-01

Full Text Available The neuropathology of Parkinson's disease (PD includes loss of dopaminergic neurons in the substantia nigra, nitrated alpha-synuclein (N-alpha-Syn enriched intraneuronal inclusions or Lewy bodies and neuroinflammation. While the contribution of innate microglial inflammatory activities to disease are known, evidence for how adaptive immune mechanisms may affect the course of PD remains obscure. We reasoned that PD-associated oxidative protein modifications create novel antigenic epitopes capable of peripheral adaptive T cell responses that could affect nigrostriatal degeneration.Nitrotyrosine (NT-modified alpha-Syn was detected readily in cervical lymph nodes (CLN from 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP intoxicated mice. Antigen-presenting cells within the CLN showed increased surface expression of major histocompatibility complex class II, initiating the molecular machinery necessary for efficient antigen presentation. MPTP-treated mice produced antibodies to native and nitrated alpha-Syn. Mice immunized with the NT-modified C-terminal tail fragment of alpha-Syn, but not native protein, generated robust T cell proliferative and pro-inflammatory secretory responses specific only for the modified antigen. T cells generated against the nitrated epitope do not respond to the unmodified protein. Mice deficient in T and B lymphocytes were resistant to MPTP-induced neurodegeneration. Transfer of T cells from mice immunized with N-alpha-Syn led to a robust neuroinflammatory response with accelerated dopaminergic cell loss.These data show that NT modifications within alpha-Syn, can bypass or break immunological tolerance and activate peripheral leukocytes in draining lymphoid tissue. A novel mechanism for disease is made in that NT modifications in alpha-Syn induce adaptive immune responses that exacerbate PD pathobiology. These results have implications for both the pathogenesis and treatment of this disabling neurodegenerative disease.

Nitrate leaching index

Science.gov (United States)

The Nitrate Leaching Index is a rapid assessment tool that evaluates nitrate (NO3) leaching potential based on basic soil and climate information. It is the basis for many nutrient management planning efforts, but it has considerable limitations because of : 1) an oversimplification of the processes...

Direct transcriptional activation of BT genes by NLP transcription factors is a key component of the nitrate response in Arabidopsis.

Science.gov (United States)

Sato, Takeo; Maekawa, Shugo; Konishi, Mineko; Yoshioka, Nozomi; Sasaki, Yuki; Maeda, Haruna; Ishida, Tetsuya; Kato, Yuki; Yamaguchi, Junji; Yanagisawa, Shuichi

2017-01-29

Nitrate modulates growth and development, functioning as a nutrient signal in plants. Although many changes in physiological processes in response to nitrate have been well characterized as nitrate responses, the molecular mechanisms underlying the nitrate response are not yet fully understood. Here, we show that NLP transcription factors, which are key regulators of the nitrate response, directly activate the nitrate-inducible expression of BT1 and BT2 encoding putative scaffold proteins with a plant-specific domain structure in Arabidopsis. Interestingly, the 35S promoter-driven expression of BT2 partially rescued growth inhibition caused by reductions in NLP activity in Arabidopsis. Furthermore, simultaneous disruption of BT1 and BT2 affected nitrate-dependent lateral root development. These results suggest that direct activation of BT1 and BT2 by NLP transcriptional activators is a key component of the molecular mechanism underlying the nitrate response in Arabidopsis. Copyright © 2016 Elsevier Inc. All rights reserved.

The nitrate time bomb: a numerical way to investigate nitrate storage and lag time in the unsaturated zone.

Science.gov (United States)

Wang, L; Butcher, A S; Stuart, M E; Gooddy, D C; Bloomfield, J P

2013-10-01

Nitrate pollution in groundwater, which is mainly from agricultural activities, remains an international problem. It threatens the environment, economics and human health. There is a rising trend in nitrate concentrations in many UK groundwater bodies. Research has shown it can take decades for leached nitrate from the soil to discharge into groundwater and surface water due to the 'store' of nitrate and its potentially long travel time in the unsaturated and saturated zones. However, this time lag is rarely considered in current water nitrate management and policy development. The aim of this study was to develop a catchment-scale integrated numerical method to investigate the nitrate lag time in the groundwater system, and the Eden Valley, UK, was selected as a case study area. The method involves three models, namely the nitrate time bomb-a process-based model to simulate the nitrate transport in the unsaturated zone (USZ), GISGroundwater--a GISGroundwater flow model, and N-FM--a model to simulate the nitrate transport in the saturated zone. This study answers the scientific questions of when the nitrate currently in the groundwater was loaded into the unsaturated zones and eventually reached the water table; is the rising groundwater nitrate concentration in the study area caused by historic nitrate load; what caused the uneven distribution of groundwater nitrate concentration in the study area; and whether the historic peak nitrate loading has reached the water table in the area. The groundwater nitrate in the area was mainly from the 1980s to 2000s, whilst the groundwater nitrate in most of the source protection zones leached into the system during 1940s-1970s; the large and spatially variable thickness of the USZ is one of the major reasons for unevenly distributed groundwater nitrate concentrations in the study area; the peak nitrate loading around 1983 has affected most of the study area. For areas around the Bowscar, Beacon Edge, Low Plains, Nord Vue

Effect of ammonium and nitrate on ferric chelate reductase and nitrate reductase in Vaccinium species.

Science.gov (United States)

Poonnachit, U; Darnell, R

2004-04-01

Most Vaccinium species have strict soil requirements for optimal growth, requiring low pH, high iron availability and nitrogen primarily in the ammonium form. These soils are limited and are often located near wetlands. Vaccinium arboreum is a wild species adapted to a wide range of soils, including high pH, low iron, and nitrate-containing soils. This broader soil adaptation in V. arboreum may be related to increased efficiency of iron or nitrate uptake compared with the cultivated Vaccinium species. Nitrate, ammonium and iron uptake, and nitrate reductase (NR) and ferric chelate reductase (FCR) activities were compared in two Vaccinium species grown hydroponically in either nitrate or ammonia, with or without iron. The species studied were the wild V. arboreum and the cultivated V. corymbosum interspecific hybrid, which exhibits the strict soil requirements of most Vaccinium species. Ammonium uptake was significantly greater than nitrate uptake in both species, while nitrate uptake was greater in the wild species, V. arboreum, compared with the cultivated species, V. corymbosum. The increased nitrate uptake in V. arboreum was correlated with increased root NR activity compared with V. corymbosum. The lower nitrate uptake in V. corymbosum was reflected in decreased plant dry weight in this species compared with V. arboreum. Root FCR activity increased significantly in V. corymbosum grown under iron-deficient conditions, compared with the same species grown under iron-sufficient conditions or with V. arboreum grown under either iron condition. V. arboreum appears to be more efficient in acquiring nitrate compared with V. corymbosum, possibly due to increased NR activity and this may partially explain the wider soil adaptation of V. arboreum.

Agricultural nitrate pollution

DEFF Research Database (Denmark)

Anker, Helle Tegner

2015-01-01

Despite the passing of almost 25 years since the adoption of the EU Nitrates Directive, agricultural nitrate pollution remains a major concern in most EU Member States. This is also the case in Denmark, although a fairly strict regulatory regime has resulted in almost a 50 per cent reduction...

Negative feedback loops leading to nitrate homeostasis and oscillatory nitrate assimilation in plants and fungi.

OpenAIRE

Huang, Yongshun

2011-01-01

Master's thesis in Biological Chemistry Nitrate is an important nutrient for plants and fungi. For plants it has been shown that cytosolic nitrate levels are under homeostatic control. Here we describe two networks that can obtain robust, i.e. perturbation independent, homeostatic behavior in cytosolic nitrate concentration. One of the networks, a member in the family of outflow controllers, is based on a negative feedback loop containing a nitrate-induced activation of a controller molecu...

Improvement in the 111In-DTPA-TYR3-octreotide and 177Lu-DOTATYR3- octreotate production

International Nuclear Information System (INIS)

Souza, Adriano A.; Herrerias, Rosana; Pires, Jose A.; Alves, Geraldo P.; Fukumori, Neuza T.O.; Matsuda, Margareth M.N.; Almeida, Erika V.; Mengatti, Jair; Barboza, Marycel F. de

2009-01-01

Recent advances in receptor mediated-tumor imaging have resulted in the development of somatostatin analogues, the biomolecular basis for the clinical use of these compounds in nuclear medicine for diagnostic and peptide receptor radionuclide therapy (PRRT). PRRT is a very good therapeutic option for patients with metastatic neuroendocrine gastroenteropancreatic (GEP) tumour. Clinical studies with different sst-positives tumors proved advantages of [ 177 Lu-DOTA-Tyr 3 ]octreotate (DOTATATE) for therapy. The aim of this work is to establish and validate the labeling, the quality control procedures of DTPA-Tyr 3 -Octreotide (DTPA-Oct) and DOTA-Tyr 3 -Octreotate (DOTATATE) labeled with In-111 and Lu-177, respectively, for routine production at Radiopharmacy Directory (DIRF) Brazil. Labeling were performed in a 'glove-box' using 111 InCl 3 (Nordion) and in hot-cell with 177 LuCl 3 (IDB-Holland) at pH 4.5; using DTPA-Oct (Pichem) and DOTATATE (IDBHolland) at room temperature and at 82-85 deg C for 30 minutes, respectively. The radiochemical purity was determined by ITLC-SG in 0.1 mol L -1 sodium citrate, pH 5.5 and by Sep-Pak silica cartridge. Sterility was performed by the microbiology procedures and pyrogen tests by the 'in-vitro' Limulus test (LAL). The stability of both radiolabeled peptides was high even 72 hours under refrigeration. The radiochemical purities of the labeled compounds were confirmed by HPLC. Sterility and pyrogen tests were negative in all delivered vials. The efficient procedure to obtain 111 In-DTPA-Oct and 177 Lu-DOTATATE was confirmed in the first comparative clinical groups. The methods were validated and 46.287 GMBq of 111 In-DTPA-Oct and 1,193 GBq of 177 Lu-DOTATATE were distributed in 2008, to nuclear medicine services in Brazil. (author)

Mutation of the Arabidopsis NRT1.5 nitrate transporter causes defective root-to-shoot nitrate transport.

Science.gov (United States)

Lin, Shan-Hua; Kuo, Hui-Fen; Canivenc, Geneviève; Lin, Choun-Sea; Lepetit, Marc; Hsu, Po-Kai; Tillard, Pascal; Lin, Huey-Ling; Wang, Ya-Yun; Tsai, Chyn-Bey; Gojon, Alain; Tsay, Yi-Fang

2008-09-01

Little is known about the molecular and regulatory mechanisms of long-distance nitrate transport in higher plants. NRT1.5 is one of the 53 Arabidopsis thaliana nitrate transporter NRT1 (Peptide Transporter PTR) genes, of which two members, NRT1.1 (CHL1 for Chlorate resistant 1) and NRT1.2, have been shown to be involved in nitrate uptake. Functional analysis of cRNA-injected Xenopus laevis oocytes showed that NRT1.5 is a low-affinity, pH-dependent bidirectional nitrate transporter. Subcellular localization in plant protoplasts and in planta promoter-beta-glucuronidase analysis, as well as in situ hybridization, showed that NRT1.5 is located in the plasma membrane and is expressed in root pericycle cells close to the xylem. Knockdown or knockout mutations of NRT1.5 reduced the amount of nitrate transported from the root to the shoot, suggesting that NRT1.5 participates in root xylem loading of nitrate. However, root-to-shoot nitrate transport was not completely eliminated in the NRT1.5 knockout mutant, and reduction of NRT1.5 in the nrt1.1 background did not affect root-to-shoot nitrate transport. These data suggest that, in addition to that involving NRT1.5, another mechanism is responsible for xylem loading of nitrate. Further analyses of the nrt1.5 mutants revealed a regulatory loop between nitrate and potassium at the xylem transport step.

Stimulation effect of synthetic cytokinins on the uptake and incorporation of nitrogen-15-labelled ammonium nitrate and urea in wheat leaves

International Nuclear Information System (INIS)

Iglewski, S.M.; Szarvas, T.; Pozsar, B.I.

1977-01-01

The turnover of different labelled nitrogen sources in wheat leaves has been investigated using the isotopic tracer technique. The 15 N at.% was determined in free ammonium ion, in the nitrate and the nitrite levels, and also in the non-disintegrated urea. The accumulation and the incorporation of stable nitrogen was measured in the TCA insoluble protein fraction. According to the experimental data the intensity of incorporation of urea nitrogen is relatively higher than that of the different inorganic compounds. The utilization of ammonium ion was 76% compared with the urea, whereas that of the nitrate nitrogen was 60% in the wheat leaves. The incorporation rate of the two nitrogen atoms from ammonium nitrate was 32% lower than that of the urea nitrogen, in the leaf protein of Bezostaia-1 wheat variety. The turnover of urea through the transamination was very rapid, the amination with ammonium ion was slower, and the first phase of the nitrate reduction was relatively more retarded than the nitrite reduction. The endogenous cytokinin-like biological activity and some synthetic cytokinins (kinetin, benzyladenine) have a remarkably stimulating effect on the incorporation of the different 15 N-labelled nitrogen sources into the leaf protein fraction. (author)

Identification of nitrate sources and discharge-depending nitrate dynamics in a mesoscale catchment

Science.gov (United States)

Mueller, Christin; Strachauer, Ulrike; Brauns, Mario; Musolff, Andreas; Kunz, Julia Vanessa; Brase, Lisa; Tarasova, Larisa; Merz, Ralf; Knöller, Kay

2017-04-01

During the last decades, nitrate concentrations in surface and groundwater have increased due to land use change and accompanying application of fertilizer in agriculture as well as increased atmospheric deposition. To mitigate nutrient impacts on downstream aquatic ecosystems, it is important to quantify potential nitrate sources, instream nitrate processing and its controls in a river system. The objective of this project is to characterize and quantify (regional) scale dynamics and trends in water and nitrogen fluxes of the entire Holtemme river catchment in central Germany making use of isotopic fingerprinting methods. Here we compare two key date sampling campaigns in 2014 and 2015, with spatially highly resolved measurements of discharge at 23 sampling locations including 11 major tributaries and 12 locations at the main river. Additionally, we have data from continuous runoff measurements at 10 locations operated by the local water authorities. Two waste water treatment plants contribute nitrogen to the Holtemme stream. This contribution impacts nitrate loads and nitrate isotopic signatures depending on the prevailing hydrological conditions. Nitrogen isotopic signatures in the catchment are mainly controlled by different sources (nitrified soil nitrogen in the headwater and manure/ effluents from WWTPs in the lowlands) and increase with raising nitrate concentrations along the main river. Nitrate loads at the outlet of the catchment are extremely different between both sampling campaigns (2014: NO3- = 97 t a-1, 2015: NO3- = 5 t a-1) which is associated with various runoff (2014: 0.8 m3 s-1, 2015: 0.2 m3 s-1). In 2015, the inflow from WWTP's raises the NO3- loads and enriches δ18O-NO3 values. Generally, oxygen isotope signatures from nitrate are more variable and are controlled by biogeochemical processes in concert with the oxygen isotopic composition of the ambient water. Elevated δ18O-NO3 in 2015 are most likely due to higher temperatures and lower

Nitrate induces a type 1 diabetic profile in alligator hatchlings.

Science.gov (United States)

Edwards, Thea M; Hamlin, Heather J; Freymiller, Haley; Green, Stephen; Thurman, Jenna; Guillette, Louis J

2018-01-01

Type 1 diabetes (T1D) is a chronic autoimmune disease that affects 1 in 300 children by age 18. T1D is caused by inflammation-induced loss of insulin-producing pancreatic beta cells, leading to high blood glucose and a host of downstream complications. Although multiple genes are associated with T1D risk, only 5% of genetically susceptible individuals actually develop clinical disease. Moreover, a growing number of T1D cases occur in geographic clusters and among children with low risk genotypes. These observations suggest that environmental factors contribute to T1D etiology. One potential factor, supported primarily by epidemiological studies, is the presence of nitrate and nitrite in drinking water. To test this hypothesis, female hatchling alligators were exposed to environmentally relevant concentrations of nitrate in their tank water (reference, 10mg/L, or 100mg/L NO 3 -N) from hatch through 5 weeks or 5 months of age. At each time point, endpoints related to T1D were investigated: plasma levels of glucose, triglycerides, testosterone, estradiol, and thyroxine; pancreas, fat body, and thyroid weights; weight gain or loss; presence of immune cells in the pancreas; and pancreatic beta cell number, assessed by antibody staining of nkx6.1 protein. Internal dosing of nitrate was confirmed by measuring plasma and urine nitrate levels and whole blood methemoglobin. Cluster analysis indicated that high nitrate exposure (most animals exposed to 100mg/L NO3-N and one alligator exposed to 10mg/L NO3-N) induced a profile of endpoints consistent with early T1D that could be detected after 5 weeks and was more strongly present after 5 months. Our study supports epidemiological data correlating elevated nitrate with T1D onset in humans, and highlights nitrate as a possible environmental contributor to the etiology of T1D, possibly through its role as a nitric oxide precursor. Copyright © 2017 Elsevier Inc. All rights reserved.

Nitrate removal from high strength nitrate-bearing wastes in granular sludge sequencing batch reactors.

Science.gov (United States)

Krishna Mohan, Tulasi Venkata; Renu, Kadali; Nancharaiah, Yarlagadda Venkata; Satya Sai, Pedapati Murali; Venugopalan, Vayalam Purath

2016-02-01

A 6-L sequencing batch reactor (SBR) was operated for development of granular sludge capable of denitrification of high strength nitrates. Complete and stable denitrification of up to 5420 mg L(-1) nitrate-N (2710 mg L(-1) nitrate-N in reactor) was achieved by feeding simulated nitrate waste at a C/N ratio of 3. Compact and dense denitrifying granular sludge with relatively stable microbial community was developed during reactor operation. Accumulation of large amounts of nitrite due to incomplete denitrification occurred when the SBR was fed with 5420 mg L(-1) NO3-N at a C/N ratio of 2. Complete denitrification could not be achieved at this C/N ratio, even after one week of reactor operation as the nitrite levels continued to accumulate. In order to improve denitrification performance, the reactor was fed with nitrate concentrations of 1354 mg L(-1), while keeping C/N ratio at 2. Subsequently, nitrate concentration in the feed was increased in a step-wise manner to establish complete denitrification of 5420 mg L(-1) NO3-N at a C/N ratio of 2. The results show that substrate concentration plays an important role in denitrification of high strength nitrate by influencing nitrite accumulation. Complete denitrification of high strength nitrates can be achieved at lower substrate concentrations, by an appropriate acclimatization strategy. Copyright © 2015 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Catalyzed reduction of nitrate in aqueous solutions

International Nuclear Information System (INIS)

Haas, P.A.

1994-08-01

Sodium nitrate and other nitrate salts in wastes is a major source of difficulty for permanent disposal. Reduction of nitrate using aluminum metal has been demonstrated, but NH 3 , hydrazine, or organic compounds containing oxygen would be advantageous for reduction of nitrate in sodium nitrate solutions. Objective of this seed money study was to determine minimum conditions for reduction. Proposed procedure was batchwise heating of aqueous solutions in closed vessels with monitoring of temperatures and pressures. A simple, convenient apparatus and procedure were demonstrated for observing formation of gaseous products and collecting samples for analyses. The test conditions were 250 degree C and 1000 psi max. Any useful reduction of sodium nitrate to sodium hydroxide as the primary product was not found. The nitrate present at pHs 3 or NH 4 NO 3 is easily decomposed, and the effect of nitromethane at these low pHs was confirmed. When acetic acid or formic acid was added, 21 to 56% of the nitrate in sodium nitrate solutions was reduced by methanol or formaldehyde. With hydrazine and acetic acid, 73 % of the nitrate was decomposed to convert NaNO 3 to sodium acetate. With hydrazine and formic acid, 36% of the nitrate was decomposed. If these products are more acceptable for final disposal than sodium nitrate, the reagents are cheap and the conversion conditions would be practical for easy use. Ammonium acetate or formate salts did not significantly reduce nitrate in sodium nitrate solutions

Photobinding of tiaprofenic acid and suprofen to proteins and cells: a combined study using radiolabeling, antibodies and laser flash photolysis of model bichromophores.

Science.gov (United States)

Castell, J V; Hernández, D; Gómez-Lechón, M J; Lahoz, A; Miranda, M A; Morera, I M; Pérez-Prieto, J; Sarabia, Z

1998-11-01

Drug photoallergy is a matter of current concern. It involves the formation of drug-protein photoadducts (photoantigens) that may ultimately trigger an immunological response. Tyrosine residues appear to be key binding sites in proteins. The present work has investigated the photobinding of tiaprofenic and (TPA) and the closely related isomer suprofen (SUP) to proteins and cells by means of radioactive labelling and drug-directed antibodies. To ascertain whether preassociation with the protein may play a role in photoreactivity, two model bichromophoric compounds (TPA-Tyr and SUP-Tyr) have been prepared and studied by laser flash photolysis. The results of this work show that (a) TPA and SUP photobind to proteins with similar efficiencies, (b) both drugs form photoadducts that share a basic common structure, as they are recognized by the same antibody and (c) drug-protein preassociation must play a key role in photoreactivity, as indicated by the dramatic decrease in the triplet state lifetimes of the model bichromophores compared to the parent drugs.

Modeled Wet Nitrate Deposition

Data.gov (United States)

U.S. Environmental Protection Agency — Modeled data on nitrate wet deposition was obtained from Dr. Jeff Grimm at Penn State Univ. Nitrate wet depostion causes acidification and eutrophication of surface...

The mechanism of the tyrosine transporter TyrP supports a proton motive tyrosine decarboxylation pathway in Lactobacillus brevis

NARCIS (Netherlands)

Wolken, WAM; Lucas, PM; Lonvaud-Funel, A; Lolkema, JS; Wolken, Wout A.M.; Lucas, Patrick M.

The tyrosine decarboxylase operon of Lactobacillus brevis IOEB9809 contains, adjacent to the tyrosine decarboxylase gene, a gene for TyrP, a putative tyrosine transporter. The two genes potentially form a proton motive tyrosine decarboxylation pathway. The putative tyrosine transporter gene of L.

Evaluation of nitrate destruction methods

International Nuclear Information System (INIS)

Taylor, P.A.; Kurath, D.E.; Guenther, R.

1993-01-01

A wide variety of high nitrate-concentration aqueous mixed [radioactive and Resource Conservation and Recovery Act (RCRA) hazardous] wastes are stored at various US Department of Energy (DOE) facilities. These wastes will ultimately be solidified for final disposal, although the waste acceptance criteria for the final waste form is still being determined. Because the nitrates in the wastes will normally increase the volume or reduce the integrity of all of the waste forms under consideration for final disposal, nitrate destruction before solidification of the waste will generally be beneficial. This report describes and evaluates various technologies that could be used to destroy the nitrates in the stored wastes. This work was funded by the Department of Energy's Office of Technology Development, through the Chemical/Physical Technology Support Group of the Mixed Waste Integrated Program. All the nitrate destruction technologies will require further development work before a facility could be designed and built to treat the majority of the stored wastes. Several of the technologies have particularly attractive features: the nitrate to ammonia and ceramic (NAC) process produces an insoluble waste form with a significant volume reduction, electrochemical reduction destroys nitrates without any chemical addition, and the hydrothermal process can simultaneously treat nitrates and organics in both acidic and alkaline wastes. These three technologies have been tested using lab-scale equipment and surrogate solutions. At their current state of development, it is not possible to predict which process will be the most beneficial for a particular waste stream

Tritium labelling of highly selective probes for. delta. -opioid receptors: ( sup 3 H)Tyr-D-Ser(O-t-Bu)-Gly-Phe-Leu-Thr(DSTBULET) and ( sup 3 H)Tyr-D-Ser(O-t-Bu)-Gly-Phe-Leu-Thr(O-t-Bu)(BUBU)

Energy Technology Data Exchange (ETDEWEB)

Fellion, E.; Gacel, G.; Roques, B.P. (Institut National de la Sante et de la Recherche Medicale (INSERM), 75 - Paris (France)); Roy, J.; Morgat, J.L. (CEA Centre d' Etudes Nucleaires de Saclay, 91 - Gif-sur-Yvette (France). Service de Biochimie)

1990-08-01

The introduction of bulky residue(s) in linear enkephalin-related hexapeptides represents a new approach in the design of selective probes for {delta}-opioid receptors, displaying the appropriate criteria to investigate biological and pharmacological properties of the assumed binding site ({delta}) of endogenous enkephalins. The selectivities and high affinities of Tyr-D-Ser(O-t-Bu)-Gly-Phe-Leu-Thr(DSTBULET) and especially Tyr-D-Ser(O-t-Bu)Gly-Phe-Leu-Thr(O-t-Bu) (BUBU) associated with a satisfactory resistance to peptidases, make them the most suitable {delta}-probes reported to date. In the present paper, we report the synthesis of DSTBULET and BUBU under tritiated forms with high specific radioactivities. These radio-labelled probes will enable extensive in vitro and in vivo investigations of {delta}-opioid receptors properties to be carried out. (author).

[Effects of nitrogen application level on soil nitrate accumulation and ammonia volatilization in high-yielding wheat field].

Science.gov (United States)

Wang, Dong; Yu, Zhenwen; Yu, Wenming; Shi, Yu; Zhou, Zhongxin

2006-09-01

The study showed that during the period from sowing to pre-wintering, the soil nitrate in high-yielding wheat field moved down to deeper layers, and accumulated in the layers below 140 cm. An application rate of 96-168 kg N x hm(-2) increased the nitrate content in 0-60 cm soil layer and the wheat grain yield and its protein content, and decreased the proportion of apparent N loss to applied N and the ammonia volatilization loss from basal nitrogen. Applying 240 kg N x hm(-2) promoted the downward movement of soil nitrate and its accumulation in deeper layers, increased the proportion of apparent N loss to applied N and the ammonia volatilization loss from basal nitrogen, had no significant effect on the protein content of wheat grain, but decreased the grain yield. The appropriate application rate of nitrogen on high-yielding wheat field was 132-204 kg N x hm(-2).

An artificial self-sufficient cytochrome P450 directly nitrates fluorinated tryptophan analogs with a different regio-selectivity.

Science.gov (United States)

Zuo, Ran; Zhang, Yi; Huguet-Tapia, Jose C; Mehta, Mishal; Dedic, Evelina; Bruner, Steven D; Loria, Rosemary; Ding, Yousong

2016-05-01

Aromatic nitration is an immensely important industrial process to produce chemicals for a variety of applications, but it often suffers from multiple unsolved challenges. Enzymes as biocatalysts have been increasingly used for organic chemistry synthesis due to their high selectivity and environmental friendliness, but nitration has benefited minimally from the development of biocatalysis. In this work, we aimed to develop TxtE as practical biocatalysts for aromatic nitration. TxtE is a unique class I cytochrome P450 enzyme that nitrates the indole of l-tryptophan. To develop cost-efficient nitration processes, we fused TxtE with the reductase domains of CYP102A1 (P450BM3) and of P450RhF to create class III self-sufficient biocatalysts. The best engineered fusion protein was comparable with wild type TxtE in terms of nitration performance and other key biochemical properties. To demonstrate the application potential of the fusion enzyme, we nitrated 4-F-dl-tryptophan and 5-F-l-tryptophan in large scale enzymatic reactions. Tandem MS/MS and NMR analyses of isolated products revealed altered nitration sites. To our knowledge, these studies represent the first practice in developing biological nitration approaches and lay a solid basis to the use of TxtE-based biocatalysts for the production of valuable nitroaromatics. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The influence of nitrate concentrations and acidity on the electrocatalytic reduction of nitrate on platinum

NARCIS (Netherlands)

Groot, de M.T.; Koper, M.T.M.

2004-01-01

A study was performed to determine the influence of nitrate concentration and acidity on the reaction rate and selectivity of the electrocatalytic nitrate reduction on platinum. There are two different nitrate reduction mechanisms on platinum: a direct mechanism (0.4–0.1 V vs. SHE) and an indirect

Pre-therapeutic dosimetry and biodistribution of {sup 86}Y-DOTA-Phe{sup 1}-Tyr{sup 3}-octreotide versus {sup 111}In-pentetreotide in patients with advanced neuroendocrine tumours

Energy Technology Data Exchange (ETDEWEB)

Helisch, Andreas; Foerster, Gregor J.; Reber, Helmut; Buchholz, Hans-Georg; Bartenstein, Peter [University of Mainz, Department of Nuclear Medicine, Mainz (Germany); Arnold, Rudolf [Philips University, Division of Gastroenterology and Endocrinology, Department of Internal Medicine, Marburg (Germany); Goeke, Burkhard [Ludwig-Maximilians-University, Department of Internal Medicine II, Klinikum Grosshadern, Munich (Germany); Weber, Matthias M. [University of Mainz, Division of Endocrinology and Metabolism, Department of Internal Medicine, Mainz (Germany); Wiedenmann, Bertram [Campus Virchow Clinic, Department of Hepatology and Gastroenterology, Charite Medical School, Berlin (Germany); Pauwels, Stanislas [Catholic University of Louvain, Center of Nuclear Medicine, Brussels (Belgium); Haus, Ulrike [Novartis Pharmaceuticals, Nuremberg (Germany); Bouterfa, Hakim [Novartis Pharmaceuticals, Basel (Switzerland)

2004-10-01

For the internal radiotherapy of neuroendocrine tumours, the somatostatin analogue DOTATOC labelled with {sup 90}Y is frequently used [{sup 90}Y-DOTA-Phe{sup 1}-Tyr{sup 3}-octreotide (SMT487-OctreoTher)]. Radiation exposure to the kidneys is critical in this therapy as it may result in renal failure. The aim of this study was to compare cumulative organ and tumour doses based upon dosimetric data acquired with the chemically identical {sup 86}Y-DOTA-Phe{sup 1}-Tyr{sup 3}-octreotide (considered as the gold standard) and the commercially available {sup 111}In-pentetreotide. The cumulative organ and tumour doses for the therapeutic administration of 13.32 GBq {sup 90}Y-DOTA-Phe{sup 1}-Tyr{sup 3}-octreotide (three cycles, each of 4.44 GBq) were estimated based on the MIRD concept (MIRDOSE 3.1 and IMEDOSE). Patients with a cumulative kidney dose exceeding 27 Gy had to be excluded from subsequent therapy with {sup 90}Y-DOTA-Phe{sup 1}-Tyr{sup 3}-octreotide, in accordance with the directives of the German radiation protection authorities. The range of doses (mGy/MBq {sup 90}Y-DOTA-Phe{sup 1}-Tyr{sup 3}-octreotide) for kidneys, spleen, liver and tumour masses was 0.6-2.8, 1.5-4.2, 0.3-1.3 and 2.1-29.5 ({sup 86}Y-DOTA-Phe{sup 1}-Tyr{sup 3}-octreotide), respectively, versus 1.3-3.0, 1.8-4.4, 0.2-0.8 and 1.4-19.7 ({sup 111}In-pentetreotide), with wide inter-subject variability. Despite renal protection with amino acid infusions, estimated cumulative kidney doses in two patients exceeded 27 Gy. Compared with {sup 86}Y-DOTA-Phe{sup 1}-Tyr{sup 3}-octreotide, dosimetry with {sup 111}In-pentetreotide overestimated doses to kidneys and spleen, whereas the radiation dose to the tumour-free liver was underestimated. However, both dosimetric approaches detected the two patients with an exceptionally high radiation burden to the kidneys that carried a potential risk of renal failure following radionuclide therapy. (orig.)

Insights into the smooth-to-rough transitioning in Mycobacterium bolletii unravels a functional Tyr residue conserved in all mycobacterial MmpL family members.

Science.gov (United States)

Bernut, Audrey; Viljoen, Albertus; Dupont, Christian; Sapriel, Guillaume; Blaise, Mickaël; Bouchier, Christiane; Brosch, Roland; de Chastellier, Chantal; Herrmann, Jean-Louis; Kremer, Laurent

2016-03-01

In mycobacteria, MmpL proteins represent key components that participate in the biosynthesis of the complex cell envelope. Whole genome analysis of a spontaneous rough morphotype variant of Mycobacterium abscessus subsp. bolletii identified a conserved tyrosine that is crucial for the function of MmpL family proteins. Isogenic smooth (S) and rough (R) variants differed by a single mutation linked to a Y842H substitution in MmpL4a. This mutation caused a deficiency in glycopeptidolipid production/transport in the R variant and a gain in the capacity to produce cords in vitro. In zebrafish, increased virulence of the M. bolletii R variant over the parental S strain was found, involving massive production of serpentine cords, abscess formation and rapid larval death. Importantly, this finding allowed us to demonstrate an essential role of Tyr842 in several different MmpL proteins, including Mycobacterium tuberculosis MmpL3. Structural homology models of MmpL4a and MmpL3 identified two additional critical residues located in the transmembrane regions TM10 and TM4 that are facing each other. We propose that these central residues are part of the proton-motive force that supplies the energy for substrate transport. Hence, we provide important insights into mechanistic/structural aspects of MmpL proteins as lipid transporters and virulence determinants in mycobacteria. © 2015 John Wiley & Sons Ltd.

Mortality of nitrate fertiliser workers.

Science.gov (United States)

Al-Dabbagh, S; Forman, D; Bryson, D; Stratton, I; Doll, R

1986-01-01

An epidemiological cohort study was conducted to investigate the mortality patterns among a group of workers engaged in the production of nitrate based fertilisers. This study was designed to test the hypothesis that individuals exposed to high concentrations of nitrates might be at increased risk of developing cancers, particularly gastric cancer. A total of 1327 male workers who had been employed in the production of fertilisers between 1946 and 1981 and who had been occupationally exposed to nitrates for at least one year were followed up until 1 March 1981. In total, 304 deaths were observed in this group and these were compared with expected numbers calculated from mortality rates in the northern region of England, where the factory was located. Analysis was also carried out separately for a subgroup of the cohort who had been heavily exposed to nitrates--that is, working in an environment likely to contain more than 10 mg nitrate/m3 for a year or longer. In neither the entire cohort nor the subgroup was any significant excess observed for all causes of mortality or for mortality from any of five broad categories of cause or from four specific types of cancer. A small excess of lung cancer was noted more than 20 years after first exposure in men heavily exposed for more than 10 years. That men were exposed to high concentrations of nitrate was confirmed by comparing concentrations of nitrates in the saliva of a sample of currently employed men with control men, employed at the same factory but not in fertiliser production. The men exposed to nitrate had substantially raised concentrations of nitrate in their saliva compared with both controls within the industry and with men in the general population and resident nearby. The results of this study therefore weight against the idea that exposure to nitrates in the environment leads to the formation in vivo of material amounts of carcinogens. PMID:3015194

Identification of mRNA transcript and screening of amino acids in response to interaction of salinity and nitrate in aquatic fern Azolla caroliniana.

Science.gov (United States)

Tammam, A A; Mostafa, E M

2012-06-01

The mechanisms by which Azolla caroliniana respond to salt stress in absence and presence of nitrate is investigated. Screening of amino acid and differential display is used to compare overall differences in gene expression between salinity-stressed and unstressed Azolla caroliniana by quantitative reverse transcriptase polymerase chain reaction (RT-PC R). Results showed that under saline conditions, aspartic acid, glutamic acid, alanine and leucine were the amino acids found to be abundant in Azolla caroliniana, accounting for 11.26%, 8.66%, 9.43%, and 12.36%, respectively. Following salinity stress, a decrease in free glutamate concomitant with a parallel decrease in free proline was indeed evident. Interaction between nitrate and salinity stress increased proline content significantly. By screening a cDNA library, we have identified protein products by homology with known proteins. The RNA transcripts encoding protein influencing secondary metabolites and vacuolar Na+/H+ antiporter that facilitate the transport system. The databasematched under interaction of nitrate and 50 mM NaCl were associated with wall biosynthesis, disease resistance, metabolite transport and protein regulator, other gene for metabolism of steroids and secondary transport. Results obtained from this research could represent a key step in understanding the molecular mechanism of salt tolerance of Azolla caroliniana in the presence and absence of nitrate.

The intrinsically disordered structural platform of the plant defence hub protein RPM1-interacting protein 4 provides insights into its mode of action in the host-pathogen interface and evolution of the nitrate-induced domain protein family.

Science.gov (United States)

Sun, Xiaolin; Greenwood, David R; Templeton, Matthew D; Libich, David S; McGhie, Tony K; Xue, Bin; Yoon, Minsoo; Cui, Wei; Kirk, Christopher A; Jones, William T; Uversky, Vladimir N; Rikkerink, Erik H A

2014-09-01

Arabidopsis thaliana (At) RPM1-interacting protein 4 (RIN4), targeted by many defence-suppressing bacterial type III effectors and monitored by several resistance proteins, regulates plant immune responses to pathogen-associated molecular patterns and type III effectors. Little is known about the overall protein structure of AtRIN4, especially in its unbound form, and the relevance of structure to its diverse biological functions. AtRIN4 contains two nitrate-induced (NOI) domains and is a member of the NOI family. Using experimental and bioinformatic approaches, we demonstrate that the unbound AtRIN4 is intrinsically disordered under physiological conditions. The intrinsically disordered polypeptide chain of AtRIN4 is interspersed with molecular recognition features (MoRFs) and anchor-identified long-binding regions, potentially allowing it to undergo disorder-to-order transitions upon binding to partner(s). A poly-l-proline II structure, often responsible for protein recognition, is also identified in AtRIN4. By performing bioinformatics analyses on RIN4 homologues from different plant species and the NOI proteins from Arabidopsis, we infer the conservation of intrinsic disorder, MoRFs and long-binding regions of AtRIN4 in other plant species and the NOI family. Intrinsic disorder and MoRFs could provide RIN4 proteins with the binding promiscuity and plasticity required to act as hubs in a pivotal position within plant defence signalling cascades. © 2014 FEBS.

Derivation of an empirical formula for determining water content of mixed uranyl nitrate-thorium nitrate solutions

International Nuclear Information System (INIS)

Min, Duck Kee; Choi, Byung Il; Ro, Seung Gy; Eom, Tae Yoon; Kim, Zong Goo

1986-01-01

Densities of a large number of mixed uranyl nitrate-thorium nitrate solutions were measured with pycnometer. By the least squares analysis of the experimental result, an empirical formula for determining water content of mixed uranyl nitrate-thorium nitrate solutions as functions of uranium concentration, thorium concentration and nitric acid normality is derived; W=1.0-0.3580 C u -0.4538 C Th -0.0307H + where W, C u , C Th , and H + stand for water content(g/cc), uranium concentration (g/cc), thorium concentration(g/cc), and nitric acid normality, respectively. Water contents of the mixed uranyl nitrate-thorium nitrate solutions are calculated by using the empirical formular, and compared with the values calculated by Bouly's equation in which an additional data, solution density, is required. The two results show good agreements within 2.7%. (Author)

CUB-domain-containing protein 1 overexpression in solid cancers promotes cancer cell growth by activating Src family kinases.

Science.gov (United States)

Leroy, C; Shen, Q; Strande, V; Meyer, R; McLaughlin, M E; Lezan, E; Bentires-Alj, M; Voshol, H; Bonenfant, D; Alex Gaither, L

2015-10-29

The transmembrane glycoprotein, CUB (complement C1r/C1s, Uegf, Bmp1) domain-containing protein 1 (CDCP1) is overexpressed in several cancer types and is a predictor of poor prognosis for patients on standard of care therapies. Phosphorylation of CDCP1 tyrosine sites is induced upon loss of cell adhesion and is thought to be linked to metastatic potential of tumor cells. Using a tyrosine-phosphoproteomics screening approach, we characterized the phosphorylation state of CDCP1 across a panel of breast cancer cell lines. We focused on two phospho-tyrosine pTyr peptides of CDCP1, containing Tyr707 and Tyr806, which were identified in all six lines, with the human epidermal growth factor 2-positive HCC1954 cells showing a particularly high phosphorylation level. Pharmacological modulation of tyrosine phosphorylation indicated that, the Src family kinases (SFKs) were found to phosphorylate CDCP1 at Tyr707 and Tyr806 and play a critical role in CDCP1 activity. We demonstrated that CDCP1 overexpression in HEK293 cells increases global phosphotyrosine content, promotes anchorage-independent cell growth and activates several SFK members. Conversely, CDCP1 downregulation in multiple solid cancer cell lines decreased both cell growth and SFK activation. Analysis of primary human tumor samples demonstrated a correlation between CDCP1 expression, SFK and protein kinase C (PKC) activity. Taken together, our results suggest that CDCP1 overexpression could be an interesting therapeutic target in multiple solid cancers and a good biomarker to stratify patients who could benefit from an anti-SFK-targeted therapy. Our data also show that multiple tyrosine phosphorylation sites of CDCP1 are important for the functional regulation of SFKs in several tumor types.

Nitrate Reductases Are Relocalized to the Nucleus by AtSIZ1 and Their Levels Are Negatively Regulated by COP1 and Ammonium

Directory of Open Access Journals (Sweden)

Joo Yong Kim

2018-04-01

Full Text Available Nitrate reductases (NRs catalyze the first step in the reduction of nitrate to ammonium. NR activity is regulated by sumoylation through the E3 ligase activity of AtSIZ1. However, it is not clear how NRs interact with AtSIZ1 in the cell, or how nitrogen sources affect NR levels and their cellular localization. Here, we show that the subcellular localization of NRs is modulated by the E3 SUMO (Small ubiquitin-related modifier ligase AtSIZ1 and that NR protein levels are regulated by nitrogen sources. Transient expression analysis of GFP fusion proteins in onion epidermal cells showed that the NRs NIA1 and NIA2 localize to the cytoplasmic membrane, and that AtSIZ1 localizes to the nucleoplasm, including nuclear bodies, when expressed separately, whereas NRs and AtSIZ1 localize to the nucleus when co-expressed. Nitrate did not affect the subcellular localization of the NRs, but it caused AtSIZ1 to move from the nucleus to the cytoplasm. NRs were not detected in ammonium-treated cells, whereas the localization of AtSIZ1 was not altered by ammonium treatment. NR protein levels increased in response to nitrate but decreased in response to ammonium. In addition, NR protein levels increased in response to a 26S proteasome inhibitor and in cop1-4 and DN-COP1-overexpressing transgenic plants. NR protein degradation occurred later in cop1-4 than in the wild-type, although the NR proteins did not interact with COP1. Therefore, AtSIZ1 controls nuclear localization of NR proteins, and ammonium negatively regulates their levels. The function and stability of NR proteins might be post-translationally modulated by ubiquitination.

Vasodilator Therapy: Nitrates and Nicorandil.

Science.gov (United States)

Tarkin, Jason M; Kaski, Juan Carlos

2016-08-01

Nitrates have been used to treat symptoms of chronic stable angina for over 135 years. These drugs are known to activate nitric oxide (NO)-cyclic guanosine-3',-5'-monophasphate (cGMP) signaling pathways underlying vascular smooth muscle cell relaxation, albeit many questions relating to how nitrates work at the cellular level remain unanswered. Physiologically, the anti-angina effects of nitrates are mostly due to peripheral venous dilatation leading to reduction in preload and therefore left ventricular wall stress, and, to a lesser extent, epicardial coronary artery dilatation and lowering of systemic blood pressure. By counteracting ischemic mechanisms, short-acting nitrates offer rapid relief following an angina attack. Long-acting nitrates, used commonly for angina prophylaxis are recommended second-line, after beta-blockers and calcium channel antagonists. Nicorandil is a balanced vasodilator that acts as both NO donor and arterial K(+) ATP channel opener. Nicorandil might also exhibit cardioprotective properties via mitochondrial ischemic preconditioning. While nitrates and nicorandil are effective pharmacological agents for prevention of angina symptoms, when prescribing these drugs it is important to consider that unwanted and poorly tolerated hemodynamic side-effects such as headache and orthostatic hypotension can often occur owing to systemic vasodilatation. It is also necessary to ensure that a dosing regime is followed that avoids nitrate tolerance, which not only results in loss of drug efficacy, but might also cause endothelial dysfunction and increase long-term cardiovascular risk. Here we provide an update on the pharmacological management of chronic stable angina using nitrates and nicorandil.

Electrochemical determination of nitrate with nitrate reductase-immobilized electrodes under ambient air.

Science.gov (United States)

Quan, De; Shim, Jun Ho; Kim, Jong Dae; Park, Hyung Soo; Cha, Geun Sig; Nam, Hakhyun

2005-07-15

Nitrate monitoring biosensors were prepared by immobilizing nitrate reductase derived from yeast on a glassy carbon electrode (GCE, d = 3 mm) or screen-printed carbon paste electrode (SPCE, d = 3 mm) using a polymer (poly(vinyl alcohol)) entrapment method. The sensor could directly determine the nitrate in an unpurged aqueous solution with the aid of an appropriate oxygen scavenger: the nitrate reduction reaction driven by the enzyme and an electron-transfer mediator, methyl viologen, at -0.85 V (GCE vs Ag/AgCl) or at -0.90 V (SPCE vs Ag/AgCl) exhibited no oxygen interference in a sulfite-added solution. The electroanalytical properties of optimized biosensors were measured: the sensitivity, linear response range, and detection limit of the sensors based on GCE were 7.3 nA/microM, 15-300 microM (r2 = 0.995), and 4.1 microM (S/N = 3), respectively, and those of SPCE were 5.5 nA/microM, 15-250 microM (r2 = 0.996), and 5.5 microM (S/N = 3), respectively. The disposable SPCE-based biosensor with a built-in well- or capillary-type sample cell provided high sensor-to-sensor reproducibility (RSD sensor system was demonstrated by determining nitrate in real samples.

The use and control of nitrate and nitrite for the processing of meat products.

Science.gov (United States)

Honikel, Karl-Otto

2008-01-01

Nitrate and nitrite are used for the purpose of curing meat products. In most countries the use of both substances, usually added as potassium or sodium salts, is limited. Either the ingoing or the residual amounts are regulated by laws. The effective substance is nitrite acting primarily as an inhibitor for some microorganisms. Nitrite added to a batter of meat is partially oxidized to nitrate by sequestering oxygen - thus it acts as an antioxidant - a part of nitrite is bound to myoglobin, forming the heat stable NO-myoglobin, a part is bound to proteins or other substances in meat. Nitrate may be reduced to nitrite in raw meat products by microorganisms. As oxidation and reduction may occur the concentrations of nitrite plus nitrate in a product has to be controlled and measured especially if the residual amounts are regulated. This sum of both compounds is important for the human body. Intake of nitrate with food leads to its absorption over the digestive tract into the blood. In the oral cavity nitrate appears again where it is reduced to nitrite. With the saliva the nitrite is mixed with food, having the same effect as nitrite in a batter (inhibiting growth of some pathogenic microorganisms) and swallowed. In the stomach nitrite can eventually form carcinogenic nitrosamines in the acidic environment.

Nitration Study of Cyclic Ladder Polyphenylsilsesquioxane

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LIANG Jia-xiang

2017-05-01

Full Text Available Several nitration reagents including fuming nitric acid, HNO3-H2SO4, KNO3-H2SO4, HNO3-KNO3, CH3COOH-KNO3, (CH3CO2O-HNO3 were used to nitrate cyclic ladder polyphenylsilsesquioxane (CL-PPSQ in different conditions in order to enhance the compatibility of the CL-PPSQ in polymers, the NO2-PPSQ was obtained. FTIR, element analysis, GPC, TGA and 1H NMR were used to characterize the structures of the nitrated products. The results show that the nitrating abilities of the fuming nitric acid, HNO3-H2SO4 and KNO3-H2SO4 are very strong. Many nitro groups can be linked with phenyl groups in CL-PPSQ, but with low molecular mass, fracture occurs in siloxane segment. However, the Mn of the product NO2-PPSQ sharply drops by 50% compared with that of CL-PPSQ, so the nitration reagents can break the cyclic structure of CL-PPSQ. The nitrating reagents of HNO3-KNO3 and CH3COOH-KNO3 have no nitration effects on CL-PPSQ. At last, NO2-CL-PPSQ was prepared using (CH3CO2O-HNO3 because of the moderate nitration process and ability. The cyclic structure of PPSQ is remained, although the number of —NO2 group is not too much. At the same time, the nitration mechanism using different nitration reagents was analyzed. A certain amount of NO2+, which is a kind of activator owning strong nitration ability, can be found in the fuming nitric acid and H2SO4-HNO3(KNO3 systems. As to the (CH3CO2O-HNO3 system, the main activator is CH3COONO2.

Effect of dietary nitrate level on enteric methane production, hydrogen emission, rumen fermentation, and nutrient digestibility in dairy cows

DEFF Research Database (Denmark)

Olijhoek, Dana; Hellwing, Anne Louise Frydendahl; Brask, Maike

2016-01-01

, microbial protein synthesis, and blood methemoglobin. In a 4 × 4 Latin square design 4 lactating Danish Holstein dairy cows fitted with rumen, duodenal, and ileal cannulas were assigned to 4 calcium ammonium nitrate addition levels: control, low, medium, and high [0, 5.3, 13.6, and 21.1 g of nitrate....../kg of dry matter (DM), respectively]. Diets were made isonitrogenous by replacing urea. Cows were fed ad libitum and, after a 6-d period of gradual introduction of nitrate, adapted to the corn-silage-based total mixed ration (forage:concentrate ratio 50:50 on DM basis) for 16 d before sampling. Digesta...... of nitrate increased hydrogen emissions (L/kg of DMI) quadratically by a factor of 2.5, 3.4, and 3.0 (as L/kg of DMI) for the low, medium, and high diets, respectively, compared with the control. Blood methemoglobin levels and nitrate concentrations in milk and urine increased with increasing nitrate intake...

Nitrate removal from alkaline high nitrate effluent by in situ generation of hydrogen using zinc dust

International Nuclear Information System (INIS)

Rajagopal, S.; Chitra, S.; Paul, Biplob

2016-01-01

Alkaline radioactive low level waste generated in Nuclear Fuel Cycle contains substantial amount of nitrate and needs to be treated to meet Central Pollution Control Board discharge limits of 90 mg/L in marine coastal area. Several denitrification methods like chemical treatment, electrochemical reduction, biological denitrification, ion exchange, reverse osmosis, photochemical reduction etc are followed for removal of nitrate. In effluent treatment plants where chemical treatment is carried out, chemical denitrification can be easily adapted without any additional set up. Reducing agents like zinc and aluminum are suitable for reducing nitrate in alkaline solution. Study on denitrification with zinc dust was taken up in this work. Not much work has been done with zinc dust on reduction of nitrate to nitrogen in alkaline waste with high nitrate content. In the present work, nitrate is reduced by nascent hydrogen generated in situ, caused by reaction between zinc dust and sodium hydroxide

Continuous flow nitration in miniaturized devices

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Amol A. Kulkarni

2014-02-01

Full Text Available This review highlights the state of the art in the field of continuous flow nitration with miniaturized devices. Although nitration has been one of the oldest and most important unit reactions, the advent of miniaturized devices has paved the way for new opportunities to reconsider the conventional approach for exothermic and selectivity sensitive nitration reactions. Four different approaches to flow nitration with microreactors are presented herein and discussed in view of their advantages, limitations and applicability of the information towards scale-up. Selected recent patents that disclose scale-up methodologies for continuous flow nitration are also briefly reviewed.

Nitrate photolysis in salty snow

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Donaldson, D. J.; Morenz, K.; Shi, Q.; Murphy, J. G.

2016-12-01

Nitrate photolysis from snow can have a significant impact on the oxidative capacity of the local atmosphere, but the factors affecting the release of gas phase products are not well understood. Here, we report the first systematic study of the amounts of NO, NO2, and total nitrogen oxides (NOy) emitted from illuminated snow samples as a function of both nitrate and total salt (NaCl and Instant Ocean) concentration. We show that the release of nitrogen oxides to the gas phase is directly related to the expected nitrate concentration in the brine at the surface of the snow crystals, increasing to a plateau value with increasing nitrate, and generally decreasing with increasing NaCl or Instant Ocean (I.O.). In frozen mixed nitrate (25 mM) - salt (0-500 mM) solutions, there is an increase in gas phase NO2 seen at low added salt amounts: NO2 production is enhanced by 35% at low prefreezing [NaCl] and by 70% at similar prefreezing [I.O.]. Raman microscopy of frozen nitrate-salt solutions shows evidence of stronger nitrate exclusion to the air interface in the presence of I.O. than with added NaCl. The enhancement in nitrogen oxides emission in the presence of salts may prove to be important to the atmospheric oxidative capacity in polar regions.

Dissimilatory nitrate reduction to nitrate, nitrous oxide, and ammonium by Pseudomonas putrefaciens.

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Samuelsson, M O

1985-10-01

The influence of redox potential on dissimilatory nitrate reduction to ammonium was investigated on a marine bacterium, Pseudomonas putrefaciens. Nitrate was consumed (3.1 mmol liter-1), and ammonium was produced in cultures with glucose and without sodium thioglycolate. When sodium thioglycolate was added, nitrate was consumed at a lower rate (1.1 mmol liter-1), and no significant amounts of nitrite or ammonium were produced. No growth was detected in glucose media either with or without sodium thioglycolate. When grown on tryptic soy broth, the production of nitrous oxide paralleled growth. In the same medium, but with sodium thioglycolate, nitrous oxide was first produced during growth and then consumed. Acetylene caused the nitrous oxide to accumulate. These results and the mass balance calculations for different nitrogen components indicate that P. putrefaciens has the capacity to dissimilate nitrate to ammonium as well as to dinitrogen gas and nitrous oxide (denitrification). The dissimilatory pathway to ammonium dominates except when sodium thioglycolate is added to the medium.

Enzyme-Triggered Defined Protein Nanoarrays: Efficient Light-Harvesting Systems to Mimic Chloroplasts.

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Zhao, Linlu; Zou, Haoyang; Zhang, Hao; Sun, Hongcheng; Wang, Tingting; Pan, Tiezheng; Li, Xiumei; Bai, Yushi; Qiao, Shanpeng; Luo, Quan; Xu, Jiayun; Hou, Chunxi; Liu, Junqiu

2017-01-24

The elegance and efficiency by which chloroplasts harvest solar energy and conduct energy transfer have been a source of inspiration for chemists to mimic such process. However, precise manipulation to obtain orderly arranged antenna chromophores in constructing artificial chloroplast mimics was a great challenge, especially from the structural similarity and bioaffinity standpoints. Here we reported a design strategy that combined covalent and noncovalent interactions to prepare a protein-based light-harvesting system to mimic chloroplasts. Cricoid stable protein one (SP1) was utilized as a building block model. Under enzyme-triggered covalent protein assembly, mutant SP1 with tyrosine (Tyr) residues at the designated sites can couple together to form nanostructures. Through controlling the Tyr sites on the protein surface, we can manipulate the assembly orientation to respectively generate 1D nanotubes and 2D nanosheets. The excellent stability endowed the self-assembled protein architectures with promising applications. We further integrated quantum dots (QDs) possessing optical and electronic properties with the 2D nanosheets to fabricate chloroplast mimics. By attaching different sized QDs as donor and acceptor chromophores to the negatively charged surface of SP1-based protein nanosheets via electrostatic interactions, we successfully developed an artificial light-harvesting system. The assembled protein nanosheets structurally resembled the natural thylakoids, and the QDs can achieve pronounced FRET phenomenon just like the chlorophylls. Therefore, the coassembled system was meaningful to explore the photosynthetic process in vitro, as it was designed to mimic the natural chloroplast.

Application of infrared spectroscopy for study of chemical bonds in complexes of rare earth nitrates with alkylammonium nitrates

International Nuclear Information System (INIS)

Klimov, V.D.; Chudinov, Eh.G.

1974-01-01

The IR absorption spectra for the tri-n-octylamine, methyl-di-n-octylamine, their nitrates and complexes with the rare element nitrates are obtained. The IR spectra analysis of the complexes has suggested that the degree of covalent character bond of a nitrate with a metal grows with the atomic number of the element. Based on the comparison of the obtained data with those available in literature for various rare-earth complexes a conclusion is made that the bond character of a metal with nitrate groups is influenced by all ligands constituting the inner coordinating sphere. As the donor capacity of a ligand grows the covalent character of the metal-nitrate bond is enhanced. The replacement of the outer-sphere cations (trioctylammonium or methyldioctylammonium) only slightly affects the bond character of a metal with the nitrate group. The distribution coefficients in the rare-earth series are shown to decrease as the electrostatic part in the metal-nitrate declines. The phenomenon is attributed to the competition between nitrate and water for the metal bond as concurrently with the intensification of metal-nitrate covalent bond in the organic phase the strength of metal hydrates in aqueous phase grows much faster. (author)

Nitrate signals determine the sensing of nitrogen through differential expression of genes involved in nitrogen uptake and assimilation in finger millet.

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Gupta, Alok Kumar; Gaur, Vikram Singh; Gupta, Sanjay; Kumar, Anil

2013-06-01

In order to understand the molecular basis of high nitrogen use efficiency of finger millet, five genes (EcHNRT2, EcLNRT1, EcNADH-NR, EcGS, and EcFd-GOGAT) involved in nitrate uptake and assimilation were isolated using conserved primer approaches. Expression profiles of these five genes along with the previously isolated EcDof1 was studied under increased KNO3 concentrations (0.15 to 1,500 μM) for 2 h as well as at 1.5 μM for 24 h in the roots and shoots of 25 days old nitrogen deprived two contrasting finger millet genotypes (GE-3885 and GE-1437) differing in grain protein content (13.76 and 6.15 %, respectively). Time kinetics experiment revealed that, all the five genes except EcHNRT2 in the leaves of GE-3885 were induced within 30 min of nitrate exposure indicating that there might be a greater nitrogen deficit in leaves and therefore quick transportation of nitrate signals to the leaves. Exposing the plants to increasing nitrate concentrations for 2 h showed that in roots of GE-3885, NR was strongly induced while GS was repressed; however, the pattern was found to be reversed in leaves of GE-1437 indicating that in GE-3885, most of the nitrate might be reduced in the roots but assimilated in leaves and vice-versa. Furthermore, compared with the low-protein genotype, expression of HNRT2 was strongly induced in both roots and shoots of high-protein genotype at the least nitrate concentration supplied. This further indicates that GE-3885 is a quick sensor of nitrogen compared with the low-protein genotype. Furthermore, expression of EcDof1 was also found to overlap the expression of NR, GS, and GOGAT indicating that Dof1 probably regulates the expression of these genes under different conditions by sensing the nitrogen fluctuations around the root zone.

Nitrate analogs as attractants for soybean cyst nematode.

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Hosoi, Akito; Katsuyama, Tsutomu; Sasaki, Yasuyuki; Kondo, Tatsuhiko; Yajima, Shunsuke; Ito, Shinsaku

2017-08-01

Soybean cyst nematode (SCN) Heterodera glycines Ichinohe, a plant parasite, is one of the most serious pests of soybean. In this paper, we report that SCN is attracted to nitrate and its analogs. We performed attraction assays to screen for novel attractants for SCN and found that nitrates were attractants for SCN and SCN recognized nitrate gradients. However, attraction of SCN to nitrates was not observed on agar containing nitrate. To further elucidate the attraction mechanism in SCN, we performed attraction assays using nitrate analogs ([Formula: see text], [Formula: see text], [Formula: see text]). SCN was attracted to all nitrate analogs; however, attraction of SCN to nitrate analogs was not observed on agar containing nitrate. In contrast, SCN was attracted to azuki root, irrespective of presence or absence of nitrate in agar media. Our results suggest that the attraction mechanisms differ between plant-derived attractant and nitrate.

Nitrate and ammonium lead to distinct global dynamic phosphorylation patterns when resupplied to nitrogen-starved Arabidopsis seedlings.

Science.gov (United States)

Engelsberger, Wolfgang R; Schulze, Waltraud X

2012-03-01

Nitrogen is an essential macronutrient for plant growth and development. Inorganic nitrogen and its assimilation products control various metabolic, physiological and developmental processes. Although the transcriptional responses induced by nitrogen have been extensively studied in the past, our work here focused on the discovery of candidate proteins for regulatory events that are complementary to transcriptional changes. Most signaling pathways involve modulation of protein abundance and/or activity by protein phosphorylation. Therefore, we analyzed the dynamic changes in protein phosphorylation in membrane and soluble proteins from plants exposed to rapid changes in nutrient availability over a time course of 30 min. Plants were starved of nitrogen and subsequently resupplied with nitrogen in the form of nitrate or ammonium. Proteins with maximum change in their phosphorylation level at up to 5 min after nitrogen resupply (fast responses) included GPI-anchored proteins, receptor kinases and transcription factors, while proteins with maximum change in their phosphorylation level after 10 min of nitrogen resupply (late responses) included proteins involved in protein synthesis and degradation, as well as proteins with functions in central metabolism and hormone metabolism. Resupply of nitrogen in the form of nitrate or ammonium resulted in distinct phosphorylation patterns, mainly of proteins with signaling functions, transcription factors and transporters. © 2011 The Authors. The Plant Journal © 2011 Blackwell Publishing Ltd.

Seasonal nitrate algorithms for nitrate retrieval using OCEANSAT-2 and MODIS-AQUA satellite data.

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Durairaj, Poornima; Sarangi, Ranjit Kumar; Ramalingam, Shanthi; Thirunavukarassu, Thangaradjou; Chauhan, Prakash

2015-04-01

In situ datasets of nitrate, sea surface temperature (SST), and chlorophyll a (chl a) collected during the monthly coastal samplings and organized cruises along the Tamilnadu and Andhra Pradesh coast between 2009 and 2013 were used to develop seasonal nitrate algorithms. The nitrate algorithms have been built up based on the three-dimensional regressions between SST, chl a, and nitrate in situ data using linear, Gaussian, Lorentzian, and paraboloid function fittings. Among these four functions, paraboloid was found to be better with the highest co-efficient of determination (postmonsoon: R2=0.711, n=357; summer: R2=0.635, n=302; premonsoon: R2=0.829, n=249; and monsoon: R2=0.692, n=272) for all seasons. Based on these fittings, seasonal nitrate images were generated using the concurrent satellite data of SST from Moderate Resolution Imaging Spectroradiometer (MODIS) and chlorophyll (chl) from Ocean Color Monitor (OCM-2) and MODIS. The best retrieval of modeled nitrate (R2=0.527, root mean square error (RMSE)=3.72, and mean normalized bias (MNB)=0.821) was observed for the postmonsoon season due to the better retrieval of both SST MODIS (28 February 2012, R2=0.651, RMSE=2.037, and MNB=0.068) and chl OCM-2 (R2=0.534, RMSE=0.317, and MNB=0.27). Present results confirm that the chl OCM-2 and SST MODIS retrieve nitrate well than the MODIS-derived chl and SST largely due to the better retrieval of chl by OCM-2 than MODIS.

RNA-seq analyses reveal insights into the function of respiratory nitrate reductase of the diazotroph Herbaspirillum seropedicae.

Science.gov (United States)

Bonato, Paloma; Batista, Marcelo B; Camilios-Neto, Doumit; Pankievicz, Vânia C S; Tadra-Sfeir, Michelle Z; Monteiro, Rose Adele; Pedrosa, Fabio O; Souza, Emanuel M; Chubatsu, Leda S; Wassem, Roseli; Rigo, Liu Un

2016-09-01

Herbaspirillum seropedicae is a nitrogen-fixing β-proteobacterium that associates with roots of gramineous plants. In silico analyses revealed that H. seropedicae genome has genes encoding a putative respiratory (NAR) and an assimilatory nitrate reductase (NAS). To date, little is known about nitrate metabolism in H. seropedicae, and, as this bacterium cannot respire nitrate, the function of NAR remains unknown. This study aimed to investigate the function of NAR in H. seropedicae and how it metabolizes nitrate in a low aerated-condition. RNA-seq transcriptional profiling in the presence of nitrate allowed us to pinpoint genes important for nitrate metabolism in H. seropedicae, including nitrate transporters and regulatory proteins. Additionally, both RNA-seq data and physiological characterization of a mutant in the catalytic subunit of NAR (narG mutant) showed that NAR is not required for nitrate assimilation but is required for: (i) production of high levels of nitrite, (ii) production of NO and (iii) dissipation of redox power, which in turn lead to an increase in carbon consumption. In addition, wheat plants showed an increase in shoot dry weight only when inoculated with H. seropedicae wild type, but not with the narG mutant, suggesting that NAR is important to H. seropedicae-wheat interaction. © 2016 Society for Applied Microbiology and John Wiley & Sons Ltd.

Temporal pattern changes in duodenal protein tyrosine nitration events in response to Eimeria acervulina infection in chickens.

Science.gov (United States)

Elsasser, Ted H; Miska, Kate; Kahl, Stanislaw; Fetterer, Raymond H; Martínez Ramirez, Alfredo

2018-06-04

Intracellular generation of nitric oxide (NO) and superoxide anion (SOA) can result in the formation of 3'-nitrotyrosine proteins (NTp). Nitrated proteins usually are associated with significant perturbation in protein function, apoptosis, autophagy, and cell death. We undertook the present study to establish the temporal dynamics of NTp generation in cytokeratin-18-positive epithelial cells (ETCs) of broiler chickens in response to infection with Eimeria acervulina. Duodenal tissue was harvested from noninfected (NOI) and infected (INF) broilers on days (d) 1, 3, 6, 7, and 10 postinfection (PI) and fixed, embedded, and sectioned for quantitative image analysis, immunohistochemistry with antibodies specific to NTp and the SOA-generating enzyme xanthine oxidase (XO). The pixel density characteristics for NTp and XO representative of ETCs demonstrated that NTp and XO increased in intestinal villi as early as d1 PI (P ETCs through d6 PI. For XO, increases in cell content increased only through d3. On d6 and d7 PI, high levels of NTp were present in immune infiltrating cells (IIC) where no XO was detected. The increases in ETC NTp occurred in a defined pattern, significant by villus-to-crypt location for day of infection, initiating in the distal villus and progressing down into the crypts. Two NTp patterns were observed for ETCs: a high level associated with ETCs harboring parasites and a low-level increase in ETCs not containing Eimeria but in proximity to such. The data suggest that NTp and XO responses may mediate some of the processes through which ETCs respond to Eimeria to limit the extent of infection by this pathogen.

FIP1 Plays an Important Role in Nitrate Signaling and Regulates CIPK8 and CIPK23 Expression in Arabidopsis

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Chao Wang

2018-05-01

Full Text Available Unraveling the molecular mechanisms of nitrate regulation and deciphering the underlying genetic network is vital for elucidating nitrate uptake and utilization in plants. Such knowledge could lead to the improvement of nitrogen-use efficiency in agriculture. Here, we report that the FIP1 gene (factor interacting with poly(A polymerase 1 plays an important role in nitrate signaling in Arabidopsis thaliana. FIP1 encodes a putative core component of the polyadenylation factor complex. We found that FIP1 interacts with the cleavage and polyadenylation specificity factor 30-L (CPSF30-L, which is also an essential player in nitrate signaling. The induction of nitrate-responsive genes following nitrate treatment was inhibited in the fip1 mutant. The nitrate content was also reduced in fip1 seedlings due to their decreased nitrate uptake activity. Furthermore, the nitrate content was higher in the roots but lower in the roots of fip1, which may result from the downregulation of NRT1.8 and the upregulation of the nitrate assimilation genes. In addition, qPCR analyses revealed that FIP1 negatively regulated the expression of CIPK8 and CIPK23, two protein kinases involved in nitrate signaling. In the fip1 mutant, the increased expression of CIPK23 may affect nitrate uptake, resulting in its lower nitrate content. Genetic and molecular evidence suggests that FIP1 and CPSF30-L function in the same nitrate-signaling pathway, with FIP1 mediating signaling through its interaction with CPSF30-L and its regulation of CIPK8 and CIPK23. Analysis of the 3′-UTR of NRT1.1 showed that the pattern of polyadenylation sites was altered in the fip1 mutant. These findings add a novel component to the nitrate regulation network and enhance our understanding of the underlying mechanisms for nitrate signaling.

Bio nitrate Project: a new technology for water nitrate elimination by means of ionic exchange resins

International Nuclear Information System (INIS)

Arellano Ortiz, J.

2009-01-01

The use of ion exchange resins for nitrate elimination from water generates a waste containing a sodium chloride mixture plus the retained nitrates. this waste must be correctly disposed. In this project, the resin ionic form is modified to be regenerated with other compounds, different from the common salt, which are interesting because of the presence of mineral nutrition. So, with Bio nitrate Project, nitrates are recovered and the regeneration waste is apt to be use as fertilizer, for agricultural uses, or as complementary contribution of nutrients in biological water treatment. (Author) 27 refs.

Impact of ammonium nitrate and sodium nitrate on tadpoles of Alytes obstetricans.

Science.gov (United States)

Garriga, Núria; Montori, A; Llorente, G A

2017-07-01

The presence of pesticides, herbicides and fertilisers negatively affect aquatic communities in general, and particularly amphibians in their larval phase, even though sensitivity to pollutants is highly variable among species. The Llobregat Delta (Barcelona, Spain) has experienced a decline of amphibian populations, possibly related to the reduction in water quality due to the high levels of farming activity, but also to habitat loss and alteration. We studied the effects of increasing ammonium nitrate and sodium nitrate levels on the survival and growth rate of Alytes obstetricans tadpoles under experimental conditions. We exposed larvae to increasing concentrations of nitrate and ammonium for 14 days and then exposed them to water without pollutants for a further 14 days. Only the higher concentrations of ammonium (>33.75 mg/L) caused larval mortality. The growth rate of larvae was reduced at ≥22.5 mg/L NH 4 + , although individuals recovered and even increased their growth rate once exposure to the pollutant ended. The effect of nitrate on growth rate was detected at ≥80 mg/L concentrations, and the growth rate reduction in tadpoles was even observed during the post-exposure phase. The concentrations of ammonium with adverse effects on larvae are within the range levels found in the study area, while the nitrate concentrations with some adverse effect are close to the upper range limit of current concentrations in the study area. Therefore, only the presence of ammonium in the study area is likely to be considered of concern for the population of this species, even though the presence of nitrate could cause some sublethal effects. These negative effects could have an impact on population dynamics, which in this species is highly sensitive to larval mortality due to its small clutch size and prolonged larval period compared to other anuran amphibians.

EXTRACTION OF URANYL NITRATE FROM AQUEOUS SOLUTIONS

Science.gov (United States)

Furman, N.H.; Mundy, R.J.

1957-12-10

An improvement in the process is described for extracting aqueous uranyl nitrate solutions with an organic solvent such as ether. It has been found that the organic phase will extract a larger quantity of uranyl nitrate if the aqueous phase contains in addition to the uranyl nitrate, a quantity of some other soluble nitrate to act as a salting out agent. Mentioned as suitable are the nitrates of lithium, calcium, zinc, bivalent copper, and trivalent iron.

The impact of nitration on the structure and immunogenicity of the major birch pollen allergen Bet v 1.0101.

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Chloé Ackaert

Full Text Available Allergy prevalence has increased in industrialized countries. One contributing factor could be pollution, which can cause nitration of allergens exogenously (in the air or endogenously (in inflamed lung tissue. We investigated the impact of nitration on both the structural and immunological behavior of the major birch pollen allergen Bet v 1.0101 to determine whether nitration might be a factor in the increased incidence of allergy. Bet v 1.0101 was nitrated with tetranitromethane. Immune effects were assessed by measuring the proliferation of specific T-cell lines (TCLs upon stimulation with different concentrations of nitrated and unmodified allergen, and by measurement of cytokine release of monocyte-derived dendritic cells (moDCs and primary DCs (primDCs stimulated with nitrated versus unmodified allergen. HPLC-MS, crystallography, gel electrophoresis, amino acid analysis, size exclusion chromatography and molecular dynamics simulation were performed to characterize structural changes after nitration of the allergen. The proliferation of specific TCLs was higher upon stimulation with the nitrated allergen in comparison to the unmodified allergen. An important structural consequence of nitration was oligomerization. Moreover, analysis of the crystal structure of nitrated Bet v 1.0101 showed that amino acid residue Y83, located in the hydrophobic cavity, was nitrated to 100%. Both moDCs and primDCs showed decreased production of TH1-priming cytokines, thus favoring a TH2 response. These results implicate that nitration of Bet v 1.0101 might be a contributing factor to the observed increase in birch pollen allergy, and emphasize the importance of protein modifications in understanding the molecular basis of allergenicity.

Drug-protein interactions assessed by fluorescence measurements in the real complexes and in model dyads

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Vayá, Ignacio; Pérez-Ruiz, Raúl; Lhiaubet-Vallet, Virginie; Jiménez, M. Consuelo; Miranda, Miguel A.

2010-02-01

In the present work, a systematic fluorescence study on supramolecular systems using two serum albumins (HSA or BSA) as hosts and the nonsteroidal antiinflammatory drugs carprofen (CPF) or naproxen (NPX) as guests has been undertaken. In parallel, model dyads containing Tyr or Trp covalently linked to CPF or NPX have also been investigated. In HSA/(S)-CPF and BSA/(S)-CPF ( λexc = 266 nm), at 1:1 M ratio, an important degree (more than 40%) of singlet-singlet energy transfer (SSET) was observed to take place. The distance ( r) calculated for energy transfer from the SAs to (S)-CPF through a FRET mechanism was found to be ca. 21 Å. In the case of HSA/(S)-NPX and BSA/(S)-NPX, energy transfer occurred to a lower extent (ca. 7%), and r was determined as ca. 24 Å. In order to investigate the possible excited state interactions between bound ligands and the relevant amino acids present in the protein binding sites, four pairs of model dyads were designed and synthesised, namely ( S, S)-TyrCPF, ( S, R)-TyrCPF, ( S, S)-TrpCPF, ( S, R)-TrpCPF, ( S, S)-TyrNPX, ( S, R)-TyrNPX, ( S, S)-TrpNPX and ( S, R)-TrpNPX. A complete SSET was observed from Tyr or Trp to CPF, since no contribution from the amino acids was present in the emission of the dyads. Likewise, a very efficient Tyr or Trp to NPX energy transfer was observed. Remarkably, in ( S, S)-TrpNPX and ( S, R)-TrpNPX a configuration-dependent reduction in the emission intensity was observed, revealing a strong and stereoselective intramolecular quenching. This effect can be attributed to exciplex formation and is dynamic in nature, as the fluorescence lifetimes were much shorter in ( S, R)- and ( S, S)-TrpNPX (1.5 and 3.1 ns, respectively) than in (S)-NPX (11 ns).

Diagnostic utility of 99mTc-EDDA-tricine-HYNIC-Tyr3-octreotate SPECT for differentiation of active from inactive pulmonary tuberculosis.

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Ahmadihosseini, Hossein; Abedi, Javad; Ghodsi Rad, Mohammad A; Zakavi, Seyed R; Knoll, Peter; Mirzaei, Siroos; Sadeghi, Ramin

2014-12-01

The current study was performed to evaluate the impact of Tc-EDDA-tricine-HYNIC-Tyr-octreotate in the differentiation of active from inactive pulmonary tuberculosis lesions. Ten consecutive patients (six male and four female, age range 24-83 years) with proven pulmonary tuberculosis (with a positive smear or culture) were enrolled in the study. At 120 min after injection of 740 MBq of Tc-EDDA-tricine-HYNIC-Tyr-octreotate, planar and single-photon emission computed tomography (SPECT) images of the thorax were taken. A semiquantitative evaluation of lesion and nonlesion areas was performed. The scan was repeated following the same protocol after standard treatment for tuberculosis after a negative sputum culture. Semiquantitative evaluation of the lesions showed a statistically significant higher uptake before treatment in both planar and SPECT images (P=0.005 and 0.007, respectively). Lesion-to-nonlesion ratios were also higher in the pretreatment sets on both planar and SPECT images (1.4±0.2 vs. 1.19±0.15, P=0.001, for planar images and 2.32±0.55 vs. 1.32±0.32, P=0.0001, for SPECT images). Tc-EDDA-tricine-HYNIC-Tyr-octreotate scintigraphy may help to differentiate between active and inactive pulmonary tuberculosis. SPECT imaging and semiquantitative evaluation are indispensable for increasing the diagnostic yield of this method. Larger studies are needed to corroborate our results.

[Effect of electroacupuncture on phosphorylation of NR2B at Tyr 1742 site in the spinal dorsal horn of CFA rats].

Science.gov (United States)

Liang, Yi; Fang, Jian-Qiao; Fang, Jun-Fan; Du, Jun-Ying; Qiu, Yu-Jie; Liu, Jin

2013-10-01

To observe the effect of electroacupuncture (EA) on phosphorylation of spinal NR2B at Tyr 1742 site in complete Freund's adjuvant (CFA) induced inflammatory pain rats. METHods Forty male Sprague Dawley rats were randomly divided into normal group (N group, n = 10), the model group (CFA group, n = 15), and the EA group (n = 15). The inflammatory pain model was established by subcutaneous injecting CFA (0.1 mL per rat) into the right hind paw. Paw withdrawal thresholds (PWTs) were measured before CFA injection (as the base), as well as at 24 h, 25 h, 3rd day, and 7th day after CFA injection. Phosphorylation of NR2B at Tyr 1742 site in the ispilateral spinal dorsal horn at the 3rd day post-injection were detected using immunohistochemical assay. PWTs in the CFA group were significantly lower than those of the N group at every detective time point post-injection (P CFA group at 25 h and 3rd day post-injection (P CFA group was up-regulated. Compared with the CFA group, the ratio of p-NR2B positive cells in the ispilateral spinal dorsal horn of rats showed a decreasing tendency in the EA group. EA might effectively inhibit CFA-induced inflammatory pain possibly associated with down-regulating phosphorylation of NR2B at Tyr 1742 site in the ispilateral spinal dorsal horn.

Interaction of Yna1 and Yna2 Is Required for Nuclear Accumulation and Transcriptional Activation of the Nitrate Assimilation Pathway in the Yeast Hansenula polymorpha.

Science.gov (United States)

Silvestrini, Lucia; Rossi, Beatrice; Gallmetzer, Andreas; Mathieu, Martine; Scazzocchio, Claudio; Berardi, Enrico; Strauss, Joseph

2015-01-01

A few yeasts, including Hansenula polymorpha are able to assimilate nitrate and use it as nitrogen source. The genes necessary for nitrate assimilation are organised in this organism as a cluster comprising those encoding nitrate reductase (YNR1), nitrite reductase (YNI1), a high affinity transporter (YNT1), as well as the two pathway specific Zn(II)2Cys2 transcriptional activators (YNA1, YNA2). Yna1p and Yna2p mediate induction of the system and here we show that their functions are interdependent. Yna1p activates YNA2 as well as its own (YNA1) transcription thus forming a nitrate-dependent autoactivation loop. Using a split-YFP approach we demonstrate here that Yna1p and Yna2p form a heterodimer independently of the inducer and despite both Yna1p and Yna2p can occupy the target promoter as mono- or homodimer individually, these proteins are transcriptionally incompetent. Subsequently, the transcription factors target genes containing a conserved DNA motif (termed nitrate-UAS) determined in this work by in vitro and in vivo protein-DNA interaction studies. These events lead to a rearrangement of the chromatin landscape on the target promoters and are associated with the onset of transcription of these target genes. In contrast to other fungi and plants, in which nuclear accumulation of the pathway-specific transcription factors only occur in the presence of nitrate, Yna1p and Yna2p are constitutively nuclear in H. polymorpha. Yna2p is needed for this nuclear accumulation and Yna1p is incapable of strictly positioning in the nucleus without Yna2p. In vivo DNA footprinting and ChIP analyses revealed that the permanently nuclear Yna1p/Yna2p heterodimer only binds to the nitrate-UAS when the inducer is present. The nitrate-dependent up-regulation of one partner protein in the heterodimeric complex is functionally similar to the nitrate-dependent activation of nuclear accumulation in other systems.

Activation of the protein tyrosine phosphatase SHP2 via the interleukin-6 signal transducing receptor protein gp130 requires tyrosine kinase Jak1 and limits acute-phase protein expression.

Science.gov (United States)

Schaper, F; Gendo, C; Eck, M; Schmitz, J; Grimm, C; Anhuf, D; Kerr, I M; Heinrich, P C

1998-11-01

Stimulation of the interleukin-6 (IL-6) signalling pathway occurs via the IL-6 receptor-glycoprotein 130 (IL-6R-gp130) receptor complex and results in the regulation of acute-phase protein genes in liver cells. Ligand binding to the receptor complex leads to tyrosine phosphorylation and activation of Janus kinases (Jak), phosphorylation of the signal transducing subunit gp130, followed by recruitment and phosphorylation of the signal transducer and activator of transcription factors STAT3 and STAT1 and the src homology domain (SH2)-containing protein tyrosine phosphatase (SHP2). The tyrosine phosphorylated STAT factors dissociate from the receptor, dimerize and translocate to the nucleus where they bind to enhancer sequences of IL-6 target genes. Phosphorylated SHP2 is able to bind growth factor receptor bound protein (grb2) and thus might link the Jak/STAT pathway to the ras/raf/mitogen-activated protein kinase pathway. Here we present data on the dose-dependence, kinetics and kinase requirements for SHP2 phosphorylation after the activation of the signal transducer, gp130, of the IL-6-type family receptor complex. When human fibrosarcoma cell lines deficient in Jak1, Jak2 or tyrosine kinase 2 (Tyk2) were stimulated with IL-6-soluble IL-6R complexes it was found that only in Jak1-, but not in Jak 2- or Tyk2-deficient cells, SHP2 activation was greatly impaired. It is concluded that Jak1 is required for the tyrosine phosphorylation of SHP2. This phosphorylation depends on Tyr-759 in the cytoplasmatic domain of gp130, since a Tyr-759-->Phe exchange abrogates SHP2 activation and in turn leads to elevated and prolonged STAT3 and STAT1 activation as well as enhanced acute-phase protein gene induction. Therefore, SHP2 plays an important role in acute-phase gene regulation.

Functional properties and differential mode of regulation of the nitrate transporter from a plant symbiotic ascomycete

Science.gov (United States)

Montanini, Barbara; Viscomi, Arturo R.; Bolchi, Angelo; Martin, Yusé; Siverio, José M.; Balestrini, Raffaella; Bonfante, Paola; Ottonello, Simone

2005-01-01

Nitrogen assimilation by plant symbiotic fungi plays a central role in the mutualistic interaction established by these organisms, as well as in nitrogen flux in a variety of soils. In the present study, we report on the functional properties, structural organization and distinctive mode of regulation of TbNrt2 (Tuber borchii NRT2 family transporter), the nitrate transporter of the mycorrhizal ascomycete T. borchii. As revealed by experiments conducted in a nitrate-uptake-defective mutant of the yeast Hansenula polymorpha, TbNrt2 is a high-affinity transporter (Km=4.7 μM nitrate) that is bispecific for nitrate and nitrite. It is expressed in free-living mycelia and in mycorrhizae, where it preferentially accumulates in the plasma membrane of root-contacting hyphae. The TbNrt2 mRNA, which is transcribed from a single-copy gene clustered with the nitrate reductase gene in the T. borchii genome, was specifically up-regulated following transfer of mycelia to nitrate- (or nitrite)-containing medium. However, at variance with the strict nitrate-dependent induction commonly observed in other organisms, TbNrt2 was also up-regulated (at both the mRNA and the protein level) following transfer to a nitrogen-free medium. This unusual mode of regulation differs from that of the adjacent nitrate reductase gene, which was expressed at basal levels under nitrogen deprivation conditions and required nitrate for induction. The functional and expression properties, described in the present study, delineate TbNrt2 as a versatile transporter that may be especially suited to cope with the fluctuating (and often low) mineral nitrogen concentrations found in most natural, especially forest, soils. PMID:16201972

Research studies on in vitro and ex vivo yield of the miconazole nitrate from oral biomucoadhesive tablets.

Science.gov (United States)

Birsan, Magdalena; Cojocaru, Ileana; Scutariu, Mihaela Monica; Popovici, Iuliana

2014-01-01

Among the various routes of drug administration, the oral mucosa is perhaps the most often preferred by patients and medical staff. However, oral administration of drugs has disadvantages, which may limit or prevent oral administration of some drugs, especially peptides and proteins, little when they are inserted in special administration systems for the colon. The disaggregation of some oral biomucoadhesive tablets and the in vitro yield of the miconazole nitrate was evaluated and in parallel with this, the evaluation of the in vivo yield of the antifungal from the pharmaceutical form. Thus, for a clear determination of the oral mucobioadhesive tablets' disintegration with miconazole nitrate, it was necessary to implement a method to simulate the conditions of the oral cavity at a flow of solution (artificial saliva) similar to that of the human one. miconazole nitrate. The determination of disintegration time according to method A (FRX); the disaggregation of oral biomucoadhesive tablets with miconazole nitrate by means of simulation methods of in vitro conditions; the quantitative determination of the miconazole nitrate by means of HPLC method, after the in vitro dissolution test; the study of miconazole nitrate's yield in dynamic condition from biomucoadhesive tablets in the presence of artificial saliva (AFNOR). The yield profile of the miconazole nitrate in the disintegration solutions by means of classical method from FR X, by HPLC dosage was researched. The release of miconazole nitrate from the oral mucobioadhesive tablets was determined, that varies in time, depending on the type and relation of matrix forming polymers; a low yield speed of the miconazole nitrate from the tablets was determined; the yield profile of miconazole nitrate in disintegration solutions by means of the new suggested method was researched. The release of miconazole nitrate from the formulated biomucoadhesive tablets is of swelling and erosion.

Sustainable nitrate-contaminated water treatment using multi cycle ion-exchange/bioregeneration of nitrate selective resin.

Science.gov (United States)

Ebrahimi, Shelir; Roberts, Deborah J

2013-11-15

The sustainability of ion-exchange treatment processes using high capacity single use resins to remove nitrate from contaminated drinking water can be achieved by regenerating the exhausted resin and reusing it multiple times. In this study, multi cycle loading and bioregeneration of tributylamine strong base anion (SBA) exchange resin was studied. After each cycle of exhaustion, biological regeneration of the resin was performed using a salt-tolerant, nitrate-perchlorate-reducing culture for 48 h. The resin was enclosed in a membrane to avoid direct contact of the resin with the culture. The results show that the culture was capable of regenerating the resin and allowing the resin to be used in multiple cycles. The concentrations of nitrate in the samples reached a peak in first 0.5-1h after placing the resin in medium because of desorption of nitrate from resin with desorption rate of 0.099 ± 0.003 hr(-1). After this time, since microorganisms began to degrade the nitrate in the aqueous phase, the nitrate concentration was generally non-detectable after 10h. The average of calculated specific degradation rate of nitrate was -0.015 mg NO3(-)/mg VSS h. Applying 6 cycles of resin exhaustion/regeneration shows resin can be used for 4 cycles without a loss of capacity, after 6 cycles only 6% of the capacity was lost. This is the first published research to examine the direct regeneration of a resin enclosed in a membrane, to allow reuse without any disinfection or cleaning procedures. Copyright © 2013 Elsevier B.V. All rights reserved.

Silver and Nitrate Oppositely Modulate Antimony Susceptibility through Aquaglyceroporin 1 in Leishmania (Viannia) Species.

Science.gov (United States)

Andrade, Juvana M; Baba, Elio H; Machado-de-Avila, Ricardo A; Chavez-Olortegui, Carlos; Demicheli, Cynthia P; Frézard, Frédéric; Monte-Neto, Rubens L; Murta, Silvane M F

2016-08-01

Antimony (Sb) resistance in leishmaniasis chemotherapy has become one of the major challenges to the control of this spreading worldwide public health problem. Since the plasma membrane pore-forming protein aquaglyceroporin 1 (AQP1) is the major route of Sb uptake in Leishmania, functional studies are relevant to characterize drug transport pathways in the parasite. We generated AQP1-overexpressing Leishmania guyanensis and L. braziliensis mutants and investigated their susceptibility to the trivalent form of Sb (Sb(III)) in the presence of silver and nitrate salts. Both AQP1-overexpressing lines presented 3- to 4-fold increased AQP1 expression levels compared with those of their untransfected counterparts, leading to an increased Sb(III) susceptibility of about 2-fold. Competition assays using silver nitrate, silver sulfadiazine, or silver acetate prior to Sb(III) exposure increased parasite growth, especially in AQP1-overexpressing mutants. Surprisingly, Sb(III)-sodium nitrate or Sb(III)-potassium nitrate combinations showed significantly enhanced antileishmanial activities compared to those of Sb(III) alone, especially against AQP1-overexpressing mutants, suggesting a putative nitrate-dependent modulation of AQP1 activity. The intracellular level of antimony quantified by graphite furnace atomic absorption spectrometry showed that the concomitant exposure to Sb(III) and nitrate favors antimony accumulation in the parasite, increasing the toxicity of the drug and culminating with parasite death. This is the first report showing evidence of AQP1-mediated Sb(III) susceptibility modulation by silver in Leishmania and suggests the potential antileishmanial activity of the combination of nitrate salts and Sb(III). Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Vascular effects of dietary nitrate (as found in green leafy vegetables and beetroot) via the nitrate-nitrite-nitric oxide pathway.

Science.gov (United States)

Lidder, Satnam; Webb, Andrew J

2013-03-01

The discovery that dietary (inorganic) nitrate has important vascular effects came from the relatively recent realization of the 'nitrate-nitrite-nitric oxide (NO) pathway'. Dietary nitrate has been demonstrated to have a range of beneficial vascular effects, including reducing blood pressure, inhibiting platelet aggregation, preserving or improving endothelial dysfunction, enhancing exercise performance in healthy individuals and patients with peripheral arterial disease. Pre-clinical studies with nitrate or nitrite also show the potential to protect against ischaemia-reperfusion injury and reduce arterial stiffness, inflammation and intimal thickness. However, there is a need for good evidence for hard endpoints beyond epidemiological studies. Whilst these suggest reduction in cardiovascular risk with diets high in nitrate-rich vegetables (such as a Mediterranean diet), others have suggested possible small positive and negative associations with dietary nitrate and cancer, but these remain unproven. Interactions with other nutrients, such as vitamin C, polyphenols and fatty acids may enhance or inhibit these effects. In order to provide simple guidance on nitrate intake from different vegetables, we have developed the Nitrate 'Veg-Table' with 'Nitrate Units' [each unit being 1 mmol of nitrate (62 mg)] to achieve a nitrate intake that is likely to be sufficient to derive benefit, but also to minimize the risk of potential side effects from excessive ingestion, given the current available evidence. The lack of data concerning the long term effects of dietary nitrate is a limitation, and this will need to be addressed in future trials. © 2012 The Authors. British Journal of Clinical Pharmacology © 2012 The British Pharmacological Society.

Tyrosol and its metabolites as antioxidative and anti-inflammatory molecules in human endothelial cells.

Science.gov (United States)

Muriana, Francisco J G; Montserrat-de la Paz, Sergio; Lucas, Ricardo; Bermudez, Beatriz; Jaramillo, Sara; Morales, Juan C; Abia, Rocio; Lopez, Sergio

2017-08-01

Tyrosol (Tyr) is a phenolic compound found in virgin olive oil. After ingestion, Tyr undergoes extensive first pass intestinal/hepatic metabolism. However, knowledge about the biological effects of Tyr metabolites is scarce. We chemically synthesized Tyr glucuronate (Tyr-GLU) and sulphate (Tyr-SUL) metabolites and explored their properties against oxidative stress and inflammation in TNF-α-treated human umbilical vein endothelial cells (hECs). Tyr and Tyr-SUL prevented the rise of reactive oxygen species, the depletion of glutathione, and the down-regulation of glutathione peroxidase 1, glutamate-cysteine ligase catalytic subunit, and heme oxygenase-1 genes. Tyr-SUL and to a lower extent Tyr and Tyr-GLU prevented the phosphorylation of NF-κB signaling proteins. Tyr-GLU and Tyr-SUL also prevented the over-expression of adhesion molecules at gene, protein, and secretory levels, and the adhesion (Tyr-SUL > Tyr-GLU) of human monocytes to hECs. In vivo, Tyr, and most notably Tyr-SUL in a dose-dependent manner, ameliorated plantar and ear edemas in mice models of acute and chronic inflammation. This study demonstrates the antioxidant and/or anti-inflammatory properties of Tyr metabolites, with Tyr-SUL being the most effective.

The nitrate time bomb : a numerical way to investigate nitrate storage and lag time in the unsaturated zone

OpenAIRE

Wang, L.; Butcher, A.S.; Stuart, M.E.; Gooddy, D.C.; Bloomfield, J.P.

2013-01-01

Nitrate pollution in groundwater, which is mainly from agricultural activities, remains an international problem. It threatens the environment, economics and human health. There is a rising trend in nitrate concentrations in many UK groundwater bodies. Research has shown it can take decades for leached nitrate from the soil to discharge into groundwater and surface water due to the ‘store’ of nitrate and its potentially long travel time in the unsaturated and satura...

A comparison of organic and inorganic nitrates/nitrites.

Science.gov (United States)

Omar, Sami A; Artime, Esther; Webb, Andrew J

2012-05-15

Although both organic and inorganic nitrates/nitrites mediate their principal effects via nitric oxide, there are many important differences. Inorganic nitrate and nitrite have simple ionic structures and are produced endogenously and are present in the diet, whereas their organic counterparts are far more complex, and, with the exception of ethyl nitrite, are all medicinally synthesised products. These chemical differences underlie the differences in pharmacokinetic properties allowing for different modalities of administration, particularly of organic nitrates, due to the differences in their bioavailability and metabolic profiles. Whilst the enterosalivary circulation is a key pathway for orally ingested inorganic nitrate, preventing an abrupt effect or toxic levels of nitrite and prolonging the effects, this is not used by organic nitrates. The pharmacodynamic differences are even greater; while organic nitrates have potent acute effects causing vasodilation, inorganic nitrite's effects are more subtle and dependent on certain conditions. However, in chronic use, organic nitrates are considerably limited by the development of tolerance and endothelial dysfunction, whereas inorganic nitrate/nitrite may compensate for diminished endothelial function, and tolerance has not been reported. Also, while inorganic nitrate/nitrite has important cytoprotective effects against ischaemia-reperfusion injury, continuous use of organic nitrates may increase injury. While there are concerns that inorganic nitrate/nitrite may induce carcinogenesis, direct evidence of this in humans is lacking. While organic nitrates may continue to dominate the therapeutic arena, this may well change with the increasing recognition of their limitations, and ongoing discovery of beneficial effects and specific advantages of inorganic nitrate/nitrite. Copyright © 2012 Elsevier Inc. All rights reserved.

Mitigation of a nitrate reducing Pseudomonas aeruginosa biofilm and anaerobic biocorrosion using ciprofloxacin enhanced by D-tyrosine.

Science.gov (United States)

Jia, Ru; Yang, Dongqing; Xu, Dake; Gu, Tingyue

2017-07-31

Pseudomonas aeruginosa (PA) is a ubiquitous microbe. It can form recalcitrant biofilms in clinical and industrial settings. PA biofilms cause infections in patients. They also cause biocorrosion of medical implants. In this work, D-tyrosine (D-tyr) was investigated as an antimicrobial enhancer for ciprofloxacin (CIP) against a wild-type PA biofilm (strain PAO1) on C1018 carbon steel in a strictly anaerobic condition. Seven-day biofilm prevention test results demonstrated that 2 ppm (w/w) D-tyr enhanced 30 ppm CIP by achieving extra 2-log sessile cell reduction compared with the 30 ppm CIP alone treatment. The cocktail of 30 ppm CIP + 2 ppm D-tyr achieved similar efficacy as the 80 ppm CIP alone treatment in the biofilm prevention test. Results also indicated that the enhanced antimicrobial treatment reduced weight loss and pitting corrosion. In the 3-hour biofilm removal test, the cocktail of 80 ppm CIP + 5 ppm D-tyr achieved extra 1.5-log reduction in sessile cell count compared with the 80 ppm CIP alone treatment. The cocktail of 80 ppm CIP + 5 ppm D-tyr achieved better efficacy than the 150 ppm CIP alone treatment in the biofilm removal test.

21 CFR 172.160 - Potassium nitrate.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Potassium nitrate. 172.160 Section 172.160 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN... Preservatives § 172.160 Potassium nitrate. The food additive potassium nitrate may be safely used as a curing...

Genetic basis for nitrate resistance in Desulfovibrio strains

Directory of Open Access Journals (Sweden)

Hannah eKorte

2014-04-01

Full Text Available Nitrate is an inhibitor of sulfate-reducing bacteria (SRB. In petroleum production sites, amendments of nitrate and nitrite are used to prevent SRB production of sulfide that causes souring of oil wells. A better understanding of nitrate stress responses in the model SRB, Desulfovibrio vulgaris Hildenborough and Desulfovibrio alaskensis G20, will strengthen predictions of environmental outcomes. Nitrate inhibition of SRB has historically been considered to result from the generation of small amounts of nitrite, to which SRB are quite sensitive. Here we explored the possibility that nitrate might inhibit SRB by a mechanism other than through nitrite inhibition. We found that nitrate-stressed D. vulgaris cultures grown in lactate-sulfate conditions eventually grew in the presence of high concentrations of nitrate, and their resistance continued through several subcultures. Nitrate consumption was not detected over the course of the experiment, suggesting adaptation to nitrate. With high-throughput genetic approaches employing TnLE-seq for D. vulgaris and a pooled mutant library of D. alaskensis, we determined the fitness of many transposon mutants of both organisms in nitrate stress conditions. We found that several mutants, including homologs present in both strains, had a greatly increased ability to grow in the presence of nitrate but not nitrite. The mutated genes conferring nitrate resistance included the gene encoding the putative Rex transcriptional regulator (DVU0916/Dde_2702, as well as a cluster of genes (DVU0251-DVU0245/Dde_0597-Dde_0605 that is poorly annotated. Follow-up studies with individual D. vulgaris transposon and deletion mutants confirmed high-throughput results. We conclude that, in D. vulgaris and D. alaskensis, nitrate resistance in wild-type cultures is likely conferred by spontaneous mutations. Furthermore, the mechanisms that confer nitrate resistance may be different from those that confer nitrite resistance.

Association between receptor protein-tyrosine phosphatase RPTPalpha and the Grb2 adaptor. Dual Src homology (SH) 2/SH3 domain requirement and functional consequences

DEFF Research Database (Denmark)

Su, J; Yang, L T; Sap, J

1996-01-01

domain in Grb2 (, ). We show here that association of Grb2 with RPTPalpha also involves a critical function for the C-terminal SH3 domain of Grb2. Furthermore, Grb2 SH3 binding peptides interfere with RPTPalpha-Grb2 association in vitro, and the RPTPalpha protein can dissociate the Grb2-Sos complex...... in vivo. These observations constitute a novel mode of Grb2 association and suggest a model in which association with a tyrosine-phosphorylated protein restricts the repertoire of SH3 binding proteins with which Grb2 can simultaneously interact. The function of the Tyr798 tyrosine phosphorylation/Grb2...... binding site in RPTPalpha was studied further by expression of wild type or mutant RPTPalpha proteins in PC12 cells. In these cells, wild type RPTPalpha interferes with acidic fibroblast growth factor-induced neurite outgrowth; this effect requires both the catalytic activity and the Grb2 binding Tyr798...

Relationship between nitrate reductase and nitrate uptake in phytoplankton in the Peru upwelling region

International Nuclear Information System (INIS)

Blasco, D.; MacIsaac, J.J.; Packard, T.T.; Dugdale, R.C.

1984-01-01

Nitrate reductase (NR) activity and 15 NO 3 - uptake in phytoplankton were compared under different environmental conditions on two cruises in the upwelling region off Peru. The NR activity and NO 3 - uptake rates responded differently to light and nutrients and the differences led to variations in the uptake: reductase ratio. Analysis of these variations suggests that the re-equilibration time of the two processes in response to environmental perturbation is an important source of variability. The nitrate uptake system responds faster than the nitrate reductase system. Considering these differences in response time the basic differences in the two processes, and the differences in their measurement, the authors conclude that the Nr activity measures the current nitrate-reducing potential, which reflects NO 3 - assimilation before the sampling time, while 15 NO 3 - uptake measures NO 3 - assimilation in the 6-h period following sampling

Nitrate Removal from Ground Water: A Review

Directory of Open Access Journals (Sweden)

Archna

2012-01-01

Full Text Available Nitrate contamination of ground water resources has increased in Asia, Europe, United States, and various other parts of the world. This trend has raised concern as nitrates cause methemoglobinemia and cancer. Several treatment processes can remove nitrates from water with varying degrees of efficiency, cost, and ease of operation. Available technical data, experience, and economics indicate that biological denitrification is more acceptable for nitrate removal than reverse osmosis and ion exchange. This paper reviews the developments in the field of nitrate removal processes which can be effectively used for denitrifying ground water as well as industrial water.

The Nitrate/(Per)Chlorate Relationship on Mars

Science.gov (United States)

Stern, Jennifer C.; Sutter, Brad; Jackson, W. Andrew; Navarro-Gonzalez, Rafael; McKay, Christopher P.; Ming, Douglas W.; Archer, P. Douglas; Mahaffy, Paul R.

2017-01-01

Nitrate was recently detected in Gale Crater sediments on Mars at abundances up to approximately 600 mg/kg, confirming predictions of its presence at abundances consistent with models based on impact-generated nitrate and other sources of fixed nitrogen. Terrestrial Mars analogs, Mars meteorites, and other solar system materials help establish a context for interpreting in situ nitrate measurements on Mars, particularly in relation to other cooccuring salts. We compare the relative abundance of nitrates to oxychlorine (chlorate and/or perchlorate, hereafter (per)chlorate) salts on Mars and Earth. The nitrate/(per)chlorate ratio on Mars is greater than 1, significantly lower than on Earth (nitrate/(per)chlorate greater than 10(exp.3)), suggesting not only the absence of biological activity but also different (per)chlorate formation mechanisms on Mars than on Earth.

Dietary nitrates, nitrites, and cardiovascular disease.

Science.gov (United States)

Hord, Norman G

2011-12-01

Dietary nitrate (NO(3)), nitrite (NO(2)), and arginine can serve as sources for production of NO(x) (a diverse group of metabolites including nitric oxide, nitrosothiols, and nitroalkenes) via ultraviolet light exposure to skin, mammalian nitrate/nitrite reductases in tissues, and nitric oxide synthase enzymes, respectively. NO(x) are responsible for the hypotensive, antiplatelet, and cytoprotective effects of dietary nitrates and nitrites. Current regulatory limits on nitrate intakes, based on concerns regarding potential risk of carcinogenicity and methemoglobinemia, are exceeded by normal daily intakes of single foods, such as soya milk and spinach, as well as by some recommended dietary patterns such as the Dietary Approaches to Stop Hypertension diet. This review includes a call for regulatory bodies to consider all available data on the beneficial physiologic roles of nitrate and nitrite in order to derive rational bases for dietary recommendations.

Preparation of fluorescence quenched libraries containing interchain disulphide bonds for studies of protein disulphide isomerases

DEFF Research Database (Denmark)

Spetzler, J C; Westphal, V; Winther, Jakob R.

1998-01-01

Protein disulphide isomerase is an enzyme that catalyses disulphide redox reactions in proteins. In this paper, fluorogenic and interchain disulphide bond containing peptide libraries and suitable substrates, useful in the study of protein disulphide isomerase, are described. In order to establish...... the quenching chromophore (Tyr(NO2)) and Cys(pNpys) activated for reaction with a second thiol. The formation and cleavage of the interchain disulphide bonds in the library were monitored under a fluorescence microscope. Substrates to investigate the properties of protein disulphide isomerase in solution were...

Open-Source Photometric System for Enzymatic Nitrate Quantification.

Science.gov (United States)

Wittbrodt, B T; Squires, D A; Walbeck, J; Campbell, E; Campbell, W H; Pearce, J M

2015-01-01

Nitrate, the most oxidized form of nitrogen, is regulated to protect people and animals from harmful levels as there is a large over abundance due to anthropogenic factors. Widespread field testing for nitrate could begin to address the nitrate pollution problem, however, the Cadmium Reduction Method, the leading certified method to detect and quantify nitrate, demands the use of a toxic heavy metal. An alternative, the recently proposed Environmental Protection Agency Nitrate Reductase Nitrate-Nitrogen Analysis Method, eliminates this problem but requires an expensive proprietary spectrophotometer. The development of an inexpensive portable, handheld photometer will greatly expedite field nitrate analysis to combat pollution. To accomplish this goal, a methodology for the design, development, and technical validation of an improved open-source water testing platform capable of performing Nitrate Reductase Nitrate-Nitrogen Analysis Method. This approach is evaluated for its potential to i) eliminate the need for toxic chemicals in water testing for nitrate and nitrite, ii) reduce the cost of equipment to perform this method for measurement for water quality, and iii) make the method easier to carryout in the field. The device is able to perform as well as commercial proprietary systems for less than 15% of the cost for materials. This allows for greater access to the technology and the new, safer nitrate testing technique.

Comparative evaluation of nitrate removal technologies

International Nuclear Information System (INIS)

Darbi, A.; Viraraghavan, T.; Butler, R.; Corkal, D.

2002-01-01

Due to the extensive application of artificial nitrogen-based fertilizers and animal manure on land, many water agencies face problems of increasing concentrations of nitrate in groundwater. The contamination of groundwater by nitrate may pose a significant public health problem. The threat of methemoglobinemia is well documented and reflected in the U.S. drinking water standard of 10 mg/L as nitrate-nitrogen. Approximately 45% of Saskatchewan's population use groundwater for drinking purposes, out of which, approximately 23% (230,000) are rural residents. The water used is made available from over 48,000 privately owned wells in regions where there is an extensive application of chemical fertilizers. Biological denitrification, ion exchange and reveres osmosis (RO) processes were selected for further study. Field studies were conducted on these processes. The sulfur/limestone autotrophic denitrification (SLAD) process was selected to achieve biological removal of nitrate from groundwater. The feasibility of the system was evaluated under anaerobic conditions. An ion exchange study was conducted using Ionac A554 which is strong anion exchange resins. In the case of groundwater containing low sulfate concentrations, A554 offered high nitrate removal. However, the disposal of regenerant brine can be a problem. A reverse osmosis unit with Filmtec membrane elements (FT30-Element Family) was used in the study on nitrate removal. The unit effluent average nitrate concentration was less than the maximum allowable concentration. (author)

Determination of the total nitrate content of thorium nitrate solution with a selective electrode

International Nuclear Information System (INIS)

Wirkner, F.M.

1979-01-01

The nitrate content of thorium nitrate solutions is determined with a liquid membrane nitrate selective electrode utilizing the known addition method in 0.1 M potassium fluoride medium as ionic strength adjustor. It is studied the influence of pH and the presence of chloride, sulphate, phosphate, meta-silicate, thorium, rare earths, iron, titanium, uranium and zirconium at the same concentrations as for the aqueous feed solutions in the thorium purification process. The method is tested in synthetic samples and in samples proceeding from nitric dissolutions of thorium hidroxide and thorium oxicarbonate utilized as thorium concentrates to be purified [pt

Nitrate Removal from Ground Water: A Review

OpenAIRE

Archna; Sharma, Surinder K.; Sobti, Ranbir Chander

2012-01-01

Nitrate contamination of ground water resources has increased in Asia, Europe, United States, and various other parts of the world. This trend has raised concern as nitrates cause methemoglobinemia and cancer. Several treatment processes can remove nitrates from water with varying degrees of efficiency, cost, and ease of operation. Available technical data, experience, and economics indicate that biological denitrification is more acceptable for nitrate removal than reverse osmosis and ion ex...

Functional mapping of protein-protein interactions in an enzyme complex by directed evolution.

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Kathrin Roderer

Full Text Available The shikimate pathway enzyme chorismate mutase converts chorismate into prephenate, a precursor of Tyr and Phe. The intracellular chorismate mutase (MtCM of Mycobacterium tuberculosis is poorly active on its own, but becomes >100-fold more efficient upon formation of a complex with the first enzyme of the shikimate pathway, 3-deoxy-d-arabino-heptulosonate-7-phosphate synthase (MtDS. The crystal structure of the enzyme complex revealed involvement of C-terminal MtCM residues with the MtDS interface. Here we employed evolutionary strategies to probe the tolerance to substitution of the C-terminal MtCM residues from positions 84-90. Variants with randomized positions were subjected to stringent selection in vivo requiring productive interactions with MtDS for survival. Sequence patterns identified in active library members coincide with residue conservation in natural chorismate mutases of the AroQδ subclass to which MtCM belongs. An Arg-Gly dyad at positions 85 and 86, invariant in AroQδ sequences, was intolerant to mutation, whereas Leu88 and Gly89 exhibited a preference for small and hydrophobic residues in functional MtCM-MtDS complexes. In the absence of MtDS, selection under relaxed conditions identifies positions 84-86 as MtCM integrity determinants, suggesting that the more C-terminal residues function in the activation by MtDS. Several MtCM variants, purified using a novel plasmid-based T7 RNA polymerase gene expression system, showed that a diminished ability to physically interact with MtDS correlates with reduced activatability and feedback regulatory control by Tyr and Phe. Mapping critical protein-protein interaction sites by evolutionary strategies may pinpoint promising targets for drugs that interfere with the activity of protein complexes.

Functional mapping of protein-protein interactions in an enzyme complex by directed evolution.

Science.gov (United States)

Roderer, Kathrin; Neuenschwander, Martin; Codoni, Giosiana; Sasso, Severin; Gamper, Marianne; Kast, Peter

2014-01-01

The shikimate pathway enzyme chorismate mutase converts chorismate into prephenate, a precursor of Tyr and Phe. The intracellular chorismate mutase (MtCM) of Mycobacterium tuberculosis is poorly active on its own, but becomes >100-fold more efficient upon formation of a complex with the first enzyme of the shikimate pathway, 3-deoxy-d-arabino-heptulosonate-7-phosphate synthase (MtDS). The crystal structure of the enzyme complex revealed involvement of C-terminal MtCM residues with the MtDS interface. Here we employed evolutionary strategies to probe the tolerance to substitution of the C-terminal MtCM residues from positions 84-90. Variants with randomized positions were subjected to stringent selection in vivo requiring productive interactions with MtDS for survival. Sequence patterns identified in active library members coincide with residue conservation in natural chorismate mutases of the AroQδ subclass to which MtCM belongs. An Arg-Gly dyad at positions 85 and 86, invariant in AroQδ sequences, was intolerant to mutation, whereas Leu88 and Gly89 exhibited a preference for small and hydrophobic residues in functional MtCM-MtDS complexes. In the absence of MtDS, selection under relaxed conditions identifies positions 84-86 as MtCM integrity determinants, suggesting that the more C-terminal residues function in the activation by MtDS. Several MtCM variants, purified using a novel plasmid-based T7 RNA polymerase gene expression system, showed that a diminished ability to physically interact with MtDS correlates with reduced activatability and feedback regulatory control by Tyr and Phe. Mapping critical protein-protein interaction sites by evolutionary strategies may pinpoint promising targets for drugs that interfere with the activity of protein complexes.

Nitrat i drikkevandet og vores sundhed

DEFF Research Database (Denmark)

Hansen, Birgitte; Schullehner, Jörg; Sigsgaard, Torben

2014-01-01

Nitrat i drikkevandet er uønsket, da det kan påvirke vores sundhed negativt. Den øvre grænse for hvor meget nitrat der tillades i drikkevandet er fastsat i forhold til risikoen for akut forgiftning med nitrit og blå børn-syndromet. Men nitrat i drikkevandet mistænkes også for at være medvirkende...

Efficient Multiple Genome Modifications Induced by the crRNAs, tracrRNA and Cas9 Protein Complex in Zebrafish

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Ohga, Rie; Ota, Satoshi; Kawahara, Atsuo

2015-01-01

The type II clustered regularly interspaced short palindromic repeats (CRISPR) associated with Cas9 endonuclease (CRISPR/Cas9) has become a powerful genetic tool for understanding the function of a gene of interest. In zebrafish, the injection of Cas9 mRNA and guide-RNA (gRNA), which are prepared using an in vitro transcription system, efficiently induce DNA double-strand breaks (DSBs) at the targeted genomic locus. Because gRNA was originally constructed by fusing two short RNAs CRISPR RNA (crRNA) and trans-activating crRNA (tracrRNA), we examined the effect of synthetic crRNAs and tracrRNA with Cas9 mRNA or Cas9 protein on the genome editing activity. We previously reported that the disruption of tyrosinase (tyr) by tyr-gRNA/Cas9 mRNA causes a retinal pigment defect, whereas the disruption of spns2 by spns2-gRNA1/Cas9 mRNA leads to a cardiac progenitor migration defect in zebrafish. Here, we found that the injection of spns2-crRNA1, tyr-crRNA and tracrRNA with Cas9 mRNA or Cas9 protein simultaneously caused a migration defect in cardiac progenitors and a pigment defect in retinal epithelial cells. A time course analysis demonstrated that the injection of crRNAs and tracrRNA with Cas9 protein rapidly induced genome modifications compared with the injection of crRNAs and tracrRNA with Cas9 mRNA. We further show that the crRNA-tracrRNA-Cas9 protein complex is functional for the visualization of endogenous gene expression; therefore, this is a very powerful, ready-to-use system in zebrafish. PMID:26010089

Changes in cell wall properties coincide with overexpression of extensin fusion proteins in suspension cultured tobacco cells.

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Tan, Li; Pu, Yunqiao; Pattathil, Sivakumar; Avci, Utku; Qian, Jin; Arter, Allison; Chen, Liwei; Hahn, Michael G; Ragauskas, Arthur J; Kieliszewski, Marcia J

2014-01-01

Extensins are one subfamily of the cell wall hydroxyproline-rich glycoproteins, containing characteristic SerHyp4 glycosylation motifs and intermolecular cross-linking motifs such as the TyrXaaTyr sequence. Extensins are believed to form a cross-linked network in the plant cell wall through the tyrosine-derivatives isodityrosine, pulcherosine, and di-isodityrosine. Overexpression of three synthetic genes encoding different elastin-arabinogalactan protein-extensin hybrids in tobacco suspension cultured cells yielded novel cross-linking glycoproteins that shared features of the extensins, arabinogalactan proteins and elastin. The cell wall properties of the three transgenic cell lines were all changed, but in different ways. One transgenic cell line showed decreased cellulose crystallinity and increased wall xyloglucan content; the second transgenic cell line contained dramatically increased hydration capacity and notably increased cell wall biomass, increased di-isodityrosine, and increased protein content; the third transgenic cell line displayed wall phenotypes similar to wild type cells, except changed xyloglucan epitope extractability. These data indicate that overexpression of modified extensins may be a route to engineer plants for bioenergy and biomaterial production.

Extraction of nitric acid, uranyl nitrate, and bismuth nitrate from aqueous nitric acid solutions with CMPO

International Nuclear Information System (INIS)

Spencer, B.B.

1995-08-01

DOE sponsored development of the transuranium extraction (TRUEX) process for removing actinides from radioactive wastes. The solvent is a mixture of CMPO and TBP. Since the extraction characteristics of CMPO are not as well understood as those of TBP, the extraction of nitric acid, uranyl nitrate, and bismuth nitrate with CMPO (dissolved in n-dodecane) were studied. Results indicate that CMPO extracts nitric acid with a 1:1 stoichiometry; equilibrium constant is 2. 660±0.092 at 25 C, and extraction enthalpy is -5. 46±0.46 kcal/mol. Slope analysis indicates that uranyl nitrate extracts with a mixed equilibria of 1:1 and 2:1 stoichiometries in nearly equal proportion. Equil. constant of the 2: 1 extraction was 1.213 x 10 6 ±3.56 x 10 4 at 25 C; reaction enthalpy was -9.610±0.594 kcal/mol. Nitration complexation constant is 8.412±0.579, with an enthalpy of -10.72±1.87 kcal/mol. Bismuth nitrate also extracts with a mixed equilibria of (perhaps) 1:1 and 2:1 stoichiometries. A 2:1 extraction equilibrium and a nitrate complexation adequately model the data. Kinetics and enthalpies were also measured

Extraction of nitric acid, uranyl nitrate, and bismuth nitrate from aqueous nitric acid solutions with CMPO

Energy Technology Data Exchange (ETDEWEB)

Spencer, B.B.

1995-08-01

DOE sponsored development of the transuranium extraction (TRUEX) process for removing actinides from radioactive wastes. The solvent is a mixture of CMPO and TBP. Since the extraction characteristics of CMPO are not as well understood as those of TBP, the extraction of nitric acid, uranyl nitrate, and bismuth nitrate with CMPO (dissolved in n-dodecane) were studied. Results indicate that CMPO extracts nitric acid with a 1:1 stoichiometry; equilibrium constant is 2. 660{plus_minus}0.092 at 25 C, and extraction enthalpy is -5. 46{plus_minus}0.46 kcal/mol. Slope analysis indicates that uranyl nitrate extracts with a mixed equilibria of 1:1 and 2:1 stoichiometries in nearly equal proportion. Equil. constant of the 2: 1 extraction was 1.213 {times} 10{sup 6}{plus_minus}3.56 {times} 10{sup 4} at 25 C; reaction enthalpy was -9.610{plus_minus}0.594 kcal/mol. Nitration complexation constant is 8.412{plus_minus}0.579, with an enthalpy of -10.72{plus_minus}1.87 kcal/mol. Bismuth nitrate also extracts with a mixed equilibria of (perhaps) 1:1 and 2:1 stoichiometries. A 2:1 extraction equilibrium and a nitrate complexation adequately model the data. Kinetics and enthalpies were also measured.

Does the evidence about health risks associated with nitrate ingestion warrant an increase of the nitrate standard for drinking water?

Science.gov (United States)

van Grinsven, Hans JM; Ward, Mary H; Benjamin, Nigel; de Kok, Theo M

2006-01-01

Several authors have suggested that it is safe to raise the health standard for nitrate in drinking water, and save money on measures associated with nitrate pollution of drinking water resources. The major argument has been that the epidemiologic evidence for acute and chronic health effects related to drinking water nitrate at concentrations near the health standard is inconclusive. With respect to the chronic effects, the argument was motivated by the absence of evidence for adverse health effects related to ingestion of nitrate from dietary sources. An interdisciplinary discussion of these arguments led to three important observations. First, there have been only a few well-designed epidemiologic studies that evaluated ingestion of nitrate in drinking water and risk of specific cancers or adverse reproductive outcomes among potentially susceptible subgroups likely to have elevated endogenous nitrosation. Positive associations have been observed for some but not all health outcomes evaluated. Second, the epidemiologic studies of cancer do not support an association between ingestion of dietary nitrate (vegetables) and an increased risk of cancer, because intake of dietary nitrate is associated with intake of antioxidants and other beneficial phytochemicals. Third, 2–3 % of the population in Western Europe and the US could be exposed to nitrate levels in drinking water exceeding the WHO standard of 50 mg/l nitrate, particularly those living in rural areas. The health losses due to this exposure cannot be estimated. Therefore, we conclude that it is not possible to weigh the costs and benefits from changing the nitrate standard for drinking water and groundwater resources by considering the potential consequences for human health and by considering the potential savings due to reduced costs for nitrate removal and prevention of nitrate pollution. PMID:16989661

Does the evidence about health risks associated with nitrate ingestion warrant an increase of the nitrate standard for drinking water?

Science.gov (United States)

van Grinsven, Hans J M; Ward, Mary H; Benjamin, Nigel; de Kok, Theo M

2006-09-21

Several authors have suggested that it is safe to raise the health standard for nitrate in drinking water, and save money on measures associated with nitrate pollution of drinking water resources. The major argument has been that the epidemiologic evidence for acute and chronic health effects related to drinking water nitrate at concentrations near the health standard is inconclusive. With respect to the chronic effects, the argument was motivated by the absence of evidence for adverse health effects related to ingestion of nitrate from dietary sources. An interdisciplinary discussion of these arguments led to three important observations. First, there have been only a few well-designed epidemiologic studies that evaluated ingestion of nitrate in drinking water and risk of specific cancers or adverse reproductive outcomes among potentially susceptible subgroups likely to have elevated endogenous nitrosation. Positive associations have been observed for some but not all health outcomes evaluated. Second, the epidemiologic studies of cancer do not support an association between ingestion of dietary nitrate (vegetables) and an increased risk of cancer, because intake of dietary nitrate is associated with intake of antioxidants and other beneficial phytochemicals. Third, 2-3 % of the population in Western Europe and the US could be exposed to nitrate levels in drinking water exceeding the WHO standard of 50 mg/l nitrate, particularly those living in rural areas. The health losses due to this exposure cannot be estimated. Therefore, we conclude that it is not possible to weigh the costs and benefits from changing the nitrate standard for drinking water and groundwater resources by considering the potential consequences for human health and by considering the potential savings due to reduced costs for nitrate removal and prevention of nitrate pollution.

Does the evidence about health risks associated with nitrate ingestion warrant an increase of the nitrate standard for drinking water?

Directory of Open Access Journals (Sweden)

Benjamin Nigel

2006-09-01

Full Text Available Abstract Several authors have suggested that it is safe to raise the health standard for nitrate in drinking water, and save money on measures associated with nitrate pollution of drinking water resources. The major argument has been that the epidemiologic evidence for acute and chronic health effects related to drinking water nitrate at concentrations near the health standard is inconclusive. With respect to the chronic effects, the argument was motivated by the absence of evidence for adverse health effects related to ingestion of nitrate from dietary sources. An interdisciplinary discussion of these arguments led to three important observations. First, there have been only a few well-designed epidemiologic studies that evaluated ingestion of nitrate in drinking water and risk of specific cancers or adverse reproductive outcomes among potentially susceptible subgroups likely to have elevated endogenous nitrosation. Positive associations have been observed for some but not all health outcomes evaluated. Second, the epidemiologic studies of cancer do not support an association between ingestion of dietary nitrate (vegetables and an increased risk of cancer, because intake of dietary nitrate is associated with intake of antioxidants and other beneficial phytochemicals. Third, 2–3 % of the population in Western Europe and the US could be exposed to nitrate levels in drinking water exceeding the WHO standard of 50 mg/l nitrate, particularly those living in rural areas. The health losses due to this exposure cannot be estimated. Therefore, we conclude that it is not possible to weigh the costs and benefits from changing the nitrate standard for drinking water and groundwater resources by considering the potential consequences for human health and by considering the potential savings due to reduced costs for nitrate removal and prevention of nitrate pollution.

Management of Nitrate m Groundwater: A Simulation Study

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M. Ahmed

2001-01-01

Full Text Available Agriculture may cause nitrate and other chemicals to enter into groundwater systems. Nitrate in drinking water is considered a health hazard. A study was conducted to assess the extent of nitrate pollution of groundwater caused by agriculture and to evaluate the possibility of using the LEACHN model to manage nitrate entry into groundwater of agricultural areas of Al-Batinah, which is the most important agricultural region of Oman. Groundwater samples were collected and analyzed to assess the problem and to detect possible trends. Soil sampling and analyses were done to demonstrate the difference in the nitrate concentration in agricultural and non-agricultural soils. A questionnaire survey was conducted to gather information on agricultural practices, fertilizer input, and other possible sources of nitrate pollution. Results from the study show that 23% of groundwater samples have a concentration of nitrate-N concentration of 10 mg/l and 34% samples exceed 8 mg/l. Agricultural soils have higher levels of nitrate compared to non- agricultural soils. Results also demonstrate that nitrate levels in groundwater in Al-Batinah are rising. Application of the ‘LEACHN’ model demonstrated its suitability for use as a management tool to reduce nitrate leaching to groundwater by controlling fertilizer and water input.

Isolation and characterization of a reserve protein from the seeds of Opuntia ficus-indica (Cactaceae

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Uchoa A.F.

1998-01-01

Full Text Available We describe here the isolation and characterization of a major albumin from the seeds of Opuntia ficus-indica (Cactaceae. This protein has a molecular mass of 6.5 kDa and was isolated by a combination of gel filtration chromatography and reverse-phase HPLC. The amino acid composition of this protein was determined and it was shown to have similarities with the amino acid composition of several proteins from the 2S albumin storage protein family. The N-terminal amino acid sequence of this protein is Asp-Pro-Tyr-Trp-Glu-Gln-Arg.

High dose potassium-nitrate chemical dosimeter

International Nuclear Information System (INIS)

Dorda de Cancio, E.M.; Munoz, S.S.

1982-01-01

This dosimeter is used to control 10 kGY-order doses (1 Mrad). Nitrate suffers a radiolitic reduction phenomena, which is related to the given dose. The method to use potassium nitrate as dosimeter is described, as well as effects of the temperature of irradiation, pH, nitrate concentration and post-irradiation stability. Nitrate powder was irradiated at a Semi-Industrial Plant, at Centro Atomico Ezeiza, and also in a Gammacell-220 irradiator. The dose rates used were 2,60 and 1,80 KGY/hour, and the given doses varied between 1,0 and 150 KGY. The uncertainty was +-3% in all the range. (author) [es

The Effect of Nitrate Levels and Harvest Times on Fe, Zn, Cu, and K, Concentrations and Nitrate Reductase Activity in Lettuce and Spinach

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Z. Gheshlaghi

2015-09-01

Full Text Available Leafy vegetables are considered as the main sources of nitrate in the human diet. In order to investigate the effect of nitrate levels and harvest times on nitrate accumulation, nitrate reductase activity, concentrations of Fe, Zn, Cu and K in Lettuce and Spinach and their relation to nitrate accumulation in these leafy vegetables, two harvest times (29 and 46 days after transplanting, two vegetable species of lettuce and spinach and two concentrations of nitrate (10 and 20 mM were used in a hydroponics greenhouse experiment with a completely randomized design and 3 replications. Modified Hoagland and Arnon nutrient solutions were used for the experiment. The results indicated that by increasing nitrate concentration of solution, nitrate accumulation in roots and shoots of lettuce and spinach increased significantly (P ≤ 0.05, and the same trend was observed for the nitrate reductase activity in the shoots of the two species. Increasing the nitrate concentrations of solution, reduced the shoot dry weight and the concentration of Fe and Cu in both species, where as it increased the K and Zn concentrations in the shoots of the two species in each both harvest times, the nitrate accumulation increased, but the nitrate reductase activity decreased in the shoots of the two species over the course of the growth. The Concentration of Fe, Cu and K decreased in the shoots of lettuce and the spinach with the time, despite the increase in Zn concentration in the shoots. The results also indicated that increasing nitrate concentrations of solution to the levels greater than the plant capacity for reduction and net uptake of nitrate, leads to the nitrate accumulation in the plants. Nitrate accumulation in plant tissue led to decreases in fresh shoot yield and Fe and Cu concentrations and nitrate reductase activities in both lettuce and spinach.

Expression and Purification of Soluble STAT5b/STAT3 Proteins for SH2 Domain Binding Assay.

Science.gov (United States)

Asai, Akira; Takakuma, Kazuyuki

2017-01-01

When a large hydrophobic full-length protein is expressed in bacteria, it is often challenging to obtain recombinant proteins in the soluble fraction. One way to overcome this challenge is expression of deletion mutants that have improved solubility while maintaining biological activity. In this chapter, we describe a protocol for expression of truncated forms of STAT5b and STAT3 proteins that are soluble and retain SH2-mediated activity for phospho-Tyr peptide recognition.

Nitrate leaching from short-hydroperiod floodplain soils

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B. Huber

2012-11-01

Full Text Available Numerous studies have shown the importance of riparian zones to reduce nitrate (NO₃⁻ contamination coming from adjacent agricultural land. Much less is known about nitrogen (N transformations and nitrate fluxes in riparian soils with short hydroperiods (1–3 days of inundation and there is no study that could show whether these soils are a N sink or source. Within a restored section of the Thur River in NE Switzerland, we measured nitrate concentrations in soil solutions as an indicator of the net nitrate production. Samples were collected along a quasi-successional gradient from frequently inundated gravel bars to an alluvial forest, at three different depths (10, 50 and 100 cm over a one-year period. Along this gradient we quantified N input (atmospheric deposition and sedimentation and N output (leaching to create a nitrogen balance and assess the risk of nitrate leaching from the unsaturated soil to the groundwater. Overall, the main factor explaining the differences in nitrate concentrations was the field capacity (FC. In subsoils with high FCs and VWC near FC, high nitrate concentrations were observed, often exceeding the Swiss and EU groundwater quality criterions of 400 and 800 μmol L⁻¹, respectively. High sedimentation rates of river-derived nitrogen led to apparent N retention up to 200 kg N ha⁻¹ yr⁻¹ in the frequently inundated zones. By contrast, in the mature alluvial forest, nitrate leaching exceeded total N input most of the time. As a result of the large soil N pools, high amounts of nitrate were produced by nitrification and up to 94 kg N-NO₃⁻ ha⁻¹ yr⁻¹ were leached into the groundwater. Thus, during flooding when water fluxes are high, nitrate from soils can contribute up to 11% to the total nitrate load in groundwater.

CARBON-BASED REACTIVE BARRIER FOR NITRATE ...

Science.gov (United States)

Nitrate (NO3-) is a common ground water contaminant related to agricultural activity, waste water disposal, leachate from landfills, septic systems, and industrial processes. This study reports on the performance of a carbon-based permeable reactive barrier (PRB) that was constructed for in-situ bioremediation of a ground water nitrate plume caused by leakage from a swine CAFO (concentrated animal feeding operation) lagoon. The swine CAFO, located in Logan County, Oklahoma, was in operation from 1992-1999. The overall site remediation strategy includes an ammonia recovery trench to intercept ammonia-contaminated ground water and a hay straw PRB which is used to intercept a nitrate plume caused by nitrification of sorbed ammonia. The PRB extends approximately 260 m to intercept the nitrate plume. The depth of the trench averages 6 m and corresponds to the thickness of the surficial saturated zone; the width of the trench is 1.2 m. Detailed quarterly monitoring of the PRB began in March, 2004, about 1 year after construction activities ended. Nitrate concentrations hydraulically upgradient of the PRB have ranged from 23 to 77 mg/L N, from 0 to 3.2 mg/L N in the PRB, and from 0 to 65 mg/L N hydraulically downgradient of the PRB. Nitrate concentrations have generally decreased in downgradient locations with successive monitoring events. Mass balance considerations indicate that nitrate attenuation is dominantly from denitrification but with some component of

Molecular imaging of reduced renal uptake of radiolabelled [DOTA{sup 0},Tyr{sup 3}]octreotate by the combination of lysine and Gelofusine in rats

Energy Technology Data Exchange (ETDEWEB)

Rolleman, E.J.; Bernard, B.F.; Breeman, W.A.P.; Forrer, F.; Blois, E. de; Krenning, E.P.; Jong, M. de [Dept. of Nuclear Medicine, Erasmus MC Rotterdam, Nijmegen (Netherlands); Hoppin, J. [Bioscan Inc., Washington, DC (United States); Gotthardt, M.; Boerman, O.C. [Dept. of Nuclear Medicine, Radboud Univ. Hospital, Nijmegen (Netherlands)

2008-07-01

Aim: in peptide receptor radionuclide therapy (PRRT) using radiolabelled somatostatin analogues, kidney uptake of radiolabelled compound is the major dose-limiting factor. We studied the effects of Gelofusine (20 mg) and lysine (100 mg) and the combination of both after injection of therapeutic doses of radiolabelled [DOTA{sup 0},Tyr{sup 3}]octreotate (60 MBq {sup 111}In or 555 MBq {sup 177}Lu labelled to 15 {mu}g peptide) in male Lewis rats. Methods: kidney uptake was measured by single photon emission computed tomography (SPECT) scans with a four-headed multi-pinhole camera (NanoSPECT) at 24 h, 5 and 7 days p. i. and was quantified by volume of interest analysis. For validation the activity concentration in the dissected kidneys was also determined ex vivo using a gamma counter and a dose calibrator. Results: Gelofusine and lysine both reduced kidney uptake of [{sup 177}Lu-DOTA{sup 0},Tyr{sup 3}]octreotate significantly by about 40% at all time points. The combination of Gelofusine and lysine resulted in a 62% inhibition of kidney uptake (p < 0.01 vs. lysine alone). A weak but significant dose-response relationship for Gelofusine, but not for lysine, was found. In a study with [{sup 111}In-DOTA{sup 0},Tyr{sup 3}]octreotate, conclusions drawn from NanoSPECT data were confirmed by biodistribution data. Conclusions: we conclude that rat kidney uptake of radiolabelled somatostatin analogues can be monitored for a longer period in the same animal using animal SPECT. Gelofusine and lysine had equal potential to reduce kidney uptake of therapeutic doses of [{sup 177}Lu-DOTA{sup 0},Tyr{sup 3}]octreotate. The combination of these compounds caused a significantly larger reduction than lysine or Gelofusine alone and may therefore offer new possibilities in PRRT. The NanoSPECT data were validated by standard biodistribution experiments. (orig.)

Combinatorial function of velvet and AreA in transcriptional regulation of nitrate utilization and secondary metabolism.

Science.gov (United States)

López-Berges, Manuel S; Schäfer, Katja; Hera, Concepción; Di Pietro, Antonio

2014-01-01

Velvet is a conserved protein complex that functions as a regulator of fungal development and secondary metabolism. In the soil-inhabiting pathogen Fusarium oxysporum, velvet governs mycotoxin production and virulence on plant and mammalian hosts. Here we report a previously unrecognized role of the velvet complex in regulation of nitrate metabolism. F. oxysporum mutants lacking VeA or LaeA, two key components of the complex, were impaired in growth on the non-preferred nitrogen sources nitrate and nitrite. Both velvet and the general nitrogen response GATA factor AreA were required for transcriptional activation of nitrate (nit1) and nitrite (nii1) reductase genes under de-repressing conditions, as well as for the nitrate-triggered increase in chromatin accessibility at the nit1 locus. AreA also contributed to chromatin accessibility and expression of two velvet-regulated gene clusters, encoding biosynthesis of the mycotoxin beauvericin and of the siderophore ferricrocin. Thus, velvet and AreA coordinately orchestrate primary and secondary metabolism as well as virulence functions in F. oxysporum. Copyright © 2013 Elsevier Inc. All rights reserved.

Evaluating Ecosystem Services for Reducing Groundwater Nitrate Contamination: Nitrate Attenuation in the Unsaturated and Saturated Zones

Science.gov (United States)

Wang, J.

2013-12-01

Nitrates are the most common type of groundwater contamination in agricultural regions. Environmental policies targeting nitrates have focused on input control (e.g., restricted fertilizer application), intermediate loads control (e.g., reduce nitrate leached from crop fields), and final loads control (e.g., reduce catchment nitrate loads). Nitrate loads can be affected by hydrological processes in both unsaturated and saturated zones. Although many of these processes have been extensively investigated in literature, they are commonly modeled as exogenous to farm management. A couple of recent studies by scientists from the Lawrence Livermore National Laboratory show that in some situations nitrate attenuation processes in the unsaturated/saturated zone, particularly denitrification, can be intensified by certain management practices to mitigate nitrate loads. Therefore, these nitrate attenuation processes can be regarded as a set of ecosystem services that farmers can take advantage of to reduce their cost of complying with environmental policies. In this paper, a representative California dairy farm is used as a case study to show how such ecosystem attenuation services can be framed within the farm owner's decision-making framework as an option for reducing groundwater nitrate contamination. I develop an integrated dynamic model, where the farmer maximizes discounted net farm profit over multiple periods subject to environmental regulations. The model consists of three submodels: animal-waste-crop, hydrologic, and economic model. In addition to common choice variables such as irrigation, fertilization, and waste disposal options, the farmer can also endogenously choose from three water sources: surface water, deep groundwater (old groundwater in the deep aquifer that is not affected by farm effluent in the short term), and shallow groundwater (drainage water that can be recycled via capture wells at the downstream end of the farm). The capture wells not only

Ion dynamics in moltmolti melkaltal nitrates

International Nuclear Information System (INIS)

Kamiyama, Takashi; Nakamura, Yoshio; Shibata, Kaoru; Suzuki, Kenji.

1993-01-01

Quasielastic neutron scattering experiments have been performed on simple molten alkali metal nitrates, RbNO 3 and LiNO 3 . The experiments were carried out by the medium resolution inverted geometry spectrometer LAM-40 at KENS neutron scattering facility in Japan. The measured spectra are composed of narrow and broad quasielastic spectra. We assigned that the broad component corresponds to the fast intra-ionic motions in a nitrate ion. From momentum dependence of integrated intensity for this component it is found that the motion of nitrate ions in RbNO 3 melt is mainly the librational one centered C 3 axis on the ion. On the other hand the intra-ionic motion in LiNo 3 is the librational motion cnetered C 3 axis on the nitrate ion which amplitude is smaller than in RbNO 3 melt. This fact shows that the motion of nitrate ions in LiNO 3 is restricted strongly by surrounding cations. (author)

The UK Nitrate Time Bomb (Invited)

Science.gov (United States)

Ward, R.; Wang, L.; Stuart, M.; Bloomfield, J.; Gooddy, D.; Lewis, M.; McKenzie, A.

2013-12-01

The developed world has benefitted enormously from the intensification of agriculture and the increased availability and use of synthetic fertilizers during the last century. However there has also been unintended adverse impact on the natural environment (water and ecosystems) with nitrate the most significant cause of water pollution and ecosystem damage . Many countries have introduced controls on nitrate, e.g. the European Union's Water Framework and Nitrate Directives, but despite this are continuing to see a serious decline in water quality. The purpose of our research is to investigate and quantify the importance of the unsaturated (vadose) zone pathway and groundwater in contributing to the decline. Understanding nutrient behaviour in the sub-surface environment and, in particular, the time lag between action and improvement is critical to effective management and remediation of nutrient pollution. A readily-transferable process-based model has been used to predict temporal loading of nitrate at the water table across the UK. A time-varying nitrate input function has been developed based on nitrate usage since 1925. Depth to the water table has been calculated from groundwater levels based on regional-scale observations in-filled by interpolated river base levels and vertical unsaturated zone velocities estimated from hydrogeological properties and mapping. The model has been validated using the results of more than 300 unsaturated zone nitrate profiles. Results show that for about 60% of the Chalk - the principal aquifer in the UK - peak nitrate input has yet to reach the water table and concentrations will continue to rise over the next 60 years. The implications are hugely significant especially where environmental objectives must be achieved in much shorter timescales. Current environmental and regulatory management strategies rarely take lag times into account and as a result will be poorly informed, leading to inappropriate controls and conflicts

Knock-down of a tonoplast localized low-affinity nitrate transporter OsNPF7.2 affects rice growth under high nitrate ssupply

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Rui Hu

2016-10-01

Full Text Available The large nitrate transporter 1/peptide transporter family (NPF has been shown to transport diverse substrates, including nitrate, amino acids, peptides, phytohormones, and glucosinolates. However, the rice (Oryza sativa root-specific expressed member OsNPF7.2 has not been characterized. Here, our data show that OsNPF7.2 is a tonoplast localized low-affinity nitrate transporter, and affects rice growth under high nitrate supply. The expression analysis showed that OsNPF7.2 was mainly expressed in the elongation and maturation zones of roots, especially in the root sclerenchyma, cortex and stele. It was also induced by high concentrations of nitrate. Subcellular localization analysis showed that OsNPF7.2 was localized on the tonoplast of large and small vacuoles. Heterogenous expression in Xenopus laevis oocytes suggested that OsNPF7.2 was a low-affinity nitrate transporter. Knock-down of OsNPF7.2 retarded rice growth under high concentrations of nitrate. Therefore, we deduce that OsNPF7.2 plays a role in intracellular allocation of nitrate in roots, and thus influences rice growth under high nitrate supply.

Increase in the biomass of some green algae species in nitrate and ammonium mediums depending on auto-, mixo- or heterotrophic conditions

Directory of Open Access Journals (Sweden)

Stefan Gumiński

2014-01-01

Full Text Available The increase in total dry mass and protein in cultures of Chlorella pyrenoidosa, Scenedesmus quadricauda and Ankistrodesmus acicularis was studied. Under autotrophic conditions, increases in dry mass were, as a rule, larger in the nitrate medium than in the ammonium one, under mixotrophic conditions the situation was reversed and in the case of heterotrophy, the individual species reacted differently. The dependence ot the protein content increase on the nitrate or ammonium form of the medium was not clear. Changes in time of the pH and rH of the mediums were followed and the interdependence of these changes with the production of biomass is discussed.

High-nitrate vegetable diet increases plasma nitrate and nitrite concentrations and reduces blood pressure in healthy women.

Science.gov (United States)

Ashworth, Ann; Mitchell, Klaus; Blackwell, Jamie R; Vanhatalo, Anni; Jones, Andrew M

2015-10-01

Epidemiological studies suggest that green leafy vegetables, which are high in dietary nitrate, are protective against CVD such as stroke. High blood pressure (BP) is a major risk factor for stroke and inorganic nitrate has been shown to reduce BP. The objective of the present study was to test the hypothesis that diets containing high-nitrate (HN) vegetables would increase plasma nitrate and nitrite concentrations and reduce BP in healthy women. A randomized, crossover trial, where participants received HN vegetables (HN diet) or avoided HN vegetables (Control diet) for 1 week. Before and after each intervention, resting BP and plasma nitrate and nitrite concentrations were measured. University of Exeter, UK. Nineteen healthy women (mean age 20 (sd 2) years; mean BMI 22·5 (sd 3·8) kg/m2). The HN diet significantly increased plasma nitrate concentration (before HN diet: mean 24·4 (sd 5·6) µmol/l; after HN diet: mean 61·0 (sd 44·1) µmol/l, Pdiet: mean 98 (sd 91) nmol/l; after HN diet: mean 185 (sd 34) nmol/l, Pdiet. The HN diet significantly reduced resting systolic BP (before HN diet: mean 107 (sd 9) mmHg; after HN diet: mean 103 (sd 6) mmHg, Pdiet (before Control diet: mean 106 (sd 8) mmHg; after Control diet: mean 106 (sd 8) mmHg). Consumption of HN vegetables significantly increased plasma nitrate and nitrite concentrations and reduced BP in normotensive women.

Evidence for a plasma-membrane-bound nitrate reductase involved in nitrate uptake of Chlorella sorokiniana

Science.gov (United States)

Tischner, R.; Ward, M. R.; Huffaker, R. C.

1989-01-01

Anti-nitrate-reductase (NR) immunoglobulin-G (IgG) fragments inhibited nitrate uptake into Chlorella cells but had no affect on nitrate uptake. Intact anti-NR serum and preimmune IgG fragments had no affect on nitrate uptake. Membrane-associated NR was detected in plasma-membrane (PM) fractions isolated by aqueous two-phase partitioning. The PM-associated NR was not removed by sonicating PM vesicles in 500 mM NaCl and 1 mM ethylenediaminetetraacetic acid and represented up to 0.8% of the total Chlorella NR activity. The PM NR was solubilized by Triton X-100 and inactivated by Chlorella NR antiserum. Plasma-membrane NR was present in ammonium-grown Chlorella cells that completely lacked soluble NR activity. The subunit sizes of the PM and soluble NRs were 60 and 95 kDa, respectively, as determined by sodium-dodecyl-sulfate electrophoresis and western blotting.

Ammonium nitrate with 15 wt % potassium nitrate-ethylenediamine dinitrate-nitroguanidine system

Energy Technology Data Exchange (ETDEWEB)

Spencer, W.; Cady, H.H.

1981-01-01

The phase diagram for the ternary system ammonium nitrate(AN) with 15 wt % potassium nitrate(AN:15KN)-ethylenediamine dinitrate(EDD)-nitroguanidine(NQ) has been determined from room temperature to the melting point. The ternary eutectic temperature, measured for a mixture containing 67.24, 25.30, and 7.46 mole % of AN:15KN, EDD, and NQ, respectively, was found to be 98.9/sup 0/C. The binary phase diagrams for the systems AN:15KN-EDD, AN:15KN-NQ, and EDD-NQ were also determined.

Quantifying an aquifer nitrate budget and future nitrate discharge using field data from streambeds and well nests

Science.gov (United States)

Gilmore, Troy E.; Genereux, David P.; Solomon, D. Kip; Farrell, Kathleen M.; Mitasova, Helena

2016-11-01

Novel groundwater sampling (age, flux, and nitrate) carried out beneath a streambed and in wells was used to estimate (1) the current rate of change of nitrate storage, dSNO3/dt, in a contaminated unconfined aquifer, and (2) future [NO3-]FWM (the flow-weighted mean nitrate concentration in groundwater discharge) and fNO3 (the nitrate flux from aquifer to stream). Estimates of dSNO3/dt suggested that at the time of sampling (2013) the nitrate storage in the aquifer was decreasing at an annual rate (mean = -9 mmol/m2yr) equal to about one-tenth the rate of nitrate input by recharge. This is consistent with data showing a slow decrease in the [NO3-] of groundwater recharge in recent years. Regarding future [NO3-]FWM and fNO3, predictions based on well data show an immediate decrease that becomes more rapid after ˜5 years before leveling out in the early 2040s. Predictions based on streambed data generally show an increase in future [NO3-]FWM and fNO3 until the late 2020s, followed by a decrease before leveling out in the 2040s. Differences show the potential value of using information directly from the groundwater—surface water interface to quantify the future impact of groundwater nitrate on surface water quality. The choice of denitrification kinetics was similarly important; compared to zero-order kinetics, a first-order rate law levels out estimates of future [NO3-]FWM and fNO3 (lower peak, higher minimum) as legacy nitrate is flushed from the aquifer. Major fundamental questions about nonpoint-source aquifer contamination can be answered without a complex numerical model or long-term monitoring program.

An updated model for nitrate uptake modelling in plants. I. Functional component: cross-combination of flow–force interpretation of nitrate uptake isotherms, and environmental and in planta regulation of nitrate influx

Science.gov (United States)

Le Deunff, Erwan; Malagoli, Philippe

2014-01-01

Background and Aims In spite of major breakthroughs in the last three decades in the identification of root nitrate uptake transporters in plants and the associated regulation of nitrate transport activities, a simplified and operational modelling approach for nitrate uptake is still lacking. This is due mainly to the difficulty in linking the various regulations of nitrate transport that act at different levels of time and on different spatial scales. Methods A cross-combination of a Flow–Force approach applied to nitrate influx isotherms and experimentally determined environmental and in planta regulation is used to model nitrate in oilseed rape, Brassica napus. In contrast to ‘Enzyme–Substrate’ interpretations, a Flow–Force modelling approach considers the root as a single catalytic structure and does not infer hypothetical cellular processes among nitrate transporter activities across cellular layers in the mature roots. In addition, this approach accounts for the driving force on ion transport based on the gradient of electrochemical potential, which is more appropriate from a thermodynamic viewpoint. Key Results and Conclusions Use of a Flow–Force formalism on nitrate influx isotherms leads to the development of a new conceptual mechanistic basis to model more accurately N uptake by a winter oilseed rape crop under field conditions during the whole growth cycle. This forms the functional component of a proposed new structure–function mechanistic model of N uptake. PMID:24638820

Alpha autoradiography by cellulose nitrate layer

International Nuclear Information System (INIS)

Simonovic, J.; Vukovic, J.; Antanasijevic, R.

1977-01-01

From domestic cellulose nitrate bulk material thin layers for α-particle autoradiography were prepared. An artificial test specimen of a uniformly alpha labelled grid source was used. The efficiency of autoradiography by cellulose nitrate was calculated comparing with data from an Ilford K2 nuclear emulsion exposed under the same conditions as the cellulose nitrate film. The resolution was determined as the distance from grid pitch edge at which the track density fell considerably. (Auth.)

Some physiological aspects of nitrate reductase-deficient Nicotiana plumbaginifolia plants

International Nuclear Information System (INIS)

Saux, C.; Morot-Gaudry, J.F.; Lemoine, Y.; Caboche, M.

1986-01-01

Chlorate-resistant Nicotiana plumbaginifolia (cv. Viviani) mutants were found to be defective in the nitrate reductase apoprotein (NR - nia). Because they could not grow with nitrate as sole nitrogen source, they were cultivated as graftings on wild type Nicotiana tabacum. The grafts of NR - plants were found to contain less malate but more amino acids, sugars and starch than the wild type. Moreover they were chlorotic, with a slight increase of the proportion of LH Chl a/b protein complexes and they exhibited a lowering of the efficiency of energy transfer between the light-harvesting complexes and the active centers. After 14 CO 2 pulse and chase experiments. The total 14 C incorporation of the mutant leaves was approximately 20% of that of the control. The NR - leaves mainly accumulated 14 C in the whole intermediates of the Calvin-cycle and in sucrose. In the most deficient NR leaves, chloroplasts were stuffed with large starch inclusions disorganizing the lamellar system

Some physiological aspects of nitrate reductase-deficient Nicotiana plumbaginifolia plants

Energy Technology Data Exchange (ETDEWEB)

Saux, C.; Morot-Gaudry, J.F.; Lemoine, Y.; Caboche, M.

1986-04-01

Chlorate-resistant Nicotiana plumbaginifolia (cv. Viviani) mutants were found to be defective in the nitrate reductase apoprotein (NR/sup -/ nia). Because they could not grow with nitrate as sole nitrogen source, they were cultivated as graftings on wild type Nicotiana tabacum. The grafts of NR/sup -/ plants were found to contain less malate but more amino acids, sugars and starch than the wild type. Moreover they were chlorotic, with a slight increase of the proportion of LH Chl a/b protein complexes and they exhibited a lowering of the efficiency of energy transfer between the light-harvesting complexes and the active centers. After /sup 14/CO/sub 2/ pulse and chase experiments. The total /sup 14/C incorporation of the mutant leaves was approximately 20% of that of the control. The NR/sup -/ leaves mainly accumulated /sup 14/C in the whole intermediates of the Calvin-cycle and in sucrose. In the most deficient NR leaves, chloroplasts were stuffed with large starch inclusions disorganizing the lamellar system.

Relationship between nitrate reductase and nitrate uptake in phytoplankton in the Peru upwelling region

International Nuclear Information System (INIS)

Blasco, D.; MacIsaac, J.J.; Packard, T.T.; Dugdale, R.C.

1984-01-01

Nitrate reductase (NR) activity and 15 NO 3 - uptake in phytoplankton were compared under different environmental conditions on two cruises in the upwelling region off Peru. The NR activity and NO 3 - uptake rates responded differently to light and nutrients and the differences led to variations in the uptake:reductase ratio. Analysis of these variations suggests that the re-equilibration time of the two processes in response to environmental perturbation is an important source of variability. The nitrate uptake system responds faster than the nitrate reductase system. Considering these differences in response time, the basic differences in the two processes, and the differences in their measurement, the authors conclude that the NR activity measures the current nitrate-reducing potential, which relfects NO 3 - assimilation before the sampling time, while 15 NO 3 - uptake measures NO 3 - assimilation in the 6-h period following sampling. Thus, considering the sampling time as a point of reference, the former is a measure of the past and the latter is a measure of the future

Coordination modes of tyrosinate-ligated catalase-type heme enzymes: magnetic circular dichroism studies of Plexaura homomalla allene oxide synthase, Mycobacterium avium ssp. paratuberculosis protein-2744c, and bovine liver catalase in their ferric and ferrous states.

Science.gov (United States)

Bandara, D M Indika; Sono, Masanori; Bruce, Grant S; Brash, Alan R; Dawson, John H

2011-12-01

Bovine liver catalase (BLC), catalase-related allene oxide synthase (cAOS) from Plexaura homomalla, and a recently isolated protein from the cattle pathogen Mycobacterium avium ssp. paratuberculosis (MAP-2744c (MAP)) are all tyrosinate-ligated heme enzymes whose crystal structures have been reported. cAOS and MAP have low (enzymes in their ferric and ferrous states using magnetic circular dichroism and UV-visible absorption spectroscopy. The MAP protein shows remarkable spectral similarities to cAOS and BLC in its native Fe(III) state, but clear differences from ferric proximal heme ligand His93Tyr Mb (myoglobin) mutant, which may be attributed to the presence of an Arg(+)-N(ω)-H···¯O-Tyr (proximal heme axial ligand) hydrogen bond in the first three heme proteins. Furthermore, the spectra of Fe(III)-CN¯, Fe(III)-NO, Fe(II)-NO (except for five-coordinate MAP), Fe(II)-CO, and Fe(II)-O(2) states of cAOS and MAP, but not H93Y Mb, are also similar to the corresponding six-coordinate complexes of BLC, suggesting that a tyrosinate (Tyr-O¯) is the heme axial ligand trans to the bound ligands in these complexes. The Arg(+)-N(ω)-H to ¯O-Tyr hydrogen bond would be expected to modulate the donor properties of the proximal tyrosinate oxyanion and, combined with the subtle differences in the catalytic site structures, affect the activities of cAOS, MAP and BLC. Copyright © 2011 Elsevier Inc. All rights reserved.

Decomposition of metal nitrate solutions

International Nuclear Information System (INIS)

Haas, P.A.; Stines, W.B.

1982-01-01

Oxides in powder form are obtained from aqueous solutions of one or more heavy metal nitrates (e.g. U, Pu, Th, Ce) by thermal decomposition at 300 to 800 deg C in the presence of about 50 to 500% molar concentration of ammonium nitrate to total metal. (author)

Supercritical fluid extraction of uranium and neodymium nitrates

International Nuclear Information System (INIS)

Sujatha, K.; Sivaraman, N.; Srinivasan, T.G.; Vasudeva Rao, P.R.

2011-01-01

Supercritical fluid extraction (SFE) of uranyl nitrate and neodymium nitrate salts from a mixture was investigated in the present study using Sc-CO 2 modified with various ligands such as organophosphorous compounds, amides, and diketones. Preferential extraction of uranyl nitrate over neodymium nitrate was demonstrated using Sc-CO 2 modified with amide, di-(2ethylhexyl) isobutyramide (D2EHIBA). (author)

The Effect of Nitrate Levels and Harvest Times on Fe, Zn, Cu, and K, Concentrations and Nitrate Reductase Activity in Lettuce and Spinach

OpenAIRE

Z. Gheshlaghi; R. Khorassani; G.H. Haghnia; M. Kafi

2015-01-01

Leafy vegetables are considered as the main sources of nitrate in the human diet. In order to investigate the effect of nitrate levels and harvest times on nitrate accumulation, nitrate reductase activity, concentrations of Fe, Zn, Cu and K in Lettuce and Spinach and their relation to nitrate accumulation in these leafy vegetables, two harvest times (29 and 46 days after transplanting), two vegetable species of lettuce and spinach and two concentrations of nitrate (10 and 20 mM) were used in ...

Alpha autoradiography by cellulose nitrate layer

International Nuclear Information System (INIS)

Simonovic, J.; Vukovic, J.; Antanasijevic, R.

1976-01-01

From domestic cellulose nitrate bulk material thin layers for α-particle autoradiography were prepared. An artifical test specimen of a uniformly alpha labelled grid source was used. The efficiency of autoradiographs by cellulose nitrate was calculated comparing with data from an Ilford K2 nuclear emulsion exposed under the same conditions as the cellulose nitrate film. The resolution was determined as the distance from grid pitch edge at which the track density fell considerably. (orig.) [de

The changing trend in nitrate concentrations in major aquifers due to historical nitrate loading from agricultural land across England and Wales from 1925 to 2150

Energy Technology Data Exchange (ETDEWEB)

Wang, L., E-mail: lei.wang@bgs.ac.uk [British Geological Survey, Keyworth, Nottingham NG12 5GG (United Kingdom); Stuart, M.E.; Lewis, M.A. [British Geological Survey, Maclean Building, Wallingford, Oxfordshire OX10 8BB (United Kingdom); Ward, R.S. [British Geological Survey, Keyworth, Nottingham NG12 5GG (United Kingdom); Skirvin, D. [ADAS UK Ltd., Pendeford House, Pendeford Business Park, Wobaston Road, Wolverhampton WV9 5AP (United Kingdom); Naden, P.S. [Centre for Ecology and Hydrology, Maclean Building, Wallingford, Oxfordshire OX10 8BB (United Kingdom); Collins, A.L. [Sustainable Soils and Grassland Systems Department, Rothamsted Research, North Wyke, Okehampton EX20 2SB (United Kingdom); Ascott, M.J. [British Geological Survey, Maclean Building, Wallingford, Oxfordshire OX10 8BB (United Kingdom)

2016-01-15

Nitrate is necessary for agricultural productivity, but can cause considerable problems if released into aquatic systems. Agricultural land is the major source of nitrates in UK groundwater. Due to the long time-lag in the groundwater system, it could take decades for leached nitrate from the soil to discharge into freshwaters. However, this nitrate time-lag has rarely been considered in environmental water management. Against this background, this paper presents an approach to modelling groundwater nitrate at the national scale, to simulate the impacts of historical nitrate loading from agricultural land on the evolution of groundwater nitrate concentrations. An additional process-based component was constructed for the saturated zone of significant aquifers in England and Wales. This uses a simple flow model which requires modelled recharge values, together with published aquifer properties and thickness data. A spatially distributed and temporally variable nitrate input function was also introduced. The sensitivity of parameters was analysed using Monte Carlo simulations. The model was calibrated using national nitrate monitoring data. Time series of annual average nitrate concentrations along with annual spatially distributed nitrate concentration maps from 1925 to 2150 were generated for 28 selected aquifer zones. The results show that 16 aquifer zones have an increasing trend in nitrate concentration, while average nitrate concentrations in the remaining 12 are declining. The results are also indicative of the trend in the flux of groundwater nitrate entering rivers through baseflow. The model thus enables the magnitude and timescale of groundwater nitrate response to be factored into source apportionment tools and to be taken into account alongside current planning of land-management options for reducing nitrate losses. - Highlights: • An approach to modelling groundwater nitrate at the national scale is presented. • The long time-lag for nitrate in the

The changing trend in nitrate concentrations in major aquifers due to historical nitrate loading from agricultural land across England and Wales from 1925 to 2150

International Nuclear Information System (INIS)

Wang, L.; Stuart, M.E.; Lewis, M.A.; Ward, R.S.; Skirvin, D.; Naden, P.S.; Collins, A.L.; Ascott, M.J.

2016-01-01

Nitrate is necessary for agricultural productivity, but can cause considerable problems if released into aquatic systems. Agricultural land is the major source of nitrates in UK groundwater. Due to the long time-lag in the groundwater system, it could take decades for leached nitrate from the soil to discharge into freshwaters. However, this nitrate time-lag has rarely been considered in environmental water management. Against this background, this paper presents an approach to modelling groundwater nitrate at the national scale, to simulate the impacts of historical nitrate loading from agricultural land on the evolution of groundwater nitrate concentrations. An additional process-based component was constructed for the saturated zone of significant aquifers in England and Wales. This uses a simple flow model which requires modelled recharge values, together with published aquifer properties and thickness data. A spatially distributed and temporally variable nitrate input function was also introduced. The sensitivity of parameters was analysed using Monte Carlo simulations. The model was calibrated using national nitrate monitoring data. Time series of annual average nitrate concentrations along with annual spatially distributed nitrate concentration maps from 1925 to 2150 were generated for 28 selected aquifer zones. The results show that 16 aquifer zones have an increasing trend in nitrate concentration, while average nitrate concentrations in the remaining 12 are declining. The results are also indicative of the trend in the flux of groundwater nitrate entering rivers through baseflow. The model thus enables the magnitude and timescale of groundwater nitrate response to be factored into source apportionment tools and to be taken into account alongside current planning of land-management options for reducing nitrate losses. - Highlights: • An approach to modelling groundwater nitrate at the national scale is presented. • The long time-lag for nitrate in the

Lysobacter capsici AZ78 produces cyclo(L-Pro-L-Tyr), a 2,5-diketopiperazine with toxic activity against sporangia of Phytophthora infestans and Plasmopara viticola.

Science.gov (United States)

Puopolo, G; Cimmino, A; Palmieri, M C; Giovannini, O; Evidente, A; Pertot, I

2014-10-01

To investigate low molecular weight compounds produced in vitro by Lysobacter capsici AZ78 and their toxic activity against sporangia of plant pathogenic oomycetes. Assays carried out in vitro showed that L. capsici AZ78 drastically inhibits the growth of plant pathogenic oomycetes. Accordingly, the preventive application of culture filtrates of L. capsici AZ78 on grapevine and tomato plants reduced the infections, respectively, caused by Plasmopara (Pl.) viticola and Phytophthora infestans. The subsequent chemical analysis of the culture filtrates of L. capsici AZ78 by spectroscopic (essentially 1D and 2D (1)H NMR and (13)C NMR and ESI MS spectra) and optical methods led to the identification of the 2,5-diketopiperazine cyclo(L-Pro-L-Tyr) that inhibited the development of P. infestans sporangia in vitro and on tomato leaves. Furthermore, a genomic region with high sequence identity with genes coding for a hybrid polyketide synthase and nonribosomal peptide synthetase was detected in L. capsici AZ78. Lysobacter capsici AZ78 produces cyclo(L-Pro-L-Tyr) in vitro that was effective in killing the sporangia of P. infestans and Pl. viticola in vitro. Moreover, this low molecular weight compound prevents the occurrence of late blight lesions when applied on tomato leaves. The application of L. capsici AZ78 cells or its own culture filtrates effectively controls both P. infestans and Pl. viticola. Cyclo(L-Pro-L-Tyr) produced by L. capsici AZ78 is toxic against sporangia of both these oomycetes. These data enforce the potential in the use of Lysobacter members for the control of plant pathogenic oomycetes and provide the basis for the development of new low-impact fungicides based on cyclo(L-Pro-L-Tyr). © 2014 The Society for Applied Microbiology.

Uptake kinetics of the somatostatin receptor ligand [86Y]DOTA-dPhe1-Tyr3-octreotide ([86Y]SMT487) using positron emission tomography in non-human primates and calculation of radiation doses of the 90Y-labelled analogue

International Nuclear Information System (INIS)

Roesch, F.; Brockmann, J.; Koehle, M.

1999-01-01

[ 90 Y]DOTA-dPhe 1 -Tyr 3 -octreotide ([ 90 Y]-SMT487) has been suggested as a promising radiotherapeutic agent for somatostatin receptor-expressing tumours. In order to quantify the in vivo parameters of this compound and the radiation doses delivered to healthy organs, the analogue [ 86 Y]DOTA-dPhe 1 -Tyr 3 -octreotide was synthesised and its uptake measured in baboons using positron emission tomography (PET). [ 86 Y]DOTA-dPhe 1 -Tyr 3 -octreotide was administered at two different peptide concentrations, namely 2 and 100 μg peptide per m 2 body surface. The latter concentration corresponded to a radiotherapeutic dose. In a third protocol [ 86 Y]DOTA-dPhe 1 -Tyr 3 -octreotide was injected in conjunction with a simultaneous infusion of an amino acid solution that was high in l-lysine in order to lower the renal uptake of radioyttrium. Quantitative whole-body PET scans were recorded to measure the uptake kinetics for kidneys, liver, lung and bone. The individual absolute uptake kinetics were used to calculate the radiation doses for [ 90 Y]DOTA-dPhe 1 -Tyr 3 -octreotide according to the MIRD recommendations extrapolated to a 70-kg human. The highest radiation dose was received by the kidneys, with 2.1-3.3 mGy per MBq [ 90 Y]DOTA-dPhe 1 -Tyr 3 -octreotide injected. For the 100 μg/m 2 SMT487 protocol with amino acid co-infusion this dose was about 20%-40% lower than for the other two treatment protocols. The liver and the red bone marrow received doses ranging from 0.32 to 0.53 mGy and 0.03 to 0.07 mGy per MBq [ 90 Y]DOTA-dPhe 1 -Tyr 3 -octreotide, respectively. The average effective dose equivalent amounted to 0.23-0.32 mSv/MBq. The comparatively low estimated radiation doses to normal organs support the initiation of clinical phase I trials with [ 90 Y]DOTA-dPhe 1 -Tyr 3 -octreotide in patients with somatostatin receptor-expressing tumours. (orig.)

Synthesis of a new energetic nitrate ester

Energy Technology Data Exchange (ETDEWEB)

Chavez, David E [Los Alamos National Laboratory

2008-01-01

Nitrate esters have been known as useful energetic materials since the discovery of nitroglycerin by Ascanio Sobrero in 1846. The development of methods to increase the safety and utility of nitroglycerin by Alfred Nobel led to the revolutionary improvement in the utility of nitroglycerin in explosive applications in the form of dynamite. Since then, many nitrate esters have been prepared and incorporated into military applications such as double-based propellants, detonators and as energetic plasticizers. Nitrate esters have also been shown to have vasodilatory effects in humans and thus have been studied and used for treatments of ailments such as angina. The mechanism of the biological response towards nitrate esters has been elucidated recently. Interestingly, many of the nitrate esters used for military purposes are liquids (ethylene glycol dinitrate, propylene glycol dinitrate, etc). Pentaerythritol tetranitrate (PETN) is one of the only solid nitrate esters, besides nitrocellulose, that is used in any application. Unfortunately, PETN melting point is above 100 {sup o}C, and thus must be pressed as a solid for detonator applications. A more practical material would be a melt-castable explosive, for potential simplification of manufacturing processes. Herein we describe the synthesis of a new energetic nitrate ester (1) that is a solid at ambient temperatures, has a melting point of 85-86 {sup o}C and has the highest density of any known nitrate ester composed only of carbon, hydrogen, nitrogen and oxygen. We also describe the chemical, thermal and sensitivity properties of 1 as well as some preliminary explosive performance data.

Radiation-induced nitration of organic compounds in aqueous solutions

International Nuclear Information System (INIS)

Ershov, B.G.; Gordeev, A.V.; Bykov, G.L.; Moisy, P.

2012-01-01

Radiolysis in aqueous nitrate and acetic acid solutions and nitrate/nitric acid and phenol was studied. The radiolysis of these solutes occurs with circle NO 2 radical, which is the active nitrating agent. Accumulation of nitromethane and nitrite was determined during γ-irradiation of aqueous solutions containing acetic and nitrate solutions. Irradiation of aqueous phenol-nitrate/nitric acid solutions results in the formation of 2- and 4-nitrophenols.

Radiation-induced nitration of organic compounds in aqueous solutions

Energy Technology Data Exchange (ETDEWEB)

Ershov, B.G.; Gordeev, A.V.; Bykov, G.L. [Russian Academy of Sciences, Moscow (Russian Federation). Frumkin Inst. of Physical Chemistry and Electrochemistry; Moisy, P. [CEA, Bagnols sur Ceze (France). Nuclear Energy Div.

2012-07-01

Radiolysis in aqueous nitrate and acetic acid solutions and nitrate/nitric acid and phenol was studied. The radiolysis of these solutes occurs with {sup circle} NO{sup 2} radical, which is the active nitrating agent. Accumulation of nitromethane and nitrite was determined during {gamma}-irradiation of aqueous solutions containing acetic and nitrate solutions. Irradiation of aqueous phenol-nitrate/nitric acid solutions results in the formation of 2- and 4-nitrophenols.

Carbon-11 tyrosine PET for visualization and protein synthesis rate assessment of laryngeal and hypopharyngeal carcinomas

Energy Technology Data Exchange (ETDEWEB)

Boer, Jurjan R. de; Laan, Bernard F.A.M. van der; Albers, Frans W.J. [Department of Otorhinolaryngology and Head and Neck Surgery, Groningen University Hospital, Hanzeplein 1, 9700 RB Groningen (Netherlands); Pruim, Jan; Que, Tjin H.; Willemsen, Antoon T.M.; Vaalburg, Willem [PET Center of Groningen University Hospital, Groningen (Netherlands); Burlage, Fred [Department of Radiotherapy, Groningen University Hospital, Groningen (Netherlands); Krikke, Allard [Department of Radiology, Groningen University Hospital, Groningen (Netherlands); Tiebosch, Anton T.M.G. [Department of Pathology, Groningen University Hospital, Groningen (Netherlands)

2002-09-01

Accurate assessment of tumour extent and lymph node involvement in squamous cell carcinomas of the head and neck region is essential for therapy planning. Unfortunately, conventional diagnostic examination and imaging techniques, which monitor tumours on the basis of anatomical parameters, have drawbacks in clinical practice. The aim of this study was to investigate the feasibility of L-[1-{sup 11}C]-tyrosine (TYR) positron emission tomography (PET) for visualisation of squamous cell carcinoma of the larynx and hypopharynx and quantification of tumour activity by assessment of protein synthesis rate (PSR). Dynamic TYR PET was performed on 31 patients with T1-T4 laryngeal or hypopharyngeal carcinoma before therapy. Plasma activity of TYR, {sup 11}CO{sub 2} and {sup 11}C-protein levels were measured, and PSRs were calculated for primary malignancies. All 31 laryngeal and hypopharyngeal tumours were visualised as a hotspot (sensitivity 100%). The median PSR of the tumours (2.06 nmol ml{sup -1} min{sup -1}; range 0.72-6.96) was significantly higher (P<0.001) than that of non-tumour (background) tissue (0.51 nmol ml{sup -1} min{sup -1}; range 0.22-0.89). L-[1-{sup 11}C]-Tyrosine PET appears to be a potential method for visualisation of primary laryngeal and hypopharyngeal tumours. In vivo quantification of tumour activity by assessment of PSR is possible and may have a future role in the therapy planning and therapy evaluation of laryngeal and hypopharyngeal tumours. (orig.)

Assimilation of 15N-labelled urea nitrogen and ammonium nitrate nitrogen by plants in case of root and non-root fertilization

International Nuclear Information System (INIS)

Muravin, Eh.A.; Kozhemyachko, V.A.; Vernichenko, I.V.

1974-01-01

Assimilation of 15 N labeled urea and ammonium nitrate in root and foliar application by spring wheat and barley has been studied during 1970-1973 period in a series of vegetative experiments at the Department of Agrochemistry, Timiryazev Agricultural Academy, and at D.N. Pryanishnikov Experimental Agrochemical Station. Additional fertilizer nitrogen applied at later ontogenesis stages (flowering and milky ripeness) is utilized mostly for protein synthesis in developing grains, thus leading to a significant increase in the relative grain protein content. A transfer of a part of nitrogen from the main ortion of fertilizer at later stages of nitrition results, at the same time, in a lower yield. Nitrogen utilization degree of urea and ammonium nitrate, when introduced before sowing or at the flowering stage is similar but in the latter case, however, additional assimilation of soil nitrogen is lower. The assimilation rate of nitrogen in root application is the lower the later the fertilizer is applied. When ammonium nitrate is additionally applied as nutrition to barley at the milky ripeness stage, ammonia and nitrate nitrogen are assimilated at the same rate and to the same extent but ammonia nitrogen is more rapidly used for protein synthesis and the rate of its transfer to the developing grains is higher. The rate of nitrogen assimilation at plant is much higher in foliar than in root application. Wheat utilizes more urea nitrogen at the flowering stage when root application is used but at the milky ripeness stage foliar application is more effective