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Sample records for 15n resonance assignments

  1. Sequence-specific sup 1 H and sup 15 N resonance assignments for human dihydrofolate reductase in solution

    SciTech Connect

    Stockman, B.J.; Nirmala, N.R.; Wagner, G. ); Delcamp, T.J.; DeYarman, M.T.; Freisheim, J.H. )

    1992-01-14

    Dihydrofolate reductase is an intracellular target enzyme for folate antagonists, including the anticancer drug methotrexate. In order to design novel drugs with altered binding properties, a detailed description of protein-drug interactions in solution is desirable to understand the specificity of drug binding. As a first step in this process, heteronuclear three-dimensional NMR spectroscopy has been used to make sequential resonance assignments for more than 90% of the residues in human dihydrofolate reductase complexed with methotrexate. Uniform enrichment of the 21.5-kDa protein with {sup 15}N was required to obtain the resonance assignments via heteronuclear 3D NMR spectroscopy since homonuclear 2D spectra did not provide sufficient {sup 1}H resonance dispersion. Medium- and long-range NOE's have been used to characterize the secondary structure of the binary ligand-enzyme complex in solution.

  2. (1)H, (13)C, and (15)N resonance assignments for the pro-inflammatory cytokine interleukin-36α.

    PubMed

    Goradia, Nishit; Wißbrock, Amelie; Wiedemann, Christoph; Bordusa, Frank; Ramachandran, Ramadurai; Imhof, Diana; Ohlenschläger, Oliver

    2016-10-01

    Interleukin-36α (IL-36α) is a recently characterised member of the interleukin-1 superfamily. It is involved in the pathogenesis of inflammatory arthritis in one third of psoriasis patients. By binding of IL-36α to its receptor IL-36R via the NF-κB pathway other cytokines involved in inflammatory and apoptotic cascade are activated. The efficacy of complex formation is controlled by N-terminal processing. To obtain a more detailed view on the structure function relationship we performed a heteronuclear multidimensional NMR investigation and here report the (1)H, (13)C, and (15)N resonance assignments for the backbone and side chain nuclei of the pro-inflammatory cytokine interleukin-36α.

  3. 1H, 15N and 13C backbone resonance assignments of the archetypal serpin α1-antitrypsin.

    PubMed

    Nyon, Mun Peak; Kirkpatrick, John; Cabrita, Lisa D; Christodoulou, John; Gooptu, Bibek

    2012-10-01

    Alpha(1)-antitrypsin is a 45-kDa (394-residue) serine protease inhibitor synthesized by hepatocytes, which is released into the circulatory system and protects the lung from the actions of neutrophil elastase via a conformational transition within a dynamic inhibitory mechanism. Relatively common point mutations subvert this transition, causing polymerisation of α(1)-antitrypsin and deficiency of the circulating protein, predisposing carriers to severe lung and liver disease. We have assigned the backbone resonances of α(1)-antitrypsin using multidimensional heteronuclear NMR spectroscopy. These assignments provide the starting point for a detailed solution state characterization of the structural properties of this highly dynamic protein via NMR methods.

  4. Sequence-specific 1H, 15N and 13C resonance assignments of Art v 1: a proline-rich allergen of Artemisia vulgaris pollen.

    PubMed

    Razzera, Guilherme; Gadermaier, Gabriele; Almeida, Marcius S; Ferreira, Fatima; Almeida, Fabio C L; Valente, Ana Paula

    2009-06-01

    Art v 1 is the major allergen of Artemisia vulgaris. The IgE raised against Art v 1 not only can cross-react with other proteins from the Asteraceae family members but also with components of various forms of food. Art v 1 is an important target for immunotherapy strategies, including vaccination with hypoallergenic derivatives or chimeras. We report the (1)H, (13)C, and (15)N resonance assignments of the recombinant Art v 1 and identification of secondary structures based on (13)C chemical shifts.

  5. 1H, 13C, and 15N backbone and side chain resonance assignments of thermophilic Geobacillus kaustophilus cyclophilin-A

    SciTech Connect

    Holliday, Michael; Zhang, Fengli; Isern, Nancy G.; Armstrong, Geoffrey S.; Eisenmesser, Elan Z.

    2014-04-01

    Cyclophilins catalyze the reversible peptidyl-prolyl isomerization of their substrates and are present across all kingdoms of life from humans to bacteria. Although numerous biological roles have now been discovered for cyclophilins, their function was initially ascribed to their chaperone-like activity in protein folding where they catalyze the often rate-limiting step of proline isomerization. This chaperone-like activity may be especially important under extreme conditions where cyclophilins are often over expressed, such as in tumors for human cyclophilins {Lee, 2010 #1167}, but also in organisms that thrive under extreme conditions, such as theromophilic bacteria. Moreover, the reversible nature of the peptidyl-prolyl isomerization reaction catalyzed by cyclophilins has allowed these enzymes to serve as model systems for probing the role of conformational changes during catalytic turnover {Eisenmesser, 2002 #20;Eisenmesser, 2005 #203}. Thus, we present here the resonance assignments of a thermophilic cyclophilin from Geobacillus kaustophilus derived from deep-sea sediment {Takami, 2004 #1384}. This thermophilic cyclophilin may now be studied at a variety of temperatures to provide insight into the comparative structure, dynamics, and catalytic mechanism of cyclophilins.

  6. 1H, 13C and 15N resonance assignment of the cytosolic dithiol glutaredoxin 1 from the pathogen Trypanosoma brucei.

    PubMed

    Stefani, Monica; Sturlese, Mattia; Manta, Bruno; Löhr, Frank; Mammi, Stefano; Comini, Marcelo; Bellanda, Massimo

    2016-04-01

    Trypanosomatids are parasites responsible for several tropical and subtropical diseases, such as Chaga's disease, sleeping sickness and Leishmaniasis. In contrast to the mammalian host, the thiol-redox metabolism of these pathogens depends on trypanothione [bis-glutathionylspermidine, T(SH)2] instead of glutathione (GSH) providing a set of lineage-specific proteins as drug target candidates. Glutaredoxins (Grx) are ubiquitous small thiol-disulfide oxidoreductases that belong to the thioredoxin-fold family. They play a central role in redox homeostasis and iron sulfur-cluster biogenesis. Each species, including trypanosomes, possesses its own set of isoforms distributed in different subcellular compartments. The genome of trypanosomatids encodes for two class I (dithiolic) Grxs named 2-C-Grx1 and 2-C-Grx2. Both proteins were shown to efficiently reduce different disulfides at the expenses of T(SH)2 using a mechanism that involves the two cysteines in the active site. Moreover, the cytosolic Trypanosoma brucei 2-C-Grx1 but not the mitochondrial 2-C-Grx2 was able to coordinate an iron-sulfur cluster with T(SH)2 or GSH as ligand. As a first step to unravel the structural basis for the specificity observed in the trypanosomal glutaredoxins, we present here the NMR resonance assignment of 2-C-Grx1 from the parasite T. brucei brucei.

  7. 1H, 13C, and 15N resonance assignments for Escherichia coli ytfP, a member of the broadly conserved UPF0131 protein domain family

    SciTech Connect

    Aramini, James M.; Swapna, G.V.T.; Huang, Yuanpeng; Rajan, Paranji K.; Xiao, Rong; Shastry, Ritu; Acton, Thomas; Cort, John R.; Kennedy, Michael A.; Montelione, Gaetano

    2005-11-01

    Protein ytfP from Escherichia coli (Swiss-Prot ID: YTFP-ECOLI; NESG target ID: ER111; Wunderlich et al., 2004) is a 113-residue member of the UPF0131 protein family (Pfam ID: PF03674) of unknown function. This domain family is found in organisms from all three kingdoms, archaea, eubacteria and eukaryotes. Using triple resonance NMR techniques, we have determined 97% of backbone and 91% of side chain 1H, 13C, and 15N resonance assignments. The chemical shift and 3J(HN?Ha) scalar coupling data reveal a mixed a/b topology,????????. BMRB deposit with Accession No. 6448. Reference: Wunderlich et al. (2004) Proteins, 56, 181?187.

  8. 1H, 13C, and 15N resonance assignments for the protein coded by gene locus BB0938 of Bordetella bronchiseptica

    SciTech Connect

    Rossi, Paolo; Ramelot, Theresa A.; Xiao, Rong; Ho, Chi K.; Ma, LiChung; Acton, Thomas; Kennedy, Michael A.; Montelione, Gaetano

    2005-11-01

    The product of gene locus BB0938 from Bordetella bronchiseptica (Swiss-Prot ID: Q7WNU7-BORBR; NESG target ID: BoR11; Wunderlich et al., 2004; Pfam ID: PF03476) is a 128-residue protein of unknown function. This broadly conserved protein family is found in eubacteria and eukaryotes. Using triple resonance NMR techniques, we have determined 98% of backbone and 94% of side chain 1H, 13C, and 15N resonance assignments. The chemical shift and 3J(HN?Ha) scalar coupling data reveal a b topology with a seven-residue helical insert, ??????????. BMRB deposit with accession number 6693. Reference: Wunderlich et al. (2004) Proteins, 56, 181?187.

  9. Near-complete 1H, 13C, 15N resonance assignments of dimethylsulfoxide-denatured TGFBIp FAS1-4 A546T.

    PubMed

    Kulminskaya, Natalia V; Yoshimura, Yuichi; Runager, Kasper; Sørensen, Charlotte S; Bjerring, Morten; Andreasen, Maria; Otzen, Daniel E; Enghild, Jan J; Nielsen, Niels Chr; Mulder, Frans A A

    2016-04-01

    The transforming growth factor beta induced protein (TGFBIp) is a major protein component of the human cornea. Mutations occurring in TGFBIp may cause corneal dystrophies, which ultimately lead to loss of vision. The majority of the disease-causing mutations are located in the C-terminal domain of TGFBIp, referred as the fourth fascilin-1 (FAS1-4) domain. In the present study the FAS1-4 Ala546Thr, a mutation that causes lattice corneal dystrophy, was investigated in dimethylsulfoxide using liquid-state NMR spectroscopy, to enable H/D exchange strategies for identification of the core formed in mature fibrils. Isotope-labeled fibrillated FAS1-4 A546T was dissolved in a ternary mixture 95/4/1 v/v/v% dimethylsulfoxide/water/trifluoroacetic acid, to obtain and assign a reference 2D (1)H-(15)N HSQC spectrum for the H/D exchange analysis. Here, we report the near-complete assignments of backbone and aliphatic side chain (1)H, (13)C and (15)N resonances for unfolded FAS1-4 A546T at 25 °C.

  10. (1)H, (13)C and (15)N resonance assignments of the RodA hydrophobin from the opportunistic pathogen Aspergillus fumigatus.

    PubMed

    Pille, Ariane; Kwan, Ann H; Cheung, Ivan; Hampsey, Matthew; Aimanianda, Vishukumar; Delepierre, Muriel; Latge, Jean-Paul; Sunde, Margaret; Guijarro, J Iñaki

    2015-04-01

    Hydrophobins are fungal proteins characterised by their amphipathic properties and an idiosyncratic pattern of eight cysteine residues involved in four disulphide bridges. The soluble form of these proteins spontaneously self-assembles at hydrophobic/hydrophilic interfaces to form an amphipathic monolayer. The RodA hydrophobin of the opportunistic pathogen Aspergillus fumigatus forms an amyloid layer with a rodlet morphology that covers the surface of fungal spores. This rodlet layer bestows hydrophobicity to the spores facilitating their dispersal in the air and rendering the conidia inert relative to the human immune system. As a first step in the analysis of the solution structure and self-association of RodA, we report the (1)H, (13)C and (15)N resonance assignments of the soluble monomeric form of RodA.

  11. Natural-abundance 15N NMR studies of Turkey ovomucoid third domain. Assignment of peptide 15N resonances to the residues at the reactive site region via proton-detected multiple-quantum coherence

    NASA Astrophysics Data System (ADS)

    Ortiz-Polo, Gilberto; Krishnamoorthi, R.; Markley, John L.; Live, David H.; Davis, Donald G.; Cowburn, David

    Heteronuclear two-dimensional 1H{ 15N} multiple-quantum (MQ) spectroscopy has been applied to a protein sample at natural abundance: ovomucoid third domain from turkey ( Meleagris gallopavo), a serine proteinase inhibitor of 56 amino acid residues. Peptide amide 1H NMR assignments obtained by two-dimensional 1H{ 1H} NMR methods (R. Krishnamoorthi and J. L. Markley, unpublished data) led to identification of the corresponding 1H{ 15N} MQ coherence cross peaks. From these, 15N NMR chemical shifts were determined for several specific backbone amide groups of amino acid residues located around the reactive site region of the inhibitor. The results suggest that amide 15N chemical shifts, which are readily obtained in this way, may serve as sensitive probes for conformational studies of proteins.

  12. Backbone (1)H, (13)C, and (15)N NMR resonance assignments of the Krüppel-like factor 4 activation domain.

    PubMed

    Conroy, Brigid S; Weiss, Emma R; Smith, Steven P; Langelaan, David N

    2017-04-01

    Krüppel-like factor 4 (KLF4) is a transcription factor involved in diverse biological processes, including development, cellular differentiation and proliferation, and maintenance of tissue homeostasis. KLF4 has also been associated with disease states, such as cardiovascular disease and several cancers. KLF4 contains an activation domain, repression domain, and a structurally characterized C-terminal zinc finger domain that mediates its binding to DNA. The structurally uncharacterized KLF4 activation domain is critical for transactivation by KLF4 and mediates its binding to the transcriptional coactivator CBP/p300. Here, we report the (1)H, (15)N, (13)CO, (13)Cα and (13)Cβ NMR chemical shift assignments of KLF41-130, which contains the KLF4 activation domain. Narrow chemical shift dispersion in the (1)H dimension of the (1)H-(15)N HSQC spectrum suggests that the KLF41-130 fragment is intrinsically disordered.

  13. 1H, 15N and 13C resonance assignments of light organ-associated fatty acid-binding protein of Taiwanese fireflies.

    PubMed

    Tseng, Kai-Li; Lee, Yi-Zong; Chen, Yun-Ru; Lyu, Ping-Chiang

    2016-04-01

    Fatty acid-binding proteins (FABPs) are a family of proteins that modulate the transfer of various fatty acids in the cytosol and constitute a significant portion in many energy-consuming cells. The ligand binding properties and specific functions of a particular type of FABP seem to be diverse and depend on the respective binding cavity as well as the cell type from which this protein is derived. Previously, a novel FABP (lcFABP; lc: Luciola cerata) was identified in the light organ of Taiwanese fireflies. The lcFABP was proved to possess fatty acids binding capabilities, especially for fatty acids of length C14-C18. However, the structural details are unknown, and the structure-function relationship has remained to be further investigated. In this study, we finished the (1)H, (15)N and (13)C chemical shift assignments of (15)N/(13)C-enriched lcFABP by solution NMR spectroscopy. In addition, the secondary structure distribution was revealed based on the backbone N, H, Cα, Hα, C and side chain Cβ assignments. These results can provide the basis for further structural exploration of lcFABP.

  14. (1)H, (13)C and (15)N resonance assignments and secondary structure analysis of translation initiation factor 1 from Pseudomonas aeruginosa.

    PubMed

    Bernal, Alejandra; Hu, Yanmei; Palmer, Stephanie O; Silva, Aaron; Bullard, James; Zhang, Yonghong

    2016-10-01

    Pseudomonas aeruginosa is a Gram-negative opportunistic pathogen and a primary cause of infection in humans. P. aeruginosa can acquire resistance against multiple groups of antimicrobial agents, including β-lactams, aminoglycosides and fluoroquinolones, and multidrug resistance is increasing in this organism which makes treatment of the infections difficult and expensive. This has led to the unmet need for discovery of new compounds distinctly different from present antimicrobials. Protein synthesis is an essential metabolic process and a validated target for the development of new antibiotics. Translation initiation factor 1 from P. aeruginosa (Pa-IF1) is the smallest of the three initiation factors that acts to establish the 30S initiation complex to initiate translation during protein biosynthesis, and its structure is unknown. Here we report the (1)H, (13)C and (15)N chemical shift assignments of Pa-IF1 as the basis for NMR structure determination and interaction studies. Secondary structure analyses deduced from the NMR chemical shift data have identified five β-strands with an unusually extended β-strand at the C-terminal end of the protein and one short α-helix arranged in the sequential order β1-β2-β3-α1-β4-β5. This is further supported by (15)N-{(1)H} hetero NOEs. These secondary structure elements suggest the Pa-IF1 adopts the typical β-barrel structure and is composed of an oligomer-binding motif.

  15. Sequence-specific {sup 1}H, {sup 13}C, and {sup 15}N resonance assignments for intestinal fatty-acid-binding protein complexed with palmitate (15.4 kDA)

    SciTech Connect

    Hodsdon, M.E.; Toner, J.J.; Cistola, D.P.

    1994-12-01

    Intestinal fatty-acid-binding protein (I-FABP) belongs to a family of soluble, cytoplasmic proteins that are thought to function in the intracellular transport and trafficking of polar lipids. Individual members of this protein family have distinct specificities and affinities for fatty acids, cholesterol, bile salts, and retinoids. We are comparing several retinol- and fatty-acid-binding proteins from intestine in order to define the factors that control molecular recognition in this family of proteins. We have established sequential resonance assignments for uniformly {sup 13}C/{sup 15}N-enriched I-FABP complexed with perdeuterated palmitate at pH7.2 and 37{degrees}C. The assignment strategy was similar to that introduced for calmodulin. We employed seven three-dimensional NMR experiments to establish scalar couplings between backbone and sidechain atoms. Backbone atoms were correlated using triple-resonance HNCO, HNCA, TOCSY-HMQC, HCACO, and HCA(CO)N experiments. Sidechain atoms were correlated using CC-TOCSY, HCCH-TOCSY, and TOCSY-HMQC. The correlations of peaks between three-dimensional spectra were established in a computer-assisted manner using NMR COMPASS (Molecular Simulations, Inc.) Using this approach, {sup 1}H, {sup 13}C, and {sup 15}N resonance assignments have been established for 120 of the 131 residues of I-FABP. For 18 residues, amide {sup 1}H and {sup 15}N resonances were unobservable, apparently because of the rapid exchange of amide protons with bulk water at pH 7.2. The missing amide protons correspond to distinct amino acid patterns in the protein sequence, which will be discussed. During the assignment process, several sources of ambiguity in spin correlations were observed. To overcome this ambiguity, the additional inter-residue correlations often observed in the HNCA experiment were used as cross-checks for the sequential backbone assignments.

  16. 1H, 13C and 15N resonance assignments and secondary structure analysis of CmPI-II, a serine protease inhibitor isolated from marine snail Cenchritis muricatus.

    PubMed

    Cabrera-Muñoz, Aymara; Rojas, Laritza; Alonso-del-Rivero Antigua, Maday; Pires, José Ricardo

    2016-04-01

    A protease inhibitor (CmPI-II) (UNIPROT: IPK2_CENMR) from the marine mollusc Cenchritis muricatus, has been isolated and characterized. It is the first member of a new group (group 3) of non-classical Kazal-type inhibitors. CmPI-II is a tight-binding inhibitor of serine proteases: trypsin, human neutrophil elastase (HNE), subtilisin A and pancreatic elastase. This specificity is exceptional in the members of Kazal-type inhibitor family. Several models of three-dimensional structure of CmPI-II have been constructed by homology with other inhibitors of the family but its structure has not yet been solved experimentally. Here we report the (1)H, (15)N and (13)C chemical shift assignments of CmPI-II as basis for NMR structure determination and interaction studies. Secondary structure analyses deduced from the NMR chemical shift data have identified three β-strands β1: residues 14-19, β2: 23-35 and β3: 43-45 and one helix α1: 28-37 arranged in the sequential order β1-β2-α1-β3. These secondary structure elements suggest that CmPI-II adopts the typical scaffold of a Kazal-type inhibitor.

  17. 1H, 13C and 15N resonance assignments and second structure information of Fag s 1: Fagales allergen from Fagus sylvatica.

    PubMed

    Moraes, A H; Asam, C; Batista, A; Almeida, F C L; Wallner, M; Ferreira, F; Valente, A P

    2016-04-01

    Fagales allergens belonging to the Bet v 1 family account responsible for the majority of spring pollinosis in the temperate climate zones in the Northern hemisphere. Among them, Fag s 1 from beech pollen is an important trigger of Fagales pollen associated allergic reactions. The protein shares high similarity with birch pollen Bet v 1, the best-characterized member of this allergen family. Of note, recent work on Bet v 1 and its homologues found in Fagales pollen demonstrated that not all allergenic members of this family have the capacity to induce allergic sensitization. Fag s 1 was shown to bind pre-existing IgE antibodies most likely primarily directed against other members of this multi-allergen family. Therefore, it is especially interesting to compare the structures of Bet v 1-like pollen allergens, which have the potential to induce allergic sensitization with allergens that are mainly cross-reactive. This in the end will help to identify allergy eliciting molecular pattern on Bet v 1-like allergens. In this work, we report the (1)H, (15)N and (13)C NMR assignment of beech pollen Fag s 1 as well as the secondary structure information based on backbone chemical shifts.

  18. 1H, 13C and 15N resonance assignments and secondary structure analysis of translation initiation factor 1 from Pseudomonas aeruginosa

    PubMed Central

    Bernal, Alejandra; Hu, Yanmei; Palmer, Stephanie O.; Silva, Aaron; Bullard, James; Zhang, Yonghong

    2016-01-01

    Pseudomonas aeruginosa is a Gram-negative opportunistic pathogen and a primary cause of infection in humans. P. aeruginosa can acquire resistance against multiple groups of antimicrobial agents, including β-lactams, aminoglycosides and fluoroquinolones, and multidrug resistance is increasing in this organism which makes treatment of the infections difficult and expensive. This has led to the unmet need for discovery of new compounds distinctly different from present antimicrobials. Protein synthesis is an essential metabolic process and a validated target for the development of new antibiotics. Translation initiation factor 1 from P. aeruginosa (Pa-IF1) is the smallest of the three initiation factors that acts to establish the 30S initiation complex to initiate translation during protein biosynthesis, and its structure is unknown. Here we report the 1H, 13C and 15N chemical shift assignments of Pa-IF1 as the basis for NMR structure determination and interaction studies. Secondary structure analyses deduced from the NMR chemical shift data have identified five β-strands with an unusually extended β-strand at the C-terminal end of the protein and one short α-helix arranged in the sequential order β1–β2–β3–α1–β4–β5. This is further supported by 15N–{1H} hetero NOEs. These secondary structure elements suggest the Pa-IF1 adopts the typical β-barrel structure and is composed of an oligomer-binding motif. PMID:26983940

  19. 1H, 15N, 13C resonance assignment of 9.7 M urea-denatured state of the GTPase effector domain (GED) of dynamin.

    PubMed

    Chugh, Jeetender; Sharma, Shilpy; Kumar, Dinesh; Hosur, Ramakrishna V

    2009-06-01

    The GTPase effector domain (GED) of dynamin, a multi-domain protein involved in endocytosis, forms a megadalton-sized self-assembly (even at micromolar concentrations) in native conditions in vitro. While such large assemblies have remained inaccessible to detailed NMR structural characterization, till date, a significant recent achievement has been the elucidation of the GED association pathway starting from a Gdn-HCl denatured monomer. Since, the nature of the denaturant has a strong influence on the conformational preferences in the denatured states, and hence on the association pathways, or even on the final assembly, we report here the NMR resonance assignment of 9.7 M urea-denatured GED from Homo sapiens. This will form the basis for the characterization of the association pathways and the final assembly driven by urea dilution.

  20. NMR study of non-structural proteins--part I: (1)H, (13)C, (15)N backbone and side-chain resonance assignment of macro domain from Mayaro virus (MAYV).

    PubMed

    Melekis, Efstathios; Tsika, Aikaterini C; Lichière, Julie; Chasapis, Christos T; Margiolaki, Irene; Papageorgiou, Nicolas; Coutard, Bruno; Bentrop, Detlef; Spyroulias, Georgios A

    2015-04-01

    Macro domains are ADP-ribose-binding modules present in all eukaryotic organisms, bacteria and archaea. They are also found in non-structural proteins of several positive strand RNA viruses such as alphaviruses. Here, we report the high yield expression and preliminary structural analysis through solution NMR spectroscopy of the macro domain from New World Mayaro Alphavirus. The recombinant protein was well-folded and in a monomeric state. An almost complete sequence-specific assignment of its (1)H, (15)N and (13)C resonances was obtained and its secondary structure determined by TALOS+.

  1. (1)H, (15)N, (13)C backbone resonance assignments of human soluble catechol O-methyltransferase in complex with S-adenosyl-L-methionine and 3,5-dinitrocatechol.

    PubMed

    Czarnota, Sylwia; Baxter, Nicola J; Cliff, Matthew J; Waltho, Jonathan P; Scrutton, Nigel S; Hay, Sam

    2017-04-01

    Catechol O-methyltransferase (COMT) is an enzyme that plays a major role in catechol neurotransmitter deactivation. Inhibition of COMT can increase neurotransmitter levels, which provides a means of treatment for Parkinson's disease, schizophrenia and depression. COMT exists as two isozymes: a soluble cytoplasmic form (S-COMT), expressed in the liver and kidneys and a membrane-bound form (MB-COMT), found mostly in the brain. Here we report the backbone (1)H, (15)N and (13)C chemical shift assignments of S-COMT in complex with S-adenosyl-L-methionine, 3,5-dinitrocatechol and Mg(2+). Assignments were obtained by heteronuclear multidimensional NMR spectroscopy. In total, 97 % of all backbone resonances were assigned in the complex, with 205 out of a possible 215 residues assigned in the (1)H-(15)N TROSY spectrum. Prediction of solution secondary structure from a chemical shift analysis using the TALOS+ webserver is in good agreement with published X-ray crystal structures.

  2. (1)H, (13)C, and (15)N backbone resonance assignments of the full-length 40 kDa S. acidocaldarius Y-family DNA polymerase, dinB homolog.

    PubMed

    Moro, Sean L; Cocco, Melanie J

    2015-10-01

    The dinB homolog (Dbh) is a member of the Y-family of translesion DNA polymerases, which are specialized to accurately replicate DNA across from a wide variety of lesions in living cells. Lesioned bases block the progression of high-fidelity polymerases and cause detrimental replication fork stalling; Y-family polymerases can bypass these lesions. The active site of the translesion synthesis polymerase is more open than that of a replicative polymerase; consequently Dbh polymerizes with low fidelity. Bypass polymerases also have low processivity. Short extension past the lesion allows the high-fidelity polymerase to switch back onto the site of replication. Dbh and the other Y-family polymerases have been used as structural models to investigate the mechanisms of DNA polymerization and lesion bypass. Many high-resolution crystal structures of Y-family polymerases have been reported. NMR dynamics studies can complement these structures by providing a measure of protein motions. Here we report the (15)N, (1)H, and (13)C backbone resonance assignments at two temperatures (35 and 50 °C) for Sulfolobus acidocaldarius Dbh polymerase. Backbone resonance assignments have been obtained for 86 % of the residues. The polymerase active site is assigned as well as the majority of residues in each of the four domains.

  3. (1)H, (13)C, (15)N backbone and side chain NMR resonance assignments for E73 from Sulfolobus spindle-shaped virus ragged hills, a hyperthermophilic crenarchaeal virus from Yellowstone National Park.

    PubMed

    Schlenker, Casey; Menon, Smita; Lawrence, C Martin; Copié, Valérie

    2009-12-01

    Crenarchaeal viruses are commonly found in hyperthermal acidic environments such as those of Yellowstone National Park. These remarkable viruses not only exhibit unusual morphologies, but also display extreme genetic diversity. However, little is known about crenarchaeal viral life cycles, virus-host interactions, and their adaptation to hyperthermophilic environments. In an effort to better understand the functions of crenarchaeal viruses and the proteins encoded by their genomes, we have undertaken detailed structural and functional studies of gene products encoded in the open reading frames of Sulfolobus spindle-shaped virus ragged hills. Herein, we report ((15)N, (13)C, (1)H) resonance assignments of backbone and side chain atoms of a 19.1 kDa homodimeric E73 protein of SSVRH.

  4. NMR study of non-structural proteins--part II: (1)H, (13)C, (15)N backbone and side-chain resonance assignment of macro domain from Venezuelan equine encephalitis virus (VEEV).

    PubMed

    Makrynitsa, Garyfallia I; Ntonti, Dioni; Marousis, Konstantinos D; Tsika, Aikaterini C; Lichière, Julie; Papageorgiou, Nicolas; Coutard, Bruno; Bentrop, Detlef; Spyroulias, Georgios A

    2015-10-01

    Macro domains consist of 130-190 amino acid residues and appear to be highly conserved in all kingdoms of life. Intense research on this field has shown that macro domains bind ADP-ribose and other similar molecules, but their exact function still remains intangible. Macro domains are highly conserved in the Alphavirus genus and the Venezuelan equine encephalitis virus (VEEV) is a member of this genus that causes fatal encephalitis to equines and humans. In this study we report the high yield recombinant expression and preliminary solution NMR study of the macro domain of VEEV. An almost complete sequence-specific assignment of its (1)H, (15)N and (13)C resonances was obtained and its secondary structure predicted by TALOS+. The protein shows a unique mixed α/β-fold.

  5. 1H, 13C, and 15N resonance assignments of an enzymatically active domain from the catalytic component (CDTa, residues 216-420) of a binary toxin from Clostridium difficile.

    PubMed

    Roth, Braden M; Godoy-Ruiz, Raquel; Varney, Kristen M; Rustandi, Richard R; Weber, David J

    2016-04-01

    Clostridium difficile is a bacterial pathogen and is the most commonly reported source of nosocomial infection in industrialized nations. Symptoms of C. difficile infection (CDI) include antibiotic-associated diarrhea, pseudomembranous colitis, sepsis and death. Over the last decade, rates and severity of hospital infections in North America and Europe have increased dramatically and correlate with the emergence of a hypervirulent strain of C. difficile characterized by the presence of a binary toxin, CDT (C. difficile toxin). The binary toxin consists of an enzymatic component (CDTa) and a cellular binding component (CDTb) that together form the active binary toxin complex. CDTa harbors a pair of structurally similar but functionally distinct domains, an N-terminal domain (residues 1-215; (1-215)CDTa) that interacts with CDTb and a C-terminal domain (residues 216-420; (216-420)CDTa) that harbors the intact ADP-ribosyltransferase (ART) active site. Reported here are the (1)H, (13)C, and (15)N backbone resonance assignments of the 23 kDa, 205 amino acid C-terminal enzymatic domain of CDTa, termed (216-420)CDTa. These NMR resonance assignments for (216-420)CDTa represent the first for a family of ART binary toxins and provide the framework for detailed characterization of the solution-state protein structure determination, dynamic studies of this domain, as well as NMR-based drug discovery efforts.

  6. Sequential assignment of 1H, 15N, 13C resonances and secondary structure of human calmodulin-like protein determined by NMR spectroscopy.

    PubMed Central

    Qian, H.; Rogers, M. S.; Schleucher, J.; Edlund, U.; Strehler, E. E.; Sethson, I.

    1998-01-01

    Human calmodulin-like protein (CLP) is closely related to vertebrate calmodulin, yet its unique cell specific expression pattern, overlapping but divergent biochemical properties, and specific target proteins suggest that it is not an isoform of calmodulin. To gain insight into the structural differences that may underlie the difference target specificities and biochemical properties of CLP when compared to calmodulin, we determined the sequential backbone assignment and associated secondary structure of 144 out of the 148 residues of Ca2+-CLP by using multinuclear multidimensional NMR spectroscopy. Despite a very high overall degree of structural similarity between CLP and calmodulin, a number of significant differences were found mainly in the length of alpha-helices and in the central nonhelical flexible region. Interestingly, the regions of greatest primary sequence divergence between CLP and calmodulin in helices III and VIII displayed only minor secondary structure differences. The data suggest that the distinct differences in target specificity and biochemical properties of CLP and calmodulin result from the sum of several minor structural and side-chain changes spread over multiple domains in these proteins. PMID:9828009

  7. 1H, 13C, 15N resonance assignments of the extracellular loop 1 domain (ECL1) of Streptococcus pneumoniae D39 FtsX, an essential cell division protein

    PubMed Central

    Fu, Yue; Bruce, Kevin E.; Rued, Britta; Winkler, Malcolm E.; Giedroc, David P.

    2015-01-01

    FtsX is an integral membrane protein from Streptococcus pneumoniae (pneumococcus) that harbors an extracellular loop 1 domain (FtsXECL1Spn) that interacts with PcsB, an peptidoglycan hydrolase that is essential for cell growth and division. Here, we report nearly complete backbone and side chain resonance assignments and a secondary structural analysis of FtsXECL1Spn (residues 47–168 of FtsX) as first steps toward structure determination of FtsXECL1Spn. PMID:26370567

  8. High-throughput backbone resonance assignment of small 13C, 15N-labeled proteins by a triple-resonance experiment with four sequential connectivity pathways using chemical shift-dependent, apparent 1J ( 1H, 13C): HNCACB codedHAHB

    NASA Astrophysics Data System (ADS)

    Pegan, Scott; Kwiatkowski, Witek; Choe, Senyon; Riek, Roland

    2003-12-01

    The proposed three-dimensional triple-resonance experiment HNCACB codedHAHB correlates sequential 15N, 1H moieties via the chemical shifts of 13C α, 13C β, 1H α, and 1H β. The four sequential correlation pathways are achieved by the incorporation of the concept of chemical shift-coding [J. Biomol. NMR 25 (2003) 281] to the TROSY-HNCACB experiment. The monitored 1H α and 1H β chemical shifts are then coded in the line shape of the cross-peaks of 13C α, 13C β along the 13C dimension through an apparent residual scalar coupling, the size of which depends on the attached hydrogen chemical shift. The information of four sequential correlation pathways enables a rapid backbone assignment. The HNCACB codedHAHB experiment was applied to ˜85% labeled 13C, 15N-labeled amino-terminal fragment of Vaccinia virus DNA topoisomerase I comprising residues 1-77. After one day of measurement on a Bruker Avance 700 MHz spectrometer and 8 h of manual analysis of the spectrum 93% of the backbone assignment was achieved.

  9. Ner protein of phage Mu: Assignments using {sup 13}C/{sup 15}N-labeled protein

    SciTech Connect

    Strzelecka, T.; Gronenborn, A.M.; Clore, G.M.

    1994-12-01

    The Ner protein is a small (74-amino acid) DNA-binding protein that regulates a switch between the lysogenic and lytic stages of phage Mu. It inhibits expression of the C repressor gene and down-regulates its own expression. Two-dimensional NMR experiments on uniformly {sup 15}N-labeled protein provided most of the backbone and some of the sidechain proton assignments. The secondary structure determination using two-dimensional NOESY experiments showed that Ner consists of five {alpha}-helices. However, because most of the sidechain protons could not be assigned, the full structure was not determined. Using uniformly {sup 13}C/{sup 15}N-labeled Ner and a set of three-dimensional experiments, we were able to assign all of the backbone and 98% of the sidechain protons. In particular, the CBCANH and CBCA(CO)NH experiments were used to sequentially assign the C{alpha} and C{beta} resonances; the HCCH-CTOCSY and HCCH-COSY were used to assign sidechain carbon and proton resonances.

  10. Stereospecific assignments of glycine in proteins by stereospecific deuteration and {sup 15}N labeling

    SciTech Connect

    Hansen, A.P.; Curley, R.W. Jr.; Panigot, M.J.; Fesik, S.W.

    1994-12-01

    Stereospecific assignments are important for accurately determining the three-dimensional structures of proteins through the use of multidimensional NMR techniques. It is especially important to stereospecifically assign the glycine {alpha}-protons in proteins because of the potential for different backbone conformations of this residue. These stereospecific assignments are critical for interpreting the {sup 3}J{sub NH,{alpha}H} coupling constants and NOEs involving the glycine {alpha}-protons that determine the conformation of this part of the protein. However, it is often difficult to unambiguously obtain the stereospecific assignments for glycine residues by using only NOE data. In this poster, we present a method for unambiguous, stereospecific assignment of the {alpha}-protons of glycine residues. This method involves synthesis of stereo-specifically deuterated and {sup 15}N-labeled Gly using a slightly modified procedure originally described by Woodard and coworkers for the stereoselective deuteration of glycine. The stereospecifically deuterated and {sup 15}N-labeled Gy has been incorporated into recombinant proteins expressed in both bacterial systems (FKBP) and mammalian cells (u-PA). Two- and three-dimensional isotope-filtered and isotope-edited NMR experiments were used to obtain the stereospecific assignments of the glycine {alpha}-protons for these proteins.

  11. 1H, 13C and 15N chemical shift assignments of the thioredoxin from the obligate anaerobe Desulfovibrio vulgaris Hildenborough.

    PubMed

    Garcin, Edwige B; Bornet, Olivier; Pieulle, Laetitia; Guerlesquin, Françoise; Sebban-Kreuzer, Corinne

    2011-10-01

    Thioredoxins are ubiquitous key antioxidant enzymes which play an essential role in cell defense against oxidative stress. They maintain the redox homeostasis owing to the regulation of thiol-disulfide exchange. In the present paper, we report the full resonance assignments of (1)H, (13)C and (15)N atoms for the reduced and oxidized forms of Desulfovibrio vulgaris Hildenborough thioredoxin 1 (Trx1). 2D and 3D heteronuclear NMR experiments were performed using uniformly (15)N-, (13)C-labelled Trx1. Chemical shifts of 97% of the backbone and 90% of the side chain atoms were obtained for the oxidized and reduced form (BMRB deposits with accession number 17299 and 17300, respectively).

  12. 1H, 13C and 15N NMR assignments of phenazopyridine derivatives.

    PubMed

    Burgueño-Tapia, Eleuterio; Mora-Pérez, Yolanda; Morales-Ríos, Martha S; Joseph-Nathan, Pedro

    2005-03-01

    Phenazopyridine hydrochloride (1), a drug in clinical use for many decades, and some derivatives were studied by one- and two-dimensional (1)H, (13)C and (15)N NMR methodology. The assignments, combined with DFT calculations, reveal that the preferred protonation site of the drug is the pyridine ring nitrogen atom. The chemoselective acetylation of phenazopyridine (2) and its influence on the polarization of the azo nitrogen atoms were evidenced by the (15)N NMR spectra. Molecular calculations of the phenazopyridines 2-4 show that the pyridine and phenyl groups are oriented in an antiperiplanar conformation with intramolecular hydrogen bonding between the N-b atom and the C-2 amino group preserving the E-azo stereochemistry.

  13. MUSIC in triple-resonance experiments: amino acid type-selective (1)H-(15)N correlations

    PubMed

    Schubert; Smalla; Schmieder; Oschkinat

    1999-11-01

    Amino acid type-selective triple-resonance experiments can be of great help for the assignment of protein spectra, since they help to remove ambiguities in either manual or automated assignment procedures. Here, modified triple-resonance experiments that yield amino acid type-selective (1)H-(15)N correlations are presented. They are based on novel coherence transfer schemes, the MUSIC pulse sequence elements, that replace the initial INEPT transfer and are selective for XH(2) or XH(3) (X can be (15)N or (13)C). The desired amino acid type is thereby selected based on the topology of the side chain. Experiments for Gly (G-HSQC); Ala (A-HSQC); Thr, Val, Ile, and Ala (TAVI-HSQC); Thr and Ala (TA-HSQC), as well as Asn and Gln (N-HSQC and QN-HSQC), are described. The new experiments are recorded as two-dimensional experiments and therefore need only small amounts of spectrometer time. The performance of the experiments is demonstrated with the application to two protein domains. Copyright 1999 Academic Press.

  14. MUSIC in Triple-Resonance Experiments: Amino Acid Type-Selective 1H- 15N Correlations

    NASA Astrophysics Data System (ADS)

    Schubert, Mario; Smalla, Maika; Schmieder, Peter; Oschkinat, Hartmut

    1999-11-01

    Amino acid type-selective triple-resonance experiments can be of great help for the assignment of protein spectra, since they help to remove ambiguities in either manual or automated assignment procedures. Here, modified triple-resonance experiments that yield amino acid type-selective 1H-15N correlations are presented. They are based on novel coherence transfer schemes, the MUSIC pulse sequence elements, that replace the initial INEPT transfer and are selective for XH2 or XH3 (X can be 15N or 13C). The desired amino acid type is thereby selected based on the topology of the side chain. Experiments for Gly (G-HSQC); Ala (A-HSQC); Thr, Val, Ile, and Ala (TAVI-HSQC); Thr and Ala (TA-HSQC), as well as Asn and Gln (N-HSQC and QN-HSQC), are described. The new experiments are recorded as two-dimensional experiments and therefore need only small amounts of spectrometer time. The performance of the experiments is demonstrated with the application to two protein domains.

  15. Combining automated peak tracking in SAR by NMR with structure-based backbone assignment from 15N-NOESY

    PubMed Central

    2012-01-01

    Background Chemical shift mapping is an important technique in NMR-based drug screening for identifying the atoms of a target protein that potentially bind to a drug molecule upon the molecule's introduction in increasing concentrations. The goal is to obtain a mapping of peaks with known residue assignment from the reference spectrum of the unbound protein to peaks with unknown assignment in the target spectrum of the bound protein. Although a series of perturbed spectra help to trace a path from reference peaks to target peaks, a one-to-one mapping generally is not possible, especially for large proteins, due to errors, such as noise peaks, missing peaks, missing but then reappearing, overlapped, and new peaks not associated with any peaks in the reference. Due to these difficulties, the mapping is typically done manually or semi-automatically, which is not efficient for high-throughput drug screening. Results We present PeakWalker, a novel peak walking algorithm for fast-exchange systems that models the errors explicitly and performs many-to-one mapping. On the proteins: hBclXL, UbcH5B, and histone H1, it achieves an average accuracy of over 95% with less than 1.5 residues predicted per target peak. Given these mappings as input, we present PeakAssigner, a novel combined structure-based backbone resonance and NOE assignment algorithm that uses just 15N-NOESY, while avoiding TOCSY experiments and 13C-labeling, to resolve the ambiguities for a one-to-one mapping. On the three proteins, it achieves an average accuracy of 94% or better. Conclusions Our mathematical programming approach for modeling chemical shift mapping as a graph problem, while modeling the errors directly, is potentially a time- and cost-effective first step for high-throughput drug screening based on limited NMR data and homologous 3D structures. PMID:22536902

  16. Through-space (19) F-(15) N couplings for the assignment of stereochemistry in flubenzimine.

    PubMed

    Ghiviriga, Ion; Rubinski, Miles A; Dolbier, William R

    2016-07-01

    Through-space (19) F-(15) N couplings revealed the configuration of flubenzimine, with the CF3 group on N4 pointing towards the lone pair of N5. The (19) F-(15) N coupling constants were measured at natural abundance using a spin-state selective indirect-detection pulse sequence. As (15) N-labelled proteins are routinely synthesized for NMR studies, through-space (19) F-(15) N couplings have the potential to probe the stereochemistry of these proteins by (19) F labelling of some amino acids or can reveal the site of docking of fluorine-containing drugs. Copyright © 2016 John Wiley & Sons, Ltd.

  17. 1H, 13C and 15N NMR assignments of a calcium-binding protein from Entamoeba histolytica.

    PubMed

    Verma, Deepshikha; Bhattacharya, Alok; Chary, Kandala V R

    2016-04-01

    We report almost complete sequence specific (1)H, (13)C and (15)N NMR assignments of a 150-residue long calmodulin-like calcium-binding protein from Entamoeba histolytica (EhCaBP6), as a prelude to its structural and functional characterization.

  18. RNA-PAIRS: RNA probabilistic assignment of imino resonance shifts

    PubMed Central

    Bahrami, Arash; Clos, Lawrence J.; Markley, John L.; Butcher, Samuel E.

    2012-01-01

    The significant biological role of RNA has further highlighted the need for improving the accuracy, efficiency and the reach of methods for investigating RNA structure and function. Nuclear magnetic resonance (NMR) spectroscopy is vital to furthering the goals of RNA structural biology because of its distinctive capabilities. However, the dispersion pattern in the NMR spectra of RNA makes automated resonance assignment, a key step in NMR investigation of biomolecules, remarkably challenging. Herein we present RNA Probabilistic Assignment of Imino Resonance Shifts (RNA-PAIRS), a method for the automated assignment of RNA imino resonances with synchronized verification and correction of predicted secondary structure. RNA-PAIRS represents an advance in modeling the assignment paradigm because it seeds the probabilistic network for assignment with experimental NMR data, and predicted RNA secondary structure, simultaneously and from the start. Subsequently, RNA-PAIRS sets in motion a dynamic network that reverberates between predictions and experimental evidence in order to reconcile and rectify resonance assignments and secondary structure information. The procedure is halted when assignments and base-parings are deemed to be most consistent with observed crosspeaks. The current implementation of RNA-PAIRS uses an initial peak list derived from proton-nitrogen heteronuclear multiple quantum correlation (1H–15N 2D HMQC) and proton–proton nuclear Overhauser enhancement spectroscopy (1H–1H 2D NOESY) experiments. We have evaluated the performance of RNA-PAIRS by using it to analyze NMR datasets from 26 previously studied RNAs, including a 111-nucleotide complex. For moderately sized RNA molecules, and over a range of comparatively complex structural motifs, the average assignment accuracy exceeds 90%, while the average base pair prediction accuracy exceeded 93%. RNA-PAIRS yielded accurate assignments and base pairings consistent with imino resonances for a majority

  19. Determination of γ -ray widths in 15N using nuclear resonance fluorescence

    NASA Astrophysics Data System (ADS)

    Szücs, T.; Bemmerer, D.; Caciolli, A.; Fülöp, Zs.; Massarczyk, R.; Michelagnoli, C.; Reinhardt, T. P.; Schwengner, R.; Takács, M. P.; Ur, C. A.; Wagner, A.; Wagner, L.

    2015-07-01

    Background: The stable nucleus 15N is the mirror of 15O, the bottleneck in the hydrogen burning CNO cycle. Most of the 15N level widths below the proton emission threshold are known from just one nuclear resonance fluorescence (NRF) measurement, with limited precision in some cases. A recent experiment with the AGATA demonstrator array determined level lifetimes using the Doppler shift attenuation method in 15O. As a reference and for testing the method, level lifetimes in 15N have also been determined in the same experiment. Purpose: The latest compilation of 15N level properties dates back to 1991. The limited precision in some cases in the compilation calls for a new measurement to enable a comparison to the AGATA demonstrator data. The widths of several 15N levels have been studied with the NRF method. Method: The solid nitrogen compounds enriched in 15N have been irradiated with bremsstrahlung. The γ rays following the deexcitation of the excited nuclear levels were detected with four high-purity germanium detectors. Results: Integrated photon-scattering cross sections of 10 levels below the proton emission threshold have been measured. Partial γ -ray widths of ground-state transitions were deduced and compared to the literature. The photon-scattering cross sections of two levels above the proton emission threshold, but still below other particle emission energies have also been measured, and proton resonance strengths and proton widths were deduced. Conclusions: Gamma and proton widths consistent with the literature values were obtained, but with greatly improved precision.

  20. Contribution of 19F resonances on 18O( p, α)15N reaction rate

    NASA Astrophysics Data System (ADS)

    Benmeslem, Meriem; Chafa, Azzedine; Barhoumi, Slimane; Tribeche, Mouloud

    2014-08-01

    The 18O( p, α)15N reaction influences the isotopes production such as 19F, 18O, and 15N which can be used to test the models of stellar evolution. 19F is synthesized in both asymptotic giant branch (AGB) and metal-rich Wolf-Rayet (WR) stars. Using R-matrix theory we allow new values of resonances parameters in 19F. We show that the most important contribution to the differential and total cross section at low energies, comes from the levels in 19F situated at resonances energies E R =151, 680 and 840 keV with spin and parity 1/2+. The total width of the 680 keV resonance is badly known. So, we have focused on this broad resonance corresponding to the 8.65 MeV level in 19F. We delimit the temperature range in which each resonance contribution to the total reaction rate occurs by analyzing the ratio ( N A < σν> i / N A < σν>). This allowed us to show that the 680 and 840 keV broad resonances strongly dominate the reaction rate over the stellar temperature range T 9=0.02-0.06 and T 9=0.5-5. Finally, these results were compared to NACRE and Iliadis astrophysical compilations.

  1. HCN, a triple-resonance NMR technique for selective observation of histidine and tryptophan side chains in 13C/15N-labeled proteins.

    PubMed

    Sudmeier, J L; Ash, E L; Günther, U L; Luo, X; Bullock, P A; Bachovchin, W W

    1996-12-01

    HCN, a new 3D NMR technique for stepwise coherence transfer from 1H to 13C to 15N and reverse through direct spin couplings 1JCH and 1JCN, is presented as a method for detection and assignment of histidine and tryptophan side-chain 1H, 13C, and 15N resonances in uniformly 13C/15N-labeled proteins. Product-operator calculations of cross-peak volumes vs adjustable delay tau 3 were employed for determination of optimal tau 3. For the phosphatidylinositol 3-kinase (PI3K SH3 domain, MW = 9.6 kD) at pH 6, H(C)N, the 1H/15N projection, produced observable cross peaks within 20 min. and was completely selective for the single tryptophan and single histidine. The 3D HCN experiment yielded well-defined cross peaks in 20 h for the 13C/15N-labeled origin-specific DNA binding domain from simian virus 40 T-antigen (T-ag-OBD131-259, MW = 15.4 kD) at pH 5.5. Resonances from all six histidines in T-ag-OBD were observed, and 11 of the 12 1H and 13C chemical shifts and 10 of the 12 15N chemical shifts were determined. The 13C dimension proved essential in assignment of the multiply overlapping 1H and 15N resonances. From the spectra recorded at a single pH, three of the imidazoles were essentially neutral and the other three were partially protonated (22-37%). HCN yielded strong cross peaks after 18 h on a 2.0 mM sample of phenylmethanesulfonyl fluoride (PMSF)-inhibited alpha-lytic protease (MW = 19.8 kD) at pH 4.4. No spectra have been obtained, however, of native or boronic acid-inhibited alpha-lytic protease after 18 h at various temperatures ranging from 5 to 55 degrees C, probably due to efficient relaxation of active-site imidazole 1H and/or 15N nuclei.

  2. Contact replacement for NMR resonance assignment

    PubMed Central

    Xiong, Fei; Pandurangan, Gopal; Bailey-Kellogg, Chris

    2008-01-01

    Motivation: Complementing its traditional role in structural studies of proteins, nuclear magnetic resonance (NMR) spectroscopy is playing an increasingly important role in functional studies. NMR dynamics experiments characterize motions involved in target recognition, ligand binding, etc., while NMR chemical shift perturbation experiments identify and localize protein–protein and protein–ligand interactions. The key bottleneck in these studies is to determine the backbone resonance assignment, which allows spectral peaks to be mapped to specific atoms. This article develops a novel approach to address that bottleneck, exploiting an available X-ray structure or homology model to assign the entire backbone from a set of relatively fast and cheap NMR experiments. Results: We formulate contact replacement for resonance assignment as the problem of computing correspondences between a contact graph representing the structure and an NMR graph representing the data; the NMR graph is a significantly corrupted, ambiguous version of the contact graph. We first show that by combining connectivity and amino acid type information, and exploiting the random structure of the noise, one can provably determine unique correspondences in polynomial time with high probability, even in the presence of significant noise (a constant number of noisy edges per vertex). We then detail an efficient randomized algorithm and show that, over a variety of experimental and synthetic datasets, it is robust to typical levels of structural variation (1–2 AA), noise (250–600%) and missings (10–40%). Our algorithm achieves very good overall assignment accuracy, above 80% in α-helices, 70% in β-sheets and 60% in loop regions. Availability: Our contact replacement algorithm is implemented in platform-independent Python code. The software can be freely obtained for academic use by request from the authors. Contact: gopal@cs.purdue.edu; cbk@cs.dartmouth.edu PMID:18586716

  3. HCN, A Triple-Resonance NMR Technique for Selective Observation of Histidine and Tryptophan Side Chains in 13C/ 15N-Labeled Proteins

    NASA Astrophysics Data System (ADS)

    Sudmeier, James L.; Ash, Elissa L.; Günther, Ulrich L.; Luo, Xuelian; Bullock, Peter A.; Bachovchin, William W.

    1996-12-01

    HCN, a new 3D NMR technique for stepwise coherence transfer from1H to13C to15N and reverse through direct spin couplings1JCHand1JCN, is presented as a method for detection and assignment of histidine and tryptophan side-chain1H,13C, and15N resonances in uniformly13C/15N-labeled proteins. Product-operator calculations of cross-peak volumes vs adjustable delay τ3were employed for determination of optimal τ3. For the phosphatidylinositol 3-kinase (PI3K SH3 domain, MW = 9.6 kD) at pH 6, H(C)N, the1H/15N projection, produced observable cross peaks within 20 min. and was completely selective for the single tryptophan and single histidine. The 3D HCN experiment yielded well-defined cross peaks in 20 h for the13C/15N-labeled origin-specific DNA binding domain from simian virus 40 T-antigen (T-ag-OBD131-259, MW = 15.4 kD) at pH 5.5. Resonances from all six histidines in T-ag-OBD were observed, and 11 of the 121H and13C chemical shifts and 10 of the 1215N chemical shifts were determined. The13C dimension proved essential in assignment of the multiply overlapping1H and15N resonances. From the spectra recorded at a single pH, three of the imidazoles were essentially neutral and the other three were partially protonated (22-37%). HCN yielded strong cross peaks after 18 h on a 2.0 mMsample of phenylmethanesulfonyl fluoride (PMSF)-inhibited α-lytic protease (MW = 19.8 kD) at pH 4.4. No spectra have been obtained, however, of native or boronic acid-inhibited α-lytic protease after 18 h at various temperatures ranging from 5 to 55°C, probably due to efficient relaxation of active-site imidazole1H and/or15N nuclei.

  4. 1H, 15N and 13C assignments of an intramolecular Lmo2-LIM2/Ldb1-LID complex.

    PubMed

    Wilkinson-White, Lorna E; Dastmalchi, Siavoush; Kwan, Ann H; Ryan, Daniel P; Mackay, Joel P; Matthews, Jacqueline M

    2010-10-01

    Lmo2 is a LIM-only protein involved in hematopoiesis and the development of T-cell acute lymphoblastic leukaemia. Here we report backbone and side chain NMR assignments for an engineered intramolecular complex of the C-terminal LIM domain from Lmo2 tethered to the LIM interaction domain (LID) from LIM domain binding protein 1 (Ldb1).

  5. Ratios of 15N/12C and 4He/12C inclusive electroproduction cross sections in the nucleon resonance region

    SciTech Connect

    Peter Bosted; Robert Fersch

    2007-12-14

    The ratio of inclusive electron scattering cross sections for 15N/12C was determined in the kinematic range 0.8<2 GeV and 0.2<1 GeV2 using 2.285 GeV electrons and the CLAS detector at Jefferson Lab. The ratio exhibits only slight resonance structure, as predicted by a phenomenological model, and also by quark-hadron duality. Within the super-scaling quasi-elastic model, slight evidence is found for a 1 MeV lower effective nucleon binding energy in 15N than in 12C. Ratios of 4He/12C using 1.6 to 2.5 GeV electrons are in good agreement with the phenomenological model.

  6. Conformation of alamethicin in oriented phospholipid bilayers determined by (15)N solid-state nuclear magnetic resonance.

    PubMed Central

    Bak, M; Bywater, R P; Hohwy, M; Thomsen, J K; Adelhorst, K; Jakobsen, H J; Sørensen, O W; Nielsen, N C

    2001-01-01

    The conformation of the 20-residue antibiotic ionophore alamethicin in macroscopically oriented phospholipid bilayers has been studied using (15)N solid-state nuclear magnetic resonance (NMR) spectroscopy in combination with molecular modeling and molecular dynamics simulations. Differently (15)N-labeled variants of alamethicin and an analog with three of the alpha-amino-isobutyric acid residues replaced by alanines have been investigated to establish experimental structural constraints and determine the orientation of alamethicin in hydrated phospholipid (dimyristoylphosphatidylcholine) bilayers and to investigate the potential for a major kink in the region of the central Pro(14) residue. From the anisotropic (15)N chemical shifts and (1)H-(15)N dipolar couplings determined for alamethicin with (15)N-labeling on the Ala(6), Val(9), and Val(15) residues and incorporated into phospholipid bilayer with a peptide:lipid molar ratio of 1:8, we deduce that alamethicin has a largely linear alpha-helical structure spanning the membrane with the molecular axis tilted by 10-20 degrees relative to the bilayer normal. In particular, we find compatibility with a straight alpha-helix tilted by 17 degrees and a slightly kinked molecular dynamics structure tilted by 11 degrees relative to the bilayer normal. In contrast, the structural constraints derived by solid-state NMR appear not to be compatible with any of several model structures crossing the membrane with vanishing tilt angle or the earlier reported x-ray diffraction structure (Fox and Richards, Nature. 300:325-330, 1982). The solid-state NMR-compatible structures may support the formation of a left-handed and parallel multimeric ion channel. PMID:11509381

  7. Electron paramagnetic resonance of the N2V- defect in 15N -doped synthetic diamond

    NASA Astrophysics Data System (ADS)

    Green, B. L.; Dale, M. W.; Newton, M. E.; Fisher, D.

    2015-10-01

    Nitrogen is the dominant impurity in the majority of natural and synthetic diamonds, and the family of nitrogen vacancy-type (NnV ) defects are crucial in our understanding of defect dynamics in these diamonds. A significant gap is the lack of positive identification of N2V- , the dominant charge state of N2V in diamond that contains a significant concentration of electron donors. In this paper, we employ isotopically-enriched diamond to identify the EPR spectrum associated with 15N2V- and use the derived spin Hamiltonian parameters to identify 14N2V- in a natural isotopic abundance sample. The electronic wave function of the N2V- ground state and previous lack of identification is discussed. The N2V- EPR spectrum intensity is shown to correlate with the H2 optical absorption over an order of magnitude in concentration.

  8. In Vivo Fluxes in the Ammonium-Assimilatory Pathways in Corynebacterium glutamicum Studied by 15N Nuclear Magnetic Resonance

    PubMed Central

    Tesch, M.; de Graaf, A. A.; Sahm, H.

    1999-01-01

    Glutamate dehydrogenase (GDH) and glutamine synthetase (GS)–glutamine 2-oxoglutarate-aminotransferase (GOGAT) represent the two main pathways of ammonium assimilation in Corynebacterium glutamicum. In this study, the ammonium assimilating fluxes in vivo in the wild-type ATCC 13032 strain and its GDH mutant were quantitated in continuous cultures. To do this, the incorporation of 15N label from [15N]ammonium in glutamate and glutamine was monitored with a time resolution of about 10 min with in vivo 15N nuclear magnetic resonance (NMR) used in combination with a recently developed high-cell-density membrane-cyclone NMR bioreactor system. The data were used to tune a standard differential equation model of ammonium assimilation that comprised ammonia transmembrane diffusion, GDH, GS, GOGAT, and glutamine amidotransferases, as well as the anabolic incorporation of glutamate and glutamine into biomass. The results provided a detailed picture of the fluxes involved in ammonium assimilation in the two different C. glutamicum strains in vivo. In both strains, transmembrane equilibration of 100 mM [15N]ammonium took less than 2 min. In the wild type, an unexpectedly high fraction of 28% of the NH4+ was assimilated via the GS reaction in glutamine, while 72% were assimilated by the reversible GDH reaction via glutamate. GOGAT was inactive. The analysis identified glutamine as an important nitrogen donor in amidotransferase reactions. The experimentally determined amount of 28% of nitrogen assimilated via glutamine is close to a theoretical 21% calculated from the high peptidoglycan content of C. glutamicum. In the GDH mutant, glutamate was exclusively synthesized over the GS/GOGAT pathway. Its level was threefold reduced compared to the wild type. PMID:10049869

  9. Natural abundance 15N NMR assignments delineate structural differences between intact and reactive-site hydrolyzed Cucurbita maxima trypsin inhibitor III.

    PubMed

    Krishnamoorthi, R; Nemmers, S; Tobias, B

    1992-06-15

    15N NMR assignments were made to the backbone amide nitrogen atoms at natural isotopic abundance of intact and reactive-site (Arg5-Ile6) hydrolyzed Cucurbita maxima trypsin inhibitor III (CMTI-III and CMTI-III*, respectively) by means of 2D proton-detected heteronuclear single bond chemical shift correlation (HSBC) spectroscopy, utilizing the previously made sequence-specific 1H NMR assignments (Krishnamoorthi et al. (1992) Biochemistry 31, 898-904). Comparison of the 15N chemical shifts of the two forms of the inhibitor molecule revealed significant changes not only for residues located near the reactive-site region, but also for those distantly located. Residues Cys3, Arg5, Leu7, Met8, Cys10, Cys16, Glu19, His25, Tyr27, Cys28 and Gly29 showed significant chemical shift changes ranging from 0.3 to 6.1 ppm, thus indicating structural perturbations that were transmitted throughout the molecule. These findings confirm the earlier conclusions based on 1H NMR investigations.

  10. Triosephosphate isomerase: 15N and 13C chemical shift assignments and conformational change upon ligand binding by magic-angle spinning solid-state NMR spectroscopy.

    PubMed

    Xu, Yimin; Lorieau, Justin; McDermott, Ann E

    2010-03-19

    Microcrystalline uniformly (13)C,(15)N-enriched yeast triosephosphate isomerase (TIM) is sequentially assigned by high-resolution solid-state NMR (SSNMR). Assignments are based on intraresidue and interresidue correlations, using dipolar polarization transfer methods, and guided by solution NMR assignments of the same protein. We obtained information on most of the active-site residues involved in chemistry, including some that were not reported in a previous solution NMR study, such as the side-chain carbons of His95. Chemical shift differences comparing the microcrystalline environment to the aqueous environment appear to be mainly due to crystal packing interactions. Site-specific perturbations of the enzyme's chemical shifts upon ligand binding are studied by SSNMR for the first time. These changes monitor proteinwide conformational adjustment upon ligand binding, including many of the sites probed by solution NMR and X-ray studies. Changes in Gln119, Ala163, and Gly210 were observed in our SSNMR studies, but were not reported in solution NMR studies (chicken or yeast). These studies identify a number of new sites with particularly clear markers for ligand binding, paving the way for future studies of triosephosphate isomerase dynamics and mechanism.

  11. Automated resonance assignment of the 21 kDa stereo-array isotope labeled thioldisulfide oxidoreductase DsbA

    NASA Astrophysics Data System (ADS)

    Schmidt, Elena; Ikeya, Teppei; Takeda, Mitsuhiro; Löhr, Frank; Buchner, Lena; Ito, Yutaka; Kainosho, Masatsune; Güntert, Peter

    2014-12-01

    The automated chemical shift assignment algorithm FLYA has been extended for use with stereo-array isotope labeled (SAIL) proteins to determine the sequence-specific resonance assignments of large proteins. Here we present the assignment of the backbone and sidechain chemical shifts of the 21 kDa thioldisulfide oxidoreductase DsbA from Escherichia coli that were determined with the SAIL-FLYA algorithm in conjunction with automated peak picking. No manual corrections of peak lists or assignments were applied. The assignments agreed with manually determined reference assignments in 95.4% of the cases if 16 input spectra were used, 94.1% if only 3D 13C/15N-resolved NOESY, CBCA(CO)NH, and 2D [13C/15N,1H]-HSQC were used, and 86.8% if exclusively 3D 13C/15N-resolved NOESY spectra were used. Considering only the assignments that are classified as reliable by the SAIL-FLYA algorithm, the degrees of agreement increased to 97.5%, 96.5%, and 94.2%, respectively. With our approach it is thus possible to automatically obtain almost complete and correct assignments of proteins larger than 20 kDa.

  12. 1H, 15N, and 13C chemical shift assignments of cyanobacteriochrome NpR6012g4 in the red-absorbing dark state.

    PubMed

    Yu, Qinhong; Lim, Sunghyuk; Rockwell, Nathan C; Martin, Shelley S; Clark Lagarias, J; Ames, James B

    2016-04-01

    Cyanobacteriochrome (CBCR) photosensory proteins are phytochrome homologs using bilin chromophores for light sensing across the visible spectrum. NpR6012g4 is a CBCR from Nostoc punctiforme that serves as a model for a widespread CBCR subfamily with red/green photocycles. We report NMR chemical shift assignments for both the protein backbone and side-chain resonances of the red-absorbing dark state of NpR6012g4 (BMRB no. 26582).

  13. Identifying the African Wintering Grounds of Hybrid Flycatchers Using a Multi–Isotope (δ2H, δ13C, δ15N) Assignment Approach

    PubMed Central

    Van Wilgenburg, Steven L.; Hobson, Keith A.; Folmer, Eelke; Font, Laura; Klaassen, Marcel

    2014-01-01

    Migratory routes and wintering grounds can have important fitness consequences, which can lead to divergent selection on populations or taxa differing in their migratory itinerary. Collared (Ficedula albicollis) and pied (F. hypoleuca) flycatchers breeding in Europe and wintering in different sub-Saharan regions have distinct migratory routes on the eastern and western sides of the Sahara desert, respectively. In an earlier paper, we showed that hybrids of the two species did not incur reduced winter survival, which would be expected if their migration strategy had been a mix of the parent species' strategies potentially resulting in an intermediate route crossing the Sahara desert to different wintering grounds. Previously, we compared isotope ratios and found no significant difference in stable-nitrogen isotope ratios (δ15N) in winter-grown feathers between the parental species and hybrids, but stable-carbon isotope ratios (δ13C) in hybrids significantly clustered only with those of pied flycatchers. We followed up on these findings and additionally analyzed the same feathers for stable-hydrogen isotope ratios (δ2H) and conducted spatially explicit multi-isotope assignment analyses. The assignment results overlapped with presumed wintering ranges of the two species, highlighting the efficacy of the method. In contrast to earlier findings, hybrids clustered with both parental species, though most strongly with pied flycatcher. PMID:24847717

  14. 1H, 13C, and 15N backbone, side-chain, and heme chemical shift assignments for oxidized and reduced forms of the monoheme c-type cytochrome ApcA isolated from the acidophilic metal-reducing bacterium Acidiphilium cryptum.

    SciTech Connect

    Cort, John R.; Swenson, Michael; Magnuson, Timothy S.

    2011-03-04

    We report the 1H, 13C, and 15N chemical shift assignments of both oxidized and reduced forms of an abundant periplasmic c-type cytochrome, designated ApcA, from the acidophilic gram-negative facultatively anaerobic metal-reducing alpha-proteobacterium Acidiphilium cryptum. These resonance assignments prove that ApcA is a monoheme cytochrome c2 and the product of the Acry_2099 gene. An absence of resonance peaks in the NMR spectra for the 21 N-terminal residues suggests that a predicted N-terminal signal sequence is cleaved. We also describe the preparation and purification of the protein in labeled form from laboratory cultures of A. cryptum growing on 13C- and 15N- labeled substrates.

  15. A Set of Efficient nD NMR Protocols for Resonance Assignments of Intrinsically Disordered Proteins.

    PubMed

    Wiedemann, Christoph; Bellstedt, Peter; Häfner, Sabine; Herbst, Christian; Bordusa, Frank; Görlach, Matthias; Ohlenschläger, Oliver; Ramachandran, Ramadurai

    2016-07-04

    The RF pulse scheme RN[N-CA HEHAHA]NH, which provides a convenient approach to the acquisition of different multidimensional chemical shift correlation NMR spectra leading to backbone resonance assignments, including those of the proline residues of intrinsically disordered proteins (IDPs), is experimentally demonstrated. Depending on the type of correlation data required, the method involves the generation of in-phase ((15) N)(x) magnetisation via different magnetisation transfer pathways such as H→N→CO→N, HA→CA→CO→N, H→N→CA→N and H→CA→N, the subsequent application of (15) N-(13) C(α) heteronuclear Hartmann-Hahn mixing over a period of ≈100 ms, chemical-shift labelling of relevant nuclei before and after the heteronuclear mixing step and amide proton detection in the acquisition dimension. It makes use of the favourable relaxation properties of IDPs and the presence of (1) JCαN and (2) JCαN couplings to achieve efficient correlation of the backbone resonances of each amino acid residue "i" with the backbone amide resonances of residues "i-1" and "i+1". It can be implemented in a straightforward way through simple modifications of the RF pulse schemes commonly employed in protein NMR studies. The efficacy of the approach is demonstrated using a uniformly ((15) N,(13) C) labelled sample of α-synuclein. The different possibilities for obtaining the amino-acid-type information, simultaneously with the connectivity data between the backbone resonances of sequentially neighbouring residues, have also been outlined.

  16. 15N solid-state nuclear magnetic resonance study of pyrolyzed metal-polyaniline cathode catalysts for oxygen reduction in fuel cells

    NASA Astrophysics Data System (ADS)

    Kuroki, Shigeki; Hosaka, Yo; Yamauchi, Chiharu; Nagata, Shinsuke; Sonoda, Mayu

    2015-09-01

    The oxygen reduction reaction (ORR) activity of pyrolyzed metal-free and metal (Mn, Fe, Co, Ni and Cu)-containing polyaniline (PANI) in polymer electrolyte fuel cell (PEFC) was studied. The metal-free PANI800 shows quite poor ORR catalytic activity, whilst the metal-containing PANIMe800 display a better ORR activity. The 15N CP/MAS NMR spectra of PANINi800 and PANICu800 show one weak peak at 118 ppm and there is no peak observed in PANIFe800, against that of PANI800, PANIMn800, PANICo800 and PANINi800 show two peaks at 273 and 118 ppm assigned to the pyridinic and pyridinium nitrogens. It is because of the paramagnetic effect of metal ions. The 15N spin-echo NMR spectra of PANIMe800 with fast recycle delay show the peaks at 140 and 270 ppm assigned to the graphitic and pyridinic nitrogens, against that of PANI800 shows no peak. The spectra of PANIMn800, PANICo800, PANINi800 and PANICu600 also contain a very broaden peak at 430 ppm assigned to the nitrogen with Fermi-contact effect from metal ions. The spectra of PANIFe800 show some spinning side bands and the average Fe3+-15N distance can be calculated. The some amount of iron ion are relieved and average Fe3+-15N distance increase after acid washing and the ORR activity decreases.

  17. Measurement of the 20 and 90 keV Resonances in the {sup 18}O(p,{alpha}){sup 15}N Reaction via the Trojan Horse Method

    SciTech Connect

    La Cognata, M.; Spitaleri, C.; Cherubini, S.; Crucilla, V.; Gulino, M.; Lamia, L.; Pizzone, R. G.; Puglia, S. M. R.; Rapisarda, G. G.; Romano, S.; Sergi, M. L.; Tumino, A.; Mukhamedzhanov, A. M.; Tribble, R. E.; Banu, A.; Goldberg, V. Z.; Tabacaru, G.; Trache, L.; Irgaziev, B.; Coc, A.

    2008-10-10

    The {sup 18}O(p,{alpha}){sup 15}N reaction is of primary importance in several astrophysical scenarios, including fluorine nucleosynthesis inside asymptotic giant branch stars as well as oxygen and nitrogen isotopic ratios in meteorite grains. Thus the indirect measurement of the low energy region of the {sup 18}O(p,{alpha}){sup 15}N reaction has been performed to reduce the nuclear uncertainty on theoretical predictions. In particular the strength of the 20 and 90 keV resonances has been deduced and the change in the reaction rate evaluated.

  18. Statistical Assignment of Neutron Orbital Angular Momentum to a Resonance

    SciTech Connect

    Oh, Soo-Youl; Chang, Jonghwa; Leal, Luiz C.

    2004-09-15

    We have derived formulas in a general form for suggesting the neutron orbital angular momentum quantum number l to each neutron resonance if it is not identified experimentally. By assuming the (2J + 1) law of level density, these general formulas are reduced to the formulas found in previous works. The suggestion of l is based on the probability that a resonance having a certain value of g{gamma}{sub n} is an l-wave resonance. The probability is calculated from the Bayes theorem on conditional probability. For each l, the probability density function (pdf) of g{gamma}{sub n} was derived from the {chi}{sup 2} distribution proposed by Porter and Thomas. The pdf takes into account two possible channel spins that result in the same total spin for a given l larger than zero. Meanwhile, regardless of the resolution of measurement, we suggest adopting the level density as the prior probability in the Bayesian approach, as Gyulassy et al. did. As a sample problem, we presented the result of l-assignment for {sup 109}Ag resonances. The SUGGEL code, in which the methodology is incorporated, correctly assigned l's for 67 among 70 resonances for which l's had been determined experimentally. The other test for {sup 27}Al showed the applicability of the code as a preanalysis tool, even though such applicability is limited to a certain extent for light nuclides. The use of the code SUGGEL is expected to reduce the number of repeated runs of a fitting code such as SAMMY, thus reducing time and effort for the extraction of resonance parameters from measurements.

  19. Resonance strengths in the {sup 14}N(p,gamma){sup 15}O and {sup 15}N(p,alphagamma){sup 12}C reactions

    SciTech Connect

    Marta, Michele; Trompler, Erik; Bemmerer, Daniel; Beyer, Roland; Grosse, Eckart; Hannaske, Roland; Junghans, Arnd R.; Nair, Chithra; Schwengner, Ronald; Wagner, Andreas; Yakorev, Dmitry; Broggini, Carlo; Caciolli, Antonio; Erhard, Martin; Menegazzo, Roberto; Fueloep, Zsolt; Gyuerky, Gyoergy; Szuecs, Tamas; Vezzu, Simone

    2010-05-15

    The {sup 14}N(p,gamma){sup 15}O reaction is the slowest reaction of the carbon-nitrogen-oxygen cycle of hydrogen burning in stars. As a consequence, it determines the rate of the cycle. The {sup 15}N(p,alphagamma){sup 12}C reaction is frequently used in inverse kinematics for hydrogen depth profiling in materials. The {sup 14}N(p,gamma){sup 15}O and {sup 15}N(p,alphagamma){sup 12}C reactions have been studied simultaneously, using titanium nitride targets of natural isotopic composition and a proton beam. The strengths of the resonances at E{sub p} = 1058 keV in {sup 14}N(p,gamma){sup 15}O and at E{sub p} = 897 and 430 keV in {sup 15}N(p,alphagamma){sup 12}C have been determined with improved precision, relative to the well-known resonance at E{sub p} = 278 keV in {sup 14}N(p,gamma){sup 15}O. The new recommended values are omegagamma=0.353+-0.018, 362+-20, and 21.9+-1.0 eV for their respective strengths. In addition, the branching ratios for the decay of the E{sub p} = 1058 keV resonance in {sup 14}N(p,gamma){sup 15}O have been redetermined. The data reported here should facilitate future studies of off-resonant capture in the {sup 14}N(p,gamma){sup 15}O reaction that are needed for an improved R-matrix extrapolation of the cross section. In addition, the data on the 430 keV resonance in {sup 15}N(p,alphagamma){sup 12}C may be useful for hydrogen depth profiling.

  20. A Monte Carlo/simulated annealing algorithm for sequential resonance assignment in solid state NMR of uniformly labeled proteins with magic-angle spinning

    NASA Astrophysics Data System (ADS)

    Tycko, Robert; Hu, Kan-Nian

    2010-08-01

    We describe a computational approach to sequential resonance assignment in solid state NMR studies of uniformly 15N, 13C-labeled proteins with magic-angle spinning. As input, the algorithm uses only the protein sequence and lists of 15N/ 13C α crosspeaks from 2D NCACX and NCOCX spectra that include possible residue-type assignments of each crosspeak. Assignment of crosspeaks to specific residues is carried out by a Monte Carlo/simulated annealing algorithm, implemented in the program MC_ASSIGN1. The algorithm tolerates substantial ambiguity in residue-type assignments and coexistence of visible and invisible segments in the protein sequence. We use MC_ASSIGN1 and our own 2D spectra to replicate and extend the sequential assignments for uniformly-labeled HET-s(218-289) fibrils previously determined manually by Siemer et al. (J. Biomol. NMR, 34 (2006) 75-87) from a more extensive set of 2D and 3D spectra. Accurate assignments by MC_ASSIGN1 do not require data that are of exceptionally high quality. Use of MC_ASSIGN1 (and its extensions to other types of 2D and 3D data) is likely to alleviate many of the difficulties and uncertainties associated with manual resonance assignments in solid state NMR studies of uniformly labeled proteins, where spectral resolution and signal-to-noise are often sub-optimal.

  1. iHADAMAC: A complementary tool for sequential resonance assignment of globular and highly disordered proteins

    NASA Astrophysics Data System (ADS)

    Feuerstein, Sophie; Plevin, Michael J.; Willbold, Dieter; Brutscher, Bernhard

    2012-01-01

    An experiment, iHADAMAC, is presented that yields information on the amino-acid type of individual residues in a protein by editing the 1H- 15N correlations into seven different 2D spectra, each corresponding to a different class of amino-acid types. Amino-acid type discrimination is realized via a Hadamard encoding scheme based on four different spin manipulations as recently introduced in the context of the sequential HADAMAC experiment. Both sequential and intra-residue HADAMAC experiments yield highly complementary information that greatly facilitate resonance assignment of proteins with high frequency degeneracy, as demonstrated here for a 188-residue intrinsically disordered protein fragment of the hepatitis C virus protein NS5A.

  2. 15N electron nuclear double resonance of the primary donor cation radical P+.865 in reaction centers of Rhodopseudomonas sphaeroides: additional evidence for the dimer model.

    PubMed Central

    Lubitz, W; Isaacson, R A; Abresch, E C; Feher, G

    1984-01-01

    Four 15N hyperfine coupling constants, including signs, have been measured by electron nuclear double resonance (ENDOR) and electron nuclear nuclear triple resonance (TRIPLE) for the bacteriochlorophyll a radical cation, BChla+., in vitro and for the light-induced primary donor radical cation, P+.865, in reaction centers of Rhodopseudomonas sphaeroides R-26. A comparison of the data shows that the hyperfine coupling constants have the same sign in both radicals and are, on the average, smaller by a factor of 2 in P+.865. These results provide additional evidence that P+.865 is a bacteriochlorophyll dimer and are in contradiction with the monomer structure of P+.865 recently proposed by O'Malley and Babcock. The reduction factors of the individual 15N couplings, together with the evidence from proton ENDOR data and molecular orbital calculations, indicate a dimer structure in which only two rings (either I and I or III and III) of the bacteriochlorophyll macrocycles overlap. PMID:6096857

  3. Detection and classification of hyperfine-shifted 1H, 2H, and 15N resonances of the Rieske ferredoxin component of toluene 4-monooxygenase.

    PubMed

    Xia, B; Pikus, J D; Xia, W; McClay, K; Steffan, R J; Chae, Y K; Westler, W M; Markley, J L; Fox, B G

    1999-01-12

    T4MOC is a 12.3 kDa soluble Rieske ferredoxin that is obligately required for electron transfer between the oxidoreductase and diiron hydroxylase components of toluene 4-monooxygenase from Pseudomonas mendocina KR1. Our preliminary 1H NMR studies of oxidized and reduced T4MOC [Markley, J. L., Xia, B., Chae, Y. K., Cheng, H., Westler, W. M., Pikus, J. D., and Fox, B. G. (1996) in Protein Structure Function Relationships (Zaidi, Z., and Smith, D., Eds.) pp 135-146, Plenum Press, London] revealed the presence of hyperfine-shifted 1H resonances whose short relaxation times made it impractical to use nuclear Overhauser effect (NOE) measurements for assignment purposes. We report here the use of selective isotopic labeling to analyze the hyperfine-shifted 1H, 2H, and 15N signals from T4MOC. Selective deuteration led to identification of signals from the four Hbeta atoms of cluster ligands C45 and C64 in the oxidized and reduced forms of T4MOC. In the reduced state, the Curie temperature dependence of the Hbeta protons corresponded to that predicted from the simple vector spin-coupling model for nuclei associated with the localized ferric site. The signal at 25.5 ppm in the 1H spectrum of reduced T4MOC was assigned on the basis of selective 2H labeling to the His Hepsilon1 atom of one of the cluster ligands (H47 or H67). This assignment was corroborated by a one bond 1H-13C correlation (at 25.39 ppm 1H and 136.11 ppm 13C) observed in spectra of [U-13C]T4MOC with a 1H-13C coupling constant of approximately 192 Hz. The carbon chemical shift and one bond coupling constant are those expected for 1Hepsilon1-13Cepsilon1 in the imidazolium ring of histidine and are inconsistent with values expected for cysteine 1Halpha-13Calpha. The His Hepsilon1 proton exhibited weak Curie temperature dependence from 283 to 303 K, contrary to the anti-Curie temperature dependence predicted from the spin coupling model for nuclei associated with the localized ferrous site. A 1H peak at -12.3 ppm

  4. Efficient Resonance Assignment of Proteins in MAS NMR by Simultaneous Intra- and Inter-residue 3D Correlation Spectroscopy

    PubMed Central

    Daviso, Eugenio; Eddy, Matthew T.; Andreas, Loren B.; Griffin, Robert G.; Herzfeld, Judith

    2013-01-01

    Resonance assignment is the first step in NMR structure determination. For magic angle spinning NMR, this is typically achieved with a set of heteronuclear correlation experiments (NCaCX, NCOCX, CONCa) that utilize SPECIFIC-CP 15N-13C transfers. However, the SPECIFIC-CP transfer efficiency is often compromised by molecular dynamics and probe performance. Here we show that one-bond ZF-TEDOR 15N-13C transfers provide simultaneous NCO and NCa transfers with at least as much sensitivity as SPECIFIC-CP for some non-crystalline samples. Furthermore, a 3D TEDOR-CC experiment provides heteronuclear sidechains correlations and robustness with respect to proton decoupling and radiofrequency power instabilities. We demonstrate transfer efficiencies and connectivities by application of 3D ZF-TEDOR-DARR to a model microcrystalline protein, GB1, and a less ideal system, GvpA in intact gas vesicles. PMID:23334347

  5. Fermentation and Cost-Effective 13C/15N Labeling of the Nonribosomal Peptide Gramicidin S for Nuclear Magnetic Resonance Structure Analysis

    PubMed Central

    Berditsch, Marina; Afonin, Sergii; Steineker, Anna; Orel, Nataliia; Jakovkin, Igor; Weber, Christian

    2015-01-01

    Gramicidin S (GS) is a nonribosomally synthesized decapeptide from Aneurinibacillus migulanus. Its pronounced antibiotic activity is attributed to amphiphilic structure and enables GS interaction with bacterial membranes. Despite its medical use for over 70 years, the peptide-lipid interactions of GS and its molecular mechanism of action are still not fully understood. Therefore, a comprehensive structural analysis of isotope-labeled GS needs to be performed in its biologically relevant membrane-bound state, using advanced solid-state nuclear magnetic resonance (NMR) spectroscopy. Here, we describe an efficient method for producing the uniformly 13C/15N-labeled peptide in a minimal medium supplemented by selected amino acids. As GS is an intracellular product of A. migulanus, we characterized the producer strain DSM 5759 (rough-convex phenotype) and examined its biosynthetic activity in terms of absolute and biomass-dependent peptide accumulation. We found that the addition of either arginine or ornithine increases the yield only at very high supplementing concentrations (1% and 0.4%, respectively) of these expensive 13C/15N-labeled amino acids. The most cost-effective production of 13C/15N-GS, giving up to 90 mg per gram of dry cell weight, was achieved in a minimal medium containing 1% 13C-glycerol and 0.5% 15N-ammonium sulfate, supplemented with only 0.025% of 13C/15N-phenylalanine. The 100% efficiency of labeling is corroborated by mass spectrometry and preliminary solid-state NMR structure analysis of the labeled peptide in the membrane-bound state. PMID:25795666

  6. Resonance assignments and secondary structure of a phytocystatin from Sesamum indicum.

    PubMed

    Hu, Yu-Jun; Irene, Deli; Lo, Chi-Jen; Cai, Yong-Liang; Tzen, T-C; Lin, Ta-Hsien; Chyan, Chia-Lin

    2015-10-01

    A cDNA encoding a cysteine protease inhibitor, cystatin was cloned from sesame (Sesamum indicum L.) seed. This clone was constructed into an expression vector and expressed in E. coli and purified to homogeneous. The recombinant sesame cystatin (SiCYS) showed effectively inhibitory activity toward C1 cysteine proteases. In order to unravel its inhibitory action from structural point of view, multidimensional heteronuclear NMR techniques were used to characterize the structure of SiCYS. The full (1)H, (15)N, and (13)C resonances of SiCYS were assigned. The secondary structure of SiCYS was identified by using the assigned chemical shifts of (1)H(α), (13)C(α), (13)C(β), and (13)CO through the consensus chemical shift index (CSI). The results of CSI analysis of SiCYS suggest eight β-strands (residues 33-46, 51-61, 63-75, 80-87, 150-155, 157-169, 172-183, and 192-195) and two α-helices (residues 16-30, and 120-135).

  7. NMR resonance assignments of the lantibiotic immunity protein NisI from Lactococcus lactis.

    PubMed

    Hacker, Carolin; Christ, Nina Alexandra; Duchardt-Ferner, Elke; Korn, Sophie; Berninger, Lucija; Kötter, Peter; Entian, Karl-Dieter; Wöhnert, Jens

    2015-10-01

    The lantibiotic nisin is a small antimicrobial peptide which acts against a wide range of Gram-positive bacteria. Nisin-producing Lactococcus lactis strains express four genes for self-protection against their own antimicrobial compound. This immunity system consists of the lipoprotein NisI and the ABC transporter NisFEG. NisI is attached to the outside of the cytoplasmic membrane via a covalently linked diacylglycerol anchor. Both the lipoprotein and the ABC transporter are needed for full immunity but the exact immunity mechanism is still unclear. To gain insights into the highly specific immunity mechanism of nisin producing strains on a structural level we present here the backbone resonance assignment of NisI (25.8 kDa) as well as the virtually complete (1)H,(15)N,(13)C chemical shift assignments for the isolated 12.7 kDa N-terminal and 14.6 kDa C-terminal domains of NisI.

  8. Resonance assignment of PsbP: an extrinsic protein from photosystem II of Spinacia oleracea.

    PubMed

    Rathner, Adriana; Chandra, Kousik; Rathner, Petr; Horničáková, Michaela; Schlagnitweit, Judith; Kohoutová, Jaroslava; Ettrich, Rüdiger; Müller, Norbert

    2015-10-01

    PsbP (23 kDa) is an extrinsic eukaryotic protein of photosystem II found in the thylakoid membrane of higher plants and green algae. It has been proven to be indispensable for proper functioning of the oxygen evolving complex. By interaction with other extrinsic proteins (PsbQ, PsbO and PsbR), it modulates the concentration of two cofactors of the water splitting reaction, Ca(2+) and Cl(-). The crystallographic structure of PsbP from Spinacia oleracea lacks the N-terminal part as well as two inner regions which were modelled as loops. Those unresolved parts are believed to be functionally crucial for the binding of PsbP to the thylakoid membrane. In this NMR study we report (1)H, (15)N and (13)C resonance assignments of the backbone and side chain atoms of the PsbP protein. Based on these data, an estimate of the secondary structure has been made. The structural motifs found fit the resolved parts of the crystallographic structure very well. In addition, the complete assignment set provides preliminary insight into the dynamic regions.

  9. Resonance assignments and secondary structure of a phytocystatin from Ananas comosus.

    PubMed

    Irene, Deli; Chen, Bo-Jiun; Lo, Si-Hung; Liu, Ting-Hang; Tzen, Jason T-C; Chyan, Chia-Lin

    2012-04-01

    A cDNA encoding a cysteine protease inhibitor, cystatin was cloned from pineapple (Ananas comosus L.) stem. This clone was constructed into an expression vector and expressed in E. coli and purified to homogeneous. The recombinant pineapple cystatins (AcCYS) showed effectively inhibitory activity toward cysteine proteases including papain, bromelain, and cathepsin B. In order to unravel its inhibitory action from structural point of view, multidimensional heteronuclear NMR techniques were used to characterize the structure of AcCYS. The full (1)H, (15)N, and (13)C resonance assignments of AcCYS were determined. The secondary structure of AcCYS was identified by using the assigned chemical shift of (1)Hα, (13)Cα, (13)Cβ, and (13)CO through the consensus chemical shift index (CSI). The results of CSI analysis suggest 5 β-strands (residues 45-47, 84-91, 94-104, 106-117, and 123-130) and one α-helix (residues 55-73).

  10. Optimization of amino acid type-specific 13C and 15N labeling for the backbone assignment of membrane proteins by solution- and solid-state NMR with the UPLABEL algorithm.

    PubMed

    Hefke, Frederik; Bagaria, Anurag; Reckel, Sina; Ullrich, Sandra Johanna; Dötsch, Volker; Glaubitz, Clemens; Güntert, Peter

    2011-02-01

    We present a computational method for finding optimal labeling patterns for the backbone assignment of membrane proteins and other large proteins that cannot be assigned by conventional strategies. Following the approach of Kainosho and Tsuji (Biochemistry 21:6273-6279 (1982)), types of amino acids are labeled with (13)C or/and (15)N such that cross peaks between (13)CO(i - 1) and (15)NH(i) result only for pairs of sequentially adjacent amino acids of which the first is labeled with (13)C and the second with (15)N. In this way, unambiguous sequence-specific assignments can be obtained for unique pairs of amino acids that occur exactly once in the sequence of the protein. To be practical, it is crucial to limit the number of differently labeled protein samples that have to be prepared while obtaining an optimal extent of labeled unique amino acid pairs. Our computer algorithm UPLABEL for optimal unique pair labeling, implemented in the program CYANA and in a standalone program, and also available through a web portal, uses combinatorial optimization to find for a given amino acid sequence labeling patterns that maximize the number of unique pair assignments with a minimal number of differently labeled protein samples. Various auxiliary conditions, including labeled amino acid availability and price, previously known partial assignments, and sequence regions of particular interest can be taken into account when determining optimal amino acid type-specific labeling patterns. The method is illustrated for the assignment of the human G-protein coupled receptor bradykinin B2 (B(2)R) and applied as a starting point for the backbone assignment of the membrane protein proteorhodopsin.

  11. A triple resonance hyperfine sublevel correlation experiment for assignment of electron-nuclear double resonance lines

    NASA Astrophysics Data System (ADS)

    Potapov, Alexey; Epel, Boris; Goldfarb, Daniella

    2008-02-01

    A new, triple resonance, pulse electron paramagnetic resonance (EPR) sequence is described. It provides spin links between forbidden electron spin transitions (ΔMS=±1, ΔMI≠0) and allowed nuclear spin transitions (ΔMI=±1), thus, facilitating the assignment of nuclear frequencies to their respective electron spin manifolds and paramagnetic centers. It also yields the relative signs of the hyperfine couplings of the different nuclei. The technique is based on the combination of electron-nuclear double resonance (ENDOR) and electron-electron double resonance (ELDOR)-detected NMR experiments in a way similar to the TRIPLE experiment. The feasibility and the information content of the method are demonstrated first on a single crystal of Cu-doped L-histidine and then on a frozen solution of a Cu-histidine complex.

  12. 1H, 15N and 13C assignments of domain 5 of Dictyostelium discoideum gelation factor (ABP-120) in its native and 8M urea-denatured states.

    PubMed

    Hsu, Shang-Te Danny; Cabrita, Lisa D; Christodoulou, John; Dobson, Christopher M

    2009-06-01

    The gelation factor from Dictyostelium discoideum (ABP-120) is an actin binding protein consisting of six immunoglobulin (Ig) domains in the C-terminal rod domain. We have recently used the pair of domains 5 and 6 of ABP-120 as a model system for studying multi-domain nascent chain folding on the ribosome. Here we present the NMR assignments of domain 5 in its native and 8M urea-denatured states.

  13. Backbone 1H, 13C and 15N assignments of YibK and avariant containing a unique cysteine residue at C-terminus in 8 M urea-denatured states [corrected].

    PubMed

    Hsieh, Shu-Ju Micky; Mallam, Anna L; Jackson, Sophie E; Hsu, Shang-Te Danny

    2014-10-01

    YibK is a tRNA methyltransferase from Haemophilus influenzae, which forms a stable homodimer in solution and contains a deep trefoil 31 knot encompassing the C-terminal helix that threads through a long loop. It has been a model system for investigating knotted protein folding pathways. Recent data have shown that the polypeptide chain of YibK remains loosely knotted under highly denaturing conditions. Here, we report (1)H, (13)C and (15)N chemical shift assignments for YibK and its variant in the presence of 8 M urea. This work forms the basis for further analysis using NMR techniques such as paramagnetic relaxation enhancement, residual dipolar couplings and spin-relaxation dynamics analysis.

  14. Backbone resonance assignments for the PHD-Bromo dual-domain of the human chromatin reader TRIM24.

    PubMed

    Walser, Reto; Renshaw, Jonathan; Milbradt, Alexander G

    2016-04-01

    Plant homeodomains (PHD) and Bromo domains are both chromatin reader domains that recognise histone methylation degree and acetylation state, respectively. The tripartite motif protein TRIM24 is a multidomain protein carrying a PHD-Bromo motif at its C-terminus, through which it is able to bind to histone 3 (H3) N-terminal tails with a specific modification pattern, namely unmethylated at K4 and acetylated at K23 (H3-K4me0K23ac). Here we report the 1H, 13C and 15N backbone resonance assignment of this 23 kDa motif, which we have obtained by heteronuclear multidimensional NMR spectroscopy. Furthermore we show that the secondary Cα and Cβ chemical shifts are in good agreement with a previously published crystal structure.

  15. An automated framework for NMR resonance assignment through simultaneous slice picking and spin system forming.

    PubMed

    Abbas, Ahmed; Guo, Xianrong; Jing, Bing-Yi; Gao, Xin

    2014-06-01

    Despite significant advances in automated nuclear magnetic resonance-based protein structure determination, the high numbers of false positives and false negatives among the peaks selected by fully automated methods remain a problem. These false positives and negatives impair the performance of resonance assignment methods. One of the main reasons for this problem is that the computational research community often considers peak picking and resonance assignment to be two separate problems, whereas spectroscopists use expert knowledge to pick peaks and assign their resonances at the same time. We propose a novel framework that simultaneously conducts slice picking and spin system forming, an essential step in resonance assignment. Our framework then employs a genetic algorithm, directed by both connectivity information and amino acid typing information from the spin systems, to assign the spin systems to residues. The inputs to our framework can be as few as two commonly used spectra, i.e., CBCA(CO)NH and HNCACB. Different from the existing peak picking and resonance assignment methods that treat peaks as the units, our method is based on 'slices', which are one-dimensional vectors in three-dimensional spectra that correspond to certain ([Formula: see text]) values. Experimental results on both benchmark simulated data sets and four real protein data sets demonstrate that our method significantly outperforms the state-of-the-art methods while using a less number of spectra than those methods. Our method is freely available at http://sfb.kaust.edu.sa/Pages/Software.aspx.

  16. Near-complete backbone resonance assignments of acid-denatured human cytochrome c in dimethylsulfoxide: a prelude to studying interactions with phospholipids.

    PubMed

    Karsisiotis, Andreas Ioannis; Deacon, Oliver M; Macdonald, Colin; Blumenschein, Tharin M A; Moore, Geoffrey R; Worrall, Jonathan A R

    2017-03-04

    Human cytochrome c plays a central role in the mitochondrial electron transfer chain and in the intrinsic apoptosis pathway. Through the interaction with the phospholipid cardiolipin, cytochrome c triggers release of pro-apoptotic factors, including itself, from the mitochondrion into the cytosol of cells undergoing apoptosis. The cytochrome c/cardiolipin complex has been extensively studied through various spectroscopies, most recently with high-field solution and solid-state NMR spectroscopies, but there is no agreement between the various studies on key structural features of cytochrome c in its complex with cardiolipin. In the present study, we report backbone (1)H, (13)C, (15)N resonance assignments of acid-denatured human cytochrome c in the aprotic solvent dimethylsulfoxide. These have led to the assignment of a reference 2D (1)H-(15)N HSQC spectrum in which out of the 99 non-proline residues 87% of the backbone amides are assigned. These assignments are being used in an interrupted H/D exchange strategy to map the binding site of cardiolipin on human cytochrome c.

  17. Complete Assignment of (1)H-NMR Resonances of the King Cobra Neurotoxin CM-11.

    PubMed

    Pang, Yu-Xi; Liu, Wei-Dong; Liu, Ai-Zhuo; Pei, Feng-Kui

    1997-01-01

    The king cobra (Ophiophagus Hannah) neurotoxin CM-Il is long-chain peptide with 72 amino acid residues. Its complete assignment of (1)H-NMR resonances was obtained using various 2D-NMR technologies, including DQF-COSY, clean-TOCSY and NOESY.

  18. A program for semi-automatic sequential resonance assignments in protein 1H nuclear magnetic resonance spectra

    NASA Astrophysics Data System (ADS)

    Billeter, M.; Basus, V. J.; Kuntz, I. D.

    A new approach to the sequential resonance assignment of protein 1H NMR spectra based on a computer program is presented. Two main underlying concepts were used in the design of this program. First, it considers at any time all possible assignments that are consistent with the currently available data. If new information is added then assignments that have become inconsistent are eliminated. Second, the process of the assignment is split into formal steps that follow strictly from the available data and steps that involve the interpretation of ambiguous NMR data. The first kind of step is safe in the sense that it never leads to false assignments provided that the input does not contain any error; these steps are executed automatically by the program when the input files are read and whenever new data have been entered interactively. The second kind of step is left to the user: An interactive dialog provides detailed information on the current situation of the assignment and indicates what kind of new data would be most promising for further assignment. The user then provides new data to the program and restarts the automatic part which will attempt to draw logical conclusions from the joint use of the new data and the earlier available information and will eliminate assignments that have become inconsistent. Results of test problems using simulated NMR data for proteins consisting of up to 99 residues as well as the application of the program to obtain the complete assignment of α-bungarotoxin, a 74-residue snake neurotoxin, are reported.

  19. sup 13 C and sup 15 N nuclear magnetic resonance evidence of the ionization state of substrates bound to bovine dihydrofolate reductase

    SciTech Connect

    Selinsky, B.S.; Perlman, M.E.; London, R.E. ); Unkefer, C.J. ); Mitchell, J. ); Blakley, R.L. Univ. of Tennessee, Memphis )

    1990-02-06

    The state of protonation of substrates bound to mammalian dihydrofolate reductase (DHFR) has significance for the mechanism of catalysis. To investigate this, dihydrofolate and dihydropteroylpentaglutamate have been synthesized with {sup 15}N enrichment at N-5. {sup 15}N NMR studies have been performed on the binary complexes formed by bovine DHFR with these compounds and with (5-{sup 15}N)dihydrobiopterin. The results indicate that there is no protonation at N-5 in the binary complexes, and this was confirmed by {sup 13}C NMR studies with folate and dihydrofolate synthesized with {sup 13}C enrichment at C-6. The chemical shift displacements produced by complex formation are in the same direction as those which result from deprotonation of the N-3/C-4-O amide group and are consistent with at least partial loss of the proton from N-3. This would be possible if, as crystallographic data indicate, there is interaction of N-3 and the 2-amino group of the bound ligands with the carboxylate of the active site glutamate residue (Glu{sup 30}).

  20. Proton nuclear magnetic resonance studies on the variant-3 neurotoxin from Centruroides sculpturatus Ewing: Sequential assignment of resonances

    SciTech Connect

    Nettesheim, D.G.; Klevit, R.E.; Drobny, G.; Watt, D.D.; Krishna, N.R. )

    1989-02-21

    The authors report the sequential assignment of resonances to specific residues in the proton nuclear magnetic resonance spectrum of the variant-3 neurotoxin from the scorpion Centruroides sculpturatus Ewing (range southwestern U.S.A.). A combination of two-dimensional NMR experiments such as 2D-COSY, 2D-NOESY, and single- and double-RELAY coherence transfer spectroscopy has been employed on samples of the protein dissolved in D{sub 2}O and in H{sub 2}O for assignment purposes. These studies provide a basis for the determination of the solution-phase conformation of this protein and for undertaking detailed structure-function studies of these neurotoxins that modulate the flow of sodium current by binding to the sodium channels of excitable membranes.

  1. Backbone resonance assignments of an artificially engineered TEM-1/PSE-4 Class A β-lactamase chimera.

    PubMed

    Morin, Sébastien; Clouthier, Christopher M; Gobeil, Sophie; Pelletier, Joelle N; Gagné, Stéphane M

    2010-10-01

    The rapid evolution of Class A β-lactamases, which procure resistance to an increasingly broad panel of β-lactam antibiotics, underscores the urgency to better understand the relation between their sequence variation and their structural and functional features. To date, more than 300 clinically-relevant β-lactamase variants have been reported, and this number continues to increase. With the aim of obtaining insights into the evolutionary potential of β-lactamases, an artificially engineered, catalytically active chimera of the Class A TEM-1 and PSE-4 β-lactamases is under study by kinetics and NMR. Here we report the (1)H, (13)C and (15)N backbone resonance assignments for the 30 kDa chimera cTEM-17m. Despite its high molecular weight, the data provide evidence that this artificially-evolved chimeric enzyme is well folded. The hydrolytic activity of cTEM-17m was determined using the chromogenic substrate CENTA, with K (M) = 160 ± 35 μM and k (cat) = 20 ± 4 s(-1), which is in the same range as the values for TEM-1 and PSE-4 β-lactamases.

  2. Spin and parity assignments for {sup 94,95}Mo neutron resonances

    SciTech Connect

    Sheets, S. A.; Agvaanluvsan, U.; Becker, J. A.; Parker, W. E.; Wu, C. Y.; Becvar, F.; Krticka, M.; Bredeweg, T. A.; Haight, R. C.; Jandel, M.; O'Donnell, J. M.; Reifarth, R.; Rundberg, R. S.; Ullmann, J. L.; Vieira, D. J.; Wouters, J. M.; Wilhelmy, J. B.; Mitchell, G. E.; Sharapov, E. I.; Tomandl, I.

    2007-12-15

    The {gamma} rays following the {sup 94,95}Mo(n,{gamma}) reactions were measured as a function of incident neutron energy by the time-of-flight method with the DANCE (Detector for Advanced Neutron Capture Experiments) array of 160 BaF{sub 2} scintillation detectors at the Los Alamos Neutron Science Center. The targets were enriched samples: 91.59% {sup 94}Mo and 96.47% {sup 95}Mo. The {gamma}-ray multiplicities and energy spectra for different multiplicities were measured in s- and p-wave resonances up to E{sub n}=10 keV for {sup 94}Mo and up to E{sub n}=2 keV for {sup 95}Mo. Definite spins and parities were assigned in {sup 96}Mo for about 60% of the resonances, and tentative spins and parities were assigned for the remaining resonances. In {sup 95}Mo the parities were determined for the observed resonances, confirming previously known assignments.

  3. A set of 4D NMR experiments of enhanced resolution for easy resonance assignment in proteins

    NASA Astrophysics Data System (ADS)

    Zawadzka-Kazimierczuk, Anna; Kazimierczuk, Krzysztof; Koźmiński, Wiktor

    2010-01-01

    This paper presents examples of techniques based on the principle of random sampling that allows acquisition of NMR spectra featuring extraordinary resolution. This is due to increased dimensionality and maximum evolution time reached. The acquired spectra of CsPin protein and maltose binding protein were analyzed statistically with the aim to evaluate each technique. The results presented include exemplary spectral cross-sections. The spectral data provided by the proposed techniques allow easy assignment of backbone and side-chain resonances.

  4. Backbone, side chain and heme resonance assignments of cytochrome OmcF from Geobacter sulfurreducens.

    PubMed

    Dantas, Joana M; Silva E Sousa, Marta; Salgueiro, Carlos A; Bruix, Marta

    2015-10-01

    Gene knockout studies on Geobacter sulfurreducens (Gs) cells showed that the outer membrane cytochrome OmcF is involved in respiratory pathways leading to the extracellular reduction of Fe(III) citrate and U(VI) oxide. In addition, microarray analysis of OmcF-deficient mutant versus the wild-type strain revealed that many of the genes with decreased transcript level were those whose expression is upregulated in cells grown with a graphite electrode as electron acceptor. This suggests that OmcF also regulates the electron transfer to electrode surfaces and the concomitant electrical current production by Gs in microbial fuel cells. Extracellular electron transfer processes (EET) constitute nowadays the foundations to develop biotechnological applications in biofuel production, bioremediation and bioenergy. Therefore, the structural characterization of OmcF is a fundamental step to understand the mechanisms underlying EET. Here, we report the complete assignment of the heme proton signals together with (1)H, (13)C and (15)N backbone and side chain assignments of the OmcF, excluding the hydrophobic residues of the N-terminal predicted lipid anchor.

  5. Fermi resonance in CO2: Mode assignment and quantum nuclear effects from first principles molecular dynamics.

    PubMed

    Basire, Marie; Mouhat, Félix; Fraux, Guillaume; Bordage, Amélie; Hazemann, Jean-Louis; Louvel, Marion; Spezia, Riccardo; Bonella, Sara; Vuilleumier, Rodolphe

    2017-04-07

    Vibrational spectroscopy is a fundamental tool to investigate local atomic arrangements and the effect of the environment, provided that the spectral features can be correctly assigned. This can be challenging in experiments and simulations when double peaks are present because they can have different origins. Fermi dyads are a common class of such doublets, stemming from the resonance of the fundamental excitation of a mode with the overtone of another. We present a new, efficient approach to unambiguously characterize Fermi resonances in density functional theory (DFT) based simulations of condensed phase systems. With it, the spectral features can be assigned and the two resonating modes identified. We also show how data from DFT simulations employing classical nuclear dynamics can be post-processed and combined with a perturbative quantum treatment at a finite temperature to include analytically thermal quantum nuclear effects. The inclusion of these effects is crucial to correct some of the qualitative failures of the Newtonian dynamics simulations at a low temperature such as, in particular, the behavior of the frequency splitting of the Fermi dyad. We show, by comparing with experimental data for the paradigmatic case of supercritical CO2, that these thermal quantum effects can be substantial even at ambient conditions and that our scheme provides an accurate and computationally convenient approach to account for them.

  6. Backbone resonance assignments of the micro-RNA precursor binding region of human TRBP.

    PubMed

    Benoit, Matthieu P M H; Plevin, Michael J

    2013-10-01

    TAR-RNA binding protein (TRBP) is a multidomain human protein involved in micro-RNA (miRNA) biogenesis. TRBP is a component of both the Dicer complex, which processes precursor miRNAs, and the RNA-induced silencing complex-loading complex. In addition, TRBP is implicated in the human immunodeficiency virus replication cycle and interferon-protein kinase R activity. TRBP contains 3 double-stranded RNA binding domains the first two of which have been shown to interact with miRNA precursors. Here we present the backbone resonance assignments and secondary structure of residues 19-228 of human TRBP2.

  7. Structure and backbone dynamics of vanadate-bound PRL-3: comparison of 15N nuclear magnetic resonance relaxation profiles of free and vanadate-bound PRL-3.

    PubMed

    Jeong, Ki-Woong; Kang, Dong-Il; Lee, Eunjung; Shin, Areum; Jin, Bonghwan; Park, Young-Guen; Lee, Chung-Kyoung; Kim, Eun-Hee; Jeon, Young Ho; Kim, Eunice Eunkyeong; Kim, Yangmee

    2014-07-29

    Phosphatases of regenerating liver (PRLs) constitute a novel class of small, prenylated phosphatases with oncogenic activity. PRL-3 is particularly important in cancer metastasis and represents a potential therapeutic target. The flexibility of the WPD loop as well as the P-loop of protein tyrosine phosphatases is closely related to their catalytic activity. Using nuclear magnetic resonance spectroscopy, we studied the structure of vanadate-bound PRL-3, which was generated by addition of sodium orthovanadate to PRL-3. The WPD loop of free PRL-3 extended outside of the active site, forming an open conformation, whereas that of vanadate-bound PRL-3 was directed into the active site by a large movement, resulting in a closed conformation. We suggest that vanadate binding induced structural changes in the WPD loop, P-loop, helices α4-α6, and the polybasic region. Compared to free PRL-3, vanadate-bound PRL-3 has a longer α4 helix, where the catalytic R110 residue coordinates with vanadate in the active site. In addition, the hydrophobic cavity formed by helices α4-α6 with a depth of 14-15 Å can accommodate a farnesyl chain at the truncated prenylation motif of PRL-3, i.e., from R169 to M173. Conformational exchange data suggested that the WPD loop moves between open and closed conformations with a closing rate constant k(close) of 7 s(-1). This intrinsic loop flexibility of PRL-3 may be related to their catalytic rate and may play a role in substrate recognition.

  8. Spin and Parity Assignment of Neutron Resonances using Gamma-ray Multiplicity

    SciTech Connect

    Agvaanluvsan, U.; Mitchell, G. E.; Baramsai, B.; Chyzh, A.; Walker, C.; Dashdorj, D.; Becker, J. A.; Parker, W. E.; Sleaford, B.; Wu, C. Y.; Bredeweg, T. A.; Couture, A.; Haight, R. C.; Jandel, M.; Rundberg, R. S.; Ullmann, J. L.; Vieira, D. J.; Wouters, J. M.; Krticka, M.; Becvar, F.

    2009-03-31

    Decay gamma rays following neutron capture on various isotopes are collected by the Detector for Advanced Neutron Capture Experiments (DANCE) array, which is located at flight path 14 at the Lujan Neutron Scattering Center at Los Alamos National Laboratory. The high segmentation (160 detectors) and close packing of the detector array enable gamma-ray multiplicity measurements. The calorimetric properties of the DANCE array coupled with the neutron time-of-flight technique enables one to gate on a specific resonance of a given isotope in the time-of-flight spectrum and obtain the summed energy spectrum for that isotope. The singles gamma-ray spectrum for each multiplicity can be separated by their DANCE cluster multiplicity. The multiplicity distribution contains the signatures of spin and parity of the capture state. Under suitable circumstances where the difference between spins of the initial (capture) and final (ground) state is large enough, the signatures in the multiplicity distribution can be used in improving the spin assignment of the initial state. The spin assignment is applied with varying degree of success to difference isotopes and description of this application for {sup 95}Mo, {sup 151,153}Eu, and {sup 155,157}Gd is reviewed briefly.

  9. Selective excitation enables assignment of proton resonances and (1)H-(1)H distance measurement in ultrafast magic angle spinning solid state NMR spectroscopy.

    PubMed

    Zhang, Rongchun; Ramamoorthy, Ayyalusamy

    2015-07-21

    Remarkable developments in ultrafast magic angle spinning (MAS) solid-state NMR spectroscopy enabled proton-based high-resolution multidimensional experiments on solids. To fully utilize the benefits rendered by proton-based ultrafast MAS experiments, assignment of (1)H resonances becomes absolutely necessary. Herein, we propose an approach to identify different proton peaks by using dipolar-coupled heteronuclei such as (13)C or (15)N. In this method, after the initial preparation of proton magnetization and cross-polarization to (13)C nuclei, transverse magnetization of desired (13)C nuclei is selectively prepared by using DANTE (Delays Alternating with Nutations for Tailored Excitation) sequence and then, it is transferred to bonded protons with a short-contact-time cross polarization. Our experimental results demonstrate that protons bonded to specific (13)C atoms can be identified and overlapping proton peaks can also be assigned. In contrast to the regular 2D HETCOR experiment, only a few 1D experiments are required for the complete assignment of peaks in the proton spectrum. Furthermore, the finite-pulse radio frequency driven recoupling sequence could be incorporated right after the selection of specific proton signals to monitor the intensity buildup for other proton signals. This enables the extraction of (1)H-(1)H distances between different pairs of protons. Therefore, we believe that the proposed method will greatly aid in fast assignment of peaks in proton spectra and will be useful in the development of proton-based multi-dimensional solid-state NMR experiments to study atomic-level resolution structure and dynamics of solids.

  10. Selective excitation enables assignment of proton resonances and 1H-1H distance measurement in ultrafast magic angle spinning solid state NMR spectroscopy

    NASA Astrophysics Data System (ADS)

    Zhang, Rongchun; Ramamoorthy, Ayyalusamy

    2015-07-01

    Remarkable developments in ultrafast magic angle spinning (MAS) solid-state NMR spectroscopy enabled proton-based high-resolution multidimensional experiments on solids. To fully utilize the benefits rendered by proton-based ultrafast MAS experiments, assignment of 1H resonances becomes absolutely necessary. Herein, we propose an approach to identify different proton peaks by using dipolar-coupled heteronuclei such as 13C or 15N. In this method, after the initial preparation of proton magnetization and cross-polarization to 13C nuclei, transverse magnetization of desired 13C nuclei is selectively prepared by using DANTE (Delays Alternating with Nutations for Tailored Excitation) sequence and then, it is transferred to bonded protons with a short-contact-time cross polarization. Our experimental results demonstrate that protons bonded to specific 13C atoms can be identified and overlapping proton peaks can also be assigned. In contrast to the regular 2D HETCOR experiment, only a few 1D experiments are required for the complete assignment of peaks in the proton spectrum. Furthermore, the finite-pulse radio frequency driven recoupling sequence could be incorporated right after the selection of specific proton signals to monitor the intensity buildup for other proton signals. This enables the extraction of 1H-1H distances between different pairs of protons. Therefore, we believe that the proposed method will greatly aid in fast assignment of peaks in proton spectra and will be useful in the development of proton-based multi-dimensional solid-state NMR experiments to study atomic-level resolution structure and dynamics of solids.

  11. Selective excitation enables assignment of proton resonances and {sup 1}H-{sup 1}H distance measurement in ultrafast magic angle spinning solid state NMR spectroscopy

    SciTech Connect

    Zhang, Rongchun; Ramamoorthy, Ayyalusamy

    2015-07-21

    Remarkable developments in ultrafast magic angle spinning (MAS) solid-state NMR spectroscopy enabled proton-based high-resolution multidimensional experiments on solids. To fully utilize the benefits rendered by proton-based ultrafast MAS experiments, assignment of {sup 1}H resonances becomes absolutely necessary. Herein, we propose an approach to identify different proton peaks by using dipolar-coupled heteronuclei such as {sup 13}C or {sup 15}N. In this method, after the initial preparation of proton magnetization and cross-polarization to {sup 13}C nuclei, transverse magnetization of desired {sup 13}C nuclei is selectively prepared by using DANTE (Delays Alternating with Nutations for Tailored Excitation) sequence and then, it is transferred to bonded protons with a short-contact-time cross polarization. Our experimental results demonstrate that protons bonded to specific {sup 13}C atoms can be identified and overlapping proton peaks can also be assigned. In contrast to the regular 2D HETCOR experiment, only a few 1D experiments are required for the complete assignment of peaks in the proton spectrum. Furthermore, the finite-pulse radio frequency driven recoupling sequence could be incorporated right after the selection of specific proton signals to monitor the intensity buildup for other proton signals. This enables the extraction of {sup 1}H-{sup 1}H distances between different pairs of protons. Therefore, we believe that the proposed method will greatly aid in fast assignment of peaks in proton spectra and will be useful in the development of proton-based multi-dimensional solid-state NMR experiments to study atomic-level resolution structure and dynamics of solids.

  12. Backbone and side–chain heteronuclear resonance assignments and hyperfine NMR shifts in horse cytochrome c

    PubMed Central

    Liu, Weixia; Rumbley, Jon; Englander, S. Walter; Wand, A. Joshua

    2003-01-01

    The [H26N, H33N] mutant of horse heart cytochrome c was expressed in E. coli during growth on isotopically enriched minimal media. Complete resonance assignments of both the diamagnetic reduced (spin zero) and paramagnetic oxidized (spin ½) states of the protein were obtained using standard triple resonance and total correlation spectroscopy using the previously determined 1H chemical shifts of the wild-type protein as a guide. The correspondence of chemical shifts between the wild type and the mutant protein is excellent, indicating that they have nearly identical structures. The expanded library of chemical shifts for both redox states in both proteins allowed the refinement of the electron spin g-tensor of the oxidized states. The g-tensors of the oxidized states of the wild-type and [H26N, H33N] mutant proteins are closely similar, indicating that the subtle details of the ligand fields are nearly identical. The refined g-tensors were then used to probe for redox-dependent structure change in the two proteins. PMID:12931009

  13. FLAMEnGO 2.0: an enhanced fuzzy logic algorithm for structure-based assignment of methyl group resonances.

    PubMed

    Chao, Fa-An; Kim, Jonggul; Xia, Youlin; Milligan, Michael; Rowe, Nancy; Veglia, Gianluigi

    2014-08-01

    We present an enhanced version of the FLAMEnGO (Fuzzy Logic Assignment of Methyl Group) software, a structure-based method to assign methyl group resonances in large proteins. FLAMEnGO utilizes a fuzzy logic algorithm coupled with Monte Carlo sampling to obtain a probability-based assignment of the methyl group resonances. As an input, FLAMEnGO requires either the protein X-ray structure or an NMR structural ensemble including data such as methyl-methyl NOESY, paramagnetic relaxation enhancement (PRE), methine-methyl TOCSY data. Version 2.0 of this software (FLAMEnGO 2.0) has a user-friendly graphic interface and presents improved modules that enable the input of partial assignments and additional NMR restraints. We tested the performance of FLAMEnGO 2.0 on maltose binding protein (MBP) as well as the C-subunit of the cAMP-dependent protein kinase A (PKA-C). FLAMEnGO 2.0 can be used as a standalone method or to assist in the completion of partial resonance assignments and can be downloaded at www.chem.umn.edu/groups/veglia/forms/flamengo2-form.html.

  14. FLAMEnGO 2.0: An enhanced fuzzy logic algorithm for structure-based assignment of methyl group resonances

    NASA Astrophysics Data System (ADS)

    Chao, Fa-An; Kim, Jonggul; Xia, Youlin; Milligan, Michael; Rowe, Nancy; Veglia, Gianluigi

    2014-08-01

    We present an enhanced version of the FLAMEnGO (Fuzzy Logic Assignment of Methyl Group) software, a structure-based method to assign methyl group resonances in large proteins. FLAMEnGO utilizes a fuzzy logic algorithm coupled with Monte Carlo sampling to obtain a probability-based assignment of the methyl group resonances. As an input, FLAMEnGO requires either the protein X-ray structure or an NMR structural ensemble including data such as methyl-methyl NOESY, paramagnetic relaxation enhancement (PRE), methine-methyl TOCSY data. Version 2.0 of this software (FLAMEnGO 2.0) has a user-friendly graphic interface and presents improved modules that enable the input of partial assignments and additional NMR restraints. We tested the performance of FLAMEnGO 2.0 on maltose binding protein (MBP) as well as the C-subunit of the cAMP-dependent protein kinase A (PKA-C). FLAMEnGO 2.0 can be used as a standalone method or to assist in the completion of partial resonance assignments and can be downloaded at www.chem.umn.edu/groups/veglia/forms/flamengo2-form.html.

  15. Backbone assignments and secondary structure of the Escherichia coli enzyme-II mannitol A domain determined by heteronuclear three-dimensional NMR spectroscopy.

    PubMed Central

    Kroon, G. J.; Grötzinger, J.; Dijkstra, K.; Scheek, R. M.; Robillard, G. T.

    1993-01-01

    This report presents the backbone assignments and the secondary structure determination of the A domain of the Escherichia coli mannitol transport protein, enzyme-IImtl. The backbone resonances were partially assigned using three-dimensional heteronuclear 1H NOE 1H-15N single-quantum coherence (15N NOESY-HSQC) spectroscopy and three-dimensional heteronuclear 1H total correlation 1H-15N single-quantum coherence (15N TOCSY-HSQC) spectroscopy on uniformly 15N enriched protein. Triple-resonance experiments on uniformly 15N/13C enriched protein were necessary to complete the backbone assignments, due to overlapping 1H and 15N frequencies. Data obtained from three-dimensional 1H-15N-13C alpha correlation experiments (HNCA and HN(CO)CA), a three-dimensional 1H-15N-13CO correlation experiment (HNCO), and a three-dimensional 1H alpha-13C alpha-13CO correlation experiment (COCAH) were combined using SNARF software, and yielded the assignments of virtually all observed backbone resonances. Determination of the secondary structure of IIAmtl is based upon NOE information from the 15N NOESY-HSQC and the 1H alpha and 13C alpha secondary chemical shifts. The resulting secondary structure is considerably different from that reported for IIAglc of E. coli and Bacillus subtilis determined by NMR and X-ray. PMID:8401218

  16. Enzyme IIBcellobiose of the phosphoenol-pyruvate-dependent phosphotransferase system of Escherichia coli: backbone assignment and secondary structure determined by three-dimensional NMR spectroscopy.

    PubMed Central

    Ab, E.; Schuurman-Wolters, G. K.; Saier, M. H.; Reizer, J.; Jacuinod, M.; Roepstorff, P.; Dijkstra, K.; Scheek, R. M.; Robillard, G. T.

    1994-01-01

    The assignment of backbone resonances and the secondary structure determination of the Cys 10 Ser mutant of enzyme IIBcellobiose of the Escherichia coli cellobiose-specific phosphoenol-pyruvate-dependent phosphotransferase system are presented. The backbone resonances were assigned using 4 triple resonance experiments, the HNCA and HN(CO)CA experiments, correlating backbone 1H, 15N, and 13C alpha resonances, and the HN(CA)CO and HNCO experiments, correlating backbone 1H,15N and 13CO resonances. Heteronuclear 1H-NOE 1H-15N single quantum coherence (15N-NOESY-HSQC) spectroscopy and heteronuclear 1H total correlation 1H-15N single quantum coherence (15N-TOCSY-HSQC) spectroscopy were used to resolve ambiguities arising from overlapping 13C alpha and 13CO frequencies and to check the assignments from the triple resonance experiments. This procedure, together with a 3-dimensional 1H alpha-13C alpha-13CO experiment (COCAH), yielded the assignment for all observed backbone resonances. The secondary structure was determined using information both from the deviation of observed 1H alpha and 13C alpha chemical shifts from their random coil values and 1H-NOE information from the 15N-NOESY-HSQC. These data show that enzyme IIBcellobiose consists of a 4-stranded parallel beta-sheet and 5 alpha-helices. In the wild-type enzyme IIBcellobiose, the catalytic residue appears to be located at the end of a beta-strand. PMID:8003964

  17. NMR resonance assignments for the N-terminal domain of the δ subunit of the E. coli γ clamp loader complex.

    PubMed

    Alyami, Esmael M; Rizzo, Alessandro A; Beuning, Penny J; Korzhnev, Dmitry M

    2017-03-06

    The β-clamp protein and the γ clamp loader complex are essential components of bacterial DNA replication machinery. The β-clamp is a ring-shaped homodimer that encircles DNA and increases the efficiency of replication by providing a binding platform for DNA polymerases and other replication-related proteins. The β-clamp is loaded onto DNA by the five-subunit γ clamp loader complex in a multi-step ATP-dependent process. The initial steps of this process involve the cooperative binding of the β-clamp by the five subunits of ATP-bound clamp loader, which induces or traps an open conformation of the clamp. Remarkably, the δ subunit of the E. coli clamp loader, or even its 140 residue N-terminal domain (called mini-δ), alone can shift conformational equilibrium of the β-clamp towards the open state. Here we report nearly complete backbone and side-chain (1)H, (13)C and (15)N NMR resonance assignments of mini-δ that will facilitate NMR studies of the mechanisms of β-clamp opening and its loading on DNA by the clamp loader.

  18. 13C-direct detected NMR experiments for the sequential J-based resonance assignment of RNA oligonucleotides

    PubMed Central

    Richter, Christian; Kovacs, Helena; Buck, Janina; Wacker, Anna; Fürtig, Boris; Bermel, Wolfgang

    2010-01-01

    We present here a set of 13C-direct detected NMR experiments to facilitate the resonance assignment of RNA oligonucleotides. Three experiments have been developed: (1) the (H)CC-TOCSY-experiment utilizing a virtual decoupling scheme to assign the intraresidual ribose 13C-spins, (2) the (H)CPC-experiment that correlates each phosphorus with the C4′ nuclei of adjacent nucleotides via J(C,P) couplings and (3) the (H)CPC-CCH-TOCSY-experiment that correlates the phosphorus nuclei with the respective C1′,H1′ ribose signals. The experiments were applied to two RNA hairpin structures. The current set of 13C-direct detected experiments allows direct and unambiguous assignment of the majority of the hetero nuclei and the identification of the individual ribose moieties following their sequential assignment. Thus, 13C-direct detected NMR methods constitute useful complements to the conventional 1H-detected approach for the resonance assignment of oligonucleotides that is often hindered by the limited chemical shift dispersion. The developed methods can also be applied to large deuterated RNAs. Electronic supplementary material The online version of this article (doi:10.1007/s10858-010-9429-5) contains supplementary material, which is available to authorized users. PMID:20544375

  19. Nuclear magnetic resonance assignment and secondary structure of an ankyrin-like repeat-bearing protein: myotrophin.

    PubMed

    Yang, Y; Rao, N S; Walker, E; Sen, S; Qin, J

    1997-06-01

    Multidimensional heteronuclear NMR has been applied to the structural analysis of myotrophin, a novel protein identified from spontaneously hypertensive rat hearts and hypertrophic human hearts. Myotrophin has been shown to stimulate protein synthesis in myocytes and likely plays an important role in the initiation of cardiac hypertrophy, a major cause of mortality in humans. Recent cDNA cloning revealed that myotrophin has 11B amino acids containing 2.5 contiguous ANK repeats, a motif known to be involved in a wide range of macromolecular recognition. A series of two- and three-dimensional heteronuclear bond correlation NMR experiments have been performed on uniformly 15N-labeled or uniformly 15N/13C-labeled protein to obtain the 1H, 15N, and 13C chemical shift assignments. The secondary structure of myotrophin has been determined by a combination of NOEs, NH exchange data, 3JHN alpha coupling constants, and chemical shifts of 1H alpha, 13C alpha, and 13 C beta. The protein has been found to consist of seven helices, all connected by turns or loops. Six of the seven helices (all but the C-terminal helix) form three separate helix-turn-helix motifs. The two full ANK repeats in myotrophin are characteristic of multiple turns followed by a helix-turn-helix motif. A hairpin-like turn involving L32-R36 in ANK repeat #1 exhibits slow conformational averaging on the NMR time scale and appears dynamically different from the corresponding region (D65-169) of ANK repeat #2.

  20. Empirical Equation Based Chirality (n, m) Assignment of Semiconducting Single Wall Carbon Nanotubes from Resonant Raman Scattering Data

    PubMed Central

    Arefin, Md Shamsul

    2012-01-01

    This work presents a technique for the chirality (n, m) assignment of semiconducting single wall carbon nanotubes by solving a set of empirical equations of the tight binding model parameters. The empirical equations of the nearest neighbor hopping parameters, relating the term (2n− m) with the first and second optical transition energies of the semiconducting single wall carbon nanotubes, are also proposed. They provide almost the same level of accuracy for lower and higher diameter nanotubes. An algorithm is presented to determine the chiral index (n, m) of any unknown semiconducting tube by solving these empirical equations using values of radial breathing mode frequency and the first or second optical transition energy from resonant Raman spectroscopy. In this paper, the chirality of 55 semiconducting nanotubes is assigned using the first and second optical transition energies. Unlike the existing methods of chirality assignment, this technique does not require graphical comparison or pattern recognition between existing experimental and theoretical Kataura plot.

  1. Assignment of resonances in dissociative recombination of HD{sup +} ions: High-resolution measurements compared with accurate computations

    SciTech Connect

    Waffeu Tamo, F. O.; Buhr, H.; Schwalm, D.; Motapon, O.; Altevogt, S.; Andrianarijaona, V. M.; Grieser, M.; Lammich, L.; Lestinsky, M.; Motsch, M.; Novotny, S.; Orlov, D. A.; Pedersen, H. B.; Sprenger, F.; Weigel, U.; Wolf, A.; Nevo, I.; Urbain, X.; Schneider, I. F.

    2011-08-15

    The collision-energy resolved rate coefficient for dissociative recombination of HD{sup +} ions in the vibrational ground state is measured using the photocathode electron target at the heavy-ion storage ring TSR. Rydberg resonances associated with rovibrational excitation of the HD{sup +} core are scanned as a function of the electron collision energy with an instrumental broadening below 1 meV in the low-energy limit. The measurement is compared to calculations using multichannel quantum defect theory, accounting for rotational structure and interactions and considering the six lowest rotational energy levels as initial ionic states. Using thermal-equilibrium-level populations at 300 K to approximate the experimental conditions, close correspondence between calculated and measured structures is found up to the first vibrational excitation threshold of the cations near 0.24 eV. Detailed assignments, including naturally broadened and overlapping Rydberg resonances, are performed for all structures up to 0.024 eV. Resonances from purely rotational excitation of the ion core are found to have similar strengths as those involving vibrational excitation. A dominant low-energy resonance is assigned to contributions from excited rotational states only. The results indicate strong modifications in the energy dependence of the dissociative recombination rate coefficient through the rotational excitation of the parent ions, and underline the need for studies with rotationally cold species to obtain results reflecting low-temperature ionized media.

  2. Stereospecific assignment of 1H resonances through chemical shift calculation and their use in structure determination by NMR

    NASA Astrophysics Data System (ADS)

    Harvey, Timothy S.; van Gunsteren, Wilfred F.; Ikura, Mitsuhiko

    1995-04-01

    Understanding of the factors which influence proton chemical shifts in nuclear magnetic resonance (NMR) spectra of proteins has advanced steadily as the number of proteins, for which assignments in conjunction with high resolution structures have been obtained, has increased. Progress has been made in both the calculation of chemical shifts from given coordinates, both empirically for 1H (Williamson & Asakura J. Magn. Reson. (1991) 94, 557) and using ab initio approaches for calculation of 13C (De Dios et al. Science (1993) 260, 1491). Concomitantly Wishart et al. (J. Mol. Biol. (1992) 222, 311), using statistical methods have clarified the relationship between Hα chemical shift and regular secondary structure in proteins to a high degree of accuracy. We recently demonstrated the significant amount of structural information present in the Hα chemical shift through the use of chemical shift restrained molecular dynamics simulations (Harvey & van Gunsteren Techniques in Protein Chemistry IV (1993) 615, Academic Press). Here we apply a similar methodology to the stereospecific assignment of methylene and methyl proton resonances in proteins. Stereospecific assignment of such 1H resonances dramatically increases the degree of precision of ensembles of structures derived from NMR data. However, this is often a cumbersome process, requiring detailed analysis of large amounts of data. Furthermore, experimental considerations such as poor signal-to-noise ratios, spectral overlap and spin diffusion combine to make this process somewhat unreliable. We present calculations of the chemical shifts for the known structures of bovine pancreatic trypsin inhibitor (Mw 6.5 kDa) and the α-amylase inhibitor tendamistat (Mw 8 kDa), for which stereospecific assignments and high resolution structures from both NMR and crystallographic studies are available. The methods described are also applied to the ensemble of structures obtained for protein S (Mw 19 kDa) for both structure

  3. ¹H, ¹³C and ¹⁵N resonance assignments and second structure information of Gad m 1: a β-parvalbumin allergen from Atlantic cod (Gadus morhua).

    PubMed

    Moraes, A H; Ackerbauer, D; Kostadinova, M; Bublin, M; Ferreira, F; Almeida, F C L; Breiteneder, H; Valente, A P

    2013-10-01

    Gad m 1 is the major allergen from Atlantic cod. It belongs to β-parvalbumin protein family and is characterized by the presence of two calcium-binding sites so called EF-hand motifs. β-Parvalbumins such as Gad m 1 are the most important fish allergens and their high cross-reactivity is the cause of the observed polysensitization to various fish species in allergic patients. Despite extensive efforts, the complete elucidation of β-parvalbumin-IgE complexes has not been achieved yet. Allergen structural studies are essential for the development of novel immunotherapy strategies, including vaccination with hypoallergenic derivatives and chimeric molecules. Here, we report for the first time the NMR study of a β-parvalbumin: Gad m 1. This report includes: (1)H, (13)C and (15)N resonance assignments of Gad m 1 as well as the second structure information based on the (13)C chemical shifts.

  4. Solution structural characterization of cyanometmyoglobin: Resonance assignment of heme cavity residues by two-dimensional NMR

    SciTech Connect

    Emerson, S.D.; La Mar, G.N. )

    1990-02-13

    Steady-state nuclear Overhauser effects (NOE), two-dimensional (2D) nuclear Overhauser effect spectroscopy (NOESY), and 2D spin correlation spectroscopy (COSY) have been applied to the fully paramagnetic low-spin, cyanide-ligated complex of sperm whale ferric myoglobin to assign the majority of the heme pocket side-chain proton signals and the remainder of the heme signals. It is shown that the 2D NOESY map reveals essentially all dipolar connectivities observed in ordinary 1D NOE experiments and expected on the basis of crystal coordinates, albeit often more weakly than in a diamagnetic analogue. For extremely broad ({approximately}600-Hz) and rapidly relaxing (T{sub 1} {approximately} 3 ms) signals which show no NOESY peaks, the authors demonstrate that conventional steady-state NOEs obtained under very rapid pulsing conditions still allow detection of the critical dipolar connectivities that allow unambiguous assignments. Numerous critical COSY cross peaks between strongly hyperfine-shifted peaks were resolved and assigned. In all, 95% (53 of 56 signals) of the total proton sets within {approximately}7.5 {angstrom} of the iron, the region experiencing the strongest hyperfine shifts and paramagnetic relaxation, are now unambiguously assigned. Hence it is clear that the 2D methods can be profitably applied to paramagnetic proteins. The scope and limitations of such application are discussed. The resulting hyperfine shift pattern for the heme confirmed expectations based on model compounds.

  5. Complete Proton and Carbon Assignment of Triclosan via One- and Two- Dimensional Nuclear Magnetic Resonance Analysis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Students from an upper-division undergraduate spectroscopy class analyzed one- and two-dimensional 400 MHz NMR spectroscopic data from triclosan in CDCl3. Guided assignment of all proton and carbon signals was completed via 1D proton and carbon, nuclear Overhauser effect (nOe), distortionless enhanc...

  6. Optimized {gamma}-Multiplicity Based Spin Assignments of s-Wave Neutron Resonances

    SciTech Connect

    Becvar, F.; Koehler, Paul Edward; Krticka, Milan; Mitchell, G. E.; Ullmann, J. L.

    2011-01-01

    The multiplicity of -ray emission following neutron capture at isolated resonances carries valuable information on the resonance spin. Several methods utilizing this information have been developed. The latest method was recently introduced for analyzing the data from time-of-flight measurements with 4 -calorimetric detection systems. The present paper describes a generalization of this method. The goal is the separation of the -emission yields belonging to the two neutron capturing state spins of isolated (or even unresolved) s-wave neutron resonances on targets with non-zero spin. The formalism for performing this separation is described and then tested on artificially generated data. This new method was applied to the -multiplicity data obtained for the 147Sm(n, )148Sm reaction using the DANCE detector system at the LANSCE facility at Los Alamos National Laboratory. The analyzing power of the upgraded method is supported by combined dicebox and geant4 simulations of the fluctuation properties of the multiplicity distributions.

  7. Rapid analysis of protein backbone resonance assignments using cryogenic probes, a distributed Linux-based computing architecture, and an integrated set of spectral analysis tools.

    PubMed

    Monleón, Daniel; Colson, Kimberly; Moseley, Hunter N B; Anklin, Clemens; Oswald, Robert; Szyperski, Thomas; Montelione, Gaetano T

    2002-01-01

    Rapid data collection, spectral referencing, processing by time domain deconvolution, peak picking and editing, and assignment of NMR spectra are necessary components of any efficient integrated system for protein NMR structure analysis. We have developed a set of software tools designated AutoProc, AutoPeak, and AutoAssign, which function together with the data processing and peak-picking programs NMRPipe and Sparky, to provide an integrated software system for rapid analysis of protein backbone resonance assignments. In this paper we demonstrate that these tools, together with high-sensitivity triple resonance NMR cryoprobes for data collection and a Linux-based computer cluster architecture, can be combined to provide nearly complete backbone resonance assignments and secondary structures (based on chemical shift data) for a 59-residue protein in less than 30 hours of data collection and processing time. In this optimum case of a small protein providing excellent spectra, extensive backbone resonance assignments could also be obtained using less than 6 hours of data collection and processing time. These results demonstrate the feasibility of high throughput triple resonance NMR for determining resonance assignments and secondary structures of small proteins, and the potential for applying NMR in large scale structural proteomics projects.

  8. Complete assignment of the 1H nuclear magnetic resonance spectrum of French bean plastocyanin. Application of an integrated approach to spin system identification in proteins.

    PubMed

    Chazin, W J; Rance, M; Wright, P E

    1988-08-05

    The identification of the spin systems that comprise the 1H nuclear magnetic resonance spectrum of French bean Cu(I) plastocyanin (Mr 10,600) has been made using an approach that integrates a wide range of two-dimensional nuclear magnetic resonance experiments. A very large percentage of these assignments has been obtained in spectra acquired from 1H2O solution using a backbone amide-based strategy. The spin systems of 91 of the 99 residues have been assigned to the appropriate amino acid, thereby providing an ample basis for obtaining sequence-specific assignments, as described in the accompanying paper.

  9. Complete resonance assignment of the first and second apple domains of MIC4 from Toxoplasma gondii, using a new NMRView-based assignment aid.

    PubMed

    Marchant, Jan; Sawmynaden, Kovilen; Saouros, Savvas; Simpson, Peter; Matthews, Stephen

    2008-12-01

    Microneme protein 4 is involved in cell binding by the important parasite Toxoplasma gondii. We present here the backbone and side-chain assignments of the first two apple domains together with a new graphical aid for their assignment using NMRView.

  10. NMR resonance assignments for the tetramethylrhodamine binding RNA aptamer 3 in complex with the ligand 5-carboxy-tetramethylrhodamine.

    PubMed

    Duchardt-Ferner, Elke; Juen, Michael; Kreutz, Christoph; Wöhnert, Jens

    2017-04-01

    RNA aptamers are used in a wide range of biotechnological or biomedical applications. In many cases the high resolution structures of these aptamers in their ligand-complexes have revealed fundamental aspects of RNA folding and RNA small molecule interactions. Fluorescent RNA-ligand complexes in particular find applications as optical sensors or as endogenous fluorescent tags for RNA tracking in vivo. Structures of RNA aptamers and aptamer ligand complexes constitute the starting point for rational function directed optimization approaches. Here, we present the NMR resonance assignment of an RNA aptamer binding to the fluorescent ligand tetramethylrhodamine (TMR) in complex with the ligand 5-carboxy-tetramethylrhodamine (5-TAMRA) as a starting point for a high-resolution structure determination using NMR spectroscopy in solution.

  11. Photodissociation spectroscopy of stored CH+ ions: Detection, assignment, and close-coupled modeling of near-threshold Feshbach resonances

    NASA Astrophysics Data System (ADS)

    Hechtfischer, Ulrich; Williams, Carl J.; Lange, Michael; Linkemann, Joachim; Schwalm, Dirk; Wester, Roland; Wolf, Andreas; Zajfman, Daniel

    2002-11-01

    We have measured and theoretically analyzed a photodissociation spectrum of the CH+ molecular ion in which most observed energy levels lie within the fine-structure splitting of the C+ fragment and predissociate, and where the observed irregular line shapes and dipole-forbidden transitions indicate that nonadiabatic interactions lead to multichannel dynamics. The molecules were prepared in low rotational levels J''=0-9 of the vibrational ground state X 1Sigma+ (v'')=0 by storing a CH+ beam at 7.1 MeV in the heavy-ion storage ring TSR for up to 30 s, which was sufficient for the ions to rovibrationally thermalize to room temperature by spontaneous infrared emission. The internally cold molecules were irradiated with a dye laser at photon energies between 31 600-33 400 cm-1, and the resulting C+ fragments were counted with a particle detector. The photodissociation cross section displays the numerous Feshbach resonances between the two C+ fine-structure states predicted by theory for low rotation. The data are analyzed in two steps. First, from the overall structure of the spectrum, by identifying branches, and by a Le Roy-Bernstein analysis of level spacings we determine the dissociation energy D0=(32 946.7plus-or-minus1.1) cm-1 (with respect to the lower fine-structure limit) and assign the strongest features to the vibrational levels v'=11-14 of the dipole-allowed A 1Pi state. The majority of the 66 observed resonances cannot be assigned in this way. Therefore, in a second step, the complete spectrum is simulated with a close-coupling model, starting from recent ab initio Born-Oppenheimer potentials. For the long-range induction, dispersion and exchange energies, we propose an analytical expression and derive the C6 coefficients. After a systematic variation of just the vibrational defects of the four Born-Oppenheimer potentials involved, the close-coupling model yields a quantitative fit to the measured cross section in all detail, and is used to assign most of

  12. 15N Hyperpolarization by Reversible Exchange Using SABRE-SHEATH

    PubMed Central

    2016-01-01

    NMR signal amplification by reversible exchange (SABRE) is a NMR hyperpolarization technique that enables nuclear spin polarization enhancement of molecules via concurrent chemical exchange of a target substrate and parahydrogen (the source of spin order) on an iridium catalyst. Recently, we demonstrated that conducting SABRE in microtesla fields provided by a magnetic shield enables up to 10% 15N-polarization (Theis, T.; et al. J. Am. Chem. Soc.2015, 137, 1404). Hyperpolarization on 15N (and heteronuclei in general) may be advantageous because of the long-lived nature of the hyperpolarization on 15N relative to the short-lived hyperpolarization of protons conventionally hyperpolarized by SABRE, in addition to wider chemical shift dispersion and absence of background signal. Here we show that these unprecedented polarization levels enable 15N magnetic resonance imaging. We also present a theoretical model for the hyperpolarization transfer to heteronuclei, and detail key parameters that should be optimized for efficient 15N-hyperpolarization. The effects of parahydrogen pressure, flow rate, sample temperature, catalyst-to-substrate ratio, relaxation time (T1), and reversible oxygen quenching are studied on a test system of 15N-pyridine in methanol-d4. Moreover, we demonstrate the first proof-of-principle 13C-hyperpolarization using this method. This simple hyperpolarization scheme only requires access to parahydrogen and a magnetic shield, and it provides large enough signal gains to enable one of the first 15N images (2 × 2 mm2 resolution). Importantly, this method enables hyperpolarization of molecular sites with NMR T1 relaxation times suitable for biomedical imaging and spectroscopy. PMID:25960823

  13. Kinetic 15N-isotope effects on algal growth

    PubMed Central

    Andriukonis, Eivydas; Gorokhova, Elena

    2017-01-01

    Stable isotope labeling is a standard technique for tracing material transfer in molecular, ecological and biogeochemical studies. The main assumption in this approach is that the enrichment with a heavy isotope has no effect on the organism metabolism and growth, which is not consistent with current theoretical and empirical knowledge on kinetic isotope effects. Here, we demonstrate profound changes in growth dynamics of the green alga Raphidocelis subcapitata grown in 15N-enriched media. With increasing 15N concentration (0.37 to 50 at%), the lag phase increased, whereas maximal growth rate and total yield decreased; moreover, there was a negative relationship between the growth and the lag phase across the treatments. The latter suggests that a trade-off between growth rate and the ability to adapt to the high 15N environment may exist. Remarkably, the lag-phase response at 3.5 at% 15N was the shortest and deviated from the overall trend, thus providing partial support to the recently proposed Isotopic Resonance hypothesis, which predicts that certain isotopic composition is particularly favorable for living organisms. These findings confirm the occurrence of KIE in isotopically enriched algae and underline the importance of considering these effects when using stable isotope labeling in field and experimental studies. PMID:28281640

  14. Kinetic 15N-isotope effects on algal growth

    NASA Astrophysics Data System (ADS)

    Andriukonis, Eivydas; Gorokhova, Elena

    2017-03-01

    Stable isotope labeling is a standard technique for tracing material transfer in molecular, ecological and biogeochemical studies. The main assumption in this approach is that the enrichment with a heavy isotope has no effect on the organism metabolism and growth, which is not consistent with current theoretical and empirical knowledge on kinetic isotope effects. Here, we demonstrate profound changes in growth dynamics of the green alga Raphidocelis subcapitata grown in 15N-enriched media. With increasing 15N concentration (0.37 to 50 at%), the lag phase increased, whereas maximal growth rate and total yield decreased; moreover, there was a negative relationship between the growth and the lag phase across the treatments. The latter suggests that a trade-off between growth rate and the ability to adapt to the high 15N environment may exist. Remarkably, the lag-phase response at 3.5 at% 15N was the shortest and deviated from the overall trend, thus providing partial support to the recently proposed Isotopic Resonance hypothesis, which predicts that certain isotopic composition is particularly favorable for living organisms. These findings confirm the occurrence of KIE in isotopically enriched algae and underline the importance of considering these effects when using stable isotope labeling in field and experimental studies.

  15. 15N and13C NMR investigation of hydroxylamine-derivatized humic substances

    USGS Publications Warehouse

    Thorn, K.A.; Arterburn, J.B.; Mikita, M.A.

    1992-01-01

    Five fulvic and humic acid samples of diverse origins were derivatized with 15N-labeled hydroxylamine and analyzed by liquid-phase 15N NMR spectrometry. The 15N NMR spectra indicated that hydroxylamine reacted similarly with all samples and could discriminate among carbonyl functional groups. Oximes were the major derivatives; resonances attributable to hydroxamic acids, the reaction products of hydroxylamine with esters, and resonances attributable to the tautomeric equilibrium position between the nitrosophenol and monoxime derivatives of quinones, the first direct spectroscopic evidence for quinones, also were evident. The 15N NMR spectra also suggested the presence of nitriles, oxazoles, oxazolines, isocyanides, amides, and lactams, which may all be explained in terms of Beckmann reactions of the initial oxime derivatives. INEPT and ACOUSTIC 15N NMR spectra provided complementary information on the derivatized samples. 13C NMR spectra of derivatized samples indicated that the ketone/quinone functionality is incompletely derivatized with hydroxylamine. ?? 1991 American Chemical Society.

  16. Cytotoxic Properties and Complete Nuclear Magnetic Resonance Assignment of Isolated Xanthones from the Root of Garcinia cowa Roxb.

    PubMed Central

    Wahyuni, Fatma Sri; Shaari, Khozirah; Stanslas, Johnson; Lajis, Nordin HJ; Hamidi, Dachriyanus

    2016-01-01

    Objective: To isolate compounds from the roots of Garcinia cowa and to evaluated their cytotoxic activity against breast (MCF-7), prostate (DU-145), and lung (H-460) cell lines. Materials and Methods: The ground air-dried root was sequentially macerated with hexane, dichloromethane (DCM), ethyl acetate (EtOAc), and methanol. The DCM soluble extract was fractionated by vacuum liquid chromatography, column chromatography, and radial chromatography over silica gel with hexane, EtOAc and methanol as eluent in progressively increasing polarity manner; to yield three compounds. Their structures were elucidated based on their spectroscopic data and their comparison with those of the literature. The cytotoxicity of isolated compounds was carried out against human cell lines by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide colorimetric assay. The extract was added at various concentrations (0.1, 1, 10 and 100 mg/ml). The level of cytotoxicity was determined by calculating the level of IC50 that was based on the percentage of the cell death following the 24 h incubation with the extract. Results: Phytochemical study on the roots of G. cowa yielded rubraxanthone (3), cowanine (4) and 1,5-dihydroxyxanthone (5). Compound 4 with an IC50 value of 4.1 ± 1.0 μM, 5.4 ± 2.3 μM and 11.3 ± 10.0 μM against MCF-7, H-460, and DU-145, respectively while compound 3 was found to be in active. Conclusion: The results indicate that G. cowa roots could be important sources of natural cytotoxic compounds. SUMMARY Isolation of cytotoxic compounds from Garcinia cowaCowanine is the active constituent from the roots of Garcinia cowaComplete nuclear magnetic resonance assignment of isolated compoundsMS fragmentation of rubraxanthone. PMID:27041859

  17. 1H-NMR study of diamagnetic cytochrome P450cam: assignment of heme resonances and substrate dependance of one cysteinate beta proton.

    PubMed

    Mouro, C; Bondon, A; Simonneaux, G; Jung, C

    1997-09-08

    The 1H-NMR study of diamagnetic cytochrome P450cam FeII-CO has been performed for the first time. Chemical shifts of the cysteinate fifth ligand protons and of several heme protons have been assigned through 1- and 2-dimensional spectra at 500 MHz. A substrate dependance has been observed for the resonance of the cysteinate proton detected in the high-field region.

  18. NMR assignment method for amide signals with cell-free protein synthesis system.

    PubMed

    Kohno, Toshiyuki

    2010-01-01

    Nuclear magnetic resonance (NMR) methods are widely used to determine the three-dimensional structures of proteins, to estimate protein folding, and to discover high-affinity ligands for proteins. However, one of the problems to apply such NMR methods to proteins is that we should obtain mg quantities of (15)N and/or (13)C labeled pure proteins of interest. Here, we describe the method to produce dual amino acid-selective (13)C-(15)N labeled proteins for NMR study using the improved wheat germ cell-free system, which enables sequence-specific assignments of amide signals simply even for very large protein.

  19. Multinuclear magnetic resonance studies of the 2Feter dot 2S sup * ferredoxin from Anabaena species strain PCC 7120. 1. Sequence-specific hydrogen-1 resonance assignments and secondary structure in solution of the oxidized form

    SciTech Connect

    Oh, Byung-Ha; Markley, J.L. )

    1990-04-24

    Complete sequence-specific assignments were determined for the diamagnetic {sup 1}H resonances from Anabaena 7120 ferredoxin. A novel assignment procedure was followed whose first step was the identification of the {sup 13}C spin systems of the amino acids by a {sup 13}C({sup 13}C) double quantum correlation experiment. Then, the {sup 1}H spin systems of the amino acids were identified from the {sup 13}C spin systems by means of direct and relayed {sup 1}H({sup 13}C) single-bond correlations. The sequential resonance assignments were based mainly on conventional interresidue {sup 1}H{sup {alpha}}{sub i}-{sup 1}H{sup N}{sub i+1} NOE connectivities. Resonances from 18 residues were not resolved in two-dimensional {sup 1}H NMR spectra. When these residues were mapped onto the X-ray crystal structure of the homologous ferredoxin from Spirulina platensis, it was found that they correspond to amino acids close to the paramagnetic 2Fe{center dot}2S cluster. Cross peaks in two-dimensional homonuclear {sup 1}H NMR spectra were not observed for any protons closer than about 7.8 {angstrom} to both iron atoms. Secondary structural features identified in solution include two antiparallel {beta}-sheets, one parallel {beta}-sheet, and one {alpha}-helix.

  20. Proton-coupled 15N NMR spectra of neutral and protonated ethenoadenosine and ethenocytidine.

    PubMed Central

    Sierzputowska-Gracz, H; Wiewiórowski, M; Kozerski, L; von Philipsborn, W

    1984-01-01

    The 15N chemical shifts and 15N, 1H spin coupling constants were determined in the title compounds using the INEPT pulse sequence and assigned with the aid of selective proton decoupling. The delta/15N/ and J/N, H/ values are discussed in terms of involvement of the imidazole ring created by ethenobridging in the electronic structure of the whole molecule. Both spectral parameters indicate that the diligant nitrogen in this ring is the primary site of protonation in these modified nucleosides. It is concluded that 15N NMR of nucleoside bases can be largely a complementary method to 1H and 13C NMR studies and, in addition, can serve as a direct probe for studies of nitrogen environment in oligomeric fragments of nucleic acids even at moderately strong magnetic fields due to the higher spectral dispersion compared with 1H and 13C NMR spectra. PMID:6473107

  1. Resonance assignment of DVU2108 that is part of the Orange Protein complex in Desulfovibrio vulgaris Hildenborough.

    PubMed

    Neca, António J; Soares, Rui; Carepo, Marta S P; Pauleta, Sofia R

    2016-04-01

    We report the 94 % assignment of DVU2108, a protein belonging to the Orange Protein family, that in Desulfovibrio vulgaris Hildenborough forms a protein complex named the Orange Protein complex. This complex has been shown to be implicated in the cell division of this organism. DVU2108 is a conserved protein in anaerobic microorganisms and in Desulfovibrio gigas the homologous protein was isolated with a novel Mo-Cu cluster non-covalently attached to the polypeptide chain. However, the heterologously produced DVU2108 did not contain any bound metal. These assignments provide the means to characterize the interaction of DVU2108 with the proteins that form the Orange Protein complex using NMR methods.

  2. Preparation of 13C/15N-labeled oligomers using the polymerase chain reaction

    DOEpatents

    Chen, Xian; Gupta, Goutam; Bradbury, E. Morton

    2001-01-01

    Preparation of .sup.13 C/.sup.15 N-labeled DNA oligomers using the polymerase chain reaction (PCR). A PCR based method for uniform (.sup.13 C/.sup.15 N)-labeling of DNA duplexes is described. Multiple copies of a blunt-ended duplex are cloned into a plasmid, each copy containing the sequence of interest and restriction Hinc II sequences at both the 5' and 3' ends. PCR using bi-directional primers and uniformly .sup.13 C/.sup.15 N-labeled dNTP precursors generates labeled DNA duplexes containing multiple copies of the sequence of interest. Twenty-four cycles of PCR, followed by restriction and purification, gave the uniformly .sup.13 C/.sup.15 N-labeled duplex sequence with a 30% yield. Such labeled duplexes find significant applications in multinuclear magnetic resonance spectroscopy.

  3. Design of a 15N Molecular Unit to Achieve Long Retention of Hyperpolarized Spin State

    PubMed Central

    Nonaka, Hiroshi; Hirano, Masashi; Imakura, Yuki; Takakusagi, Yoichi; Ichikawa, Kazuhiro; Sando, Shinsuke

    2017-01-01

    Nuclear hyperpolarization is a phenomenon that can be used to improve the sensitivity of magnetic resonance molecular sensors. However, such sensors typically suffer from short hyperpolarization lifetime. Herein we report that [15N, D14]trimethylphenylammonium (TMPA) has a remarkably long spin–lattice relaxation time (1128 s, 14.1 T, 30 °C, D2O) on its 15N nuclei and achieves a long retention of the hyperpolarized state. [15N, D14]TMPA-based hyperpolarized sensor for carboxylesterase allowed the highly sensitive analysis of enzymatic reaction by 15N NMR for over 40 min in phophate-buffered saline (H2O, pH 7.4, 37 °C). PMID:28067292

  4. Design of a 15N Molecular Unit to Achieve Long Retention of Hyperpolarized Spin State

    NASA Astrophysics Data System (ADS)

    Nonaka, Hiroshi; Hirano, Masashi; Imakura, Yuki; Takakusagi, Yoichi; Ichikawa, Kazuhiro; Sando, Shinsuke

    2017-01-01

    Nuclear hyperpolarization is a phenomenon that can be used to improve the sensitivity of magnetic resonance molecular sensors. However, such sensors typically suffer from short hyperpolarization lifetime. Herein we report that [15N, D14]trimethylphenylammonium (TMPA) has a remarkably long spin–lattice relaxation time (1128 s, 14.1 T, 30 °C, D2O) on its 15N nuclei and achieves a long retention of the hyperpolarized state. [15N, D14]TMPA-based hyperpolarized sensor for carboxylesterase allowed the highly sensitive analysis of enzymatic reaction by 15N NMR for over 40 min in phophate-buffered saline (H2O, pH 7.4, 37 °C).

  5. Resonance assignments and secondary structure prediction of the As(III) metallochaperone ArsD in solution

    PubMed Central

    Ye, Jun; He, Yanan; Skalicky, Jack; Rosen, Barry P.; Stemmler, Timothy L.

    2012-01-01

    ArsD is a metallochaperone that delivers As(III) to the ArsA ATPase, the catalytic subunit of the ArsAB pump encoded by the arsRDABC operon of Escherichia coli plasmid R773. Conserved ArsD cysteine residues (Cys12, Cys13 and Cys18) construct the As(III) binding site of the protein, however a global structural understanding of this arsenic binding remains unclear. We have obtained NMR assignments for ArsD as a starting point for probing structural changes on the protein that occur in response to metalloid binding and upon formation of a complex with ArsA. The predicted solution structure of ArsD is in agreement with recently published crystallographic structural results. PMID:21063813

  6. Derivatives of pyrazinecarboxylic acid: 1H, 13C and 15N NMR spectroscopic investigations.

    PubMed

    Holzer, Wolfgang; Eller, Gernot A; Datterl, Barbara; Habicht, Daniela

    2009-07-01

    NMR spectroscopic studies are undertaken with derivatives of 2-pyrazinecarboxylic acid. Complete and unambiguous assignment of chemical shifts ((1)H, (13)C, (15)N) and coupling constants ((1)H,(1)H; (13)C,(1)H; (15)N,(1)H) is achieved by combined application of various 1D and 2D NMR spectroscopic techniques. Unequivocal mapping of (13)C,(1)H spin coupling constants is accomplished by 2D (delta,J) long-range INEPT spectra with selective excitation. Phenomena such as the tautomerism of 3-hydroxy-2-pyrazinecarboxylic acid are discussed.

  7. Direct assignment of EPR spectra to structurally defined iron-sulfur clusters in complex I by double electron–electron resonance

    PubMed Central

    Roessler, Maxie M.; King, Martin S.; Robinson, Alan J.; Armstrong, Fraser A.; Harmer, Jeffrey; Hirst, Judy

    2010-01-01

    In oxidative phosphorylation, complex I (NADH:quinone oxidoreductase) couples electron transfer to proton translocation across an energy-transducing membrane. Complex I contains a flavin mononucleotide to oxidize NADH, and an unusually long series of iron-sulfur (FeS) clusters, in several subunits, to transfer the electrons to quinone. Understanding coupled electron transfer in complex I requires a detailed knowledge of the properties of individual clusters and of the cluster ensemble, and so it requires the correlation of spectroscopic and structural data: This has proved a challenging task. EPR studies on complex I from Bos taurus have established that EPR signals N1b, N2 and N3 arise, respectively, from the 2Fe cluster in the 75 kDa subunit, and from 4Fe clusters in the PSST and 51 kDa subunits (positions 2, 7, and 1 along the seven-cluster chain extending from the flavin). The other clusters have either evaded detection or definitive signal assignments have not been established. Here, we combine double electron-electron resonance (DEER) spectroscopy on B. taurus complex I with the structure of the hydrophilic domain of Thermus thermophilus complex I. By considering the magnetic moments of the clusters and the orientation selectivity of the DEER experiment explicitly, signal N4 is assigned to the first 4Fe cluster in the TYKY subunit (position 5), and N5 to the all-cysteine ligated 4Fe cluster in the 75 kDa subunit (position 3). The implications of our assignment for the mechanisms of electron transfer and energy transduction by complex I are discussed. PMID:20133838

  8. Robust, integrated computational control of NMR experiments to achieve optimal assignment by ADAPT-NMR.

    PubMed

    Bahrami, Arash; Tonelli, Marco; Sahu, Sarata C; Singarapu, Kiran K; Eghbalnia, Hamid R; Markley, John L

    2012-01-01

    ADAPT-NMR (Assignment-directed Data collection Algorithm utilizing a Probabilistic Toolkit in NMR) represents a groundbreaking prototype for automated protein structure determination by nuclear magnetic resonance (NMR) spectroscopy. With a [(13)C,(15)N]-labeled protein sample loaded into the NMR spectrometer, ADAPT-NMR delivers complete backbone resonance assignments and secondary structure in an optimal fashion without human intervention. ADAPT-NMR achieves this by implementing a strategy in which the goal of optimal assignment in each step determines the subsequent step by analyzing the current sum of available data. ADAPT-NMR is the first iterative and fully automated approach designed specifically for the optimal assignment of proteins with fast data collection as a byproduct of this goal. ADAPT-NMR evaluates the current spectral information, and uses a goal-directed objective function to select the optimal next data collection step(s) and then directs the NMR spectrometer to collect the selected data set. ADAPT-NMR extracts peak positions from the newly collected data and uses this information in updating the analysis resonance assignments and secondary structure. The goal-directed objective function then defines the next data collection step. The procedure continues until the collected data support comprehensive peak identification, resonance assignments at the desired level of completeness, and protein secondary structure. We present test cases in which ADAPT-NMR achieved results in two days or less that would have taken two months or more by manual approaches.

  9. ¹⁵N, ¹³C and ¹H resonance assignments and secondary structure determination of a variable heavy domain of a heavy chain antibody.

    PubMed

    Prosser, Christine E; Waters, Lorna C; Muskett, Frederick W; Veverka, Vaclav; Addis, Philip W; Griffin, Laura M; Baker, Terry S; Lawson, Alastair D G; Wernery, Ulrich; Kinne, Jorg; Henry, Alistair J; Taylor, Richard J; Carr, Mark D

    2014-04-01

    Heavy chain antibodies differ in structure to conventional antibodies lacking both the light chain and the first heavy chain constant domain (CH1). Characteristics of the antigen-binding variable heavy domain of the heavy chain antibody (VHH) including the smaller size, high solubility and stability make them an attractive alternative to more traditional antibody fragments for detailed NMR-based structural analysis. Here we report essentially complete backbone and side chain (15)N, (13)C and (1)H assignments for a free VHH. Analysis of the backbone chemical shift data obtained indicates that the VHH is comprised predominantly of β-sheets corresponding to nearly 60% of the protein backbone.

  10. Rapid Proton-Detected NMR Assignment for Proteins with Fast Magic Angle Spinning

    PubMed Central

    2015-01-01

    Using a set of six 1H-detected triple-resonance NMR experiments, we establish a method for sequence-specific backbone resonance assignment of magic angle spinning (MAS) nuclear magnetic resonance (NMR) spectra of 5–30 kDa proteins. The approach relies on perdeuteration, amide 2H/1H exchange, high magnetic fields, and high-spinning frequencies (ωr/2π ≥ 60 kHz) and yields high-quality NMR data, enabling the use of automated analysis. The method is validated with five examples of proteins in different condensed states, including two microcrystalline proteins, a sedimented virus capsid, and two membrane-embedded systems. In comparison to contemporary 13C/15N-based methods, this approach facilitates and accelerates the MAS NMR assignment process, shortening the spectral acquisition times and enabling the use of unsupervised state-of-the-art computational data analysis protocols originally developed for solution NMR. PMID:25102442

  11. Rapid proton-detected NMR assignment for proteins with fast magic angle spinning.

    PubMed

    Barbet-Massin, Emeline; Pell, Andrew J; Retel, Joren S; Andreas, Loren B; Jaudzems, Kristaps; Franks, W Trent; Nieuwkoop, Andrew J; Hiller, Matthias; Higman, Victoria; Guerry, Paul; Bertarello, Andrea; Knight, Michael J; Felletti, Michele; Le Marchand, Tanguy; Kotelovica, Svetlana; Akopjana, Inara; Tars, Kaspars; Stoppini, Monica; Bellotti, Vittorio; Bolognesi, Martino; Ricagno, Stefano; Chou, James J; Griffin, Robert G; Oschkinat, Hartmut; Lesage, Anne; Emsley, Lyndon; Herrmann, Torsten; Pintacuda, Guido

    2014-09-03

    Using a set of six (1)H-detected triple-resonance NMR experiments, we establish a method for sequence-specific backbone resonance assignment of magic angle spinning (MAS) nuclear magnetic resonance (NMR) spectra of 5-30 kDa proteins. The approach relies on perdeuteration, amide (2)H/(1)H exchange, high magnetic fields, and high-spinning frequencies (ωr/2π ≥ 60 kHz) and yields high-quality NMR data, enabling the use of automated analysis. The method is validated with five examples of proteins in different condensed states, including two microcrystalline proteins, a sedimented virus capsid, and two membrane-embedded systems. In comparison to contemporary (13)C/(15)N-based methods, this approach facilitates and accelerates the MAS NMR assignment process, shortening the spectral acquisition times and enabling the use of unsupervised state-of-the-art computational data analysis protocols originally developed for solution NMR.

  12. Sequential NMR resonance assignment and structure determination of the Kunitz-type inhibitor domain of the Alzheimer's beta-amyloid precursor protein.

    PubMed

    Heald, S L; Tilton, R F; Hammond, L J; Lee, A; Bayney, R M; Kamarck, M E; Ramabhadran, T V; Dreyer, R N; Davis, G; Unterbeck, A

    1991-10-29

    Certain precursor proteins (APP751 and APP770) of the amyloid beta-protein (AP) present in Alzheimer's disease contain a Kunitz-type serine protease inhibitor domain (APPI). In this study, the domain is obtained as a functional inhibitor through both recombinant (APPIr) and synthetic (APPIs) methodologies, and the solution structure of APPI is determined by 1H 2D NMR techniques. Complete sequence-specific resonance assignments (except for P13 and G37 NH) for both APPIr and APPIs are achieved using standard procedures. Ambiguities arising from degeneracies in the NMR resonances are resolved by varying sample conditions. Qualitative interpretation of short- and long-range NOEs reveals secondary structural features similar to those extensively documented by NMR for bovine pancreatic trypsin inhibitor (BPTI). A more rigorous interpretation of the NOESY spectra yields NOE-derived interresidue distance restraints which are used in conjunction with dynamic simulated annealing to generate a family of APPI structures. Within this family, the beta-sheet and helical regions are in good agreement with the crystal structure of BPTI, whereas portions of the protease-binding loops deviate from those in BPTI. These deviations are consistent with those recently described in the crystal structure of APPI (Hynes et al., 1990). Also supported in the NMR study is the hydrophobic patch in the protease-binding domain created by side chain-side chain NOE contacts between M17 and F34. In addition, the NMR spectra indicate that the rotation of the W21 ring in APPI is hindered, unlike Y21 in BPTI, showing a greater than 90% preference for one orientation in the hydrophobic groove.

  13. Direct NMR resonance assignments of the active site histidine residue in Escherichia coli thioesterase I/protease I/lysophospholipase L1.

    PubMed

    Wu, Wen-Jin; Tyukhtenko, Sergiy I; Huang, Tai-Huang

    2006-11-01

    Owing to the hydrogen-bond interaction and rapid exchange rate with the bulk water, the transverse relaxation time for the N(delta1)-H proton of the catalytic histidine in Escherichia coli thioesterase I/protease I/lysophospholipase L1 (TEP-I) is rather short. Because of its catalytic importance, it is desirable to detect and assign this proton resonance. In this paper, we report the first direct NMR correlation between the short-lived N(delta1)-H proton and its covalently attached N(delta1)-nitrogen of the catalytic His157 residue in E. coli thioesterase/protease I. We have used gradient-enhanced jump-return spin-echo HMQC (GE-JR SE HMQC) to obtain a direct correlation between the short-lived N(delta1)-H proton and its covalently attached N(delta1)-nitrogen. The sensitivity of detection for the short-lived N(delta1)-H proton was enhanced substantially by improved water suppression, in particular, the suppression of radiation damping via pulsed field gradients.

  14. Resonance assignments of the periplasmic domain of a cellulose-sensing trans-membrane anti-sigma factor from Clostridium thermocellum.

    PubMed

    Ding, Xiaoke; Chen, Chao; Cui, Qiu; Li, Wenli; Feng, Yingang

    2015-10-01

    The cellulosome of Clostridium thermocellum is an elegant and efficient multi-enzyme complex for degrading lignocellulose. The cellulosome contains several dozens of carbohydrate hydrolysis enzymes, which are regulated by the presence of environmental substrates through several pairs of sigma and anti-sigma factors. The anti-sigma factors sense the presence of substrates and transduce the signals into the cell. The sigma factors are then released from the corresponding anti-sigma factors, and they recruit RNA polymerase to transcribe specific cellulosomal genes. However, it is not clear how the extracellular signals are transduced into the cell by the anti-sigma factors. The anti-sigma factors of C. thermocellum contain an N-terminal intracellular domain, a trans-membrane helix, a periplasmic domain, a proline-rich region which is probably required for crossing the cell wall, and a C-terminal carbohydrate-binding domain or glycoside hydrolase domain. The periplasmic domain may play a key role in signal transduction; however, its three-dimensional structure is still unknown. Here we report the NMR resonance assignments of the periplasmic domain of anti-sigma factor RsgI2 from C. thermocellum as a basis for further structural determination and functional studies.

  15. 4-Oxalocrotonate tautomerase, a 41-kDa homohexamer: backbone and side-chain resonance assignments, solution secondary structure, and location of active site residues by heteronuclear NMR spectroscopy.

    PubMed Central

    Stivers, J. T.; Abeygunawardana, C.; Whitman, C. P.; Mildvan, A. S.

    1996-01-01

    4-Oxalocrotonate tautomerase (4-OT), a homohexamer consisting of 62 residues per subunit, catalyzes the isomerization of unsaturated alpha-keto acids using Pro-1 as a general base (Stivers et al., 1996a, 1996b). We report the backbone and side-chain 1H, 15N, and 13C NMR assignments and the solution secondary structure for 4-OT using 2D and 3D homonuclear and heteronuclear NMR methods. The subunit secondary structure consists of an alpha-helix (residues 13-30), two beta-strands (beta 1, residues 2-8; beta 2, residues 39-45), a beta-hairpin (residues 50-57), two loops (I, residues 9-12; II, 34-38), and two turns (I, residues 30-33; II, 47-50). The remaining residues form coils. The beta 1 strand is parallel to the beta 2 strand of the same subunit on the basis of cross stand NH(i)-NH(j) NOEs in a 2D 15N-edited 1H-NOESY spectrum of hexameric 4-OT containing two 15N-labeled subunits/hexamer. The beta 1 strand is also antiparallel to another beta 1 strand from an adjacent subunit forming a subunit interface. Because only three such pairwise interactions are possible, the hexamer is a trimer of dimers. The diffusion constant, determined by dynamic light scattering, and the rotational correlation time (14.5 ns) estimated from 15N T1/T2 measurements, are consistent with the hexameric molecular weight of 41 kDa. Residue Phe-50 is in the active site on the basis of transferred NOEs to the bound partial substrate 2-oxo-1,6-hexanedioate. Modification of the general base, Pro-1, with the active site-directed irreversible inhibitor, 3-bromopyruvate, significantly alters the amide 15N and NH chemical shifts of residues in the beta-hairpin and in loop II, providing evidence that these regions change conformation when the active site is occupied. PMID:8845763

  16. Solid-State 15N NMR of 15N-Labeled Nylon 6 and Nylon 11

    DTIC Science & Technology

    1990-05-22

    S. Veeman, E. M. Menger, W. Ritchey, and E. de Boer, Macromolecules, 1979, 12, 924. 2. A. N. Garroway , W. M. Ritchey and W. B. Moniz, Macromolecules...S. Veeman and E. M. Menger, Bull. Magn. Reson., 1980, 2, 77. 26. D. L. VanderHart and A. N. Garroway , J. Chem. Phys., 1979, 71, 2773. 27. M. D

  17. Chemical shift assignments of the connexin37 carboxyl terminal domain.

    PubMed

    Li, Hanjun; Spagnol, Gaelle; Pontifex, Tasha K; Burt, Janis M; Sorgen, Paul L

    2017-03-01

    Connexin37 (Cx37) is a gap junction protein involved in cell-to-cell communication in the vasculature and other tissues. Cx37 suppresses proliferation of vascular cells involved in tissue development and repair in vivo, as well as tumor cells. Global deletion of Cx37 in mice leads to enhanced vasculogenesis in development, as well as collateralgenesis and angiogenesis in response to injury, which together support improved tissue remodeling and recovery following ischemic injury. Here we report the (1)H, (15)N, and (13)C resonance assignments for an important regulatory domain of Cx37, the carboxyl terminus (CT; C233-V333). The predicted secondary structure of the Cx37CT domain based on the chemical shifts is that of an intrinsically disordered protein. In the (1)H-(15)N HSQC, N-terminal residues S254-Y259 displayed a second weaker peak and residues E261-Y266 had significant line broadening. These residues are flanked by prolines (P250, P258, P260, and P268), suggesting proline cis-trans isomerization. Overall, these assignments will be useful for identifying the binding sites for intra- and inter-molecular interactions that affect Cx37 channel activity.

  18. Backbone and side chain chemical shift assignments of apolipophorin III from Galleria mellonella.

    PubMed

    Crowhurst, Karin A; Horn, James V C; Weers, Paul M M

    2016-04-01

    Apolipophorin III, a 163 residue monomeric protein from the greater wax moth Galleria mellonella (abbreviated as apoLp-IIIGM), has roles in upregulating expression of antimicrobial proteins as well as binding and deforming bacterial membranes. Due to its similarity to vertebrate apolipoproteins there is interest in performing atomic resolution analysis of apoLp-IIIGM as part of an effort to better understand its mechanism of action in innate immunity. In the first step towards structural characterization of apoLp-IIIGM, 99 % of backbone and 88 % of side chain (1)H, (13)C and (15)N chemical shifts were assigned. TALOS+ analysis of the backbone resonances has predicted that the protein is composed of five long helices, which is consistent with the reported structures of apolipophorins from other insect species. The next stage in the characterization of apoLp-III from G. mellonella will be to utilize these resonance assignments in solving the solution structure of this protein.

  19. 15N NMR of 1,4-dihydropyridine derivatives.

    PubMed

    Goba, Inguna; Liepinsh, Edvards

    2013-07-01

    In this article, we describe the characteristic (15)N and (1)HN NMR chemical shifts and (1)J((15)N-(1)H) coupling constants of various symmetrically and unsymmetrically substituted 1,4-dihydropyridine derivatives. The NMR chemical shifts and coupling constants are discussed in terms of their relationship to structural features such as character and position of the substituent in heterocycle, N-alkyl substitution, nitrogen lone pair delocalization within the conjugated system, and steric effects.

  20. δ15N Value Does Not Reflect Fasting in Mysticetes

    PubMed Central

    Aguilar, Alex; Giménez, Joan; Gómez–Campos, Encarna; Cardona, Luís; Borrell, Asunción

    2014-01-01

    The finding that tissue δ15N values increase with protein catabolism has led researchers to apply this value to gauge nutritive condition in vertebrates. However, its application to marine mammals has in most occasions failed. We investigated the relationship between δ15N values and the fattening/fasting cycle in a model species, the fin whale, a migratory capital breeder that experiences severe seasonal variation in body condition. We analyzed two tissues providing complementary insights: one with isotopic turnover (muscle) and one that keeps a permanent record of variations in isotopic values (baleen plates). In both tissues δ15N values increased with intensive feeding but decreased with fasting, thus contradicting the pattern previously anticipated. The apparent inconsistency during fasting is explained by the fact that a) individuals migrate between different isotopic isoscapes, b) starvation may not trigger significant negative nitrogen balance, and c) excretion drops and elimination of 15N-depleted urine is minimized. Conversely, when intensive feeding is resumed in the northern grounds, protein anabolism and excretion start again, triggering 15N enrichment. It can be concluded that in whales and other mammals that accrue massive depots of lipids as energetic reserves and which have limited access to drinking water, the δ15N value is not affected by fasting and therefore cannot be used as an indicatior of nutritive condition. PMID:24651388

  1. Sequential protein NMR assignments in the liquid state via sequential data acquisition

    NASA Astrophysics Data System (ADS)

    Wiedemann, Christoph; Bellstedt, Peter; Kirschstein, Anika; Häfner, Sabine; Herbst, Christian; Görlach, Matthias; Ramachandran, Ramadurai

    2014-02-01

    Two different NMR pulse schemes involving sequential 1H data acquisition are presented for achieving protein backbone sequential resonance assignments: (i) acquisition of 3D {HCCNH and HNCACONH} and (ii) collection of 3D {HNCOCANH and HNCACONH} chemical shift correlation spectra using uniformly 13C,15N labelled proteins. The sequential acquisition of these spectra reduces the overall experimental time by a factor of ≈2 as compared to individual acquisitions. The suitability of this approach is experimentally demonstrated for the C-terminal winged helix (WH) domain of the minichromosome maintenance (MCM) complex of Sulfolobus solfataricus.

  2. ¹H, ¹³C and ¹⁵N backbone resonance assignments of the monomeric human M-ficolin fibrinogen-like domain secreted by Brevibacillus choshinensis.

    PubMed

    Tanio, Michikazu; Kusunoki, Hideki; Kohno, Toshiyuki

    2014-04-01

    M-ficolin, which forms trimer-based multimers, is a pathogen-recognition protein in the innate immune system, and it binds to ligands through its fibrinogen-like (FBG) domain. As the first step toward the elucidation of the molecular basis for pathogen-recognition by the M-ficolin multimers, we assigned the backbone resonances of the monomeric mutant of the M-ficolin FBG domain, recombinantly expressed by Brevibacillus choshinensis. Like the wild-type trimeric FBG domain, the monomeric FBG domain also requires His251, His284 and His297 for the ligand-binding activity, as judged by mutational analyses using zonal affinity chromatography. The secondary structure predicted by the backbone resonance assignments is similar to that of the trimeric FBG domain in the crystal, indicating that the monomeric FBG domain is folded correctly to perform its function.

  3. Phenylalanine δ15N in Paleo Archives as a New Proxy for δ15N of Exported Primary Production

    NASA Astrophysics Data System (ADS)

    McCarthy, M.; Batista, F. C.; Vokhshoori, N. L.; Brown, J. T.; Guilderson, T. P.; Ravelo, A. C.; Sherwood, O.

    2012-12-01

    Compound-specific isotope analysis of individual amino acids (CSI-AA) is emerging as a powerful new tool for studying the paleo nitrogen cycle. Because most detrital organic nitrogen is composed of amino acids, CSI-AA can reveal the mechanistic basis for organic nitrogen diagenesis, preserve a record of past food web structure, and potentially reconstruct the δ15N values of past nitrate and primary production. Within the commonly measured amino acids, the δ15N value of phenylalanine (Phe) appears uniquely promising as a new proxy that reflects the nitrogen isotopic value of the original source. Phe δ15N values remain almost unchanged with trophic transfer through food webs, and also during at least the initial stages of organic matter degradation. Here we synthesize results from both bio-archives and recent sediments, which together suggest that at least in Holocene archives the Phe δ15N value does in fact record the average inorganic nitrogen δ15N value at the base of planktonic food webs. However, several important unknowns also remain. These include the extent of variation in amino acid isotopic fractionation patterns in phylogenetically distinct algal groups. The stability of Phe δ15N values in older sediments where organic matter has undergone extensive diagenesis is also an important research area, which may ultimately establish the temporal limit for application of this approach to study past geological epochs. Together, however, results to date suggest that of Phe δ15N values in paleo archives represent a novel molecular-level proxy which is not tied to any specific organism or group, but rather can provide an integrated estimate of δ15N value of exported primary production.

  4. Solvent effects on 15N NMR coordination shifts.

    PubMed

    Kleinmaier, Roland; Arenz, Sven; Karim, Alavi; Carlsson, Anna-Carin C; Erdélyi, Máté

    2013-01-01

    (15)N NMR chemical shift became a broadly utilized tool for characterization of complex structures and comparison of their properties. Despite the lack of systematic studies, the influence of solvent on the nitrogen coordination shift, Δ(15)N(coord), was hitherto claimed to be negligible. Herein, we report the dramatic impact of the local environment and in particular that of the interplay between solvent and substituents on Δ(15)N(coord). The comparative study of CDCl(3) and CD(3)CN solutions of silver(I)-bis(pyridine) and silver(I)-bis(pyridylethynyl)benzene complexes revealed the strong solvent dependence of their (15)N NMR chemical shift, with a solvent dependent variation of up to 40 ppm for one and the same complex. The primary influence of the effect of substituent and counter ion on the (15)N NMR chemical shifts is rationalized by corroborating Density-Functional Theory (nor discrete Fourier transform) calculations on the B3LYP/6-311 + G(2d,p)//B3LYP/6-31G(d) level. Cooperative effects have to be taken into account for a comprehensive description of the coordination shift and thus the structure of silver complexes in solution. Our results demonstrate that interpretation of Δ(15)N(coord) in terms of coordination strength must always consider the solvent and counter ion. The comparable magnitude of Δ(15)N(coord) for reported transition metal complexes makes the principal findings most likely general for a broad scale of complexes of nitrogen donor ligands, which are in frequent use in modern organometallic chemistry.

  5. First Experimental Measurement of the {sup 18}O(p,{alpha}){sup 15}N Reaction at Astrophysical Energies

    SciTech Connect

    La Cognata, M.; Sergi, M. L.; Spitaleri, C.; Cherubini, S.; Gulino, M.; Kiss, G.; Lamia, L.; Pizzone, R. G.; Romano, S.; Mukhamedzhanov, A.; Goldberg, V.; Tribble, R.; Coc, A.; Hammache, F.; Sereville, N. de; Tumino, A.

    2010-11-24

    The {sup 18}O(p,{alpha}){sup 15}N and {sup 17}O(p,{alpha}){sup 14}N reactions are of primary importance in several as-trophysical scenarios, including nucleosynthesis inside Asymptotic Giant Branch stars and oxygen and nitrogen isotopic ratios in meteorite grains. They are also key reactions to understand exotic systems such as R-Coronae Borealis stars and novae. Thus, the measurement of their cross sections in the low energy region can be crucial to reduce the nuclear uncertainty on theoretical predictions, because the resonance parameters are poorly determined. The Trojan Horse Method, in its newly developed form particularly suited to investigate low-energy resonances, has been applied to the {sup 2}H({sup 18}O,{alpha}{sup 15}N)n and {sup 2}H({sup 17}O,{alpha}{sup 14}N)n reactions to deduce the {sup 18}O(p,{alpha}){sup 15}N and {sup 17}O(p,{alpha}){sup 14}N cross sections at low energies. Resonances in the {sup 18}O(p,{alpha}){sup 15}N and {sup 17}O(p,{alpha}){sup 14}N excitation functions have been studied and the resonance parameters deduced.

  6. Understanding molecular formula assignment of Fourier transform ion cyclotron resonance mass spectrometry data of natural organic matter from a chemical point of view.

    PubMed

    Herzsprung, Peter; Hertkorn, Norbert; von Tümpling, Wolf; Harir, Mourad; Friese, Kurt; Schmitt-Kopplin, Philippe

    2014-12-01

    Formula assignment is one of the key challenges in evaluation of dissolved organic matter analyses using ultrahigh resolution mass spectrometry (FTICR MS). The number of possible solutions for elemental formulas grows exponentially with increasing nominal mass, especially when non-oxygen heteroatoms like N, S or P are considered. Until now, no definitive solution for finding the correct elemental formula has been given. For that reason an approach from the viewpoint of chemical feasibility was elucidated. To illustrate the new chemical formula assignment principle, a literature data set was used and evaluated by simplified chemical constraints. Only formulas containing a maximum of one sulphur and five nitrogen atoms were selected for further data processing. The resulting data table was then divided into mass peaks with unique component solutions (singlets, representing unequivocal formula assignments) and those with two or more solutions (multiple formula assignments, representing equivocal formula assignments). Based on a [double bond equivalent (DBE) versus the number of oxygen atoms (o)] frequency contour plot and a frequency versus [DBE minus o] diagram, a new assessment and decision strategy was developed to differentiate multiple formula assignments into chemically reliable and less reliable molecular formulas. Using this approach a considerable number of reliable components were identified within the equivocal part of the data set. As a control, a considerable proportion of the assigned formulas deemed to be reliable correspond to those which would have been obtained by CH 2 -based Kendrick mass defect analysis. We conclude that formula assignment in complex mixtures can be improved by group-wise decisions based on the frequency and the [DBE minus o] values of multiple formula assignments.

  7. Two-dimensional sup 1 H nuclear magnetic resonance study of AaH IT, an anti-insect toxin from the scorpion Androctonus australis Hector. Sequential resonance assignments and folding of the polypeptide chain

    SciTech Connect

    Darbon, H. ); Weber, C.; Braun, W. )

    1991-02-19

    Sequence-specific nuclear magnetic resonance assignments for the polypeptide backbone and for most of the amino acid side-chain protons, as well as the general folding of AaH IT, are described. AaH IT is a neurotoxin purified from the venom of the scorpion Androctonus australis Hector and is specifically active on the insect nervous system. The secondary structure and the hydrogen-bonding patterns in the regular secondary structure elements are deduced from nuclear Overhauser effects and the sequence locations of the slowly exchanging amide protons. The backbone folding is determined by distance geometry calculations with the DISMAN program. The regular secondary structure includes two and a half turns of {alpha}-helix running from residues 21 to 30 and a three-stranded antiparallel {beta}-sheet including peptides 3-5, 34-38, and 41-46. Two tight turns are present, one connecting the end of the {alpha}-helix to an external strand of the {beta}-sheet, i.e., turn 31-34, and another connecting this same strand to the central one, i.e., turn 38-41. The differences in the specificity of these related proteins, which are able to discriminate between mammalian and insect voltage-dependent sodium channels of excitable tissues, are most probably brought about by the position of the C-terminal peptide with regard to a hydrophobic surface common to all scorpion toxins examined thus far. Thus, the interaction of a given scorpion toxin with its receptor might well be governed by the presence of this solvent-exposed hydrophobic surface, whereas adjacent areas modulate the specificity of the interaction.

  8. 14N15N detectability in Pluto’s atmosphere

    NASA Astrophysics Data System (ADS)

    Jessup, Kandis Lea; Gladstone, G. R.; Heays, A. N.; Gibson, S. T.; Lewis, B. R.; Stark, G.

    2013-11-01

    Based on the vapor pressure behavior of Pluto’s surface ices, Pluto’s atmosphere is expected to be predominantly composed of N2 gas. Measurement of the N2 isotopologue 15N/14N ratio within Pluto’s atmosphere would provide important clues to the evolution of Pluto’s atmosphere from the time of formation to its present state. The most straightforward way of determining the N2 isotopologue 15N/14N ratio in Pluto’s atmosphere is via spectroscopic observation of the 14N15N gas species. Recent calculations of the 80-100 nm absorption behavior of the 14N2 and 14N15N isotopologues by Heays et al. (Heays, A.N. et al. [2011]. J. Chem. Phys. 135, 244301), Lewis et al. (Lewis, B.R., Heays, A.N., Gibson, S.T., Lefebvre-Brion, H., Lefebvre, R. [2008]. J. Chem. Phys. 129, 164306); Lewis et al. (Lewis, B.R., Gibson, S.T., Zhang, W., Lefebvre-Brion, H., Robbe, J.-M. [2005]. J. Chem. Phys. 122, 144302), and Haverd et al. (Haverd, V.E., Lewis, B.R., Gibson, S.T., Stark, G. [2005]. J. Chem. Phys. 123, 214304) show that the peak magnitudes of the 14N2 and 14N15N absorption bandhead cross-sections are similar, but the locations of the bandhead peaks are offset in wavelength by ∼0.05-0.1 nm. These offsets make the segregation of the 14N2 and 14N15N absorption signatures possible. We use the most recent N2 isotopologue absorption cross-section calculations and the atmospheric density profiles resulting from photochemical models developed by Krasnopolsky and Cruickshank (Krasnopolsky, V.A., Cruickshank, D.P. [1999]. J. Geophys. Res. 104, 21979-21996) to predict the level of solar light that will be transmitted through Pluto’s atmosphere as a function of altitude during a Pluto solar occultation. We characterize the detectability of the isotopic absorption signature per altitude assuming 14N15N concentrations ranging from 0.1% to 2% of the 14N2 density and instrumental spectral resolutions ranging from 0.01 to 0.3 nm. Our simulations indicate that optical depth of unity is

  9. Assignment of histidine resonances in the sup 1 H NMR (500 MHz) spectrum of subtilisin BPN prime using site-directed mutagenesis

    SciTech Connect

    Bycroft, M.; Fersht, A.R. )

    1988-09-20

    A spin-echo pulse sequence has been used to resolve the six histidine C-2H protons in the 500-MHz NMR spectrum of subtilisin BPN{prime}. Five of these residues have been substituted by site-directed mutagenesis, and this has enabled a complete assignment of these protons to be obtained. Analysis of the pH titration curves of these signals has provided microscopic pK{sub a}'s for the six histidines in this enzyme. The pK{sub a}'s of the histidine residues in subtilisin BPN{prime} have been compared with the values obtained for the histidines in the homologous enzyme from Bacillus licheniformis (subtilisin Carlsberg). Four of the five conserved histidines titrate with essentially identical pK{sub a}'s in the two enzymes. It therefore appears that the assignments made for these residues in subtilisin BPN{prime} can be transferred to subtilisin Carlsberg. On the basis of these assignments, the one histidine that titrates with a substantially different pK{sub a} in the two enzymes can be assigned to histidine-238. This difference in pK{sub a} has been attributed to a Trp to Lys substitution at position 241 in subtilisin Carlsberg.

  10. 15N and 13C NMR Determination of Allantoin Metabolism in Developing Soybean Cotyledons 1

    PubMed Central

    Coker, George T.; Schaefer, Jacob

    1985-01-01

    The metabolism of allantoin by immature cotyledons of soybean (Glycine max L. cv Elf) grown in culture was investigated using solid state 13C and 15N nuclear magnetic resonance. All of the nitrogens of allantoin were incorporated into protein in a manner similar to that of each other and to the amide nitrogen of glutamine. The C-2 of allantoin was not incorporated into cellular material; presumably it was lost as CO2. About 50% of the C-5 of allantoin was incorporated into cellular material as a methylene carbon; the other 50% was presumably also lost as CO2. The 13C-15N bonds of [5-13C;1-15N] and [2-13C;1,3-15N]allantoin were broken prior to the incorporation of the nitrogens into protein. These data are consistent with allantoin's degradation to two molecules of urea and one two-carbon fragment. Cotyledons grown on allantoin as a source of nitrogen accumulated 21% of the nitrogen of cotyledons grown on glutamine. Only 50% of the nitrogen of the degraded allantoin was incorporated into the cotyledon as organic nitrogen; the other 50% was recovered as NH4+ in the media in which the cotyledons had been grown. The latter results suggests that the lower accumulation of nitrogen by cotyledons grown on allantoin was in part due to failure to assimilate NH4+ produced from allantoin. The seed coats had a higher activity of glutamine synthetase and a higher rate of allantoin degradation than cotyledons indicating that seed coats play an important role in the assimilation and degradation of allantoin. PMID:16663995

  11. Stepwise enrichment of 15N along food chains: Further evidence and the relation between δ 15N and animal age

    NASA Astrophysics Data System (ADS)

    Minagawa, Masao; Wada, Eitaro

    1984-05-01

    The isotopic composition of nitrogen was measured in marine and fresh-water animals from the East China Sea, The Bering Sea, Lake Ashinoko and Usujiri intertidal zone. Primary producers, showed average δ15Nversus atmospheric nitrogen of +5.0%. (+3.4 to +7.5) in the Bering Sea and Lake Ashinoko, and +6.8%. (+6.0 to +7.6) in Usujiri intertidal zone. Blue green algae from the East China Sea show an average -0.55%. (-0.8 to +1.2). All consumers, Zooplankton, fish and bird exhibited Stepwise enrichment of 15N with increasing trophic level. The 15N enrichment at a single feeding process ranged from +1.3 to +5.3 averaging +3.4 ± 1.1%.. This isotopic fractionation seems to be independent of habitat. The effect of age in animals was obtained by analyzing two marine mussels. The soft tissue nitrogen showed +2.0%. enrichment relative to that of primary producers, and the magnitude was almost constant with shell ages ranging from 0 to 8 years. A similar 15N enrichment occurs in all Molluscs, Crustaceans, Insecta, Amphibia, Fish, Ave and Mammal species regardless of the difference in the form of excreted nitrogen and in laboratory cultured fish, brine shrimp and mice (+2.9 to +4.9%.). The excreted ammonia from guppy was sufficiently light to balance the concentration of 15N to animal body.

  12. ¹H, ¹³C, ¹⁵N backbone and side chain NMR resonance assignments for the N-terminal RNA recognition motif of the HvGR-RBP1 protein involved in the regulation of barley (Hordeum vulgare L.) senescence.

    PubMed

    Mason, Katelyn E; Tripet, Brian P; Parrott, David; Fischer, Andreas M; Copié, Valérie

    2014-04-01

    Leaf senescence is an important process in the developmental life of all plant species. Senescence efficiency influences important agricultural traits such as grain protein content and plant growth, which are often limited by nitrogen use. Little is known about the molecular mechanisms regulating this highly orchestrated process. To enhance our understanding of leaf senescence and its regulation, we have undertaken the structural and functional characterization of previously unknown proteins that are involved in the control of senescence in barley (Hordeum vulgare L.). Previous microarray analysis highlighted several barley genes whose transcripts are differentially expressed during senescence, including a specific gene which is greater than 40-fold up-regulated in the flag leaves of early- as compared to late-senescing near-isogenic barley lines at 14 and 21 days past flowering (anthesis). From inspection of its amino acid sequence, this gene is predicted to encode a glycine-rich RNA-binding protein herein referred to as HvGR-RBP1. HvGR-RBP1 has been expressed as a recombinant protein in Escherichia coli, and preliminary NMR data analysis has revealed that its glycine-rich C-terminal region [residues: 93-162] is structurally disordered whereas its N-terminal region [residues: 1-92] forms a well-folded domain. Herein, we report the complete (1)H, (13)C, and (15)N resonance assignments of backbone and sidechain atoms, and the secondary structural topology of the N-terminal RNA recognition motif (RRM) domain of HvGR-RBP1, as a first step to unraveling its structural and functional role in the regulation of barley leaf senescence.

  13. Marking Drosophila suzukii (Diptera: Drosophilidae) With Rubidium or 15N.

    PubMed

    Klick, J; Yang, W Q; Bruck, D J

    2015-06-01

    Drosophila suzukii Matsumura (Diptera: Drosophilidae) has caused significant economic damage to berry and stone fruit production regions. Markers that are systemic in plants and easily transferred to target organisms are needed to track D. suzukii exploitation of host resources and trophic interactions. High and low concentrations of the trace element, rubidium (Rb), and the stable isotope, 15N, were tested to mark D. suzukii larvae feeding on fruits of enriched strawberry plants grown in containers under greenhouse conditions. Fly marker content and proportion of flies marked 1, 7, and 14 d after emergence from enriched fruits and fly dry mass were analyzed. Nearly 100% of the flies analyzed 14 d after emerging from 15N-enriched plants were marked, whereas only 30-75% and 0-3% were marked 14 d after emerging from high and low Rb concentration plants, respectively. Rapid Rb decay, strong 15N persistence, and the economics of using these markers in the field to elucidate D. suzukii pest ecology are discussed.

  14. 15N chemical shift referencing in solid state NMR.

    PubMed

    Bertani, Philippe; Raya, Jésus; Bechinger, Burkhard

    2014-01-01

    Solid-state NMR spectroscopy has much advanced during the last decade and provides a multitude of data that can be used for high-resolution structure determination of biomolecules, polymers, inorganic compounds or macromolecules. In some cases the chemical shift referencing has become a limiting factor to the precision of the structure calculations and we have therefore evaluated a number of methods used in proton-decoupled (15)N solid-state NMR spectroscopy. For (13)C solid-state NMR spectroscopy adamantane is generally accepted as an external standard, but to calibrate the (15)N chemical shift scale several standards are in use. As a consequence the published chemical shift values exhibit considerable differences (up to 22 ppm). In this paper we report the (15)N chemical shift of several commonly used references compounds in order to allow for comparison and recalibration of published data and future work. We show that (15)NH4Cl in its powdered form (at 39.3 ppm with respect to liquid NH3) is a suitable external reference as it produces narrow lines when compared to other reference compounds and at the same time allows for the set-up of cross-polarization NMR experiments. The compound is suitable to calibrate magic angle spinning and static NMR experiments. Finally the temperature variation of (15)NH4Cl chemical shift is reported.

  15. A closer look at the nitrogen next door: 1H-15N NMR methods for glycosaminoglycan structural characterization

    NASA Astrophysics Data System (ADS)

    Langeslay, Derek J.; Beni, Szabolcs; Larive, Cynthia K.

    2012-03-01

    Recently, experimental conditions were presented for the detection of the N-sulfoglucosamine (GlcNS) NHSO3- or sulfamate 1H and 15N NMR resonances of the pharmaceutically and biologically important glycosaminoglycan (GAG) heparin in aqueous solution. In the present work, we explore further the applicability of nitrogen-bound proton detection to provide structural information for GAGs. Compared to the detection of 15N chemical shifts of aminosugars through long-range couplings using the IMPACT-HNMBC pulse sequence, the more sensitive two-dimensional 1H-15N HSQC-TOCSY experiments provided additional structural data. The IMPACT-HNMBC experiment remains a powerful tool as demonstrated by the spectrum measured for the unsubstituted amine of 3-O-sulfoglucosamine (GlcN(3S)), which cannot be observed with the 1H-15N HSQC-TOCSY experiment due to the fast exchange of the amino group protons with solvent. The 1H-15N HSQC-TOCSY NMR spectrum reported for the mixture of model compounds GlcNS and N-acetylglucosamine (GlcNAc) demonstrate the broad utility of this approach. Measurements for the synthetic pentasaccharide drug Arixtra® (Fondaparinux sodium) in aqueous solution illustrate the power of this NMR pulse sequence for structural characterization of highly similar N-sulfoglucosamine residues in GAG-derived oligosaccharides.

  16. Nitrogen input 15N-signatures are reflected in plant 15N natural abundances of N-rich tropical forest in China

    NASA Astrophysics Data System (ADS)

    Abdisa Gurmesa, Geshere; Lu, Xiankai; Gundersen, Per; Yunting, Fang; Mo, Jiangming

    2016-04-01

    In this study, we tested the measurement of natural abundance of 15N15N) for its ability to assess changes in N cycling due to increased N deposition in two forest types; namely, an old-growth broadleaved forest and a pine forest, in southern China. We measured δ15N values of inorganic N in input and output fluxes under ambient N deposition, and N concentration and δ15N of major ecosystem compartments under ambient and increased N deposition. Our results showed that N deposition to the forests was 15N-depleted, and was dominated by NH4-N. Plants were 15N-depleted due to imprint from the 15N-depleted atmospheric N deposition. The old-growth forest had larger N concentration and was more 15N-enriched than the pine forest. Nitrogen addition did not significantly affect N concentration, but it significantly increased δ15N values of plants, and slightly more so in the pine forest, toward the 15N signature of the added N in both forests. The result indicates that the pine forest may rely more on the 15N-depleted deposition N. Soil δ15N values were slightly decreased by the N addition. Our result suggests that ecosystem δ15N is more sensitive to the changes in ecosystem N status and N cycling than N concentration in N-saturated sub-tropical forests.

  17. ¹H, ¹³C, and ¹⁵N backbone resonance assignments of the porcine pepsin and porcine pepsin complexed with pepstatin.

    PubMed

    Horimoto, Yasumi; Wang, Shenlin; Yada, Rickey Y

    2014-04-01

    Pepsin is formed as the zymogen, pepsinogen, which includes an additional 44 residue prosegment (PS) on the N-terminus. Upon acidification (pH <3) the PS is removed, yielding active pepsin. The PS is critical to such processes as the initiation of correct folding and protein stability. In the present study, the NMR assignments of the 34.6 kDa native porcine pepsin and porcine pepsin complexed with pepstatin are reported in order to obtain structural information regarding PS-catalyzed protein folding. Such information would contribute to a better understanding of the nature of folding/unfolding energy barrier of pepsin and other aspartic proteases.

  18. Cross sections for n+{sup 14}N from an R-matrix analysis of the {sup 15}N system

    SciTech Connect

    Hale, G.M.; Young, P.G.; Chadwick, M.B.

    1994-06-01

    As part of the Hiroshima-Nagasaki Dose Reevaluation Program, a new evaluation of the neutron cross sections for {sup 14}N was made for ENDF/B-VI, based at energies below 2.5 MeV on a multichannel R-matrix analysis of reactions in the {sup 15}N system. The types of data used in the analysis, and the resulting cross sections and resonance structure for {sup 15}N are briefly described. The resonant features of the neutron cross sections were especially well determined by including precise, high-resolution neutron total cross section measurements from ORNL. While the new evaluated cross section appear to be significant improvements over the earlier ones, they still need to be tested more extensively against recent measurements of the differential elastic cross section from Oak Ridge.

  19. Assignment of selected hyperfine proton NMR resonances in the met forms of Glycera dibranchiata monomer hemoglobins and comparisons with sperm whale metmyoglobin

    SciTech Connect

    Constantinidis, I.; Satterlee, J.D.; Pandey, R.K.; Leung, H.K.; Smith, K.M.

    1988-04-19

    This work indicates a high degree of purity for our preparations of all three of the primary Glycera dibranchiata monomer hemoglobins and details assignments of the heme methyl and vinyl protons in the hyperfine shift region of the ferric (aquo.) protein forms. The assignments were carried out by reconstituting the apoproteins of each component with selectively deuteriated hemes. The results indicate that even though the individual component preparations consist of essentially a single protein, the proton NMR spectra indicate spectroscopic heterogeneity. Evidence is presented for identification and classification of major and minor protein forms that are present in solutions of each component. Finally, in contrast to previous results, a detailed analysis of the proton hyperfine shift patterns of the major and minor forms of each component, in comparison to the major and minor forms of metmyoglobin, leads to the conclusions that the corresponding forms of the proteins from each species have strikingly similar heme-globin contacts and display nearly identical heme electronic structures and coordination numbers.

  20. Fast automated protein NMR data collection and assignment by ADAPT-NMR on Bruker spectrometers

    NASA Astrophysics Data System (ADS)

    Lee, Woonghee; Hu, Kaifeng; Tonelli, Marco; Bahrami, Arash; Neuhardt, Elizabeth; Glass, Karen C.; Markley, John L.

    2013-11-01

    ADAPT-NMR (Assignment-directed Data collection Algorithm utilizing a Probabilistic Toolkit in NMR) supports automated NMR data collection and backbone and side chain assignment for [U-13C, U-15N]-labeled proteins. Given the sequence of the protein and data for the orthogonal 2D 1H-15N and 1H-13C planes, the algorithm automatically directs the collection of tilted plane data from a variety of triple-resonance experiments so as to follow an efficient pathway toward the probabilistic assignment of 1H, 13C, and 15N signals to specific atoms in the covalent structure of the protein. Data collection and assignment calculations continue until the addition of new data no longer improves the assignment score. ADAPT-NMR was first implemented on Varian (Agilent) spectrometers [A. Bahrami, M. Tonelli, S.C. Sahu, K.K. Singarapu, H.R. Eghbalnia, J.L. Markley, PLoS One 7 (2012) e33173]. Because of broader interest in the approach, we present here a version of ADAPT-NMR for Bruker spectrometers. We have developed two AU console programs (ADAPT_ORTHO_run and ADAPT_NMR_run) that run under TOPSPIN Versions 3.0 and higher. To illustrate the performance of the algorithm on a Bruker spectrometer, we tested one protein, chlorella ubiquitin (76 amino acid residues), that had been used with the Varian version: the Bruker and Varian versions achieved the same level of assignment completeness (98% in 20 h). As a more rigorous evaluation of the Bruker version, we tested a larger protein, BRPF1 bromodomain (114 amino acid residues), which yielded an automated assignment completeness of 86% in 55 h. Both experiments were carried out on a 500 MHz Bruker AVANCE III spectrometer equipped with a z-gradient 5 mm TCI probe. ADAPT-NMR is available at http://pine.nmrfam.wisc.edu/ADAPT-NMR in the form of pulse programs, the two AU programs, and instructions for installation and use.

  1. Fast automated protein NMR data collection and assignment by ADAPT-NMR on Bruker spectrometers.

    PubMed

    Lee, Woonghee; Hu, Kaifeng; Tonelli, Marco; Bahrami, Arash; Neuhardt, Elizabeth; Glass, Karen C; Markley, John L

    2013-11-01

    ADAPT-NMR (Assignment-directed Data collection Algorithm utilizing a Probabilistic Toolkit in NMR) supports automated NMR data collection and backbone and side chain assignment for [U-(13)C, U-(15)N]-labeled proteins. Given the sequence of the protein and data for the orthogonal 2D (1)H-(15)N and (1)H-(13)C planes, the algorithm automatically directs the collection of tilted plane data from a variety of triple-resonance experiments so as to follow an efficient pathway toward the probabilistic assignment of (1)H, (13)C, and (15)N signals to specific atoms in the covalent structure of the protein. Data collection and assignment calculations continue until the addition of new data no longer improves the assignment score. ADAPT-NMR was first implemented on Varian (Agilent) spectrometers [A. Bahrami, M. Tonelli, S.C. Sahu, K.K. Singarapu, H.R. Eghbalnia, J.L. Markley, PLoS One 7 (2012) e33173]. Because of broader interest in the approach, we present here a version of ADAPT-NMR for Bruker spectrometers. We have developed two AU console programs (ADAPT_ORTHO_run and ADAPT_NMR_run) that run under TOPSPIN Versions 3.0 and higher. To illustrate the performance of the algorithm on a Bruker spectrometer, we tested one protein, chlorella ubiquitin (76 amino acid residues), that had been used with the Varian version: the Bruker and Varian versions achieved the same level of assignment completeness (98% in 20 h). As a more rigorous evaluation of the Bruker version, we tested a larger protein, BRPF1 bromodomain (114 amino acid residues), which yielded an automated assignment completeness of 86% in 55 h. Both experiments were carried out on a 500 MHz Bruker AVANCE III spectrometer equipped with a z-gradient 5 mm TCI probe. ADAPT-NMR is available at http://pine.nmrfam.wisc.edu/ADAPT-NMR in the form of pulse programs, the two AU programs, and instructions for installation and use.

  2. Experimental and calculated 1H, 13C, 15N NMR spectra of famotidine

    NASA Astrophysics Data System (ADS)

    Barańska, M.; Czarniecki, K.; Proniewicz, L. M.

    2001-05-01

    Famotidine, 3-[[[2-[(aminoiminomethyl)amino]-4-thiazolyl]methyl]thio]- N-(aminosulfonyl), is a histamine H 2-receptor blocker that has been used mainly for the treatment of peptic ulcers and the Zollinger-Ellison syndrome. Its NMR spectra in different solvents were reported earlier; however, detailed interpretation has not been done thus far. In this work, experimental 1H, 13C and 15N NMR spectra of famotidine dissolved in DMSO-d 6 are shown. The assignment of observed chemical shifts is based on quantum chemical calculation at the Hartree-Fock/6-31G ∗ level. The geometry optimization of the famotidine molecule with two internal hydrogen bonds, i.e. [N(3)-H(23)⋯N(9) and N(3)⋯H(34)-N(20)], is done by using the B3LYP method with the 6-31G ∗ basis set.

  3. Resonance assignments of the myristoylated Y28F/Y67F mutant of the Mason-Pfizer monkey virus matrix protein.

    PubMed

    Doležal, Michal; Hrabal, Richard; Ruml, Tomáš; Rumlová, Michaela

    2015-10-01

    The matrix protein (MA) of the Mason-Pfizer monkey virus (M-PMV) plays a key role in the transport and budding of immature retroviral particles from the host cell. Natural N-terminal myristoylation of MA is essential for the targeting of the particles to the plasma membrane and participates in the interaction of MA with membranes phospholipids. The mutation Y28F/Y67F in MA reduces budding and thus causes the accumulation of viral particles under the cytoplasmic membrane. To investigate the impact of Y28F/Y67F mutation on the structure of MA, we prepared this protein in amount and quality suitable for NMR spectroscopy. We report backbone, side-chain and myristoyl residue assignments of the Y28F/Y67F mutant of the M-PMV matrix protein, which will be used to study the interaction with membrane phospholipids and to determine the structure of the mutant matrix protein.

  4. Backbone chemical shift assignments for Xanthomonas campestris peroxiredoxin Q in the reduced and oxidized states: a dramatic change in backbone dynamics.

    PubMed

    Buchko, Garry W; Perkins, Arden; Parsonage, Derek; Poole, Leslie B; Karplus, P Andrew

    2016-04-01

    Peroxiredoxins (Prx) are ubiquitous enzymes that reduce peroxides as part of antioxidant defenses and redox signaling. While Prx catalytic activity and sensitivity to hyperoxidative inactivation depend on their dynamic properties, there are few examples where their dynamics has been characterized by NMR spectroscopy. Here, we provide a foundation for studies of the solution properties of peroxiredoxin Q from the plant pathogen Xanthomonas campestris (XcPrxQ) by assigning the observable (1)H(N), (15)N, (13)C(α), (13)C(β), and (13)C' chemical shifts for both the reduced (dithiol) and oxidized (disulfide) states. In the reduced state, most of the backbone amide resonances (149/152, 98 %) can be assigned in the XcPrxQ (1)H-(15)N HSQC spectrum. In contrast, a remarkable 51 % (77) of these amide resonances are not visible in the (1)H-(15)N HSQC spectrum of the disulfide state of the enzyme, indicating a substantial change in backbone dynamics associated with the formation of an intramolecular C48-C84 disulfide bond.

  5. NbF{sub 5} and TaF{sub 5}: Assignment of {sup 19}F NMR resonances and chemical bond analysis from GIPAW calculations

    SciTech Connect

    Biswal, Mamata; Body, Monique; Legein, Christophe; Sadoc, Aymeric; Boucher, Florent

    2013-11-15

    The {sup 19}F isotropic chemical shifts (δ{sub iso}) of two isomorphic compounds, NbF{sub 5} and TaF{sub 5}, which involve six nonequivalent fluorine sites, have been experimentally determined from the reconstruction of 1D {sup 19}F MAS NMR spectra. In parallel, the corresponding {sup 19}F chemical shielding tensors have been calculated using the GIPAW method for both experimental and DFT-optimized structures. Furthermore, the [M{sub 4}F{sub 20}] units of NbF{sub 5} and TaF{sub 5} being held together by van der Waals interactions, the relevance of Grimme corrections to the DFT optimization processes has been evaluated. However, the semi-empirical dispersion correction term introduced by such a method does not show any significant improvement. Nonetheless, a complete and convincing assignment of the {sup 19}F NMR lines of NbF{sub 5} and TaF{sub 5} is obtained, ensured by the linearity between experimental {sup 19}F δ{sub iso} values and calculated {sup 19}F isotropic chemical shielding σ{sub iso} values. The effects of the geometry optimizations have been carefully analyzed, confirming among other matters, the inaccuracy of the experimental structure of NbF{sub 5}. The relationships between the fluorine chemical shifts, the nature of the fluorine atoms (bridging or terminal), the position of the terminal ones (opposite or perpendicular to the bridging ones), the fluorine charges, the ionicity and the length of the M–F bonds have been established. Additionally, for three of the {sup 19}F NMR lines of NbF{sub 5}, distorted multiplets, arising from {sup 1}J-coupling and residual dipolar coupling between the {sup 19}F and {sup 93}Nb nuclei, were simulated yielding to values of {sup 93}Nb–{sup 19}F {sup 1}J-coupling for the corresponding fluorine sites. - Graphical abstract: The complete assignment of the {sup 19}F NMR lines of NbF{sub 5} and TaF{sub 5} allow establishing relationships between the {sup 19}F δ{sub iso} values, the nature of the fluorine atoms

  6. CACA-TOCSY with alternate 13C–12C labeling: a 13Cα direct detection experiment for mainchain resonance assignment, dihedral angle information, and amino acid type identification

    PubMed Central

    Takeuchi, Koh; Frueh, Dominique P.; Sun, Zhen-Yu J.; Hiller, Sebastian

    2010-01-01

    We present a 13C direct detection CACA-TOCSY experiment for samples with alternate 13C–12C labeling. It provides inter-residue correlations between 13Cα resonances of residue i and adjacent Cαs at positions i − 1 and i + 1. Furthermore, longer mixing times yield correlations to Cα nuclei separated by more than one residue. The experiment also provides Cα-to-sidechain correlations, some amino acid type identifications and estimates for ψ dihedral angles. The power of the experiment derives from the alternate 13C–12C labeling with [1,3-13C] glycerol or [2-13C] glycerol, which allows utilizing the small scalar 3JCC couplings that are masked by strong 1JCC couplings in uniformly 13C labeled samples. PMID:20383561

  7. Synthesis and biosynthesis of {sup 13}C-, {sup 15}N-labeled deoxynucleosides useful for biomolecular structural determinations

    SciTech Connect

    Ashburn, D.A.; Garcia, K.; Hanners, J.L.; Silks, L.A. III; Unkefer, C.J.

    1994-12-01

    Currently, there is a great emphasis on elucidating the structure, function, and dynamics of DNA. Much of the research involved in this study uses nuclear magnetic resonance (NMR) spectroscopy. Effective use of NMR spectroscopy for DNA molecules with mw > 10,000 requires stable isotope enrichment. We present strategies for site-specific isotopic labeling of the purine bases adenosine and guanosine and the biosynthesis of (U-{sup 13}C, {sup 15}N) DNA from methylotropic bacteria. With commercially available 6-chloropurine, an effective two-step route leads to 2{prime}-deoxy-(amino-{sup 15}N)adenosine (dA). The resulting d(amino-{sup 15}N)A is used in a series of reactions to synthesize 2{prime}-deoxy-(2-{sup 13}C,1,amino-{sup 15}N{sub 2})guanosine or any combination thereof. An improved biosynthesis of labeled DNA has been accomplished using Methylobacterium extorquens AS1. Each liter of growth medium contains 4 g of methanol to yield 1 g of lyophilized cells. As much as 200 mg of RNA per liter of culture has been obtained. We are currently developing large-scale isolation protocols. General synthetic pathways to oligomeric DNA will be presented.

  8. NbF5 and TaF5: Assignment of 19F NMR resonances and chemical bond analysis from GIPAW calculations

    NASA Astrophysics Data System (ADS)

    Biswal, Mamata; Body, Monique; Legein, Christophe; Sadoc, Aymeric; Boucher, Florent

    2013-11-01

    The 19F isotropic chemical shifts (δiso) of two isomorphic compounds, NbF5 and TaF5, which involve six nonequivalent fluorine sites, have been experimentally determined from the reconstruction of 1D 19F MAS NMR spectra. In parallel, the corresponding 19F chemical shielding tensors have been calculated using the GIPAW method for both experimental and DFT-optimized structures. Furthermore, the [M4F20] units of NbF5 and TaF5 being held together by van der Waals interactions, the relevance of Grimme corrections to the DFT optimization processes has been evaluated. However, the semi-empirical dispersion correction term introduced by such a method does not show any significant improvement. Nonetheless, a complete and convincing assignment of the 19F NMR lines of NbF5 and TaF5 is obtained, ensured by the linearity between experimental 19F δiso values and calculated 19F isotropic chemical shielding σiso values. The effects of the geometry optimizations have been carefully analyzed, confirming among other matters, the inaccuracy of the experimental structure of NbF5. The relationships between the fluorine chemical shifts, the nature of the fluorine atoms (bridging or terminal), the position of the terminal ones (opposite or perpendicular to the bridging ones), the fluorine charges, the ionicity and the length of the M-F bonds have been established. Additionally, for three of the 19F NMR lines of NbF5, distorted multiplets, arising from 1J-coupling and residual dipolar coupling between the 19F and 93Nb nuclei, were simulated yielding to values of 93Nb-19F 1J-coupling for the corresponding fluorine sites.

  9. Relating the 4s{sigma}{sup -1} inner-valence photoelectron spectrum of HBr with the Br 3d{sup -1}5l{lambda} resonant Auger spectra: An approach to the assignments

    SciTech Connect

    Puettner, R.; Hu, Y. F.; Bancroft, G. M.; Kivimaeki, A.; Jurvansuu, M.; Aksela, H.; Aksela, S.

    2003-09-01

    The high resolution Br 4s{sigma}{sup -1} photoelectron spectrum of HBr is presented together with the resonant Auger spectra resulting from excitations from the 3d core levels to the low-n Rydberg orbitals 5s{sigma}, 5p{sigma}, and 5p{pi}. The very complex spectra can be broadly assigned using two observations. First, the energy splittings of the 4p{pi}{sup -2}5s and 4p{pi}{sup -2}5p states are very similar to the splittings of the 4p{pi}{sup -2}({sup 1}{sigma}{sup +},{sup 1}{delta}, and {sup 3}{sigma}{sup -}) final states seen previously in the normal Auger spectra. Second, the {sup 2}{sigma}{sup +} states, which are the dominant correlation satellites in the complex 4s{sigma}{sup -1} photoelectron spectrum, are often enhanced in the 5s{sigma} resonance Auger spectra. Electron correlation and spin-orbit interaction in the final states are important to understand all of these spectra. Unlike the normal Auger spectra, vibrational excitations play only a minor role in these spectra, showing that the 5s and 5p Rydberg orbitals have some bonding character.

  10. Application of rate equations to ELDOR and saturation recovery experiments on 14N: 15N spin-label pairs

    NASA Astrophysics Data System (ADS)

    Yin, Jun-Jie; Hyde, James S.

    Rate equations describing the time dependence of population differences of the five allowed transitions in an 14N 15N spin-label pair problem are set up. Included in the formulation are the three Heisenberg exchange rate constants and different nitrogen nuclear spin-lattice relaxation rates, electron spin-lattice relaxation rates, and populations for the 14N and 15N moieties. Using matrix algebra, stationary and time-dependent solutions are obtained in a unified theoretical framework. The calculations apply to stationary and pulse electron-electron double resonance and to saturation-recovery ESR. Particular emphasis is placed on short pulse initial excitation, where the transverse relaxation processes are sufficiently slow that only the population difference of the irradiated transition departs significantly from Boltzmann equilibrium during the excitation.

  11. Bonding in hard and elastic amorphous carbon nitride films investigated using 15N, 13C, and 1H NMR spectroscopy

    NASA Astrophysics Data System (ADS)

    Gammon, W. J.; Hoatson, G. L.; Holloway, B. C.; Vold, R. L.; Reilly, A. C.

    2003-11-01

    The nitrogen bonding in hard and elastic amorphous carbon nitride (a-CNx) films is examined with 15N, 13C, and 1H nuclear magnetic resonance (NMR) spectroscopy. Films were deposited by dc magnetron sputtering, in a pure nitrogen discharge on Si(001) substrates at 300 °C. Nanoindentation tests revealed an elastic recovery of 80%, a hardness of 5 GPa, and an elastic modulus of 47 GPa. The NMR results show that nitrogen bonding in this material is consistent with sp2 hybridized nitrogen incorporated in an aromatic carbon environment. The data also indicate that the a-CNx prepared for this study has very low hydrogen content and is hydrophilic. Specifically, analysis of 15N and 13C cross polarization magic angle spinning and 1H NMR experiments suggests that water preferentially protonates nitrogen sites.

  12. Quantitative analysis of bacterial and mammalian proteomes using a combination of cysteine affinity tags and 15N-metabolic labeling.

    PubMed

    Conrads, T P; Alving, K; Veenstra, T D; Belov, M E; Anderson, G A; Anderson, D J; Lipton, M S; Pasa-Tolić, L; Udseth, H R; Chrisler, W B; Thrall, B D; Smith, R D

    2001-05-01

    We describe the combined use of 15N-metabolic labeling and a cysteine-reactive biotin affinity tag to isolate and quantitate cysteine-containing polypeptides (Cys-polypeptides) from Deinococcus radiodurans as well as from mouse B16 melanoma cells. D. radiodurans were cultured in both natural isotopic abundance and 15N-enriched media. Equal numbers of cells from both cultures were combined and the soluble proteins extracted. This mixture of isotopically distinct proteins was derivatized using a commercially available cysteine-reactive reagent that contains a biotin group. Following trypsin digestion, the resulting modified peptides were isolated using immobilized avidin. The mixture was analyzed by capillary reversed-phase liquid chromatography (LC) online with ion trap mass spectrometry (MS) as well as Fourier transform ion cyclotron resonance (FTICR) MS. The resulting spectra contain numerous pairs of Cyspolypeptides whose mass difference corresponds to the number of nitrogen atoms present in each of the peptides. Designation of Cys-polypeptide pairs is also facilitated by the distinctive isotopic distribution of the 15N-labeled peptides versus their 14N-labeled counterparts. Studies with mouse B16 cells maintained in culture allowed the observation of hundreds of isotopically distinct pairs of peptides by LC-FTICR analysis. The ratios of the areas of the pairs of isotopically distinct peptides showed the expected 1:1 labeling of the 14N and 15N versions of each peptide. An additional benefit from the present strategy is that the 15N-labeled peptides do not display significant isotope-dependent chromatographic shifts from their 14N-labeled counterparts, therefore improving the precision for quantitating peptide abundances. The methodology presented offers an alternate, cost-effective strategy for conducting global, quantitative proteomic measurements.

  13. Characterization of Nylon 6 by 15N Solid State NMR

    DTIC Science & Technology

    1989-05-31

    M.; Ritchey, W.; de Boer, E. Macromolecules, 1979, 12, 924. 2. Garroway , A. N.; Ritchey, W. M.; Moniz, W. B.; Macromolecules, 1982, It, 1051. 3...E. Macromolecules, 1982, 15, 1406. 23. Veeman, W. S.; Menger, E. M. Bull. Magn. Reson., 1980,2,77. 24. VanderHart, D. L.; Garroway , A. N. J. Chem

  14. Complete backbone and DENQ side chain NMR assignments in proteins from a single experiment: implications to structure-function studies.

    PubMed

    Reddy, Jithender G; Hosur, Ramakrishna V

    2014-03-01

    Resonance assignment is the first and the most crucial step in all nuclear magnetic resonance (NMR) investigations on structure-function relationships in biological macromolecules. Often, the assignment exercise has to be repeated several times when specific interactions with ligands, substrates etc., have to be elucidated for understanding the functional mechanisms. While the protein backbone serves to provide a scaffold, the side chains interact directly with the ligands. Such investigations will be greatly facilitated, if there are rapid methods for obtaining exhaustive information with minimum of NMR experimentation. In this context, we present here a pulse sequence which exploits the recently introduced technique of parallel detection of multiple nuclei, e.g. (1)H and (13)C, and results in two 3D-data sets simultaneously. These yield complete backbone resonance assignment ((1)H(N), (15)N, (13)CO, (1)Hα/(13)Cα, and (1)Hβ/(13)Cβ chemical shifts) and side chain assignment of D, E, N and Q residues. Such an exhaustive assignment has the potential of yielding accurate 3D structures using one or more of several algorithms which calculate structures of the molecules very reliably on the basis of NMR chemical shifts alone. The side chain assignments of D, E, N, and Q will be extremely valuable for interaction studies with different ligands; D and E side chains are known to be involved in majority of catalytic activities. Utility of this experiment has been demonstrated with Ca(2+) bound M-crystallin, which contains largely D, E, N and Q residues at the metal binding sites.

  15. Quantitative and qualitative 1H, 13C, and 15N NMR spectroscopic investigation of the urea-formaldehyde resin synthesis.

    PubMed

    Steinhof, Oliver; Kibrik, Éléonore J; Scherr, Günter; Hasse, Hans

    2014-04-01

    Urea-formaldehyde resins are bulk products of the chemical industry. Their synthesis involves a complex reaction network. The present work contributes to its elucidation by presenting results from detailed NMR spectroscopic studies with different methods. Besides (1)H NMR and (13)C NMR, (15)N NMR spectroscopy is also applied. (15)N-enriched urea was used for the investigations. A detailed NMR signal assignment and a model of the reaction network of the hydroxymethylation step of the synthesis are presented. Because of its higher spectral dispersion and the fact that all key reactions directly involve the nitrogen centers, (15)N NMR provides a much larger amount of detail than do (1)H and (13)C NMR spectroscopy. Symmetric and asymmetric dimethylol urea can be clearly distinguished and separated from monomethylol urea, trimethylol urea, and methylene-bridged urea. The existence of hemiformals of methylol urea is confirmed. 1,3,5-Oxadiazinan-4-on (uron) and its derivatives were not found in the reaction mixtures investigated here but were prepared via alternative routes. The molar ratios of formaldehyde to urea were 1, 2, and 4, the pH values 7.5 and 8.5, and the reaction temperature 60 °C.

  16. 15N solid-state NMR provides a sensitive probe of oxidized flavin reactive sites.

    PubMed

    Koder, Ronald L; Walsh, Joseph D; Pometun, Maxim S; Dutton, P Leslie; Wittebort, Richard J; Miller, Anne-Frances

    2006-11-29

    Flavins are central to the reactivity of a wide variety of enzymes and electron transport proteins. There is great interest in understanding the basis for the different reactivities displayed by flavins in different protein contexts. We propose solid-state nuclear magnetic resonance (SS-NMR) as a tool for directly observing reactive positions of the flavin ring and thereby obtaining information on their frontier orbitals. We now report the SS-NMR signals of the redox-active nitrogens N1 and N5, as well as that of N3. The chemical shift tensor of N5 is over 720 ppm wide, in accordance with the predictions of theory and our calculations. The signal of N3 can be distinguished on the basis of coupling to 1H absent for N1 and N5, as well as the shift tensor span of only 170 ppm, consistent with N3's lower aromaticity and lack of a nonbonding lone pair. The isotropic shifts and spans of N5 and N1 reflect two opposite extremes of the chemical shift range for "pyridine-type" N's, consistent with their electrophilic and nucleophilic chemical reactivities, respectively. Upon flavin reduction, N5's chemical shift tensor contracts dramatically to a span of less than 110 ppm, and the isotropic chemical shift changes by approximately 300 ppm. Both are consistent with loss of N5's nonbonding lone pair and decreased aromaticity, and illustrate the responsiveness of the 15N chemical shift principal values to electronic structure. Thus. 15N chemical shift principal values promise to be valuable tools for understanding electronic differences that underlie variations in flavin reactivity, as well as the reactivities of other heterocyclic cofactors.

  17. NMR assignments of the N-terminal domain of Nephila clavipes spidroin 1.

    PubMed

    Parnham, Stuart; Gaines, William A; Duggan, Brendan M; Marcotte, William R; Hennig, Mirko

    2011-10-01

    The building blocks of spider dragline silk are two fibrous proteins secreted from the major ampullate gland named spidroins 1 and 2 (MaSp1, MaSp2). These proteins consist of a large central domain composed of approximately 100 tandem copies of a 35-40 amino acid repeat sequence. Non-repetitive N and C-terminal domains, of which the C-terminal domain has been implicated to transition from soluble and insoluble states during spinning, flank the repetitive core. The N-terminal domain until recently has been largely unknown due to difficulties in cloning and expression. Here, we report nearly complete assignment for all (1)H, (13)C, and (15)N resonances in the 14 kDa N-terminal domain of major ampullate spidroin 1 (MaSp1-N) of the golden orb-web spider Nephila clavipes.

  18. Distribution of 15N Among Plant Parts of Nodulating and Nonnodulating Isolines of Soybeans 1

    PubMed Central

    Shearer, Georgia; Kohl, Daniel H.; Harper, James E.

    1980-01-01

    Differences among plant parts in the natural abundance of 15N are of interest from the point of view of developing a sampling strategy for using 15N measurements to estimate the contribution of symbiotically fixed N to N2 fixing plants, and because they reflect isotopic fractionation associated with degradation, transport, and resynthesis of N-bearing molecules. This paper reports such differences in nodulating and nonnodulating isolines of soybeans (Glycine max [L] (Merrill, variety Harosoy)) grown under several different conditions. Nodules were strikingly enriched in 15N compared to other plant parts (by an average of 8.3‰ excess 15N), and the enrichment increased with time during the growing season. 15N was much more uniformly distributed among other plant parts. Although there were significant differences among other plant parts, the maximum deviation of the 15N abundance of any plant part from that of the entire plant was about 2‰ 15N excess. The 15N abundance of the seed N was most representative of the whole plant. There were significant differences between isolines in the distribution of 15N. The distribution of 15N within plants also varied with experimental conditions. The implications of these results for estimation of N2 fixation from measurements of the natural abundance of 15N are discussed. PMID:16661393

  19. The “Speedy” Synthesis of Atom-Specific 15N Imino/Amido-Labeled RNA

    PubMed Central

    Kreutz, Christoph; Micura, Ronald

    2016-01-01

    Although numerous reports on the synthesis of atom-specific 15N-labeled nucleosides exist, fast and facile access to the corresponding phosphoramidites for RNA solid-phase synthesis is still lacking. This situation represents a severe bottleneck for NMR spectroscopic investigations on functional RNAs. Here, we present optimized procedures to speed up the synthesis of 15N(1) adenosine and 15N(1) guanosine amidites, which are the much needed counterparts of the more straightforward-to-achieve 15N(3) uridine and 15N(3) cytidine amidites in order to tap full potential of 1H/15N/15N-COSY experiments for directly monitoring individual Watson–Crick base pairs in RNA. Demonstrated for two preQ1 riboswitch systems, we exemplify a versatile concept for individual base-pair labeling in the analysis of conformationally flexible RNAs when competing structures and conformational dynamics are encountered. PMID:26237536

  20. Climbing up the vibrational ladder of HC15N: High-temperature near-infrared emission measurements

    NASA Astrophysics Data System (ADS)

    Predoi-Cross, A.; Johnson, E.; Hemsing, D.; Rozario, H.; Mellau, Georg Ch.

    2013-11-01

    The near-infrared (NIR) emission spectrum of H12C15N was measured in the 6050-6500 cm-1 range at a resolution of 0.044 cm-1 using an emission setup available at the Justus-Liebig Universität, Giessen, Germany. The rich emission spectrum showing numerous overlapped spectral profiles was analyzed with the spectrum analysis software SyMath running using Mathematica as a platform. This approach allowed us to retrieve the vibrational-rotational constants for 32 bands. Many spectroscopic transitions have been observed for the first time in a lab environment. We quantified four newly observed Coriolis resonances.

  1. Estimation of nitric oxide synthase activity via LC-MS/MS determination of 15N3-citrulline in biological samples

    PubMed Central

    Shin, Beom Soo; Fung, Ho-Leung; Upadhyay, Mahesh; Shin, Soyoung

    2015-01-01

    Rationale We showed that the metabolite peaks of 15N3-citrulline (15N3-CIT) and 15N3-arginine (15N3-ARG) could be detected when 15N4-ARG was metabolized by nitric oxide synthase (NOS) in endothelial cells. The usefulness of these metabolites as potential surrogate indices of nitric oxide (NO) generation is evaluated. Methods A hydrophilic-interaction liquid chromatography electrospray tandem mass spectrometric assay (LC-MS/MS) was utilized for the simultaneous analysis of 15N4-ARG, ARG, CIT, 15N3-CIT and 15N3-ARG. 15N3-CIT and 15N3-ARG from impurities of 15N4-ARG were determined and corrected for the calculation of their concentration. 15N4-ARG-derived NO, i.e., 15NO formation was determined by analyzing 15N-nitrite accumulation by another LC-MS/MS assay. Results After EA.hy926 human endothelial cells were challenged with 15N4-ARG for 2 hours, the peak intensities of 15N3-CIT and 15N3-ARG significantly increased with 15N4-ARG concentration and positively correlated with 15N-nitrite production. The estimated Km values were independent of the metabolite (i.e., 15N3-CIT, 15N3-CIT+15N3-ARG or 15N-nitrite) used for calculation. However, after correction for its presence as a chemical contaminant of 15N4-ARG, 15N3-ARG was only a marginal contributor for the estimation of NOS activity. Conclusions These data suggest that the formation of 15N3-CIT can be used as an indicator of NOS activity when 15N4-ARG is used as a substrate. This approach may be superior to the radioactive 14C-CIT method which can be contaminated by 14C-urea, and to the 14N-nitrite method which lacks sensitivity. PMID:26349467

  2. Direct and cost-efficient hyperpolarization of long-lived nuclear spin states on universal 15N2-diazirine molecular tags

    PubMed Central

    Theis, Thomas; Ortiz, Gerardo X.; Logan, Angus W. J.; Claytor, Kevin E.; Feng, Yesu; Huhn, William P.; Blum, Volker; Malcolmson, Steven J.; Chekmenev, Eduard Y.; Wang, Qiu; Warren, Warren S.

    2016-01-01

    Conventional magnetic resonance (MR) faces serious sensitivity limitations which can be overcome by hyperpolarization methods, but the most common method (dynamic nuclear polarization) is complex and expensive, and applications are limited by short spin lifetimes (typically seconds) of biologically relevant molecules. We use a recently developed method, SABRE-SHEATH, to directly hyperpolarize 15N2 magnetization and long-lived 15N2 singlet spin order, with signal decay time constants of 5.8 and 23 minutes, respectively. We find >10,000-fold enhancements generating detectable nuclear MR signals that last for over an hour. 15N2-diazirines represent a class of particularly promising and versatile molecular tags, and can be incorporated into a wide range of biomolecules without significantly altering molecular function. PMID:27051867

  3. [15N-flow after in sacco incubation and feeding of sheep and goats with untreated wheat straw or straw treated with 15N horse urine].

    PubMed

    Schubert, R; Flachowsky, G; Bochröder, B

    1994-01-01

    Chopped wheat straw was homogeneously mixed with urine of horses (5.75 gN per 1, 16.88 atom-% 15N-excess) and airtightly stored in plastic containers for 6 months. Three rumen fistulated sheep and goats each were fed with untreated or urine treated straw. Concentrate was added to straw. Untreated and urine treated straw were given in nylon bags and incubated in the rumen of sheep and goats for 1, 3, 6, 12, 24, 48 and 72 hours. A three compartment exponential function was used to fit the measurements of 15N-excess and 15N-amount of bag content. The curves and the calculated partial Y-values of the three compartments show the inflow and outflow of 15N into or from the bags and allow conclusions about the binding of urine N. Most N of urine was not compactly bound by straw during storage. Primarily microbial N was attached to the straw in the rumen. About 6% of urine N were bound more compact to the straw. Similar curves were calculated for 15N-excess and 15N-amount of nylon bags. The curves allow conclusions about tracer flows without quantitative knowledge. There were no significant differences between animal species.

  4. Soil processes drive seasonal variation in retention of 15N tracers in a deciduous forest catchment.

    PubMed

    Goodale, Christine L; Fredriksen, Guinevere; Weiss, Marissa S; McCalley, K; Sparks, Jed P; Thomas, Steven A

    2015-10-01

    Seasonal patterns of stream nitrate concentration have long been interpreted as demonstrating the central role of plant uptake in regulating stream nitrogen loss from forested catchments. Soil processes are rarely considered as important drivers of these patterns. We examined seasonal variation in N retention in a deciduous forest using three whole-ecosystem 15N tracer additions: in late April (post-snowmelt, pre-leaf-out), late July (mid-growing- season), and late October (end of leaf-fall). We expected that plant 15N uptake would peak in late spring and midsummer, that immobilization in surface litter and soil would peak the following autumn leaf-fall, and that leaching losses would vary inversely with 15N retention. Similar to most other 15N tracer studies, we found that litter and soils dominated ecosystem retention of added 15N. However, 15N recovery in detrital pools varied tremendously by season, with > 90% retention in spring and autumn and sharply reduced 15N retention in late summer. During spring, over half of the 15N retained in soil occurred within one day in the heavy (mineral-associated) soil fraction. During summer, a large decrease in 15N retention one week after addition coincided with increased losses of 15NO3- to soil leachate and seasonal increases in soil and stream NO3- concentrations, although leaching accounted for only a small fraction of the lost 15N (< 0.2%). Uptake of 15N into roots did not vary by season and accounted for < 4% of each tracer addition. Denitrification or other processes that lead to N gas loss may have consumed the rest. These measurements of 15N movement provide strong evidence for the dominant role of soil processes in regulating seasonal N retention and losses in this catchment and perhaps others with similar soils.

  5. Nutrient Status and δ15N Values in Leaves and Soils: A Cross-Biome Comparison

    NASA Astrophysics Data System (ADS)

    Mayor, J. R.; Schuur, E. A.; Turner, B. L.; Wright, S. J.

    2011-12-01

    Stable nitrogen (N) isotope ratios (δ15N) are often assumed to provide an integrated measure of multiple nitrogen cycling processes. For instance, shifts in the bioavailability of soil N forms are thought to alter plant δ15N values. Demonstrating this relationship is important as ecosystems undergo anthropogenic disturbances. We evaluated patterns and implied mechanisms of the N cycle using ecosystem δ15N values from 16 plots in boreal black spruce (Picea mariana) forest and lowland wet tropical forest. Fertilizer N and phosphorus (P) was applied annually for five and 11 years prior to measurement of ecosystem δ15N values. Full sun canopy foliage and soil extractable nitrate, ammonium, and dissolved organic N (DON) were sampled in fertilized and control plots and analyzed for δ15N. In boreal forest, N fertilization reduced DON concentrations and caused a depletion of δ15N in foliage and fungal sporocarps. Of four species occurring in all plots in the tropical forest, one (Alseis blackiana) had increased foliar δ15N values following N fertilization, one (Tetragastris panamensis) had increased foliar δ15N values following P fertilization, and one (Oenocarpus mapora) had increased foliar δ15N following N+P fertilization. Surprisingly, soil nitrate in the boreal forest became substantially 15N-enriched under P fertilization, whereas nitrate in the tropical forest soil was enriched only under N or N+P fertilization. Collectively, nitrate enrichment is likely due to enhanced rates of soil denitrification as evidenced by elevated resin extractable soil nitrate concentrations and close correlations between δ15N and δ18O values. On average, foliar δ15N in tropical trees corresponded well with δ15N in soil nitrate in control and P fertilized plots, but was 2-3% more enriched than DON under N and N+P fertilization. In boreal forests, N and N+P fertilization increased foliar N concentration and δ15N values indicating substantial use of applied fertilizer. Taken

  6. (15)N CSA tensors and (15)N-(1)H dipolar couplings of protein hydrophobic core residues investigated by static solid-state NMR.

    PubMed

    Vugmeyster, Liliya; Ostrovsky, Dmitry; Fu, Riqiang

    2015-10-01

    In this work, we assess the usefulness of static (15)N NMR techniques for the determination of the (15)N chemical shift anisotropy (CSA) tensor parameters and (15)N-(1)H dipolar splittings in powder protein samples. By using five single labeled samples of the villin headpiece subdomain protein in a hydrated lyophilized powder state, we determine the backbone (15)N CSA tensors at two temperatures, 22 and -35 °C, in order to get a snapshot of the variability across the residues and as a function of temperature. All sites probed belonged to the hydrophobic core and most of them were part of α-helical regions. The values of the anisotropy (which include the effect of the dynamics) varied between 130 and 156 ppm at 22 °C, while the values of the asymmetry were in the 0.32-0.082 range. The Leu-75 and Leu-61 backbone sites exhibited high mobility based on the values of their temperature-dependent anisotropy parameters. Under the assumption that most differences stem from dynamics, we obtained the values of the motional order parameters for the (15)N backbone sites. While a simple one-dimensional line shape experiment was used for the determination of the (15)N CSA parameters, a more advanced approach based on the "magic sandwich" SAMMY pulse sequence (Nevzorov and Opella, 2003) was employed for the determination of the (15)N-(1)H dipolar patterns, which yielded estimates of the dipolar couplings. Accordingly, the motional order parameters for the dipolar interaction were obtained. It was found that the order parameters from the CSA and dipolar measurements are highly correlated, validating that the variability between the residues is governed by the differences in dynamics. The values of the parameters obtained in this work can serve as reference values for developing more advanced magic-angle spinning recoupling techniques for multiple labeled samples.

  7. Proton-detected 3D (15)N/(1)H/(1)H isotropic/anisotropic/isotropic chemical shift correlation solid-state NMR at 70kHz MAS.

    PubMed

    Pandey, Manoj Kumar; Yarava, Jayasubba Reddy; Zhang, Rongchun; Ramamoorthy, Ayyalusamy; Nishiyama, Yusuke

    2016-01-01

    Chemical shift anisotropy (CSA) tensors offer a wealth of information for structural and dynamics studies of a variety of chemical and biological systems. In particular, CSA of amide protons can provide piercing insights into hydrogen-bonding interactions that vary with the backbone conformation of a protein and dynamics. However, the narrow span of amide proton resonances makes it very difficult to measure (1)H CSAs of proteins even by using the recently proposed 2D (1)H/(1)H anisotropic/isotropic chemical shift (CSA/CS) correlation technique. Such difficulties due to overlapping proton resonances can in general be overcome by utilizing the broad span of isotropic chemical shifts of low-gamma nuclei like (15)N. In this context, we demonstrate a proton-detected 3D (15)N/(1)H/(1)H CS/CSA/CS correlation experiment at fast MAS frequency (70kHz) to measure (1)H CSA values of unresolved amide protons of N-acetyl-(15)N-l-valyl-(15)N-l-leucine (NAVL).

  8. Factors Controlling the Stable Nitrogen Isotopic Composition (δ15N) of Lipids in Marine Animals

    PubMed Central

    Svensson, Elisabeth; Schouten, Stefan; Hopmans, Ellen C.; Middelburg, Jack J.; Sinninghe Damsté, Jaap S.

    2016-01-01

    Lipid extraction of biomass prior to stable isotope analysis is known to cause variable changes in the stable nitrogen isotopic composition (δ15N) of residual biomass. However, the underlying factors causing these changes are not yet clear. Here we address this issue by comparing the δ15N of bulk and residual biomass of several marine animal tissues (fish, crab, cockle, oyster, and polychaete), as well as the δ15N of the extracted lipids. As observed previously, lipid extraction led to a variable offset in δ15N of biomass (differences ranging from -2.3 to +1.8 ‰). Importantly, the total lipid extract (TLE) was highly depleted in 15N compared to bulk biomass, and also highly variable (differences ranging from -14 to +0.7 ‰). The TLE consisted mainly of phosphatidylcholines, a group of lipids with one nitrogen atom in the headgroup. To elucidate the cause for the 15N-depletion in the TLE, the δ15N of amino acids was determined, including serine because it is one of the main sources of nitrogen to N-containing lipids. Serine δ15N values differed by -7 to +2 ‰ from bulk biomass δ15N, and correlated well with the 15N depletion in TLEs. On average, serine was less depleted (-3‰) than the TLE (-7 ‰), possibly due to fractionation during biosynthesis of N-containing headgroups, or that other nitrogen-containing compounds, such as urea and choline, or recycled nitrogen contribute to the nitrogen isotopic composition of the TLE. The depletion in 15N of the TLE relative to biomass increased with the trophic level of the organisms. PMID:26731720

  9. Factors Controlling the Stable Nitrogen Isotopic Composition (δ15N) of Lipids in Marine Animals.

    PubMed

    Svensson, Elisabeth; Schouten, Stefan; Hopmans, Ellen C; Middelburg, Jack J; Sinninghe Damsté, Jaap S

    2016-01-01

    Lipid extraction of biomass prior to stable isotope analysis is known to cause variable changes in the stable nitrogen isotopic composition (δ15N) of residual biomass. However, the underlying factors causing these changes are not yet clear. Here we address this issue by comparing the δ15N of bulk and residual biomass of several marine animal tissues (fish, crab, cockle, oyster, and polychaete), as well as the δ15N of the extracted lipids. As observed previously, lipid extraction led to a variable offset in δ15N of biomass (differences ranging from -2.3 to +1.8 ‰). Importantly, the total lipid extract (TLE) was highly depleted in 15N compared to bulk biomass, and also highly variable (differences ranging from -14 to +0.7 ‰). The TLE consisted mainly of phosphatidylcholines, a group of lipids with one nitrogen atom in the headgroup. To elucidate the cause for the 15N-depletion in the TLE, the δ15N of amino acids was determined, including serine because it is one of the main sources of nitrogen to N-containing lipids. Serine δ15N values differed by -7 to +2 ‰ from bulk biomass δ15N, and correlated well with the 15N depletion in TLEs. On average, serine was less depleted (-3‰) than the TLE (-7 ‰), possibly due to fractionation during biosynthesis of N-containing headgroups, or that other nitrogen-containing compounds, such as urea and choline, or recycled nitrogen contribute to the nitrogen isotopic composition of the TLE. The depletion in 15N of the TLE relative to biomass increased with the trophic level of the organisms.

  10. Elucidating the Structures of the Low- and High-pH Mo(V) Species in Respiratory Nitrate Reductase: A Combined EPR, (14,15)N HYSCORE, and DFT Study.

    PubMed

    Rendon, Julia; Biaso, Frédéric; Ceccaldi, Pierre; Toci, René; Seduk, Farida; Magalon, Axel; Guigliarelli, Bruno; Grimaldi, Stéphane

    2017-03-31

    Respiratory nitrate reductases (Nars), members of the prokaryotic Mo/W-bis Pyranopterin Guanosine dinucleotide (Mo/W-bisPGD) enzyme superfamily, are key players in nitrate respiration, a major bioenergetic pathway widely used by microorganisms to cope with the absence of dioxygen. The two-electron reduction of nitrate to nitrite takes place at their active site, where the molybdenum ion cycles between Mo(VI) and Mo(IV) states via a Mo(V) intermediate. The active site shows two distinct pH-dependent Mo(V) electron paramagnetic resonance (EPR) signals whose structure and catalytic relevance have long been debated. In this study, we use EPR and HYSCORE techniques to probe their nuclear environment in Escherichia coli Nar (EcNar). By using samples prepared at different pH and through different enrichment strategies in (98)Mo and (15)N nuclei, we demonstrate that each of the two Mo(V) species is coupled to a single nitrogen nucleus with similar quadrupole characteristics. Structure-based density functional theory calculations allow us to propose a molecular model of the low-pH Mo(V) species consistent with EPR spectroscopic data. Our results show that the metal ion is coordinated by a monodentate aspartate ligand and permit the assignment of the coupled nitrogen nuclei to the Nδ of Asn52, a residue located ∼3.9 Å to the Mo atom in the crystal structures. This is confirmed by measurements on selectively (15)N-Asn labeled EcNar. Further, we propose a Mo-O(H)···HN structure to account for the transfer of spin density onto the interacting nitrogen nucleus deduced from HYSCORE analysis. This work provides a foundation for monitoring the structure of the molybdenum active site in the presence of various substrates or inhibitors in Nars and other molybdenum enzymes.

  11. δ(15) N from soil to wine in bulk samples and proline.

    PubMed

    Paolini, Mauro; Ziller, Luca; Bertoldi, Daniela; Bontempo, Luana; Larcher, Roberto; Nicolini, Giorgio; Camin, Federica

    2016-09-01

    The feasibility of using δ(15) N as an additional isotopic marker able to link wine to its area of origin was investigated. The whole production chain (soil-leaves-grape-wine) was considered. Moreover, the research included evaluation of the effect of the fermentation process, the use of different types of yeast and white and red vinification, the addition of nitrogen adjuvants and ultrasound lysis simulating wine ageing. The δ(15) N of grapes and wine was measured in bulk samples and compounds, specifically in proline, for the first time. Despite isotopic fractionation from soil to wine, the δ(15) N values of leaves, grapes, wine and particularly must and wine proline conserved the variability of δ(15) N in the growing soil. Fermentation and ultrasound treatment did not affect the δ(15) N values of grape must, which was therefore conserved in wine. The addition of inorganic or organic adjuvants was able to influence the δ(15) N of bulk wine, depending on the amount and the difference between the δ(15) N of must and that of the adjuvant. The δ(15) N of wine proline was not influenced by adjuvant addition and is therefore the best marker for tracing the geographical origin of wine. Copyright © 2016 John Wiley & Sons, Ltd.

  12. Disturbance and topography shape nitrogen availability and δ15N over long-term forest succession

    EPA Science Inventory

    Forest disturbance and long-term succession can promote open N cycling that increases N loss and soil δ15N values. We examined soil and foliar patterns in N and δ15N, and soil N mineralization, across a topographically complex montane forest landscape influenced by human logging ...

  13. δ 15 N constraints on long-term nitrogen balances in temperate forests

    EPA Science Inventory

    Natural abundance δ15N of ecosystems integrates nitrogen (N) inputs and losses, and thus reflects factors that control the long-term development of ecosystem N balances. We here report N and carbon (C) content of forest vegetation and soils, and associated δ15N, across nine Doug...

  14. Accessible NMR Experiments Studying the Hydrodynamics of [subscript 15]N-Enriched Ubiquitin at Low Fields

    ERIC Educational Resources Information Center

    Thompson, Laura E.; Rovnyak, David

    2007-01-01

    We have recently developed and implemented two experiments in biomolecular NMR for an undergraduate-level biophysical chemistry laboratory with commercially available [subscript 15]N-enriched human ubiquitin. These experiments take advantage of [subscript 15]N direct detection of the NMR signal. The first experiment develops skills in acquiring…

  15. Accessible NMR Experiments Studying the Hydrodynamics of [superscript 15]N-Enriched Ubiquitin at Low Fields

    ERIC Educational Resources Information Center

    Thompson, Laura E.; Rovnyak, David

    2007-01-01

    We have recently developed and implemented two experiments in biomolecular NMR for an undergraduate-level biophysical chemistry laboratory with commercially available [superscript 15]N-enriched human ubiquitin. These experiments take advantage of [superscript 15]N direct detection of the NMR signal. The first experiment develops skills in…

  16. Stable Isotope Tracking of Endangered Sea Turtles: Validation with Satellite Telemetry and δ15N Analysis of Amino Acids

    PubMed Central

    Seminoff, Jeffrey A.; Benson, Scott R.; Arthur, Karen E.; Eguchi, Tomoharu; Dutton, Peter H.; Tapilatu, Ricardo F.; Popp, Brian N.

    2012-01-01

    Effective conservation strategies for highly migratory species must incorporate information about long-distance movements and locations of high-use foraging areas. However, the inherent challenges of directly monitoring these factors call for creative research approaches and innovative application of existing tools. Highly migratory marine species, such as marine turtles, regularly travel hundreds or thousands of kilometers between breeding and feeding areas, but identification of migratory routes and habitat use patterns remains elusive. Here we use satellite telemetry in combination with compound-specific isotope analysis of amino acids to confirm that insights from bulk tissue stable isotope analysis can reveal divergent migratory strategies and within-population segregation of foraging groups of critically endangered leatherback sea turtles (Dermochelys coriacea) across the Pacific Ocean. Among the 78 turtles studied, we found a distinct dichotomy in δ15N values of bulk skin, with distinct “low δ15N” and “high δ15N” groups. δ15N analysis of amino acids confirmed that this disparity resulted from isotopic differences at the base of the food chain and not from differences in trophic position between the two groups. Satellite tracking of 13 individuals indicated that their bulk skin δ15N value was linked to the particular foraging region of each turtle. These findings confirm that prevailing marine isoscapes of foraging areas can be reflected in the isotopic compositions of marine turtle body tissues sampled at nesting beaches. We use a Bayesian mixture model to show that between 82 and 100% of the 78 skin-sampled turtles could be assigned with confidence to either the eastern Pacific or western Pacific, with 33 to 66% of all turtles foraging in the eastern Pacific. Our forensic approach validates the use of stable isotopes to depict leatherback turtle movements over broad spatial ranges and is timely for establishing wise conservation efforts in

  17. Preparation and characterization of 15N-enriched, size-defined heparan sulfate precursor oligosaccharides

    PubMed Central

    Sigulinsky, Crystal; Babu, Ponnusamy; Victor, Xylophone V.; Kuberan, Balagurunathan

    2009-01-01

    We report the preparation of size-defined [15N]N-acetylheparosan oligosaccharides from Escherichia coli-derived 15N-enriched N-acetylheparosan. Optimized growth conditions of E. coli in minimal media containing 15NH4Cl yielded [15N]N-acetylheparosan on a preparative scale. Depolymerization of [15N]N-acetylheparosan by heparitinase I yielded resolvable, even-numbered oligosaccharides ranging from disaccharide to icosaccharide. Anion-exchange chromatography-assisted fractionation afforded size-defined [15N]N-acetylheparosan oligosaccharides identifiable by ESI-TOFMS. These isotopically labeled oligosaccharides will prove to be valuable research tools for the chemoenzymatic synthesis of heparin and heparan sulfate oligosaccharides and for the study of their structural biology. PMID:19945695

  18. Symbiotic nitrogen fixation in an arid ecosystem measured by sup 15 N natural abundance

    SciTech Connect

    Johnson, G.V. )

    1990-05-01

    Plants dependent on nitrogen fixation have an {sup 15}N abundance similar to the atmosphere, while non-nitrogen fixing plants usually are enriched in {sup 15}N and are similar to soil nitrogen values. The natural abundance of {sup 15}N in leaf tissues and soils was determined to evaluate symbiotic nitrogen fixation by several legumes and actinorhizal species in the Sevilleta Long-term Ecological Research area in central New Mexico. Comparison of {delta}{sup 15}N values for the legume Prosopis glandulosa (mesquite) to adjacent Atriplex canascens (fourwing saltbush) indicated that P. glandulosa obtained 66% of its nitrogen by fixation. The legume Hoffmanseggia jamesii was found to be utilizing soil nitrogen. The {delta}{sup 15}N values for the actinorhizal plants, Elaeagnus angustifolia and Cercocarpus montanus, while below values for soil nitrogen, did not differ from associated non-fixing plants.

  19. Compound-specific 15N analysis of amino acids in 15N tracer experiments provide an estimate of newly synthesised soil protein from inorganic and organic substrates

    NASA Astrophysics Data System (ADS)

    Charteris, Alice; Michaelides, Katerina; Evershed, Richard

    2015-04-01

    Organic N concentrations far exceed those of inorganic N in most soils and despite much investigation, the composition and cycling of this complex pool of SOM remains poorly understood. A particular problem has been separating more recalcitrant soil organic N from that actively cycling through the soil system; an important consideration in N cycling studies and for the soil's nutrient supplying capacity. The use of 15N-labelled substrates as stable isotope tracers has contributed much to our understanding of the soil system, but the complexity and heterogeneity of soil organic N prevents thorough compound-specific 15N analyses of organic N compounds and makes it difficult to examine any 15N-labelled organic products in any detail. As a result, a significant proportion of previous work has either simply assumed that since the majority of soil N is organic, all of the 15N retained in the soil is organic N (e.g. Sebilo et al., 2013) or subtracted 15N-labelled inorganic compounds from bulk values (e.g. Pilbeam et al., 1997). While the latter approach is more accurate, these methods only provide an estimate of the bulk 15N value of an extremely complex and non-uniformly labelled organic pool. A more detailed approach has been to use microbial biomass extraction (Brookes et al., 1985) and subsequent N isotopic analysis to determine the 15N value of biomass-N, representing the fraction of 15N assimilated by microbes or the 15N cycling through the 'living' or 'active' portion of soil organic N. However, this extraction method can only generate estimates and some lack of confidence in its validity and reliability remains. Here, we present an alternative technique to obtain a measure of the assimilation of an applied 15N substrate by the soil microbial biomass and an estimate of the newly synthesized soil protein, which is representative of the magnitude of the active soil microbial biomass. The technique uses a stable isotope tracer and compound-specific 15N analysis, but

  20. Disturbance and topography shape nitrogen availability and δ15 N over long-term forest succession

    USGS Publications Warehouse

    Perakis, Steven; Tepley, Alan J.; Compton, Jana

    2015-01-01

    Forest disturbance and long-term succession towards old-growth are thought to increase nitrogen (N) availability and N loss, which should increase soil δ15N values. We examined soil and foliar patterns in N and δ15N, and soil N mineralization, across 800 years of forest succession in a topographically complex montane landscape influenced by human logging and wildfire. In contrast to expectations, we found that disturbance caused declines in surface mineral soil δ15N values, both in logged forests measured 40–50 years after disturbance, and in unlogged forests disturbed by severe wildfire within the last 200 years. Both symbiotic N fixation and N transfers from disturbed vegetation and detritus could lower soil δ15N values after disturbance. A more important role for symbiotic N fixation is suggested by lower soil δ15N values in slow-successional sites with slow canopy closure, which favors early-successional N fixers. Soil δ15N values increased only marginally throughout 800 years of succession, reflecting soil N uptake by vegetation and strong overall N retention. Although post-disturbance N inputs lowered surface soil δ15N values, steady-state mass balance calculations suggest that wildfire combustion of vegetation and detritus can dominate long-term N loss and increase whole-ecosystem δ15N. On steeper topography, declining soil δ15N values highlight erosion and accelerated soil turnover as an additional abiotic control on N balances. We conclude for N-limited montane forests that soil δ15N and N availability are less influenced by nitrate leaching and denitrification loss than by interactions between disturbance, N fixation, and erosion.

  1. The effect of manuring on cereal and pulse amino acid δ(15)N values.

    PubMed

    Styring, Amy K; Fraser, Rebecca A; Bogaard, Amy; Evershed, Richard P

    2014-06-01

    Amino acid δ(15)N values of barley (Hordeum vulgare) and bread wheat (Triticum aestivum) grains and rachis and broad bean (Vicia faba) and pea (Pisum sativum) seeds, grown in manured and unmanured soil at the experimental farm stations of Rothamsted, UK and Bad Lauchstädt, Germany, were determined by GC-C-IRMS. Manuring was found to result in a consistent (15)N-enrichment of cereal grain amino acid δ(15)N values, indicating that manuring did not affect the metabolic routing of nitrogen (N) into cereal grain amino acids. The increase in cereal grain δ(15)N values with manuring is therefore due to a (15)N-enrichment in the δ(15)N value of assimilated inorganic-N. Greater variation was observed in the (15)N-enrichment of rachis amino acids with manuring, possibly due to enhanced sensitivity to changes in growing conditions and higher turnover of N in rachis cells compared to cereal grains. Total amino acid δ(15)N values of manured and unmanured broad beans and peas were very similar, indicating that the legumes assimilated N2 from the atmosphere rather than N from the soil, since there was no evidence for routing of (15)N-enriched manure N into any of the pulse amino acids. Crop amino acid δ(15)N values thus provide insights into the sources of N assimilated by non N2-fixing and N2-fixing crops grown on manured and unmanured soils, and reveal an effect of manure on N metabolism in different crop species and plant parts.

  2. 15N Content Reflects Development of Mycorrhizae and Nitrogen Dynamics During Primary Succession

    NASA Astrophysics Data System (ADS)

    Hobbie, E. A.; Jumpponen, A.

    2004-05-01

    Mycorrhizal fungi are ubiquitous symbionts on terrestrial plants that are particularly important for plant nitrogen nutrition. 15N content appears to be a useful marker of the mycorrhizal role in plant nitrogen supply because of an apparent fractionation against 15N during transfer of nitrogen from mycorrhizal fungi to host plants. Because plants developing during primary succession are gradually colonized by mycorrhizal fungi, such situations provide good opportunities to study interactions between mycorrhizal colonization and plant 15N content. Here, we present results of a study of nitrogen isotope patterns in ecosystem components during the first 100 years of ecosystem development after glacial retreat, and compare those patterns with those on adjacent mature terrain. Soils in primary succession were depleted in 15N relative to nitrogen-fixing plants. Nonmycorrhizal plants and plants generally colonized by ectomycorrhizal, ericoid, or arbuscular fungi showed similar 15N content very early in succession (-4 to -6‰ ), corresponding to low colonization levels of all plant species. Subsequent colonization of evergreen plants by ectomycorrhizal and ericoid fungi led to a 5-6‰ decline in 15N content, indicating transfer of 15N-depleted N from fungi to plants. The values recorded (-10 to -14‰ ) are among the lowest yet observed in vascular plants. Nonmycorrhizal plants and plants colonized by arbuscular mycorrhizal fungi did not decline in 15N content. Most ectomycorrhizal and saprotrophic fungi were similar in 15N content in early succession (-1 to -3‰ ), with the notable exception of ectomycorrhizal fungi suspected of proteolytic capabilities, which were 15N enriched relative to all other fungi. 15N contents in both plants and soil from the mature site were 5‰ greater than in recently exposed sites. We conclude that 1) the primary nitrogen source to this ecosystem must be atmospheric deposition, 2) low plant 15N content generally corresponds with greater

  3. Climate-Dependence of Plant-Soil 15N/14N Interactions Across Tropical Rainforests

    NASA Astrophysics Data System (ADS)

    Houlton, B. Z.; Sigman, D. M.; Hedin, L. O.

    2005-12-01

    In most areas of the world, the 15N/14N of bulk soils is higher than that of plant leaves, and the isotopic signatures of these two ecosystem N pools progressively diverge with increasing rainfall. However, both the cause for this isotopic trend and its implications for understanding interactions between climate and N cycles are largely unknown. We report 15N/14N measurements of nitrate, ammonium, and total dissolved N in soil extracts from a highly constrained rainfall sequence in Hawaii, across which this trend in ecosystem 15N/14N is captured, to examine the competing explanations for plant-soil 15N/14N uncouplings. While the isotopic influences of microbial transfers of N between nitrate and ammonium pools and plant-mycorrhizae interactions have been posited in plant-soil 15N/14N relationships, our data did not support an important role for either of these mechanisms. Instead, preferential regeneration of 14N during the breakdown of DON to ammonium explains why the 15N/14N of plants is lower than that of bulk soils. Fractionation at this step leads to two isotopically distinct N subcycles in each forest, a lower-15N/14N subcycle composed of ammonium, nitrate, and bulk plant biomass N that `spins' rapidly and a higher-15N/14N subcycle composed of bulk soil N and DON that is much less dynamic. The increased difference between soil and plant 15N/14N is due to changes in the impacts of nitrification and denitrification on the 15N/14N of ammonium and nitrate, coupled with a switch from nitrate to ammonium uptake by plants under the wettest conditions. For instance, the particularly large (~6 per mil) 15N/14N difference between plants and soils in the wettest sites is due to the lack of 15N-enrichment of ammonium by nitrification coupled with plant dependence on ammonium uptake only. Our results highlight the importance of interactions between DON breakdown, ecosystem N recycling, and gaseous N losses in the explaining the interactions between the 15N signatures of

  4. Alkaline Hydrolysis/Polymerization of 2,4,6-Trinitrotoluene: Characterization of Products by 13C and 15N NMR

    USGS Publications Warehouse

    Thorn, K.A.; Thorne, P.G.; Cox, L.G.

    2004-01-01

    Alkaline hydrolysis has been investigated as a nonbiological procedure for the destruction of 2,4,6-trinitrotoluene (TNT) in explosives contaminated soils and munitions scrap. Nucleophilic substitutions of the nitro and methyl groups of TNT by hydroxide ion are the initial steps in the alkaline degradation of TNT. Potential applications of the technique include both in situ surface liming and ex situ alkaline treatment of contaminated soils. A number of laboratory studies have reported the formation of an uncharacterized polymeric material upon prolonged treatment of TNT in base. As part of an overall assessment of alkaline hydrolysis as a remediation technique, and to gain a better understanding of the chemical reactions underlying the hydrolysis/polymerization process, the soluble and precipitate fractions of polymeric material produced from the calcium hydroxide hydrolysis of unlabeled and 15N-labeled TNT were analyzed by elemental analysis and 13C and 15N nuclear magnetic resonance spectroscopy. Spectra indicated that reactions leading to polymerization included nucleophilic displacement of nitro groups by hydroxide ion, formation of ketone, carboxyl, alcohol, ether, and other aliphatic carbons, conversion of methyl groups to diphenyl methylene carbons, and recondensation of aromatic amines and reduced forms of nitrite, including ammonia and possibly hydroxylamine, into the polymer. Compared to the distribution of carbons in TNT as 14% sp 3- and 86% sp2-hybridized, the precipitate fraction from hydrolysis of unlabeled TNT contained 33% sp3- and 67% sp 2-hybridized carbons. The concentration of nitrogen in the precipitate was 64% of that in TNT. The 15N NMR spectra showed that, in addition to residual nitro groups, forms of nitrogen present in the filtrate and precipitate fractions include aminohydroquinone, primary amide, indole, imine, and azoxy, among others. Unreacted nitrite was recovered in the filtrate fraction. The toxicities and susceptibilities to

  5. Nitrogen Fractionation in Protoplanetary Disks from the H13CN/HC15N Ratio

    NASA Astrophysics Data System (ADS)

    Guzmán, V. V.; Öberg, K. I.; Huang, J.; Loomis, R.; Qi, C.

    2017-02-01

    Nitrogen fractionation is commonly used to assess the thermal history of solar system volatiles. With ALMA it is for the first time possible to directly measure {}14{{N}}/{}15{{N}} ratios in common molecules during the assembly of planetary systems. We present ALMA observations of the {{{H}}}13{CN} and {{HC}}15{{N}} J=3-2 lines at 0.″5 angular resolution, toward a sample of six protoplanetary disks, selected to span a range of stellar and disk structure properties. Adopting a typical {}12{{C}}/{}13{{C}} ratio of 70, we find comet-like {}14{{N}}/{}15{{N}} ratios of 80–160 in five of the disks (3 T Tauri and 2 Herbig Ae disks) and lack constraints for one of the T Tauri disks (IM Lup). There are no systematic differences between T Tauri and Herbig Ae disks, or between full and transition disks within the sample. In addition, no correlation is observed between disk-averaged D/H and {}14{{N}}/{}15{{N}} ratios in the sample. One of the disks, V4046 Sgr, presents unusually bright HCN isotopologue emission, enabling us to model the radial profiles of {{{H}}}13{CN} and {{HC}}15{{N}}. We find tentative evidence of an increasing {}14{{N}}/{}15{{N}} ratio with radius, indicating that selective photodissociation in the inner disk is important in setting the {}14{{N}}/{}15{{N}} ratio during planet formation.

  6. Nitrogen stable isotope composition (δ15N) of vehicle-emitted NOx.

    PubMed

    Walters, Wendell W; Goodwin, Stanford R; Michalski, Greg

    2015-02-17

    The nitrogen stable isotope ratio of NOx (δ(15)N-NOx) has been proposed as a regional indicator for NOx source partitioning; however, knowledge of δ(15)N values from various NOx emission sources is limited. This study presents a detailed analysis of δ(15)N-NOx emitted from vehicle exhaust, the largest source of anthropogenic NOx. To accomplish this, NOx was collected from 26 different vehicles, including gasoline and diesel-powered engines, using a modification of a NOx collection method used by the United States Environmental Protection Agency, and δ(15)N-NOx was analyzed. The vehicles sampled in this study emitted δ(15)N-NOx values ranging from -19.1 to 9.8‰ that negatively correlated with the emitted NOx concentrations (8.5 to 286 ppm) and vehicle run time because of kinetic isotope fractionation effects associated with the catalytic reduction of NOx. A model for determining the mass-weighted δ(15)N-NOx from vehicle exhaust was constructed on the basis of average commute times, and the model estimates an average value of -2.5 ± 1.5‰, with slight regional variations. As technology improvements in catalytic converters reduce cold-start emissions in the future, it is likely to increase current δ(15)N-NOx values emitted from vehicles.

  7. Steroselective synthesis and application of L-( sup 15 N) amino acids

    SciTech Connect

    Unkefer, C.J. ); Lodwig, S.N. . Div. of Science)

    1991-01-01

    We have developed two general approaches to the stereoselective synthesis of {sup 15}N- and {sup 13}C-labeled amino acids. First, labeled serine, biosynthesized using the methylotrophic bacterium M. extorquens AM1, serves as a chiral precursor for the synthesis of other amino acids. For example, pyridoxal phosphate enzymes can be used for the conversion of L-({alpha}-{sup 15}N)serine to L-({alpha}-{sup 15}N)tyrosine, L-({alpha}-{sup 15}N)tryptophan, and L-({alpha}-{sup 15}N)cysteine. In the second approach, developed by Oppolzer and Tamura, an electrophilic amination'' reagent, 1-chloro-1-nitrosocyclohexane, was used to convert chiral enolates into L-{alpha}-amino acids. We prepared 1-chloro-1-({sup 15}N) nitrosocyclohexane and used it to aminate chiral enolates to produce L-({alpha}-{sup 15}N)amino acids. The stereoselectivity of this scheme using the Oppolzer sultam chiral auxiliary is remarkable, producing enantiomer ratios of 200 to 1. 22 refs., 4 figs.

  8. The Failed Writing Assignment.

    ERIC Educational Resources Information Center

    Swyt, Wendy

    1995-01-01

    Discusses an unsuccessful English 101 writing assignment in which students were asked to analyze a Gary Larson cartoon. Examines critically the type of assignment that seeks to address and incorporate the student writer's "local knowledge" of cultural texts, while at the same time containing what counts as knowledge within limited parameters of…

  9. Covalent binding of reduced metabolites of [{sup 15}N{sub 3}]TNT to soil organic matter during a bioremediation process analyzed by {sup 15}N NMR spectroscopy

    SciTech Connect

    Achtnich, C.; Fernandes, E.; Bollag, J.M.; Knackmuss, H.J.; Lenke, H.

    1999-12-15

    Evidence is presented for the covalent binding of biologically reduced metabolites of 2,4,6-{sup 15}N{sub 3}-trinitrotoluene (TNT) to different soil fractions, using liquid {sup 15}N NMR spectroscopy. A silylation procedure was used to release soil organic matter from humin and whole soil for spectroscopic measurements. TNT-contaminated soil was spiked with 2,4,6-{sup 15}N{sub 3}-trinitrotoluene and {sup 14}C-ring labeled TNT, before treatment in a soil slurry reactor. During the anaerobic/aerobic incubation the amount of radioactivity detected in the fulvic and humic acid fractions did not change significantly whereas the radioactivity bound to humin increased to 71%. The {sup 15}N NMR spectra of the fulvic acid samples were dominated by a large peak that corresponded to aliphatic amines or ammonia. In the early stages of incubation, {sup 15}N NMR analysis of the humic acids indicated bound azoxy compounds. The signals arising from nitro and azoxy groups disappeared with further anaerobic treatment. At the end of incubation, the NMR shifts showed that nitrogen was covalently bound to humic acid as substituted amines and amides. The NMR spectra of the silylated humin suggest formation of azoxy compounds and imine linkages. Bound metabolites possessing nitro groups were also detected. Primary amines formed during the anaerobic incubation disappeared during the aerobic treatment. Simultaneously, the amount of amides and tertiary amines increased. Nitro and azoxy groups of bound molecules were still present in humin at the end of the incubation period. Formation of azoxy compounds from partially reduced TNT followed by binding and further reduction appears to be an important mechanism for the immobilization of metabolites of TNT to soil.

  10. Carbon-rich Presolar Grains from Massive Stars: Subsolar 12C/13C and 14N/15N Ratios and the Mystery of 15N

    NASA Astrophysics Data System (ADS)

    Pignatari, M.; Zinner, E.; Hoppe, P.; Jordan, C. J.; Gibson, B. K.; Trappitsch, R.; Herwig, F.; Fryer, C.; Hirschi, R.; Timmes, F. X.

    2015-08-01

    Carbon-rich grains with isotopic anomalies compared to the Sun are found in primitive meteorites. They were made by stars, and carry the original stellar nucleosynthesis signature. Silicon carbide grains of Type X and C and low-density (LD) graphites condensed in the ejecta of core-collapse supernovae. We present a new set of models for the explosive He shell and compare them with the grains showing 12C/13C and 14N/15N ratios lower than solar. In the stellar progenitor H was ingested into the He shell and not fully destroyed before the explosion. Different explosion energies and H concentrations are considered. If the supernova shock hits the He-shell region with some H still present, the models can reproduce the C and N isotopic signatures in C-rich grains. Hot-CNO cycle isotopic signatures are obtained, including a large production of 13C and 15N. The short-lived radionuclides 22Na and 26Al are increased by orders of magnitude. The production of radiogenic 22Ne from the decay of 22Na in the He shell might solve the puzzle of the Ne-E(L) component in LD graphite grains. This scenario is attractive for the SiC grains of type AB with 14N/15N ratios lower than solar, and provides an alternative solution for SiC grains originally classified as nova grains. Finally, this process may contribute to the production of 14N and 15N in the Galaxy, helping to produce the 14N/15N ratio in the solar system.

  11. Carbon-rich presolar grains from massive stars. Subsolar 12 C/ 13 C and 14 N/ 15 N ratios and the mystery of 15 N

    DOE PAGES

    Pignatari, M.; Zinner, E.; Hoppe, P.; ...

    2015-07-30

    We compared carbon-rich grains with isotopic anomalies to the Sun are found in primitive meteorites. They were made by stars, and carry the original stellar nucleosynthesis signature. Silicon carbide grains of Type X and C and low-density (LD) graphites condensed in the ejecta of core-collapse supernovae. Furthermore, we present a new set of models for the explosive He shell and compare them with the grains showing 12C/13C and 14N/15N ratios lower than solar. In the stellar progenitor H was ingested into the He shell and not fully destroyed before the explosion. All of the explosion energies and H concentrations aremore » considered. If the supernova shock hits the He-shell region with some H still present, the models can reproduce the C and N isotopic signatures in C-rich grains. Hot-CNO cycle isotopic signatures are obtained, including a large production of 13C and 15N. The short-lived radionuclides 22Na and 26Al are increased by orders of magnitude. The production of radiogenic 22Ne from the decay of 22Na in the He shell might solve the puzzle of the Ne-E(L) component in LD graphite grains. This scenario is attractive for the SiC grains of type AB with 14N/15N ratios lower than solar, and provides an alternative solution for SiC grains originally classified as nova grains. Finally, this process may contribute to the production of 14N and 15N in the Galaxy, helping to produce the 14N/15N ratio in the solar system.« less

  12. Reduced dimensionality 3D HNCAN for unambiguous HN, CA and N assignment in proteins

    NASA Astrophysics Data System (ADS)

    Rout, Manoj Kumar; Mishra, Pushpa; Atreya, Hanudatta S.; Hosur, Ramakrishna V.

    2012-03-01

    We present here an improvisation of HNN (Panchal, Bhavesh et al., 2001) called RD 3D HNCAN for backbone (HN, CA and 15N) assignment in both folded and unfolded proteins. This is a reduced dimensionality experiment which employs CA chemical shifts to improve dispersion. Distinct positive and negative peak patterns of various triplet segments along the polypeptide chain observed in HNN are retained and these provide start and check points for the sequential walk. Because of co-incrementing of CA and 15N, peaks along one of the dimensions appear at sums and differences of the CA and 15N chemical shifts. This changes the backbone assignment protocol slightly and we present this in explicit detail. The performance of the experiment has been demonstrated using Ubiquitin and Plasmodium falciparum P2 proteins. The experiment is particularly valuable when two neighboring amino acid residues have nearly identical backbone 15N chemical shifts.

  13. Eastern oyster (Crassostrea virginica) δ15N as a bioindicator of nitrogen sources: Observations and modeling

    PubMed Central

    Fertig, B.; Carruthers, T.J.B.; Dennison, W.C.; Fertig, E.J.; Altabet, M.A.

    2013-01-01

    Stable nitrogen isotopes (δ15N) in bioindicators are increasingly employed to identify nitrogen sources in many ecosystems and biological characteristics of the eastern oyster (Crassostrea virginica) make it an appropriate species for this purpose. To assess nitrogen isotopic fractionation associated with assimilation and baseline variations in oyster mantle, gill, and muscle tissue δ15N, manipulative fieldwork in Chesapeake Bay and corresponding modeling exercises were conducted. This study (1) determined that five individuals represented an optimal sample size; (2) verified that δ15N in oysters from two locations converged after shared deployment to a new location reflecting a change in nitrogen sources; (3) identified required exposure time and temporal integration (four months for muscle, two to three months for gill and mantle); and (4) demonstrated seasonal δ15N increases in seston (summer) and oysters (winter). As bioindicators, oysters can be deployed for spatial interpolation of nitrogen sources, even in areas lacking extant populations. PMID:20381097

  14. Ecosystem N distribution and δ15N during a century of forest regrowth after agricultural abandonment

    USGS Publications Warehouse

    Compton, J.E.; Hooker, T.D.; Perakis, S.S.

    2007-01-01

    Stable isotope ratios of terrestrial ecosystem nitrogen (N) pools reflect internal processes and input–output balances. Disturbance generally increases N cycling and loss, yet few studies have examined ecosystem δ15N over a disturbance-recovery sequence. We used a chronosequence approach to examine N distribution and δ15N during forest regrowth after agricultural abandonment. Site ages ranged from 10 to 115 years, with similar soils, climate, land-use history, and overstory vegetation (white pine Pinus strobus). Foliar N and δ15N decreased as stands aged, consistent with a progressive tightening of the N cycle during forest regrowth on agricultural lands. Over time, foliar δ15N became more negative, indicating increased fractionation along the mineralization–mycorrhizal–plant uptake pathway. Total ecosystem N was constant across the chronosequence, but substantial internal N redistribution occurred from the mineral soil to plants and litter over 115 years (>25% of ecosystem N or 1,610 kg ha−1). Temporal trends in soil δ15N generally reflected a redistribution of depleted N from the mineral soil to the developing O horizon. Although plants and soil δ15N are coupled over millennial time scales of ecosystem development, our observed divergence between plants and soil suggests that they can be uncoupled during the disturbance-regrowth sequence. The approximate 2‰ decrease in ecosystem δ15N over the century scale suggests significant incorporation of atmospheric N, which was not detected by traditional ecosystem N accounting. Consideration of temporal trends and disturbance legacies can improve our understanding of the influence of broader factors such as climate or N deposition on ecosystem N balances and δ15N.

  15. Angular distributions for /sup 16/O(/gamma/,p)/sup 15/N at intermediate energies

    SciTech Connect

    Adams, G.S.; Kinney, E.R.; Matthews, J.L.; Sapp, W.W.; Soos, T.; Owens, R.O.; Turley, R.S.; Pignault, G.

    1988-12-01

    The photoproton knockout reaction on /sup 16/O leaving /sup 15/N in low-lying bound states has been observed over the photon energy range from 196 to 361 MeV. The angular distribution for the reaction populating the ground state of /sup 15/N develops sharp structure as the photon energy is increased but that for population of the excited states is smooth. The results are not explained by existing theoretical models.

  16. The First in Vivo Observation of 13C- 15N Coupling in Mammalian Brain

    NASA Astrophysics Data System (ADS)

    Kanamori, Keiko; Ross, Brian D.

    2001-12-01

    [5-13C,15N]Glutamine, with 1J(13C-15N) of 16 Hz, was observed in vivo in the brain of spontaneously breathing rats by 13C MRS at 4.7 T. The brain [5-13C]glutamine peak consisted of the doublet from [5-13C,15N]glutamine and the center [5-13C,14N]glutamine peak, resulting in an apparent triplet with a separation of 8 Hz. The time course of formation of brain [5-13C,15N]glutamine was monitored in vivo with a time resolution of 20-35 min. This [5-13C,15N]glutamine was formed by glial uptake of released neurotransmitter [5-13C]glutamate and its reaction with 15NH3 catalyzed by the glia-specific glutamine synthetase. The neurotransmitter glutamate C5 was selectively13C-enriched by intravenous [2,5-13C]glucose infusion to 13C-label whole-brain glutamate C5, followed by [12C]glucose infusion to chase 13C from the small and rapidly turning-over glial glutamate pool, leaving 13C mainly in the neurotransmitter [5-13C]glutamate pool, which is sequestered in vesicles until release. Hence, the observed [5-13C,15N]glutamine arises from a coupling between 13C of neuronal origin and 15N of glial origin. Measurement of the rate of brain [5-13C,15N]glutamine formation provides a novel noninvasive method of studying the kinetics of neurotransmitter uptake into glia in vivo, a process that is crucial for protecting the brain from glutamate excitotoxicity.

  17. Efficient Synthesis of Nicotinamide-1-15N for Ultrafast NMR Hyperpolarization Using Parahydrogen

    PubMed Central

    2016-01-01

    Nicotinamide (a vitamin B3 amide) is one of the key vitamins as well as a drug for treatment of M. tuberculosis, HIV, cancer, and other diseases. Here, an improved Zincke reaction methodology is presented allowing for straightforward and scalable synthesis of nicotinamide-1-15N with an excellent isotopic purity (98%) and good yield (55%). 15N nuclear spin label in nicotinamide-1-15N can be NMR hyperpolarized in seconds using parahydrogen gas. NMR hyperpolarization using the process of temporary conjugation between parahydrogen and to-be-hyperpolarized biomolecule on hexacoordinate iridium complex via the Signal Amplification By Reversible Exchange (SABRE) method significantly increases detection sensitivity (e.g., >20 000-fold for nicotinamide-1-15N at 9.4 T) as has been shown by Theis T. et al. (J. Am. Chem. Soc.2015, 137, 1404), and hyperpolarized in this fashion, nicotinamide-1-15N can be potentially used to probe metabolic processes in vivo in future studies. Moreover, the presented synthetic methodology utilizes mild reaction conditions, and therefore can also be potentially applied to synthesis of a wide range of 15N-enriched N-heterocycles that can be used as hyperpolarized contrast agents for future in vivo molecular imaging studies. PMID:26999571

  18. 1H and 15N NMR Analyses on Heparin, Heparan Sulfates and Related Monosaccharides Concerning the Chemical Exchange Regime of the N-Sulfo-Glucosamine Sulfamate Proton

    PubMed Central

    Pomin, Vitor H.

    2016-01-01

    Heparin and heparan sulfate are structurally related glycosaminoglycans (GAGs). Both GAGs present, although in different concentrations, N-sulfo-glucosamine (GlcNS) as one of their various composing units. The conditional fast exchange property of the GlcNS sulfamate proton in these GAGs has been pointed as the main barrier to its signal detection via NMR experiments, especially 1H-15N HSQC. Here, a series of NMR spectra is collected on heparin, heparan sulfate and related monosaccharides. The N-acetyl glucosamine-linked uronic acid types of these GAGs were properly assigned in the 1H-15N HSQC spectra. Dynamic nuclear polarization (DNP) was employed in order to facilitate 1D spectral acquisition of the sulfamate 15N signal of free GlcNS. Analyses on the multiplet pattern of scalar couplings of GlcNS 15N has helped to understand the chemical properties of the sulfamate proton in solution. The singlet peak observed for GlcNS happens due to fast chemical exchange of the GlcNS sulfamate proton in solution. Analyses on kinetics of alpha-beta anomeric mutarotation via 1H NMR spectra have been performed in GlcNS as well as other glucose-based monosaccharides. 1D 1H and 2D 1H-15N HSQC spectra recorded at low temperature for free GlcNS dissolved in a proton-rich solution showed signals from all exchangeable protons, including those belonging to the sulfamate group. This work suits well to the current grand celebration of one-century-anniversary of the discovery of heparin. PMID:27618066

  19. Highly 15N-Enriched Chondritic Clasts in the Isheyevo Meteorite

    SciTech Connect

    Bonal, L; Huss, G R; Krot, A N; Nagashima, K; Ishii, H A; Bradley, J P; Hutcheon, I D

    2009-01-14

    The metal-rich carbonaceous chondrites (CB and CH) have the highest whole-rock {sup 15}N enrichment ({delta}{sup 15}N up to +1500{per_thousand}), similar to {delta}{sup 15}N values reported in micron-sized regions (hotspots) of Interplanetary Dust Particles (IDPs) of possibly cometary origin and fine-grained matrices of unmetamorphosed chondrites. These {sup 15}N-rich hotspots are commonly attributed to low-temperature ion-molecule reactions in the protosolar molecular cloud or in the outer part of the protoplanetary disk. The nature of the whole-rock {sup 15}N enrichment of the metal-rich chondrites is not understood. We report a discovery of a unique type of primitive chondritic clasts in the CH/CB-like meteorite Isheyevo, which provides important constraints on the origin of {sup 15}N anomaly in metal-rich chondrites and nitrogen-isotope fractionation in the Solar System. These clasts contain tiny chondrules and refractory inclusions (5-15 {micro}m in size), and abundant ferromagnesian chondrule fragments (1-50 {micro}m in size) embedded in the partly hydrated, fine-grained matrix material composed of olivines, pyroxenes, poorly-organized aromatic organics, phyllosilicates and other hydrous phases. The mineralogy and oxygen isotope compositions of chondrules and refractory inclusions in the clasts are similar to those in the Isheyevo host, suggesting formation at similar heliocentric distances. In contrast to the previously known extraterrestrial samples, the fine-grained material in the clasts is highly and rather uniformly enriched in {sup 15}N, with bulk {delta}{sup 15}N values ranging between +1000 and +1300{per_thousand}; the {delta}{sup 15}N values in rare hotspots range from +1400 to +4000{per_thousand}. Since fine-grained matrices in the lithic clasts are the only component containing thermally unprocessed (during CAI and chondrule formation or during impact melting) materials that accreted into the metal rich chondrite parent body(ies), the {sup 15}N

  20. My Favorite Assignment.

    ERIC Educational Resources Information Center

    Post, Robert E.; Johnson, Jack E.

    1982-01-01

    Presents two assignments that show (1) how George Orwell's "Politics and the English Language" can be applied to business writing and (2) how structured student-teacher conferences can generate enthusiasm for oral expression in a business communication course. (AEA)

  1. Fair Package Assignment

    NASA Astrophysics Data System (ADS)

    Lahaie, Sébastien; Parkes, David C.

    We consider the problem of fair allocation in the package assignment model, where a set of indivisible items, held by single seller, must be efficiently allocated to agents with quasi-linear utilities. A fair assignment is one that is efficient and envy-free. We consider a model where bidders have superadditive valuations, meaning that items are pure complements. Our central result is that core outcomes are fair and even coalition-fair over this domain, while fair distributions may not even exist for general valuations. Of relevance to auction design, we also establish that the core is equivalent to the set of anonymous-price competitive equilibria, and that superadditive valuations are a maximal domain that guarantees the existence of anonymous-price competitive equilibrium. Our results are analogs of core equivalence results for linear prices in the standard assignment model, and for nonlinear, non-anonymous prices in the package assignment model with general valuations.

  2. Separation of Anisotropy and Exchange Broadening Using 15N CSA- 15N- 1H Dipole-Dipole Relaxation Cross-Correlation Experiments

    NASA Astrophysics Data System (ADS)

    Renner, Christian; Holak, Tad A.

    2000-08-01

    Based on the measurement of cross-correlation rates between 15N CSA and 15N-1H dipole-dipole relaxation we propose a procedure for separating exchange contributions to transverse relaxation rates (R2 = 1/T2) from effects caused by anisotropic rotational diffusion of the protein molecule. This approach determines the influence of anisotropy and chemical exchange processes independently and therefore circumvents difficulties associated with the currently standard use of T1/T2 ratios to determine the rotational diffusion tensor. We find from computer simulations that, in the presence of even small amounts of internal flexibility, fitting T1/T2 ratios tends to underestimate the anisotropy of overall tumbling. An additional problem exists when the N-H bond vector directions are not distributed homogeneously over the surface of a unit sphere, such as in helix bundles or β-sheets. Such a case was found in segment 4 of the gelation factor (ABP 120), an F-actin cross-linking protein, in which the diffusion tensor cannot be calculated from T1/T2 ratios. The 15N CSA tensor of the residues for this β-sheet protein was found to vary even within secondary structure elements. The use of a common value for the whole protein molecule therefore might be an oversimplification. Using our approach it is immediately apparent that no exchange broadening exists for segment 4 although strongly reduced T2 relaxation times for several residues could be mistaken as indications for exchange processes.

  3. Estimate of production of gaseous nitrogen in the human body based on (15)N analysis of breath N2 after administration of [(15)N2]urea.

    PubMed

    Junghans, Peter

    2013-01-01

    After oral administration of [(15)N2]urea (1.5 mmol, 95 atom% (15)N), we found that breath N2 was significantly (15)N-labelled. The result suggests that molecular nitrogen in breath must be partly produced endogenously. Based on a metabolic model, the endogenous N2 production was estimated to be 0.40±0.25 mmol kg(-1) d(-1) or 2.9±1.8 % of the total (urinary and faecal) N excretion in fasted healthy subjects (n=4). In patients infected with Helicobacter pylori (n=5), the endogenous N2 production was increased to 1.24±0.59 mmol kg(-1) d(-1) or 9.0±4.3 % of the total N excretion compared to the healthy controls (p<0.05). We conclude that N balance and gas exchange measurements may be affected by endogenously produced nitrogen, especially in metabolic situations with elevated nitrosation, for instance in oxidative and nitrosative stress-related diseases such as H. pylori infections.

  4. Importance of Nitrate Attenuation In A Small Wetland Following Forest Harvest: 18O/16O, 15N/14N in nitrate and 15N/14N) in vegetation

    NASA Astrophysics Data System (ADS)

    Spoelstra, J.; Schiff, S. L.; Semkin, R. G.; Jeffries, D. S.; Elgood, R. J.

    2004-05-01

    Forest harvest can result in elevated nitrate concentrations in streams and groundwater affecting forest regeneration and downstream aquatic ecosystems. Turkey Lakes Watershed, located near Sault Ste Marie, Ontario (TLW), exhibits relatively high nitrate export due to naturally high rates of nitrification. During a forest harvest experiment at the TLW, stable isotope techniques were used to investigate nitrate attenuation in an intermediate position natural wetland receiving high concentrations of nitrate following forest clear-cutting. Isotopic analysis of nitrate (18O/16O, 15N/14N) and vegetation (15N/14N) demonstrated that denitrification and plant uptake of nitrate resulted in significantly lower nitrate concentrations in wetland outflow compared to incoming stream water and groundwater. The 0.2-hectare forested swamp, too small to show up on standard topographic maps, retained 65 to 100 percent of upgradient nitrate inputs, elevated due to increased nitrification in soils. The 15N/14N enrichment factor associated with nitrate attenuation in wetland surface water was lower than observed during denitrification in groundwaters, suggesting that denitrification proceeded to completion in some areas of the wetland. Even small, shallow, carbon rich pockets of organic matter in topographic depressions can significantly affect biogeochemical fluxes of C, N, S and Ca. Future forest management practices designed to recognize and preserve small wetlands could significantly reduce the potentially detrimental effects of forest harvest on aquatic systems.

  5. Compound-specific δ15N amino acid measurements in littoral mussels in the California upwelling ecosystem: a new approach to generating baseline δ15N Isoscapes for coastal ecosystems.

    PubMed

    Vokhshoori, Natasha L; McCarthy, Matthew D

    2014-01-01

    We explored δ(15)N compound-specific amino acid isotope data (CSI-AA) in filter-feeding intertidal mussels (Mytilus californianus) as a new approach to construct integrated isoscapes of coastal primary production. We examined spatial δ(15)N gradients in the California Upwelling Ecosystem (CUE), determining bulk δ(15)N values of mussel tissue from 28 sites between Port Orford, Oregon and La Jolla, California, and applying CSI-AA at selected sites to decouple trophic effects from isotopic values at the base of the food web. Bulk δ(15)N values showed a strong linear trend with latitude, increasing from North to South (from ∼ 7‰ to ∼ 12‰, R(2) = 0.759). In contrast, CSI-AA trophic position estimates showed no correlation with latitude. The δ(15)N trend is therefore most consistent with a baseline δ(15)N gradient, likely due to the mixing of two source waters: low δ(15)N nitrate from the southward flowing surface California Current, and the northward transport of the California Undercurrent (CUC), with (15)N-enriched nitrate. This interpretation is strongly supported by a similar linear gradient in δ(15)N values of phenylalanine (δ(15)NPhe), the best AA proxy for baseline δ(15)N values. We hypothesize δ(15)N(Phe) values in intertidal mussels can approximate annual integrated δ(15)N values of coastal phytoplankton primary production. We therefore used δ(15)N(Phe) values to generate the first compound-specific nitrogen isoscape for the coastal Northeast Pacific, which indicates a remarkably linear gradient in coastal primary production δ(15)N values. We propose that δ(15)N(Phe) isoscapes derived from filter feeders can directly characterize baseline δ(15)N values across major biochemical provinces, with potential applications for understanding migratory and feeding patterns of top predators, monitoring effects of climate change, and study of paleo- archives.

  6. Soil N and 15N variation with time in a California annual grassland ecosystem

    USGS Publications Warehouse

    Brenner, D.L.; Amundson, Ronald; Baisden, W. Troy; Kendall, C.; Harden, J.

    2001-01-01

    The %N and ??15N values of soils and plants were measured along a chronosequence spanning 3 to 3000 Ky in a California annual grassland. Total soil N decreased with increasing soil age (1.1 to 0.4 kg N m-2) while the mean ?? 15N values of the soil N increased by several ??? from the youngest to oldest sites (+3.5 to +6.2 ???). The ?? 15N values of plants varied along the gradient, reflecting changing soil N pools and differences in the form of N uptake. The decline in total N storage with time is hypothesized to be due to a shift from N to P limitation with increasing soil age. The general increase in ?? 15N values with time is interpreted using a N mass balance model, and appears to reflect a shift toward an increasing proportional losses of inorganic mineral forms of N (vs. organic forms) with increasing soil age. We develop a quantitative index of this trend (mineral vs. organic forms of N loss) using mass balance considerations and parameters. The %N and ?? 15N values along the California age gradient were compared to the published data for a comparably aged chronosequence in Hawaii. Most striking in this comparison is the observation that the California soil and plant ?? 15N values are several ??? greater than those on comparably aged Hawaiian sites. Multiple explanations are plausible, but assuming the sites have a similar range in ?? 15N values of atmospheric inputs, the isotopic differences suggest that N may be, at least seasonally, in greater excess in the strongly seasonal, semi-arid, California grassland. Copyright ?? 2001 Elsevier Science Ltd.

  7. Sewage derive [sup 15]N in the Baltic traced in fucus

    SciTech Connect

    Hobbie, J.E.; Fry, B. ); Larsson, U.; Elmgren, R. )

    1990-01-09

    Himmerfjarden, a fjord-like bay on the eastern shore of the Baltic, receives treated sewage from 250,000 inhabitants. Because the inorganic N in the effluent is enriched in [sup 15]N through denitrification, nitrification, and ammonia volatilization, an analysis of the distribution of [sup 15]N in the Bay tells how far from the source the sewage nitrogen moves. The attached macroalga Fucus vesiculosus was collected in early May from rocky shore at 0-0.5 m depth and the [sup 15]N content of the tips of the fronds analyzed. This N represents uptake and storage during the previous six months and growth during March and April. The [delta][sup 15]N was uniformly high (11-13[per thousand]) in the main body of the Bay within 15 km from the sewage source. Beyond 15 km values decreased with distance to a low of 4.6[per thousand] at 35 km, where the Bay ends and the coastal waters begin. Using the 11-13 and 4.6[per thousand] as endmembers, the percentage of sewage N making up the Fucus at any point may be calculated. The [delta][sub 15]N of particulate organic matter in the offshore Baltic waters was around 0[per thousand] and Fucus had an [delta][sup 15]N about 1.5[per thousand] higher than the POM. From this and other evidence we conclude that there is a belt of coastal water with an elevated [delta][sup 15]N lying along the east coast of the Baltic. This presumably derives from sewage and perhaps from agriculture and is potentially of use as a tracer of coastal zone/pelagic zone interactions.

  8. 42 CFR 433.146 - Rights assigned; assignment method.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 42 Public Health 4 2010-10-01 2010-10-01 false Rights assigned; assignment method. 433.146 Section 433.146 Public Health CENTERS FOR MEDICARE & MEDICAID SERVICES, DEPARTMENT OF HEALTH AND HUMAN... Assignment of Rights to Benefits § 433.146 Rights assigned; assignment method. (a) Except as specified...

  9. Intramolecular N-Glycan/Polypeptide Interactions Observed at Multiple N-Glycan Remodeling Steps through [13C,15N]-N-Acetylglucosamine Labeling of Immunoglobulin G1

    PubMed Central

    2014-01-01

    Asparagine-linked (N) glycosylation is a common eukaryotic protein modification that affects protein folding, function, and stability through intramolecular interactions between N-glycan and polypeptide residues. Attempts to characterize the structure–activity relationship of each N-glycan are hindered by inherent properties of the glycoprotein, including glycan conformational and compositional heterogeneity. These limitations can be addressed by using a combination of nuclear magnetic resonance techniques following enzymatic glycan remodeling to simultaneously generate homogeneous glycoforms. However, widely applicable methods do not yet exist. To address this technological gap, immature glycoforms of the immunoglobulin G1 fragment crystallizable (Fc) were isolated in a homogeneous state and enzymatically remodeled with [13C,15N]-N-acetylglucosamine (GlcNAc). UDP-[13C,15N]GlcNAc was synthesized enzymatically in a one-pot reaction from [13C]glucose and [15N-amido]glutamine. Modifying Fc with recombinantly expressed glycosyltransferases (Gnt1 and Gnt2) and UDP-[13C,15N]GlcNAc resulted in complete glycoform conversion as judged by mass spectrometry. Two-dimensional heteronuclear single-quantum coherence spectra of the Gnt1 product, containing a single [13C,15N]GlcNAc residue on each N-glycan, showed that the N-glycan is stabilized through interactions with polypeptide residues. Similar spectra of homogeneous glycoforms, halted at different points along the N-glycan remodeling pathway, revealed the presence of an increased level of interaction between the N-glycan and polypeptide at each step, including mannose trimming, as the N-glycan was converted to a complex-type, biantennary form. Thus, conformational restriction increases as Fc N-glycan maturation proceeds. Gnt1 and Gnt2 catalyze fundamental reactions in the synthesis of every glycoprotein with a complex-type N-glycan; thus, the strategies presented herein can be applied to a broad range of glycoprotein

  10. Nitrogen Isotopic Ratios in Cometary NH2: Implication for 15N-fractionation in Ammonia

    NASA Astrophysics Data System (ADS)

    Shinnaka, Yoshiharu; Kawakita, Hideyo; Jehin, Emmanuël; Decock, Alice; Hutsemékers, Damien; Manfroid, Jean; Arai, Akira

    2015-11-01

    Isotopic ratios in cometary molecules are diagnostic for the physico-chemical conditions where molecules formed and are processed, from the interstellar medium to the solar nebula. Usually temperatures at the molecular formation control the fractionation of the heavier element in molecular species, e.g., D-fractionation in water.In cometary volatiles, the 14N/15N ratios in CN have been well observed (Manfroid et al. 2009, A&A, 503, 613, and reference therein) and is consistent with the ratio in HCN (a most probable parent of CN) measured in few comets (Bockelée-Morvan et al. 2008, ApJ, 679, L49). Those ratios are enriched compared to the proto-solar value by a factor of ~3. In contrast to those Nitriles, there are only few reports on 14N/15N ratios in Ammonia (as Amine) (Rousselot et al. 2014, ApJ, 780, L17; Shinnaka et al. 2014, ApJ, 782, L16). Ammonia (NH3) is usually the most abundant and HCN is the second most abundant N-bearing volatiles in cometary ice. Especially, recent observations of 15NH2 revealed the 14N/15N ratios in NH3 are comparable to those of CN. However, from the viewpoint of theoretical work, the enrichment of 15N in cometary NH3 cannot be reproduced by current chemical network models. Information about the diversity of the 14N/15N ratios in NH3 of individual comets is needed to understand the formation mechanisms/environments of NH3 in the early solar system.To clarify the diversity of the 14N/15N ratios in cometary NH3, we determine the 14N/15N ratios in NH3 for more than ten comets individually which include not only Oort cloud comets but also short period comets by using the high-resolution optical spectra of NH2. These spectra were obtained with both the UVES mounted on the VLT in Chile and the HDS on the Subaru Telescope in Hawaii.The derived 14N/15N ratios in NH3 for more than ten comets show high 15N-enrichment compared with the elemental abundances of nitrogen in the Sun by about factor of ~3 and has no large diversity depending on

  11. A new method to track seed dispersal and recruitment using 15N isotope enrichment.

    PubMed

    Carlo, Tomás A; Tewksbury, Joshua J; Martínez Del Río, Carlos

    2009-12-01

    Seed dispersal has a powerful influence on population dynamics, genetic structuring, evolutionary rates, and community ecology. Yet, patterns of seed dispersal are difficult to measure due to methodological shortcomings in tracking dispersed seeds from sources of interest. Here we introduce a new method to track seed dispersal: stable isotope enrichment. It consists of leaf-feeding plants with sprays of 15N-urea during the flowering stage such that seeds developed after applications are isotopically enriched. We conducted a greenhouse experiment with Solanum americanum and two field experiments with wild Capsicum annuum in southern Arizona, USA, to field-validate the method. First, we show that plants sprayed with 15N-urea reliably produce isotopically enriched progeny, and that delta 15N (i.e., the isotopic ratio) of seeds and seedlings is a linear function of the 15N-urea concentration sprayed on mothers. We demonstrate that three urea dosages can be used to distinctly enrich plants and unambiguously differentiate their offspring after seeds are dispersed by birds. We found that, with high urea dosages, the resulting delta 15N values in seedlings are 10(3) - 10(4) times higher than the delta 15N values of normal plants. This feature allows tracking not only where seeds arrive, but in locations where seeds germinate and recruit, because delta 15N enrichment is detectable in seedlings that have increased in mass by at least two orders of magnitude before fading to normal delta 15N values. Last, we tested a mixing model to analyze seed samples in bulk. We used the delta 15N values of batches (i.e., combined seedlings or seeds captured in seed traps) to estimate the number of enriched seeds coming from isotopically enriched plants in the field. We confirm that isotope enrichment, combined with batch-sampling, is a cheap, reliable, and user-friendly method for bulk-processing seeds and is thus excellent for the detection of rare dispersal events. This method could

  12. Using a macroalgal δ15N bioassay to detect cruise ship waste water effluent inputs.

    PubMed

    Kaldy, James

    2011-08-01

    Green macroalgae bioassays were used to determine if the δ15N signature of cruise ship waste water effluent (CSWWE) could be detected in a small harbor. Opportunistic green macroalgae (Ulva spp.) were collected, cultured under nutrient depleted conditions and characterized with regard to N content and δ15N. Samples of algae were used in controlled incubations to evaluate the direction of isotope shift from exposure to CSWWE. Algae samples exposed to CSWWE exhibited an increase of 1-2.5‰ in δ15N values indicating that the CSWWE had an enriched isotope signature. In contrast, algae samples exposed to field conditions exhibited a significant decrease in the observed δ15N indicating that a light N source was used. Isotopically light, riverine nitrogen derived from N2-fixing trees in the watershed may be a N source utilized by algae. These experiments indicate that the δ15N CSWWE signature was not detectable under the CSWWE loading conditions of this experiment.

  13. Stem injection of 15N-NH4NO3 into mature Sitka spruce (Picea sitchensis).

    PubMed

    Nair, Richard; Weatherall, Andrew; Perks, Mike; Mencuccini, Maurizio

    2014-10-01

    Stem injection techniques can be used to introduce (15)N into trees to overcome a low variation in natural abundance and label biomass with a distinct (15)N signature, but have tended to target small and young trees, of a variety of species, with little replication. We injected 98 atom% (15)N ammonium nitrate (NH4NO3) solution into 13 mature, 9- to 13-m tall edge-profile Sitka spruce trees in order to produce a large quantity of labelled litter, examining the distribution of the isotope throughout the canopy after felling in terms of both total abundance of (15)N and relative distribution of the isotope throughout individual trees. Using a simple mass balance of the canopy alone, based on observed total needle biomass and modelled branch biomass, all of the isotope injected was accounted for, evenly split between needles and branches, but with a high degree of variability both within individual trees, and among trees. Both (15)N abundance and relative within-canopy distribution were biased towards the upper and middle crown in foliage. Recovery of the label in branches was much more variable than in needles, possibly due to differences in nitrogen allocation for both growth and storage, which differ seasonally between foliage and woody biomass.

  14. The 15N isotope to evaluate fertilizer nitrogen absorption efficiency by the coffee plant.

    PubMed

    Fenilli, Tatiele A B; Reichart, Klaus; Bacchi, Osny O S; Trivelin, Paulo C O; Dourado-Neto, Durval

    2007-12-01

    The use of the 15N label for agronomic research involving nitrogen (N) cycling and the fate of fertilizer-N is well established, however, in the case of long term experimentation with perennial crops like citrus, coffee and rubber tree, there are still shortcomings mainly due to large plant size, sampling procedures, detection levels and interferences on the system. This report tries to contribute methodologically to the design and development of 15N labeled fertilizer experiments, using as an example a coffee crop fertilized with 15N labeled ammonium sulfate, which was followed for two years. The N of the plant derived from the fertilizer was studied in the different parts of the coffee plant in order to evaluate its distribution within the plant and the agronomic efficiency of the fertilizer application practice. An enrichment of the fertilizer-N of the order of 2% 15N abundance was sufficient to study N absorption rates and to establish fertilizer-N balances after one and two years of coffee cropping. The main source of errors in the estimated values lies in the inherent variability among field replicates and not in the measurements of N contents and 15N enrichments of plant material by mass-spectrometry.

  15. Rivermouth alteration of agricultural impacts on consumer tissue δ15N

    USGS Publications Warehouse

    Larson, James H.; Richardson, William B.; Vallazza, Jonathan M.; Nelson, J. C.

    2013-01-01

    Terrestrial agricultural activities strongly influence riverine nitrogen (N) dynamics, which is reflected in the δ15N of riverine consumer tissues. However, processes within aquatic ecosystems also influence consumer tissue δ15N. As aquatic processes become more important terrestrial inputs may become a weaker predictor of consumer tissue δ15N. In a previous study, this terrestrial-consumer tissue δ15N connection was very strong at river sites, but was disrupted by processes occurring in rivermouths (the ‘rivermouth effect’). This suggested that watershed indicators of N loading might be accurate in riverine settings, but could be inaccurate when considering N loading to the nearshore of large lakes and oceans. In this study, the rivermouth effect was examined on twenty-five sites spread across the Laurentian Great Lakes. Relationships between agriculture and consumer tissue δ15N occurred in both upstream rivers and at the outlets where rivermouths connect to the nearshore zone, but agriculture explained less variation and had a weaker effect at the outlet. These results suggest that rivermouths may sometimes be significant sources or sinks of N, which would cause N loading estimates to the nearshore zone that are typically made at discharge gages further upstream to be inaccurate. Identifying definitively the controls over the rivermouth effect on N loading (and other nutrients) will require integration of biogeochemical and hydrologic models.

  16. Mapping inhibitor binding modes on an active cysteine protease via nuclear magnetic resonance spectroscopy.

    PubMed

    Lee, Gregory M; Balouch, Eaman; Goetz, David H; Lazic, Ana; McKerrow, James H; Craik, Charles S

    2012-12-18

    Cruzain is a member of the papain/cathepsin L family of cysteine proteases, and the major cysteine protease of the protozoan Trypanosoma cruzi, the causative agent of Chagas disease. We report an autoinduction methodology that provides soluble cruzain in high yields (>30 mg/L in minimal medium). These increased yields provide sufficient quantities of active enzyme for use in nuclear magnetic resonance (NMR)-based ligand mapping. Using circular dichroism and NMR spectroscopy, we also examined the solution-state structural dynamics of the enzyme in complex with a covalently bound vinyl sulfone inhibitor (K777). We report the backbone amide and side chain carbon chemical shift assignments of cruzain in complex with K777. These resonance assignments were used to identify and map residues located in the substrate binding pocket, including the catalytic Cys25 and His162. Selective [(15)N]Cys, [(15)N]His, and [(13)C]Met labeling was performed to quickly assess cruzain-ligand interactions for a set of eight low-molecular weight compounds exhibiting micromolar binding or inhibition. Chemical shift perturbation mapping verified that six of the eight compounds bind to cruzain at the active site. Three different binding modes were delineated for the compounds, namely, covalent, noncovalent, and noninteracting. These results provide examples of how NMR spectroscopy can be used to screen compounds for fast evaluation of enzyme-inhibitor interactions to facilitate lead compound identification and subsequent structural studies.

  17. Nitrate reductase 15N discrimination in Arabidopsis thaliana, Zea mays, Aspergillus niger, Pichea angusta, and Escherichia coli

    PubMed Central

    Carlisle, Eli; Yarnes, Chris; Toney, Michael D.; Bloom, Arnold J.

    2014-01-01

    Stable 15N isotopes have been used to examine movement of nitrogen (N) through various pools of the global N cycle. A central reaction in the cycle involves the reduction of nitrate (NO−3) to nitrite (NO−2) catalyzed by nitrate reductase (NR). Discrimination against 15N by NR is a major determinant of isotopic differences among N pools. Here, we measured in vitro 15N discrimination by several NRs purified from plants, fungi, and a bacterium to determine the intrinsic 15N discrimination by the enzyme and to evaluate the validity of measurements made using 15N-enriched NO−3. Observed NR isotope discrimination ranged from 22 to 32‰ (kinetic isotope effects of 1.022–1.032) among the different isozymes at natural abundance 15N (0.37%). As the fractional 15N content of substrate NO−3 increased from natural abundance, the product 15N fraction deviated significantly from that expected based on substrate enrichment and 15N discrimination measured at natural abundance. Additionally, isotopic discrimination by denitrifying bacteria used to reduce NO−3 and NO−2 in some protocols became a greater source of error as 15N enrichment increased. We briefly discuss potential causes of the experimental artifacts with enriched 15N and recommend against the use of highly enriched 15N tracers to study N discrimination in plants or soils. PMID:25071800

  18. The position dependent 15N enrichment of nitrous oxide in the stratosphere.

    PubMed

    Röckmann, T; Kaiser, J; Brenninkmeijer, C A; Brand, W A; Borchers, R; Crowley, J N; Wollenhaupt, M; Crutzen, P J

    2001-01-01

    The position dependent 15N fractionation of nitrous oxide (N2O), which cannot be obtained from mass spectrometric analysis on molecular N2O itself, can be determined with high precision using isotope ratio mass spectrometry on the NO+ fragment that is formed on electron impact in the source of an isotope ratio mass spectrometer. Laboratory UV photolysis experiments show that strong position dependent 15N fractionations occur in the photolysis of N2O in the stratosphere, its major atmospheric sink. Measurements on the isotopic composition of stratospheric N2O indeed confirm the presence of strong isotope enrichments, in particular the difference in the fractionation constants for 15N14NO and 14N15NO. The absolute magnitudes of the fractionation constants found in the stratosphere are much smaller, however, than those found in the lab experiments, demonstrating the importance of dynamical and also additional chemical processes like the reaction of N2O with O(1D).

  19. Synthesis and NMR of {sup 15}N-labeled DNA fragments

    SciTech Connect

    Jones, R.A.

    1994-12-01

    DNA fragments labeled with {sup 15}N at the ring nitrogens and at the exocyclic amino groups can be used to obtain novel insight into interactions such as base pairing, hydration, drug binding, and protein binding. A number of synthetic routes to {sup 15}N-labeled pyrimidine nucleosides, purines, and purine nucleosides have been reported. Moreover, many of these labeled bases or monomers have been incorporated into nucleic acids, either by chemical synthesis or by biosynthetic procedures. The focus of this chapter will be on the preparation of {sup 15}N-labeled purine 2{prime}-deoxynucleosides, their incorporation into DNA fragments by chemical synthesis, and the results of NMR studies using these labeled DNA fragments.

  20. 15N Fractionation in Star-Forming Regions and Solar System Objects

    NASA Technical Reports Server (NTRS)

    Wirstrom, Eva; Milam, Stefanie; Adande, GIlles; Charnley, Steven; Cordiner, Martin

    2015-01-01

    A central issue for understanding the formation and evolution of matter in the early Solar System is the relationship between the chemical composition of star-forming interstellar clouds and that of primitive Solar System materials. The pristinemolecular content of comets, interplanetary dust particles and carbonaceous chondrites show significant bulk nitrogen isotopic fractionation relative to the solar value, 14N15N 440. In addition, high spatial resolution measurements in primitive materials locally show even more extreme enhancements of 14N15N 100.

  1. Reactions, characterization and uptake of ammoxidized kraft lignin labeled with 15N.

    PubMed

    Ramírez, F; Varela, G; Delgado, E; López-Dellamary, F; Zúñiga, V; González, V; Faix, O; Meier, D

    2007-05-01

    Ammoxidation of kraft lignin was carried out in a Parr reactor using (15)NH(3) as the main nitrogen source. Reaction parameters were set up until a total nitrogen content of approximately 13 wt.% in lignin was achieved, in accordance with conditions of previous studies. Analytical tools such as FTIR, Py-GC/MS, and solid state NMR were used in this research. The nature of nitrogen bondings is discussed. The incorporation of the (15)N from ammoxidized lignin was followed in pumpkins (Zucchini cucurbita pepo L.) by means of (15)N emission spectroscopy.

  2. Making Effective Assignments.

    ERIC Educational Resources Information Center

    McLeod, Alan M., Ed.

    1982-01-01

    Although the focus of this issue of the "Virginia English Bulletin" is on making effective assignments, most of the articles also emphasize the importance and power of writing. Articles deal with the following topics: (1) the use of I-search (as explained by Kenneth Macrorie in "Searching Writing") as a form of research paper…

  3. Binding of oxytocin and 8-arginine-vasopressin to neurophysin studied by /sup 15/N NMR using magnetization transfer and indirect detection via protons

    SciTech Connect

    Live, D.H.; Cowburn, D.

    1987-10-06

    NMR was used to monitor the binding to neurophysin of oxytocin and 8-arginine-vasopressin, /sup 15/N labeling being used to identify specific backbone /sup 15/N and /sup 1/H signals. The most significant effects of binding were large downfield shifts in the amino nitrogen resonance of Phe-3 of vasopressin and in its associated proton, providing evidence that the peptide bond between residues 2 and 3 of the hormones is hydrogen-bonded to the protein within hormone-neurophysin complexes. Suggestive evidence for hydrogen bonding of the amino nitrogen of Tyr-2 was also obtained in the form of decreased proton exchange rates on binding; however, the chemical shift changes of this nitrogen and its associated proton indicated that such hydrogen bonding, if present, is probably weak. Shifts in the amino nitrogen of Asn-5 and in the -NH protons of both Asn-5 and Cys-6 demonstrated that these residues are significantly perturbed by binding, suggesting conformational changes of the ring on binding and/or the presence of binding sites on the hormone outside the 1-3 region. No support was obtained for the thesis that there is a significant second binding site for vasopressin on each neutrophysin chain. The behavior of both oxytocin and vasopressin on binding was consistent with formation of 1:1 complexes in slow exchange with the free state under most pH conditions. At low pH there was evidence of an increased exchange rate. Additionally, broadening of /sup 15/N resonances in the bound state at low pH occurred without a corresponding change in the resonances of equilibrating free hormone. The results suggest significant conformational alteration in neurophysin-hormone complexes at low pH possibly associated with protonation of the carboxyl group of the hormone-protein salt bridge.

  4. Binding of oxytocin and 8-arginine-vasopressin to neurophysin studied by 15N NMR using magnetization transfer and indirect detection via protons.

    PubMed

    Live, D H; Cowburn, D; Breslow, E

    1987-10-06

    NMR was used to monitor the binding to neurophysin of oxytocin and 8-arginine-vasopressin, 15N labeling being used to identify specific backbone 15N and 1H signals. The most significant effects of binding were large downfield shifts in the amino nitrogen resonance of Phe-3 of vasopressin and in its associated proton, providing evidence that the peptide bond between residues 2 and 3 of the hormones is hydrogen-bonded to the protein within hormone-neurophysin complexes. Suggestive evidence of hydrogen bonding of the amino nitrogen of Tyr-2 was also obtained in the form of decreased proton exchange rates on binding; however, the chemical shift changes of this nitrogen and its associated proton indicated that such hydrogen bonding, if present, is probably weak. Shifts in the amino nitrogen of Asn-5 and in the -NH protons of both Asn-5 and Cys-6 demonstrated that these residues are significantly perturbed by binding, suggesting conformational changes of the ring on binding and/or the presence of binding sites on the hormone outside the 1-3 region. No support was obtained for the thesis that there is a significant second binding site for vasopressin on each neurophysin chain. The behavior of both oxytocin and vasopressin on binding was consistent with formation of 1:1 complexes in slow exchange with the free state under most pH conditions. At low pH there was evidence of an increased exchange rate. Additionally, broadening of 15N resonances in the bound state at low pH occurred without a corresponding change in the resonances of equilibrating free hormone.(ABSTRACT TRUNCATED AT 250 WORDS)

  5. The degree of urbanization across the globe is not reflected in the δ(15)N of seagrass leaves.

    PubMed

    Christiaen, Bart; Bernard, Rebecca J; Mortazavi, Behzad; Cebrian, Just; Ortmann, Alice C

    2014-06-30

    Many studies show that seagrass δ(15)N ratios increase with the amount of urbanization in coastal watersheds. However, there is little information on the relationship between urbanization and seagrass δ(15)N ratios on a global scale. We performed a meta-analysis on seagrass samples from 79 independent locations to test if seagrass δ(15)N ratios correlate with patterns of population density and fertilizer use within a radius of 10-200 km around the sample locations. Our results show that seagrass δ(15)N ratios are more influenced by intergeneric and latitudinal differences than the degree of urbanization or the amount of fertilizer used in nearby watersheds. The positive correlation between seagrass δ(15)N ratios and latitude hints at an underlying pattern in discrimination or a latitudinal gradient in the (15)N isotopic signature of nitrogen assimilated by the plants. The actual mechanisms responsible for the correlation between δ(15)N and latitude remain unknown.

  6. Production of 15N-labeled α-amanitin in Galerina marginata

    PubMed Central

    DuBois, Brandon; Sgambelluri, R. Michael; Angelos, Evan R.; Li, Xuan; Holmes, Daniel

    2015-01-01

    α-Amanitin is the major causal constituent of deadly Amanita mushrooms that account for the majority of fatal mushroom poisonings worldwide. It is also an important biochemical tool for the study of its target, RNA polymerase II. The commercial supply of this bicyclic peptide comes directly from A. phalloides, the death cap mushroom, which is collected from its natural habitat. Isotopically labeled amanitin could be useful for clinical and forensic applications, but α-amanitin has not been chemically synthesized and A. phalloides cannot be cultured on artificial medium. Using Galerina marginata, an unrelated saprobic mushroom that grows and produces α-amanitin in culture, we describe a method for producing 15N-labeled α-amanitin using growth media containing 15N as sole nitrogen source. A key to success was preparing 15N-enriched yeast extract via a novel method designated “glass bead-assisted maturation.” In the presence of the labeled yeast extract and 15N-NH4Cl, α-amanitin was produced with >97% isotope enrichment. The labeled product was confirmed by HPLC, high-resolution mass spectrometry, and NMR. PMID:26100667

  7. Production of (15)N-labeled α-amanitin in Galerina marginata.

    PubMed

    Luo, Hong; DuBois, Brandon; Sgambelluri, R Michael; Angelos, Evan R; Li, Xuan; Holmes, Daniel; Walton, Jonathan D

    2015-09-01

    α-Amanitin is the major causal constituent of deadly Amanita mushrooms that account for the majority of fatal mushroom poisonings worldwide. It is also an important biochemical tool for the study of its target, RNA polymerase II. The commercial supply of this bicyclic peptide comes from Amanita phalloides, the death cap mushroom, which is collected from the wild. Isotopically labeled amanitin could be useful for clinical and forensic applications, but α-amanitin has not been chemically synthesized and A. phalloides cannot be cultured on artificial medium. Using Galerina marginata, an unrelated saprotrophic mushroom that grows and produces α-amanitin in culture, we describe a method for producing (15)N-labeled α-amanitin using growth media containing (15)N as sole nitrogen source. A key to success was preparing (15)N-enriched yeast extract via a novel method designated "glass bead-assisted maturation." In the presence of the labeled yeast extract and (15)N-NH4Cl, α-amanitin was produced with >97% isotope enrichment. The labeled product was confirmed by HPLC, high-resolution mass spectrometry, and NMR.

  8. Monitoring the refinement of crystal structures with (15)N solid-state NMR shift tensor data.

    PubMed

    Kalakewich, Keyton; Iuliucci, Robbie; Mueller, Karl T; Eloranta, Harriet; Harper, James K

    2015-11-21

    The (15)N chemical shift tensor is shown to be extremely sensitive to lattice structure and a powerful metric for monitoring density functional theory refinements of crystal structures. These refinements include lattice effects and are applied here to five crystal structures. All structures improve based on a better agreement between experimental and calculated (15)N tensors, with an average improvement of 47.0 ppm. Structural improvement is further indicated by a decrease in forces on the atoms by 2-3 orders of magnitude and a greater similarity in atom positions to neutron diffraction structures. These refinements change bond lengths by more than the diffraction errors including adjustments to X-Y and X-H bonds (X, Y = C, N, and O) of 0.028 ± 0.002 Å and 0.144 ± 0.036 Å, respectively. The acquisition of (15)N tensors at natural abundance is challenging and this limitation is overcome by improved (1)H decoupling in the FIREMAT method. This decoupling dramatically narrows linewidths, improves signal-to-noise by up to 317%, and significantly improves the accuracy of measured tensors. A total of 39 tensors are measured with shifts distributed over a range of more than 400 ppm. Overall, experimental (15)N tensors are at least 5 times more sensitive to crystal structure than (13)C tensors due to nitrogen's greater polarizability and larger range of chemical shifts.

  9. Cold brittleness of corrosion-resistant maraging steel 08Kh15N5D2T

    NASA Astrophysics Data System (ADS)

    Makhneva, T. M.

    2012-03-01

    Results of a study of the effect of the method of remelting and of heat treatment modes on the behavior of serial curves of impact toughness and on the position of cold-shortness threshold in steel 08Kh15N5D2T are presented.

  10. Nitrate Removal in Two Relict Oxbow Urban Wetlands: A 15N Mass-balance Approach

    EPA Science Inventory

    A 15N-tracer method was used to quantify nitrogen (N) removal processes in two relict oxbow wetlands located adjacent to the Minebank Run restored stream reach in Baltimore County (Maryland, USA) during summer 2009 and early spring 2010. A mass-balance approach was used to determ...

  11. δ15N as a proxy for historic anthropogenic nitrogen loading in Charleston Harbor, SC, USA

    NASA Astrophysics Data System (ADS)

    Payne, T. N.; Andrus, C. F. T.

    2015-12-01

    Bivalve shell geochemistry can serve as a useful indicator of changes in coastal environments. There is increasing interest in developing paleoenvironmental proxies from mollusk shell organic components. Numerous studies have focused on how the δ15N obtained from bivalve tissues can be used to trace present-day wastewater input into estuaries. However, comparatively little attention has been paid to tracing the impact of anthropogenic nitrogen loading into estuaries over time. By measuring historic levels of δ15N in the organic fraction of oyster shells (Crassostrea virginica) from archaeological sites around Charleston Harbor and comparing those levels to the δ15N content of modern shells, it is possible to assess how nitrogen has fluctuated historically in the area. Whole-shell samples from the Late Archaic Period (~3000-4000 BP, Late Woodland Period (~1400-800 BP), 18th and 19th centuries, and modern controls were measured for %N and d15N. Evidence of increased anthropogenic input of N is expected to begin in the early historic period based on similar analysis in Chesapeake Bay. More ancient samples may give insight into baseline conditions prior to recent population growth and industrialization. This information could help understand how large-scale anthropogenic nitrogen loading has affected coastal ecosystems over time and guide future remediation. Furthermore, this project will help refine and improve this novel proxy of past environmental conditions.

  12. Plant delta 15N correlates with the transpiration efficiency of nitrogen acquisition in tropical trees.

    PubMed

    Cernusak, Lucas A; Winter, Klaus; Turner, Benjamin L

    2009-11-01

    Based upon considerations of a theoretical model of (15)N/(14)N fractionation during steady-state nitrate uptake from soil, we hypothesized that, for plants grown in a common soil environment, whole-plant delta(15)N (deltaP) should vary as a function of the transpiration efficiency of nitrogen acquisition (F(N)/v) and the difference between deltaP and root delta(15)N (deltaP - deltaR). We tested these hypotheses with measurements of several tropical tree and liana species. Consistent with theoretical expectations, both F(N)/v and deltaP - deltaR were significant sources of variation in deltaP, and the relationship between deltaP and F(N)/v differed between non-N(2)-fixing and N(2)-fixing species. We interpret the correlation between deltaP and F(N)/v as resulting from variation in mineral nitrogen efflux-to-influx ratios across plasma membranes of root cells. These results provide a simple explanation of variation in delta(15)N of terrestrial plants and have implications for understanding nitrogen cycling in ecosystems.

  13. (15)N natural abundance of non-fixing woody species in the Brazilian dry forest (caatinga).

    PubMed

    de Freitas, Ana Dolores Santiago; de Sa Barretto Sampaio, Everardo Valadares; Menezes, Romulo Simoes Cezar; Tiessen, Holm

    2010-06-01

    Foliar delta(15)N values are useful to calculate N(2) fixation and N losses from ecosystems. However, a definite pattern among vegetation types is not recognised and few data are available for semi-arid areas. We sampled four sites in the Brazilian caatinga, along a water availability gradient. Sites with lower annual rainfall (700 mm) but more uniform distribution (six months) had delta(15)N values of 9.4 and 10.1 per thousand, among the highest already reported, and significantly greater than those (6.5 and 6.3 per thousand) of sites with higher rainfall (800 mm) but less uniform distribution (three months). There were no significant differences at each site among species or between non-fixing legume and non-legume species, in spite of the higher N content of the first group. Therefore, they constitute ideal reference plants in estimations of legume N(2) fixation. The higher values could result from higher losses of (15)N depleted gases or lower losses of enriched (15)N material.

  14. 15N fractionation in star-forming regions and Solar System objects

    NASA Astrophysics Data System (ADS)

    Wirström, E. S.; Adande, G.; Milam, S. N.; Charnley, S. B.; Cordiner, M. A.

    2016-10-01

    We briefly review what is currently known of 14N/15N ratios in interstellar molecules. We summarize the fractionation ratios measured in HCN, HNC, CN, N2 and NH3, and compare these to theoretical predictions and to the isotopic inventory of cometary volatiles.

  15. Using a Macroalgal δ15N Bioassay to Detect Cruise Ship Waste Water Effluent Inputs

    EPA Science Inventory

    Nitrogen stable isotopes are a powerful tool for tracking sources of N to marine ecosystems. I used green macroalgae as a bioassay organism to evaluate if the δ15N signature of cruise ship waste water effluent (CSWWE) could be detected in Skagway Harbor, AK. Opportunistic green...

  16. Tracing Nitrogen through Landscapes to Coastal Wetlands using d15N of Larval Fish

    EPA Science Inventory

    Our objective was to evaluate the use of the nitrogen stable isotope value (d15N) of larval fish as an indicator of incipient anthropogenic nitrogen loading to coastal wetlands in the Great Lakes. We sampled coastal wetlands in five Lake Superior south shore tributaries that had ...

  17. Compound-Specific δ15N Amino Acid Measurements in Littoral Mussels in the California Upwelling Ecosystem: A New Approach to Generating Baseline δ15N Isoscapes for Coastal Ecosystems

    PubMed Central

    Vokhshoori, Natasha L.; McCarthy, Matthew D.

    2014-01-01

    We explored δ15N compound-specific amino acid isotope data (CSI-AA) in filter-feeding intertidal mussels (Mytilus californianus) as a new approach to construct integrated isoscapes of coastal primary production. We examined spatial δ15N gradients in the California Upwelling Ecosystem (CUE), determining bulk δ15N values of mussel tissue from 28 sites between Port Orford, Oregon and La Jolla, California, and applying CSI-AA at selected sites to decouple trophic effects from isotopic values at the base of the food web. Bulk δ15N values showed a strong linear trend with latitude, increasing from North to South (from ∼7‰ to ∼12‰, R2 = 0.759). In contrast, CSI-AA trophic position estimates showed no correlation with latitude. The δ15N trend is therefore most consistent with a baseline δ15N gradient, likely due to the mixing of two source waters: low δ15N nitrate from the southward flowing surface California Current, and the northward transport of the California Undercurrent (CUC), with15N-enriched nitrate. This interpretation is strongly supported by a similar linear gradient in δ15N values of phenylalanine (δ15NPhe), the best AA proxy for baseline δ15N values. We hypothesize δ15NPhe values in intertidal mussels can approximate annual integrated δ15N values of coastal phytoplankton primary production. We therefore used δ15NPhe values to generate the first compound-specific nitrogen isoscape for the coastal Northeast Pacific, which indicates a remarkably linear gradient in coastal primary production δ15N values. We propose that δ15NPhe isoscapes derived from filter feeders can directly characterize baseline δ15N values across major biochemical provinces, with potential applications for understanding migratory and feeding patterns of top predators, monitoring effects of climate change, and study of paleo- archives. PMID:24887109

  18. Vertical δ13C and δ15N changes during pedogenesis

    NASA Astrophysics Data System (ADS)

    Brunn, Melanie; Spielvogel, Sandra; Wells, Andrew; Condron, Leo; Oelmann, Yvonne

    2015-04-01

    The natural abundance of soil organic matter (SOM) stable C and N isotope ratios are subjected to vertical changes throughout the soil profile. This vertical distribution is a widely reported phenomenon across varieties of ecosystems and constitutes important insights of soil carbon cycling. In most ecosystems, SOM becomes enriched in heavy isotopes by several per mill in the first few centimeters of the topsoil. The enrichment of 13C in SOM with soil depth is attributed to biological and physical-chemical processes in soil e.g., plant physiological impacts, microbial decomposition, sorption and transport processes. Such vertical trends in 13C and 15N abundance have rarely been related to SOM composition during pedogenesis. The aims of our study were to investigate short and long-term δ13C and δ15N depth changes and their interrelations under progressing pedogenesis and ecosystem development. We sampled soils across the well studied fordune progradation Haast-chronosequence, a dune ridge system under super-humid climate at the West Coast of New Zealand's South Island (43° 53' S, 169° 3' E). Soils from 11 sites with five replicates each covered a time span of around 2870 yr of soil development (from Arenosol to Podzol). Vertical changes of δ13C and δ15N values of SOM were investigated in the organic layers and in 1-cm depth intervals of the upper 10 cm of the mineral soil. With increasing soil depth SOM became enriched in δ13C by 1.9 ± SE 0.1 o and in δ15N by 6.0 ± 0.4 ‰˙Litter δ13C values slightly decreased with increasing soil age (r = -0.61; p = 0.00) likely due to less efficient assimilation linked to nutrient limitations. Fractionation processes during mycorrhizal transfer appeared to affect δ15N values in the litter. We found a strong decrease of δ15N in the early succession stages ≤ 300 yr B.P. (r = -0.95; p = 0.00). Positive relations of vertical 13C and 15N enrichment with soil age might be related to decomposition and appeared to be

  19. Benchmark Theoretical and Experimental Study on (15)N NMR Shifts of Oxidatively Damaged Guanine.

    PubMed

    Dračínský, Martin; Šála, Michal; Klepetářová, Blanka; Šebera, Jakub; Fukal, Jiří; Holečková, Veronika; Tanaka, Yoshiyuki; Nencka, Radim; Sychrovský, Vladimír

    2016-02-11

    The (15)N NMR shifts of 9-ethyl-8-oxoguanine (OG) were calculated and measured in liquid DMSO and in crystal. The OG molecule is a model for oxidatively damaged 2'-deoxyguanosine that occurs owing to oxidative stress in cell. The DNA lesion is repaired with human 8-oxoguanine glycosylase 1 (hOGG1) base-excision repair enzyme, however, the exact mechanism of excision of damaged nucleobase with hOGG1 is currently unknown. This benchmark study on (15)N NMR shifts of OG aims their accurate structural interpretation and calibration of the calculation protocol utilizable in future studies on mechanism of hOGG1 enzyme. The effects of NMR reference, DFT functional, basis set, solvent, structure, and dynamics on calculated (15)N NMR shifts were first evaluated for OG in crystal to calibrate the best performing calculation method. The effect of large-amplitude motions on (15)N NMR shifts of OG in liquid was calculated employing molecular dynamics. The B3LYP method with Iglo-III basis used for B3LYP optimized geometry with 6-311++G(d,p) basis and including effects of solvent and molecular dynamic was the calculation protocol used for calculation of (15)N NMR shifts of OG. The NMR shift of N9 nitrogen of OG was particularly studied because the atom is involved in an N-glycosidic bond that is cleaved with hOGG1. The change of N9 NMR shift owing to oxidation of 9-ethylguanine (G) measured in liquid was -27.1 ppm. The calculated N9 NMR shift of OG deviated from experiment in crystal and in liquid by 0.45 and 0.65 ppm, respectively. The calculated change of N9 NMR shift owing to notable N9-pyramidalization of OG in one previously found polymorph was 20.53 ppm. We therefore assume that the pyramidal geometry of N9 nitrogen that could occur for damaged DNA within hOGG1 catalytic site might be detectable with (15)N NMR spectroscopy. The calculation protocol can be used for accurate structural interpretation of (15)N NMR shifts of oxidatively damaged guanine DNA residue.

  20. Robust and low cost uniform (15)N-labeling of proteins expressed in Drosophila S2 cells and Spodoptera frugiperda Sf9 cells for NMR applications.

    PubMed

    Meola, Annalisa; Deville, Célia; Jeffers, Scott A; Guardado-Calvo, Pablo; Vasiliauskaite, Ieva; Sizun, Christina; Girard-Blanc, Christine; Malosse, Christian; van Heijenoort, Carine; Chamot-Rooke, Julia; Krey, Thomas; Guittet, Eric; Pêtres, Stéphane; Rey, Félix A; Bontems, François

    2014-10-01

    Nuclear magnetic resonance spectroscopy is a powerful tool to study structural and functional properties of proteins, provided that they can be enriched in stable isotopes such as (15)N, (13)C and (2)H. This is usually easy and inexpensive when the proteins are expressed in Escherichiacoli, but many eukaryotic (human in particular) proteins cannot be produced this way. An alternative is to express them in insect cells. Labeled insect cell growth media are commercially available but at prohibitive prices, limiting the NMR studies to only a subset of biologically important proteins. Non-commercial solutions from academic institutions have been proposed, but none of them is really satisfying. We have developed a (15)N-labeling procedure based on the use of a commercial medium depleted of all amino acids and supplemented with a (15)N-labeled yeast autolysate for a total cost about five times lower than that of the currently available solutions. We have applied our procedure to the production of a non-polymerizable mutant of actin in Sf9 cells and of fragments of eukaryotic and viral membrane fusion proteins in S2 cells, which typically cannot be produced in E. coli, with production yields comparable to those obtained with standard commercial media. Our results support, in particular, the putative limits of a self-folding domain within a viral glycoprotein of unknown structure.

  1. Simple wavelength assignment protocol

    NASA Astrophysics Data System (ADS)

    Suryaputra, Stephen; Touch, Joseph D.; Bannister, Joseph A.

    2000-10-01

    IP routers can be coupled with wavelength-selective optical cross- connects to support existing Internet infrastructure in a wavelength division multiplexing (WDM) optical network. Because optical wavelength routing is transparent to IP, packets can bypass traditional forwarding and pass directly through the optical cross-connect, resulting in very high throughput and low delay routing. This approach shares features with label switching, but wavelengths are much more scarce resource than labels. Because optical switches have larger switching times than electronic switches, and wavelength conversions are expensive, wavelength label swapping is not easily done. Wavelength label assignments must consider these limitations to be practical in an optical environment. The performance of an instance of this approach, called Packet over Wavelengths (POW) has been simulated and studied. A new signaling protocol, Simple Wavelength Assignment Protocol (SWAP) is devised to be POW signaling protocol. SWAP takes into account the optical device limitations, and is designed to minimize wavelength conversion, utilize wavelengths with the merging of flows, and reduce the reconfiguration of optical switches. SWAP, to our knowledge, is the first approach to combine signaling and wavelength assignment in an on- line protocol. This paper describes high level SWAP design challenges, decision, and overhead.

  2. [Characteristics of urea 15N absorption, allocation, and utilization by sweet-cherry (Prunus avium L.)].

    PubMed

    Zhao, Feng-Xia; Jiang, Yuan-Mao; Peng, Fu-Tian; Gao, Xiang-Bin; Liu, Bing-Hua; Wang, Hai-Yun; Zhao, Lin

    2008-03-01

    With five-year old 'Zaodaguo' sweet-cherry (Prunus avium L.) as test material, this paper studied the characteristics of its urea 15N absorption, allocation, and utilization when applied before bud-break. The results showed that the Ndff of different organs increased gradually with time, and was higher in fine roots and storage organs at full-blooming stage. At fruit core-hardening stage, the Ndff of long shoots and leaves increased quickly, reaching to 0.72% and 0.59% , respectively. From fruit core-hardening to harvesting stage, the Ndff of fruit had a rapid increase, with the peak (1.78%) at harvesting stage. After harvest, the Ndff of neonatal organs increased slowly while that of storage organs increased quickly. At full-blooming stage, the absorbed 15N in roots was firstly allocated to storage organs, with the highest allocation rate (54.91%) in large roots. At fruit core-hardening stage, the allocation rate in fine roots and storage organs decreased from 85.43% to 55.11%, while that in neonatal branches and leaves increased to 44.89%. At harvesting stage, the allocation rate in different organs had no significant change, but after harvest, the absorbed 15N had a rapid translocation to storage organs, and the allocation rate in fine roots and storage organs reached the highest (72.26%) at flower bud differentiation stage. The 15N allocation rate in neonatal branches and leaves at flower bud differentiation stage was decreased by 19.31%, compared with that at harvesting stage. From full-blooming to flower bud differentiation stage, the utilization rate of urea 15N was increasing, and reached the peak (16.86%) at flower bud differentiation stage.

  3. Determination of 15N chemical shift anisotropy from a membrane-bound protein by NMR spectroscopy.

    PubMed

    Pandey, Manoj Kumar; Vivekanandan, Subramanian; Ahuja, Shivani; Pichumani, Kumar; Im, Sang-Choul; Waskell, Lucy; Ramamoorthy, Ayyalusamy

    2012-06-21

    Chemical shift anisotropy (CSA) tensors are essential in the structural and dynamic studies of proteins using NMR spectroscopy. Results from relaxation studies in biomolecular solution and solid-state NMR experiments on aligned samples are routinely interpreted using well-characterized CSA tensors determined from model compounds. Since CSA tensors, particularly the (15)N CSA, highly depend on a number of parameters including secondary structure, electrostatic interaction, and the amino acid sequence, there is a need for accurately determined CSA tensors from proteins. In this study, we report the backbone amide-(15)N CSA tensors for a 16.7-kDa membrane-bound and paramagnetic-heme containing protein, rabbit Cytochrome b(5) (cytb(5)), determined using the (15)N CSA/(15)N-(1)H dipolar transverse cross-correlation rates. The mean values of (15)N CSA determined for residues in helical, sheet, and turn regions are -187.9, -166.0, and -161.1 ppm, respectively, with an overall average value of -171.7 ppm. While the average CSA value determined from this study is in good agreement with previous solution NMR experiments on small globular proteins, the CSA value determined for residues in helical conformation is slightly larger, which may be attributed to the paramagnetic effect from Fe(III) of the heme unit in cytb(5). However, like in previous solution NMR studies, the CSA values reported in this study are larger than the values measured from solid-state NMR experiments. We believe that the CSA parameters reported in this study will be useful in determining the structure, dynamics, and orientation of proteins, including membrane proteins, using NMR spectroscopy.

  4. Variable δ(15)N diet-tissue discrimination factors among sharks: implications for trophic position, diet and food web models.

    PubMed

    Olin, Jill A; Hussey, Nigel E; Grgicak-Mannion, Alice; Fritts, Mark W; Wintner, Sabine P; Fisk, Aaron T

    2013-01-01

    The application of stable isotopes to characterize the complexities of a species foraging behavior and trophic relationships is dependent on assumptions of δ(15)N diet-tissue discrimination factors (∆(15)N). As ∆(15)N values have been experimentally shown to vary amongst consumers, tissues and diet composition, resolving appropriate species-specific ∆(15)N values can be complex. Given the logistical and ethical challenges of controlled feeding experiments for determining ∆(15)N values for large and/or endangered species, our objective was to conduct an assessment of a range of reported ∆(15)N values that can hypothetically serve as surrogates for describing the predator-prey relationships of four shark species that feed on prey from different trophic levels (i.e., different mean δ(15)N dietary values). Overall, the most suitable species-specific ∆(15)N values decreased with increasing dietary-δ(15)N values based on stable isotope Bayesian ellipse overlap estimates of shark and the principal prey functional groups contributing to the diet determined from stomach content analyses. Thus, a single ∆(15)N value was not supported for this speciose group of marine predatory fishes. For example, the ∆(15)N value of 3.7‰ provided the highest percent overlap between prey and predator isotope ellipses for the bonnethead shark (mean diet δ(15)N = 9‰) whereas a ∆(15)N value < 2.3‰ provided the highest percent overlap between prey and predator isotope ellipses for the white shark (mean diet δ(15)N = 15‰). These data corroborate the previously reported inverse ∆(15)N-dietary δ(15)N relationship when both isotope ellipses of principal prey functional groups and the broader identified diet of each species were considered supporting the adoption of different ∆(15)N values that reflect the predators' δ(15)N-dietary value. These findings are critical for refining the application of stable isotope modeling approaches as inferences regarding a

  5. Variable δ15N Diet-Tissue Discrimination Factors among Sharks: Implications for Trophic Position, Diet and Food Web Models

    PubMed Central

    Olin, Jill A.; Hussey, Nigel E.; Grgicak-Mannion, Alice; Fritts, Mark W.; Wintner, Sabine P.; Fisk, Aaron T.

    2013-01-01

    The application of stable isotopes to characterize the complexities of a species foraging behavior and trophic relationships is dependent on assumptions of δ15N diet-tissue discrimination factors (∆15N). As ∆15N values have been experimentally shown to vary amongst consumers, tissues and diet composition, resolving appropriate species-specific ∆15N values can be complex. Given the logistical and ethical challenges of controlled feeding experiments for determining ∆15N values for large and/or endangered species, our objective was to conduct an assessment of a range of reported ∆15N values that can hypothetically serve as surrogates for describing the predator-prey relationships of four shark species that feed on prey from different trophic levels (i.e., different mean δ15N dietary values). Overall, the most suitable species-specific ∆15N values decreased with increasing dietary-δ15N values based on stable isotope Bayesian ellipse overlap estimates of shark and the principal prey functional groups contributing to the diet determined from stomach content analyses. Thus, a single ∆15N value was not supported for this speciose group of marine predatory fishes. For example, the ∆15N value of 3.7‰ provided the highest percent overlap between prey and predator isotope ellipses for the bonnethead shark (mean diet δ15N = 9‰) whereas a ∆15N value < 2.3‰ provided the highest percent overlap between prey and predator isotope ellipses for the white shark (mean diet δ15N = 15‰). These data corroborate the previously reported inverse ∆15N-dietary δ15N relationship when both isotope ellipses of principal prey functional groups and the broader identified diet of each species were considered supporting the adoption of different ∆15N values that reflect the predators’ δ15N-dietary value. These findings are critical for refining the application of stable isotope modeling approaches as inferences regarding a species’ ecological role in their

  6. Influence of open ocean nitrogen supply on the skeletal δ15N of modern shallow-water scleractinian corals

    NASA Astrophysics Data System (ADS)

    Wang, Xingchen T.; Sigman, Daniel M.; Cohen, Anne L.; Sinclair, Daniel J.; Sherrell, Robert M.; Cobb, Kim M.; Erler, Dirk V.; Stolarski, Jarosław; Kitahara, Marcelo V.; Ren, Haojia

    2016-05-01

    The isotopic composition of skeleton-bound organic nitrogen in shallow-water scleractinian corals (hereafter, CS-δ15N) is an emerging tool for studying the marine nitrogen cycle in the past. The CS-δ15N has been shown to reflect the δ15N of nitrogen (N) sources to corals, with most applications to date focusing on the anthropogenic/terrestrial N inputs to reef environments. However, many coral reefs receive their primary N sources from the open ocean, and the CS-δ15N of these corals may provide information on past changes in the open ocean regional and global N cycle. Using a recently developed persulfate/denitrifier-based method, we measured CS-δ15N in modern shallow-water scleractinian corals from 8 sites proximal to the open ocean. At sites with low open ocean surface nitrate concentrations typical of the subtropics and tropics, measured CS-δ15N variation on seasonal and annual timescales is most often less than 2‰. In contrast, a broad range in CS-δ15N (of ∼10‰) is measured across these sites, with a strong correlation between CS-δ15N and the δ15N of the deep nitrate supply to the surface waters near the reefs. While CS-δ15N can be affected by other N sources as well and can vary in response to local reef conditions as well as coral/symbiont physiological changes, this survey indicates that, when considering corals proximal to the open ocean, the δ15N of the subsurface nitrate supply to surface waters drives most of the CS-δ15N variation across the global ocean. Thus, CS-δ15N is a promising proxy for reconstructing the open ocean N cycle in the past.

  7. Isotopic variability of cave bears (δ15N, δ13C) across Europe during MIS 3

    NASA Astrophysics Data System (ADS)

    Krajcarz, Magdalena; Pacher, Martina; Krajcarz, Maciej T.; Laughlan, Lana; Rabeder, Gernot; Sabol, Martin; Wojtal, Piotr; Bocherens, Hervé

    2016-01-01

    Collagen, the organic fraction of bone, records the isotopic parameters of consumed food for carbon (δ13C) and nitrogen (δ15N). This relationship of isotopic signature between diet and tissue is an important tool for the study of dietary preferences of modern and fossil animal species. Since the first information on the isotopic signature of cave bear was reported, numerous data from Europe have become available. The goal of this work is to track the geographical variation of cave bear collagen isotopic values in Europe during Marine Isotopic Stage 3 (about 60,000-25,000 yr BP). In this study the results of new δ13C and δ15N isotopic analyses of cave bear collagen from four Central-Eastern European sites are presented, as well as a review of all published isotopic data for cave bears of the same period. The main conclusion is a lack of geographical East-West pattern in the variations of δ13C and δ15N values of cave bear collagen. Moreover, no relationship was found between cave bear taxonomy and isotopic composition. The cave bears from Central-Eastern Europe exhibit δ13C and δ15N values near the average of the range of Central, Western and Southern European cave bears. Despite the fact that most cave bear sites follow an altitudinal gradient, separate groups of sites exhibit shift in absolute values of δ13C, what disturbs an altitude-related isotopic pattern. The most distinct groups are: high Alpine sites situated over 1500 m a.s.l. - in terms of δ13C; and two Romanian sites Peştera cu Oase and Urşilor - in case of δ15N. Although the cave bear isotopic signature is driven by altitude, the altitudinal adjustment of isotopic data is not enough to explain the isotopic dissimilarity of these cave bears. The unusually high δ15N signature of mentioned Romanian sites is an isolated case in Europe. Cave bears from relatively closely situated Central-Eastern European sites and other Romanian sites are more similar to Western European than to Romanian

  8. The origin of nitrogen on Jupiter and Saturn from the 15N/14N ratio

    NASA Astrophysics Data System (ADS)

    Fletcher, Leigh N.; Greathouse, T. K.; Orton, G. S.; Irwin, P. G. J.; Mousis, O.; Sinclair, J. A.; Giles, R. S.

    2014-08-01

    The Texas Echelon cross Echelle Spectrograph (TEXES), mounted on NASA’s Infrared Telescope Facility (IRTF), was used to map mid-infrared ammonia absorption features on both Jupiter and Saturn in February 2013. Ammonia is the principle reservoir of nitrogen on the giant planets, and the ratio of isotopologues (15N/14N) can reveal insights into the molecular carrier (e.g., as N2 or NH3) of nitrogen to the forming protoplanets, and hence the source reservoirs from which these worlds accreted. We targeted two spectral intervals (900 and 960 cm-1) that were relatively clear of terrestrial atmospheric contamination and contained close features of 14NH3 and 15NH3, allowing us to derive the ratio from a single spectrum without ambiguity due to radiometric calibration (the primary source of uncertainty in this study). We present the first ground-based determination of Jupiter’s 15N/14N ratio (in the range from 1.4×10-3 to 2.5×10-3), which is consistent with both previous space-based studies and with the primordial value of the protosolar nebula. On Saturn, we present the first upper limit on the 15N/14N ratio of no larger than 2.0×10-3 for the 900-cm-1 channel and a less stringent requirement that the ratio be no larger than 2.8×10-3 for the 960-cm-1 channel (1σ confidence). Specifically, the data rule out strong 15N-enrichments such as those observed in Titan’s atmosphere and in cometary nitrogen compounds. To the extent possible with ground-based radiometric uncertainties, the saturnian and jovian 15N/14N ratios appear indistinguishable, implying that 15N-enriched ammonia ices could not have been a substantial contributor to the bulk nitrogen inventory of either planet. This result favours accretion of primordial N2 on both planets, either in the gas phase from the solar nebula, or as ices formed at very low temperatures. Finally, spatially-resolved TEXES observations are used to derive zonal contrasts in tropospheric temperatures, phosphine and 14NH3 on both

  9. Military Personnel Assignments

    DTIC Science & Technology

    1987-01-09

    of Defense, Organization of the Joint Chiefs of Staff and the Defense Agencies," > f , February 4, 1970 (hereby canceled) (d) DoD Directive 1315.14...34DoD Components"). 2. Does not apply to service members in non-DoD activities covered by DoD Directive 1000.17 (reference ( f )). C. DEFINITIONS Terms...possible, shall be allowed to extend any assignment voluntarily beyond the prescribed tour. f . Through the grades of 0-5 for officers and E-8 for enlisted

  10. Nitrogen cycling in an extreme hyperarid environment inferred from δ15N analyses of plants, soils and herbivore diet

    PubMed Central

    Díaz, Francisca P.; Frugone, Matías; Gutiérrez, Rodrigo A.; Latorre, Claudio

    2016-01-01

    Climate controls on the nitrogen cycle are suggested by the negative correlation between precipitation and δ15N values across different ecosystems. For arid ecosystems this is unclear, as water limitation among other factors can confound this relationship. We measured herbivore feces, foliar and soil δ15N and δ13C values and chemically characterized soils (pH and elemental composition) along an elevational/climatic gradient in the Atacama Desert, northern Chile. Although very positive δ15N values span the entire gradient, soil δ15N values show a positive correlation with aridity as expected. In contrast, foliar δ15N values and herbivore feces show a hump-shaped relationship with elevation, suggesting that plants are using a different N source, possibly of biotic origin. Thus at the extreme limits of plant life, biotic interactions may be just as important as abiotic processes, such as climate in explaining ecosystem δ15N values. PMID:26956399

  11. Pathways to assignment of payees.

    PubMed

    Rosen, Marc I; Ablondi, Karen; Black, Anne C; Serowik, Kristin L; Rowe, Michael

    2014-04-01

    How clients come to be assigned representative payees and/or conservators to manage their funds is not well understood. We compared clients assigned a payee during a clinical trial of a money management-based intervention to those not assigned payees and examined antecedents to payee assignment. One year after randomization, significantly more clients assigned to the advisor teller money manager (ATM) money management intervention were assigned payees than participants in the control condition (10 of 47 vs. 2 of 43; p = .02); those assigned payees had lower baseline GAF scores and participated more in study therapies. Several ATM clients were assigned payees after third parties paid more attention to clients' finances, and others after having negotiated storage of their funds with the ATM money manager during the study. Assignment of payees appears to be influenced by whether third parties critically attend to how clients' manage funds and by clients' receptiveness to having a payee.

  12. The AGB star nucleosynthesis in the light of the recent 17O ( p ,α)14N and 18O ( p ,α)15N reaction rate determinations

    NASA Astrophysics Data System (ADS)

    Palmerini, S.; Sergi, M. L.; La Cognata, M.; Lamia, L.; Pizzone, R. G.; Spitaleri, C.

    2015-02-01

    Presolar grains form in the cold and dusty envelopes of Asymptotic Giant Branch (AGB) stars. These solides, once that have been ejected by stellar winds, come to us as inclusions in meteorites providing invaluable benchmarks and constraints for our knowledge of low temeperature H-burning in stars. The Trojan Horse Method (THM) has been used to investigate the low-energy cross sections of the 17O ( p ,α)14N and 18O ( p ,α)15N reactions. Moreover, the strength of the 65 keV resonance in the 17O ( p ,α)14N reaction, measured by means of the THM, has been used to renormalize the corresponding resonance strength in the 17O + p radiative capture channel. The new estimates of the reaction rates have been introduced into calculations of AGB star nucleosynthesis and the results have been compared with geochemical analysis of "presolar" grains to determine their impact on astrophysical environments.

  13. Natural abundance 14N and 15N solid-state NMR of pharmaceuticals and their polymorphs

    DOE PAGES

    Veinberg, Stanislav L.; Johnston, Karen E.; Jaroszewicz, Michael J.; ...

    2016-06-08

    14N ultra-wideline (UW), 1H{15N} indirectly-detected HETCOR (idHETCOR) and 15N dynamic nuclear polarization (DNP) solid-state NMR (SSNMR) experiments, in combination with plane-wave density functional theory (DFT) calculations of 14N EFG tensors, were utilized to characterize a series of nitrogen-containing active pharmaceutical ingredients (APIs), including HCl salts of scopolamine, alprenolol, isoprenaline, acebutolol, dibucaine, nicardipine, and ranitidine. Here, a case study applying these methods for the differentiation of polymorphs of bupivacaine HCl is also presented. All experiments were conducted upon samples with naturally-abundant nitrogen isotopes. For most of the APIs, it was possible to acquire frequency-stepped UW 14N SSNMR spectra of stationarymore » samples, which display powder patterns corresponding to pseudo-tetrahedral (i.e., RR'R"NH+ and RR'NH2+) or other (i.e., RNH2 and RNO2) nitrogen environments.« less

  14. Interresidue carbonyl-carbonyl polarization transfer experiments in uniformly 13C, 15N-labeled peptides and proteins

    NASA Astrophysics Data System (ADS)

    Janik, Rafal; Ritz, Emily; Gravelle, Andrew; Shi, Lichi; Peng, Xiaohu; Ladizhansky, Vladimir

    2010-03-01

    In this work, we demonstrate that Homonuclear Rotary Resonance Recoupling (HORROR) can be used to reintroduce carbonyl-carbonyl interresidue dipolar interactions and to achieve efficient polarization transfer between carbonyl atoms in uniformly 13C, 15N-labeled peptides and proteins. We show that the HORROR condition is anisotropically broadened and overall shifted to higher radio frequency intensities because of the CSA effects. These effects are analyzed theoretically using Average Hamiltonian Theory. At spinning frequencies used in this study, 22 kHz, this broadening is experimentally found to be on the order of a kilohertz at a proton field of 600 MHz. To match HORROR condition over all powder orientations, variable amplitude radio frequency (RF) fields are required, and efficient direct transfers on the order of 20-30% can be straightforwardly established. Two- and three-dimensional chemical shift correlation experiments establishing long-range interresidue connectivities (e.g., (N[i]-CO[i - 2])) are demonstrated on the model peptide N-acetyl-valine-leucine, and on the third immunoglobulin binding domain of protein G. Possible future developments are discussed.

  15. Community 15N isoscapes to resolve plant-plant-interactions at the spatial scale

    NASA Astrophysics Data System (ADS)

    Hellmann, Christine; Rascher, Katherine G.; Máguas, Cristina; Werner, Christiane

    2014-05-01

    Isoscapes have greatly improved our ability to understand biogeochemical processes on continental to global scales. However, the isoscapes framework may also have significant potential to resolve the spatial component of within-community interactions. For example, exotic plant invaders often exert strong impacts on ecosystem functioning, particularly regarding water-, carbon- and nutrient-cycles, but the spatial extent of such alterations is largely unknown. Here we show that massive N input by the N2-fixing exotic invasive Acacia longifolia to a Portuguese dune system can be traced using spatially resolved information on native plants' leaf δ15N. We found isotopic signatures of N to differ strongly between the native system (δ15N c. -10 o) and the atmospherically derived N in A. longifolia phyllodes (δ15N c. 0 o). Thus, sources of N for native plants could be readily distinguished. Leaf δ15N of a native, non-fixing species was increasingly enriched the closer the plant grew to the invader, indicating uptake of fixed N provided by A. longifolia. The enrichment was evident far beyond the stands of the invader, demonstrating that A. longifolia affected N budgets of native species up to a distance of 8 m exceeding the margin of the canopy. Furthermore, using the isoscapes approach, we were able to quantify the total area of N enrichment and could thus show that the area affected by invasion was at least 3.5 times larger than the area actually occupied by the invader. However, a native N2-fixing species had no such effects. Thus, downscaling isoscapes to the community level opens new frontiers in quantifying the spatial dimension of functional changes associated with plant invasions. Moreover, considering the feasibility and applicability of this approach, it may provide a promising tool to identify, quantify and monitor different types of functional plant-plant interactions within communities at a spatially explicit scale.

  16. Asymmetric Induction by a Nitrogen (14) N/(15) N Isotopomer in Conjunction with Asymmetric Autocatalysis.

    PubMed

    Matsumoto, Arimasa; Ozaki, Hanae; Harada, Shunya; Tada, Kyohei; Ayugase, Tomohiro; Ozawa, Hitomi; Kawasaki, Tsuneomi; Soai, Kenso

    2016-12-05

    Chirality arising from isotope substitution, especially with atoms heavier than the hydrogen isotopes, is usually not considered a source of chirality in a chemical reaction. An N(2) ,N(2) ,N(3) ,N(3) -tetramethyl-2,3-butanediamine containing nitrogen ((14) N/(15) N) isotope chirality was synthesized and it was revealed that this isotopically chiral diamine compound acts as a chiral initiator for asymmetric autocatalysis.

  17. Asymmetric Induction by a Nitrogen 14N/15N Isotopomer in Conjunction with Asymmetric Autocatalysis

    PubMed Central

    Ozaki, Hanae; Harada, Shunya; Tada, Kyohei; Ayugase, Tomohiro; Ozawa, Hitomi; Kawasaki, Tsuneomi

    2016-01-01

    Abstract Chirality arising from isotope substitution, especially with atoms heavier than the hydrogen isotopes, is usually not considered a source of chirality in a chemical reaction. An N 2 ,N 2 ,N 3 ,N 3‐tetramethyl‐2,3‐butanediamine containing nitrogen (14N/15N) isotope chirality was synthesized and it was revealed that this isotopically chiral diamine compound acts as a chiral initiator for asymmetric autocatalysis. PMID:27754589

  18. (15)N investigation of nitrogen released from tobacco-waste to be utilized by maize crop.

    PubMed

    Karaman, M Rüştü; Brohi, A Reşit; Inal, Ali; Aydeniz, Akgün

    2004-12-01

    The (15)N study aimed to estimate the portion of nitrogen released from tobacco-waste to be utilized by maize crop. Tobacco-waste at the levels of 0, 2, 4, 6 and 8 g pot(-1) and ((15)NH(4))(2)SO(4) as nitrogen fertilizer labelled with 5 at.% exc. at the levels of 0, 4, 8, 12 and 16 mg N pot(-1) together with a basal dressing of some nutrients were added to pots with Pioneer maize variety. After the harvest, dry matter yield was recorded and (15)N determinations and calculations were made. Tobacco-waste had a positive effect on the growth and on the nitrogen uptake of maize crop. Increasing the rates of tobacco-waste increased the dry matter yield of maize crop from 4.64 g pot(-1) (at control) to 7.22 g pot(-1) (at the tobacco-waste treatment of 8 g pot(-1)). The values of (15)N in the plant derived from nitrogenous fertilizer also increased with increasing nitrogen fertilizer levels, whereas they decreased from 0.427 % to 0.249 % with increasing tobacco-waste treatments from 0 to 8 g pot(-1), respectively. The average values of per cent nitrogen derived from nitrogenous fertilizer (Ndff) varied from 4.32 % to 7.95 % at the rates of 4-16 mg N pot(-1), respec-tively. However, Ndff decreased from 8.54 % to 4.99 % with increasing tobacco-waste treatments from 0 to 8 g pot(-1), respectively. Per cent nitrogen derived from tobacco-waste (Ndft) increased from 21.8 % to 38.5 % with increasing tobacco-waste treatments from 2 to 8 g pot(-1), respectively. The results have revealed that (15)N tracer technique was confidently used for the investigation of nitrogen levels released from tobacco-waste as organic waste to be utilized by maize crop.

  19. δ15N in the turtle grass from the Mexican Caribbean

    NASA Astrophysics Data System (ADS)

    Talavera-Saenz, A.; Sanchez, A.; Ortiz-Hernandez, M.

    2013-05-01

    Nutrient inputs associated with population growth threaten the integrity of coastal ecosystems. To assess the rapid increase in tourism, we compared the δ15N from Thalassia testudinum collected at sites with different levels of tourism development and population to detect the N inputs of wastewater discharge (WD) along the coast of Quintana Roo. The contributions of nitrogen enriched in 15N are directly related to the increase of WD inputs in areas of high tourism development (Nichupte Lagoon in Cancun, >3 million tourists per year from 2007 to 2011 and 0.7 million of resident population) and decreased towards Bahia Akumal and Tulum (>3 million tourists per year from 2007 to 2011 and 0.15 million of resident population). The δ15N from T. testudinum was significantly lower at Mahahual and Puerto Morelos (about 0.4 million tourists per year in 2007 to 2011 and 0.25 million of resident population) than other the sites. In areas of the lowest development and with tourist activity restricted and small population, such as the Yum Balam Reserve and Sian Ka'an Biosphere Reserve, the δ15N values were in much higher enrichment that Mahahual and Puerto Morelos. Therefore is suggested that Mahahual and Puerto Morelos may be used for baseline isotopic monitoring, over environmental pressure on the reef lagoon ecosystem, where tourist activities and population are growing very slow rate. The anthropogenic N input has the potential to impact, both environmentally and economically, the seagrass meadows and the coral reefs along the coast of Quintana Roo and the Caribbean.

  20. Mycorrhizal Fungi Provide Most of the Nitrogen for Symbiotic Arctic Plants: 15N Evidence

    NASA Astrophysics Data System (ADS)

    Hobbie, J. E.; Hobbie, E. A.

    2004-12-01

    When soil nitrogen is in short supply, most terrestrial plants form symbioses with fungi (mycorrhizae) in which fine hyphal threads take up soil nitrogen, transport it into plant roots, and in return receive plant sugars. Because the transfer rates are very difficult to measure in nature, ecologists need new tools by which to assess the role of mycorrhizal fungi in carbon and nitrogen cycling. Recent studies indicate that the natural abundance of 15N taken up from the soil by hyphae is changed during transfer of nitrogen to roots; the result is large differences among the natural abundance of 15N in soil, symbiotic plants, and symbiotic fungi that depend on the mass balance of nitrogen in the mycorrhizal symbiosis. Measurements were carried out in acidic tussock tundra at the Toolik Lake LTER site in Arctic Alaska (68\\deg N 149\\deg W). The \\delta15N of soil N was 1.5%, of soil ammonium was 1.5%, of ericoid and ectomycorrhizal plants was -5.0%, and of ectomycorrhizal fungi was 7.0 parts per mille%. The mass balance of the 15N shows that the plants received 61-86% of their nitrogen from the fungal hyphae. These values, when combined with known plant growth rates, reveal that the plants provided 7-16% of their photosynthetic carbon to the fungi for growth and respiration, or about 25% of all carbon allocated to belowground processes. This analytical technique could be readily applied to other nitrogen-limited ecosystems such as many temperate and boreal forests to quantify the importance for terrestrial carbon and nitrogen cycling of mycorrhizally mediated transfers at the plant-soil interface.

  1. Determination of the δ15N of total nitrogen in solids; RSIL lab code 2893

    USGS Publications Warehouse

    Revesz, Kinga; Qi, Haiping; Coplen, Tyler B.

    2006-01-01

    The purpose of the Reston Stable Isotope Laboratory (RSIL) lab code 2893 is to determine the δ(15N/14N), abbreviated as δ15N , of total nitrogen in solid samples. A Carlo Erba NC 2500 elemental analyzer (EA) is used to convert total nitrogen in a solid sample into N2 gas. The EA is connected to a continuous flow isotope-ratio mass spectrometer (CF-IRMS), which determines relative difference in the isotope-amount ratios of stable nitrogen isotopes (15N/14N)of the product N2 gas. The combustion is quantitative; no isotopic fractionation is involved. Samples are placed in a tin capsule and loaded into the Costech Zero Blank Autosampler of the EA. Under computer control, samples are dropped into a heated reaction tube that contains an oxidant, where the combustion takes place in a helium atmosphere containing an excess of oxygen gas. Combustion products are transported by a helium carrier through a reduction tube to remove excess oxygen and convert all nitrous oxides into N2 and through a drying tube to remove water. The gas-phase products, mainly CO2 and N2, are separated by a gas chromatograph. The gas is then introduced into the isotope-ratio mass spectrometer (IRMS) through a Finnigan MAT (now Thermo Scientific) ConFlo II interface, which also is used to inject N2 reference gas and helium for sample dilution. The IRMS is a Thermo Scientific Delta V Plus CF-IRMS. It has a universal triple collector, two wide cups with a narrow cup in the middle, capable of measuring mass/charge (m/z) 28, 29, 30, simultaneously. The ion beams from N2 are as follows: m/z 28 = N2 = 14N14N; m/z 29 = N2 = 14N15N primarily; m/z 30 = NO = 14N16O primarily, which is a sign of contamination or incomplete reduction.

  2. Food webs of two intermittently open estuaries receiving 15N-enriched sewage effluent

    NASA Astrophysics Data System (ADS)

    Hadwen, Wade L.; Arthington, Angela H.

    2007-01-01

    Carbon and nitrogen stable isotope signatures were used to assess the response of food webs to sewage effluent discharged into two small intermittently open estuaries in northern New South Wales, Australia. One of these systems, Tallows Creek, has a history of direct sewage inputs, whilst the other, Belongil Creek, receives wastewater via an extensive wetland treatment system. The food webs of both systems were driven by algal sources of carbon, reflecting high autotrophic productivity in response to the nutrients entering the system from sewage effluent. All aquatic biota collected from Tallows Creek had significantly enriched δ15N signatures relative to their conspecifics from Belongil Creek, indicating that sewage nitrogen had been assimilated and transferred throughout the Tallows Creek food web. These δ15N values were higher than those reported from studies in permanently open estuaries receiving sewage effluent. We suggest that these enriched signatures and the transfer of nitrogen throughout the entire food web reflect differences in hydrology and associated nitrogen cycling processes between permanently open and intermittently open estuaries. Although all organisms in Tallows Creek were generally 15N-enriched, isotopically light (less 15N-enriched) individuals of estuary perchlet ( Ambassis marianus) and sea mullet ( Mugil cephalus) were also collected. These individuals were most likely recent immigrants into Tallows Creek, as this system had only recently been opened to the ocean. This isotopic discrimination between resident (enriched) and immigrant (significantly less enriched) individuals can provide information on fish movement patterns and the role of heavily polluted intermittently open estuaries in supporting commercially and recreationally valuable estuarine species.

  3. Affordable uniform isotope labeling with (2)H, (13)C and (15)N in insect cells.

    PubMed

    Sitarska, Agnieszka; Skora, Lukasz; Klopp, Julia; Roest, Susan; Fernández, César; Shrestha, Binesh; Gossert, Alvar D

    2015-06-01

    For a wide range of proteins of high interest, the major obstacle for NMR studies is the lack of an affordable eukaryotic expression system for isotope labeling. Here, a simple and affordable protocol is presented to produce uniform labeled proteins in the most prevalent eukaryotic expression system for structural biology, namely Spodoptera frugiperda insect cells. Incorporation levels of 80% can be achieved for (15)N and (13)C with yields comparable to expression in full media. For (2)H,(15)N and (2)H,(13)C,(15)N labeling, incorporation is only slightly lower with 75 and 73%, respectively, and yields are typically twofold reduced. The media were optimized for isotope incorporation, reproducibility, simplicity and cost. High isotope incorporation levels for all labeling patterns are achieved by using labeled algal amino acid extracts and exploiting well-known biochemical pathways. The final formulation consists of just five commercially available components, at costs 12-fold lower than labeling media from vendors. The approach was applied to several cytosolic and secreted target proteins.

  4. Impact of seaweed beachings on dynamics of δ(15)N isotopic signatures in marine macroalgae.

    PubMed

    Lemesle, Stéphanie; Mussio, Isabelle; Rusig, Anne-Marie; Menet-Nédélec, Florence; Claquin, Pascal

    2015-08-15

    A fine-scale survey of δ(15)N, δ(13)C, tissue-N in seaweeds was conducted using samples from 17 sampling points at two sites (Grandcamp-Maisy (GM), Courseulles/Mer (COU)) along the French coast of the English Channel in 2012 and 2013. Partial triadic analysis was performed on the parameter data sets and revealed the functioning of three areas: one estuary (EstA) and two rocky areas (GM(∗), COU(∗)). In contrast to oceanic and anthropogenic reference points similar temporal dynamics characterized δ(15)N signatures and N contents at GM(∗) and COU(∗). Nutrient dynamics were similar: the N-concentrations in seawater originated from the River Seine and local coastal rivers while P-concentrations mainly from these local rivers. δ(15)N at GM(∗) were linked to turbidity suggesting inputs of autochthonous organic matter from large-scale summer seaweed beachings made up of a mixture of Rhodophyta, Phaeophyta and Chlorophyta species. This study highlights the coupling between seaweed beachings and nitrogen sources of intertidal macroalgae.

  5. A novel method for trapping and analyzing 15N in NO for tracing NO sources

    NASA Astrophysics Data System (ADS)

    Kang, Ronghua; Mulder, Jan; Dörsch, Peter

    2016-04-01

    15N isotope tracing is an effective and direct approach to investigate the biological and chemical sources of nitric oxide (NO) in soil. However, NO is highly reactive and rapidly converted to nitrogen dioxide (NO2) in the presence of ozone. Various chemical conversions of NO to the more stable solutes nitrite (NO2-) and nitrate (NO3-) have been proposed, which allow analysing the 15N abundance without major fractionation. However, NO emissions from soils are usually small, posing major challenges to conversion efficiency and background contamination. Here we present a novel method in which NO is oxidized to NO2- by chromium trioxide (CrO3) prior to conversion to NO2- and NO3- in an alkaline hydrogen peroxide (H2O2) solution. Immediately following trapping, manganese dioxide (MnO2) and 5M HCl are added to remove excess H2O2, and to adjust the pH to around 6.0-7.0, respectively. The resulting solution can be stored until analysis and is none-toxic, allowing to use a modified denitrifier method (Zhu et al., submitted), where NO2- and NO3- are reduced quantitatively to nitrous oxide (N2O). Optimum NO conversion rates of > 90% even at extremely low initial NO concentration were obtained with 4% H2O2, 0.5 M NaOH, and 0.5 L min-1 gas flow rate. In a laboratory test, using NO gas with different 15N signals produced from unlabelled and labelled NO2-, we found an overall precision of 0.4‰ for unlabelled and 49.7‰ for NO enriched with 1.0 atom% 15N, respectively. This indicates that this method can be used for both natural abundance studies of NO, as well as in labelling studies tracing NO sources. Zhu J, Yu L, Bakken LR, Mørkved PT, Mulder J, Dörsch P. Controlled induction of denitrification in Pseudomonas aureofaciens: a modified denitrifier method for 15N and 18O analysis in NO3- from natural water samples by IRMS. Submitted.

  6. Paleoenvironmental implications of taxonomic variation among δ 15 N values of chloropigments

    NASA Astrophysics Data System (ADS)

    Higgins, Meytal B.; Wolfe-Simon, Felisa; Robinson, Rebecca S.; Qin, Yelun; Saito, Mak A.; Pearson, Ann

    2011-11-01

    Natural variations in the ratios of nitrogen isotopes in biomass reflect variations in nutrient sources utilized for growth. In order to use δ 15N values of chloropigments of photosynthetic organisms to determine the corresponding δ 15N values of biomass - and by extension, surface waters - the isotopic offset between chlorophyll and biomass must be constrained. Here we examine this offset in various geologically-relevant taxa, grown using nutrient sources that may approximate ocean conditions at different times in Earth's history. Phytoplankton in this study include cyanobacteria (diazotrophic and non-diazotrophic), eukaryotic algae (red and green), and anoxygenic photosynthetic bacteria (Proteobacteria), as well as environmental samples from sulfidic lake water. Cultures were grown using N 2, NO 3-, and NH 4+ as nitrogen sources, and were examined under different light regimes and growth conditions. We find surprisingly high variability in the isotopic difference (δ 15N biomass - δ 15N chloropigment) for prokaryotes, with average values for species ranging from -12.2‰ to +11.7‰. We define this difference as ɛpor, a term that encompasses diagenetic porphyrins and chlorins, as well as chlorophyll. Negative values of ɛpor reflect chloropigments that are 15N-enriched relative to biomass. Notably, this enrichment appears to occur only in cyanobacteria. The average value of ɛpor for freshwater cyanobacterial species is -9.8 ± 1.8‰, while for marine cyanobacteria it is -0.9 ± 1.3‰. These isotopic effects group environmentally but not phylogenetically, e.g., ɛpor values for freshwater Chroococcales resemble those of freshwater Nostocales but differ from those of marine Chroococcales. Our measured values of ɛpor for eukaryotic algae (range = 4.7-8.7‰) are similar to previous reports for pure cultures. For all taxa studied, values of ɛpor do not depend on the type of nitrogen substrate used for growth. The observed environmental control of

  7. Delta15N values of tropical savanna and monsoon forest species reflect root specialisations and soil nitrogen status.

    PubMed

    Schmidt, S; Stewart, G R

    2003-03-01

    A large number of herbaceous and woody plants from tropical woodland, savanna, and monsoon forest were analysed to determine the impact of environmental factors (nutrient and water availability, fire) and biological factors (microbial associations, systematics) on plant delta(15)N values. Foliar delta(15)N values of herbaceous and woody species were not related to growth form or phenology, but a strong relationship existed between mycorrhizal status and plant delta(15)N. In woodland and savanna, woody species with ectomycorrhizal (ECM) associations and putative N(2)-fixing species with ECM/arbuscular (AM) associations had lowest foliar delta(15)N values (1.0-0.6 per thousand ), AM species had mostly intermediate delta(15)N values (average +0.6 per thousand ), while non-mycorrhizal Proteaceae had highest delta(15)N values (+2.9 to +4.1 per thousand ). Similar differences in foliar delta(15)N were observed between AM (average 0.1 and 0.2 per thousand ) and non-mycorrhizal (average +0.8 and +0.3 per thousand ) herbaceous species in woodland and savanna. Leguminous savanna species had significantly higher leaf N contents (1.8-2.5% N) than non-fixing species (0.9-1.2% N) indicating substantial N acquisition via N(2) fixation. Monsoon forest species had similar leaf N contents (average 2.4% N) and positive delta(15)N values (+0.9 to +2.4 per thousand ). Soil nitrification and plant NO(3)(-) use was substantially higher in monsoon forest than in woodland or savanna. In the studied communities, higher soil N content and nitrification rates were associated with more positive soil delta(15)N and plant delta(15)N. In support of this notion, Ficus, a high NO(3)(-) using taxa associated with NO(3)(-) rich sites in the savanna, had the highest delta(15)N values of all AM species in the savanna. delta(15)N of xylem sap was examined as a tool for studying plant delta(15)N relations. delta(15)N of xylem sap varied seasonally and between differently aged Acacia and other savanna

  8. Foliar retention of 15N-nitrate and 15N-ammonium by red maple (Acer rubrum) and white oak (Quercus alba) leaves from simulated rain

    SciTech Connect

    Garten Jr, Charles T; Hanson, Paul J

    1990-07-01

    Studies of nitrogen cycling in forests indicate that trees assimilate atmospheric nitrate and ammonium and that differences between atmospheric deposition to the forest canopy and deposition measured in forest throughfall can be attributed to the removal of these ions from rain by tree leaves. Red maple and white oak leaves were exposed to artificial rain solutions (pH 4.1) containing {sup 15}N-labeled nitrate (3.5 {micro}g N/ml) or ammonium (2.2 {micro}g N/ml). At two time intervals after exposure (2 hr and 2 days) an exposed leaf and a control (non-exposed) leaf were removed from replicate seedlings. Based on results from {sup 15}N analysis, most of the nitrate applied to tree leaves was removed by washing with water; the mean per cent removal ({+-} standard error, N = 4) was 87 {+-} 1 and 73 {+-} 4% of the {sup 15}NO-N Applied to red maple and white oak leaves, respectively. Relative retention of {sup 15}NH{sub 4}-N by the leaves was greater than that observed for {sup 15}NO{sub 3}-N. In red maple and white oak leaves, 58 {+-} 9 and 84 {+-} 7% (mean {+-} standard error, N = 4), respectively, of the applied ammonium was not removed by washing treatments. Our results show that the foliar uptake of {sup 15}NH{sub 4}{sup +} from simulated rain by deciduous tree leaves is greater than that for {sup 15}NO{sub 3}{sup -}. Greater retention of NH{sub 4}{sup +} than NO{sub 3}{sup -} ions by red maple and white oak leaves from simulated rainfall is consistent with field observations showing a preferential retention of ammonium from rainfall by forest canopies. As nitrogen chemistry and the relative importance of nitrogen compounds in the atmosphere change in response to proposed emission reductions (and possibly climate change), an improved understanding of the fate of airborne nitrogen compounds in forest biogeochemical cycles will be necessary.

  9. Plant and Soil Natural Abundance delta-15N: Indicators of Nitrogen Cycling in the Catskill Mountains, New York, USA

    NASA Astrophysics Data System (ADS)

    Templer, P. H.; Lovett, G. M.; Weathers, K.; Arthur, M. A.

    2002-12-01

    We examined the potential use of natural abundance 15N of plants and soils as an indicator of forest nitrogen (N) cycling rates within the Catskill Mountains, NY. These watersheds receive among the highest rates of N deposition in the northeastern United States and are beginning to show signs of N saturation. Many studies have shown a link between increased N cycling rates and 15N enrichment of soil and plant pools. Faster rates of N cycling processes, especially nitrification, lead to fractionation of 14/15N, creating N products that are relatively depleted in 15N. This can lead to enrichment of soil pools, as lighter 14N is lost from the system via leaching or denitrification. Plant N pools can become increasingly enriched as they take up 15N-enriched soil N. Despite similar amounts of N deposition across the Catskill Mountains, forests dominated by different tree species appear to vary in the amount of N retained or lost to nearby streams. To determine if plant and soil 15N could be used as indicators of N cycling rates, we collected foliage, wood, litterfall, organic and mineral soil, and fine roots from single species stands of American beech (Fagus grandifolia), eastern hemlock (Tsuga canadensis), red oak (Quercus rubra), and sugar maple (Acer saccharum). Fine roots and soil 15N were highest within sugar maple stands (p<0.05). Sugar maple soils also had the highest rates of net nitrification and N leaching. Therefore, soil 15N appears to correlate with forest N retention and loss. However, 15N enrichment was highest within foliage, litterfall and wood of beech trees (p<0.05). The decoupling between foliage 15N and N cycling, as well as between 15N of foliage and fine roots, illustrates that it may not be possible to use a single plant pool as an indicator of N cycling rates.

  10. Long-term 15N tracking from biological N fixation across different plant and humus components of the boreal forest

    NASA Astrophysics Data System (ADS)

    Arroniz-Crespo, Maria; Jones, David L.; Zackrisson, Olle; Nilsson, Marie-Charlotte; DeLuca, Thomas H.

    2014-05-01

    Biological N2 fixation by cyanobacteria associated with feather mosses is an important cog in the nitrogen (N) cycle of boreal forests; still, our understanding of the turnover and fate of N fixed by this association remains greatly incomplete. The 15N signature of plants and soil serves as a powerful tool to explore N dynamics in forest ecosystems. In particular, in the present study we aimed to investigate the contribution of N2 fixation to δ15N signatures of plants and humus component of the boreal forest. Here we present results from a long-term (7 years) tacking of labelled 15N2 across the humus layer, seedlings of the tree species Pinus sylvestris, two common dwarf shrub species (Empetrum hermaphroditum and Vaccinium vitis-idaea) and the feather moss Pleurozium schreibery. The enriched experiment was conducted in 2005 in a natural boreal forest in northern Sweden. Two different treatments (10% 15N2 headspace enrichment and control) were setup in nine different plots (0.5 x 0.5 m) within the forest. We observed a significant reduction of δ15N signature of the 15N-enriched moss that could be explained by a growth dilution effect. Nevertheless, after 5 years since 15N2 enrichment some of the label 15N was still detected on the moss and in particular in the dead tissue. We could not detect a clear transfer of the labelled 15N2 from the moss-cyanobacteria system to other components of the ecosystem. However, we found consistence relationship through time between increments of δ15N signature of some of the forest components in plots which exhibited higher N fixation rates in the moss. In particular, changes in natural abundance δ15N that could be associated with N fixation were more apparent in the humus layer, the dwarf shrub Vaccinium vitis-idaea and the pine seedlings when comparing across plots and years.

  11. Chiral doping effect in the B2 phase of a bent-core liquid crystal: The observation of resonant X-ray satellite peaks assigned to the 5/10 layer periodic structure

    NASA Astrophysics Data System (ADS)

    Takanishi, Yoichi; Ohtsuka, Youko; Takahashi, Yumiko; Kang, Sungmin; Iida, Atsuo

    2015-03-01

    We studied the details of a local layer structure in the B2 phase of bromo-containing bent-core liquid crystals mixed with a small amount of chiral molecules using microbeam resonant X-ray scattering. In this measurement, we detected the 1 ± 0.2 order satellite peaks, which suggest that the B2 phase of the mixture has a long-range periodic structure. Dielectric and electro-optic measurements indicate almost the same behavior as the antiferroelectric SmCAPA(B2) of pure bromo-containing bent-core liquid crystals, so that the B2 phase of the chiral mixture is also antiferroelectric, and it is concluded that the B2 phase of the chiral mixture forms a ten-layer periodic structure. Such a long-range periodic structure did not appear by mixing the racemate of the same compounds, which indicates the chiral effect on the long-range periodic structure.

  12. Structural Case Assignment in Korean

    ERIC Educational Resources Information Center

    Koak, Heeshin

    2012-01-01

    In this dissertation, I aim to provide a theory on the distribution of structural Case in Korean. I propose the following Structural Case Assignment Hypothesis (SCAH) regarding the assignment of structural Case: "Structural Case is assigned by phase heads (C: nominative; v: accusative) to every argument in the c-command domain of the phase…

  13. The signatures of stable isotopes δ 15N and δ 13C in anadromous and non-anadromous Coilia nasus living in the Yangtze River, and the adjacent sea waters

    NASA Astrophysics Data System (ADS)

    Wang, Lei; Tang, Wenqiao; Dong, Wenxia

    2015-12-01

    Stable isotopes are increasingly used to investigate seasonal migrations of aquatic organisms. This study employed stable isotopes ( δ 13C and δ 15N) for Coilia nasus from the lower Yangtze River and the adjacent East China Sea to distinguish different ecotypic groups, ascertain trophic nutrition positions, and reflect environmental influences on C. nasus. δ 13C signatures of C. nasus sampled from Zhoushan (ZS), Chongming (CM), and Jingjiang (JJ) waters were significantly higher than those from the Poyang Lake (PYL) ( P < 0.05). By contrast, δ 15N signatures of C. nasus in ZS, CM, and JJ groups were significantly lower than those in PYL group ( P < 0.05). Basing on δ 13C and δ 15N signatures, we could distinguish anadromous (ZS, CM, and JJ) and non-anadromous (PYL) groups. The trophic level (TL) of anadromous C. nasus ranged from 2.90 to 3.04, whereas that of non-anadromous C. nasus was 4.38. C. nasus occupied the middle and top nutrition positions in the marine and Poyang Lake food webs, respectively. C. nasus in Poyang Lake were significantly more enriched in δ 15N but depleted in δ 13C, suggesting that anthropogenic nutrient inputs and terrigenous organic carbon are important to the Poyang Lake food web. This study is the first to apply δ 15N and δ 13C to population assignment studies of C. nasus in the Yangtze River and its affiliated waters. Analysis of stable isotopes ( δ 15N and δ 13C) is shown to be a useful tool for discriminating anadromous and non-anadromous C. nasus.

  14. Carbon (δ13C) and Nitrogen (δ15N) Stable Isotope Signatures in Bat Fur Indicate Swarming Sites Have Catchment Areas for Bats from Different Summering Areas

    PubMed Central

    Segers, Jordi L.; Broders, Hugh G.

    2015-01-01

    Migratory patterns of bats are not well understood and traditional methods to study this, like capture-mark-recapture, may not provide enough detail unless there are many records. Stable isotope profiles of many animal species have been used to make inferences about migration. Each year Myotis lucifugus and M. septentrionalis migrate from summering roosts to swarming caves and mines in the fall, but the pattern of movement between them is not well understood. In this study, fur δ13C and δ15N values of 305 M. lucifugus and 200 M. septentrionalis were analyzed to make inferences about migration patterns between summering areas and swarming sites in Nova Scotia, Canada. We expected that there would be greater variability in δ13C and δ15N among individuals at swarming sites because it was believed that these sites are used by individuals originating from many summering areas. There was extensive overlap in the standard ellipse area, corrected for small sample sizes (SEAc), of bats at swarming sites and much less overlap in SEAc among groups sampled at summering areas. Meaningful inference could not be made on M. septentrionalis because their low variation in SEAc may have been the result of sampling only 3 summering areas. However, for M. lucifugus, swarming sites had larger SEAc than summering areas and predictive discriminant analysis assigned swarming bats to multiple summering areas, supporting the contention that swarming bats are mixed aggregations of bats from several summering areas. Together, these data support the contention that swarming sites have catchment areas for bats from multiple summering areas and it is likely that the catchment areas for swarming sites overlap. These data suggest that δ13C and δ15N profiling of bat fur offer some potential to make inferences about regional migration in bats. PMID:25923696

  15. Carbon (δ13C) and Nitrogen (δ15N) Stable Isotope Signatures in Bat Fur Indicate Swarming Sites Have Catchment Areas for Bats from Different Summering Areas.

    PubMed

    Segers, Jordi L; Broders, Hugh G

    2015-01-01

    Migratory patterns of bats are not well understood and traditional methods to study this, like capture-mark-recapture, may not provide enough detail unless there are many records. Stable isotope profiles of many animal species have been used to make inferences about migration. Each year Myotis lucifugus and M. septentrionalis migrate from summering roosts to swarming caves and mines in the fall, but the pattern of movement between them is not well understood. In this study, fur δ13C and δ15N values of 305 M. lucifugus and 200 M. septentrionalis were analyzed to make inferences about migration patterns between summering areas and swarming sites in Nova Scotia, Canada. We expected that there would be greater variability in δ13C and δ15N among individuals at swarming sites because it was believed that these sites are used by individuals originating from many summering areas. There was extensive overlap in the standard ellipse area, corrected for small sample sizes (SEAc), of bats at swarming sites and much less overlap in SEAc among groups sampled at summering areas. Meaningful inference could not be made on M. septentrionalis because their low variation in SEAc may have been the result of sampling only 3 summering areas. However, for M. lucifugus, swarming sites had larger SEAc than summering areas and predictive discriminant analysis assigned swarming bats to multiple summering areas, supporting the contention that swarming bats are mixed aggregations of bats from several summering areas. Together, these data support the contention that swarming sites have catchment areas for bats from multiple summering areas and it is likely that the catchment areas for swarming sites overlap. These data suggest that δ13C and δ15N profiling of bat fur offer some potential to make inferences about regional migration in bats.

  16. Macroalgae δ15N values in well-mixed estuaries: Indicator of anthropogenic nitrogen input or macroalgae metabolism?

    NASA Astrophysics Data System (ADS)

    Raimonet, Mélanie; Guillou, Gaël; Mornet, Françoise; Richard, Pierre

    2013-03-01

    Although nitrogen stable isotope ratio (δ15N) in macroalgae is widely used as a bioindicator of anthropogenic nitrogen inputs to the coastal zone, recent studies suggest the possible role of macroalgae metabolism in δ15N variability. Simultaneous determinations of δ15N of dissolved inorganic nitrogen (DIN) along the land-sea continuum, inter-species variability of δ15N and its sensitivity to environmental factors are necessary to confirm the efficiency of macroalgae δ15N in monitoring nitrogen origin in mixed-use watersheds. In this study, δ15N of annual and perennial macroalgae (Ulva sp., Enteromorpha sp., Fucus vesiculosus and Fucus serratus) are compared to δ15N-DIN along the Charente Estuary, after characterizing δ15N of the three main DIN sources (i.e. cultivated area, pasture, sewage treatment plant outlet). During late winter and spring, when human activities produce high DIN inputs, DIN sources exhibit distinct δ15N signals in nitrate (NO) and ammonium (NH): cultivated area (+6.5 ± 0.6‰ and +9.0 ± 11.0‰), pasture (+9.2 ± 1.8‰ and +12.4‰) and sewage treatment plant discharge (+16.9 ± 8.7‰ and +25.4 ± 5.9‰). While sources show distinct δN- in this multiple source catchment, the overall mixture of NO sources - generally >95% DIN - leads to low variations of δN-NO at the mouth of the estuary (+7.7 to +8.4‰). Even if estuarine δN-NO values are not significantly different from pristine continental and oceanic site (+7.3‰ and +7.4‰), macroalgae δ15N values are generally higher at the mouth of the estuary. This highlights high anthropogenic DIN inputs in the estuary, and enhanced contribution of 15N-depleted NH in oceanic waters. Although seasonal variations in δN-NO are low, the same temporal trends in macroalgae δ15N values at estuarine and oceanic sites, and inter-species differences in δ15N values, suggest that macroalgae δ15N values might be modified by the metabolic response of macroalgae to environmental parameters (e

  17. Sinks for nitrogen inputs in terrestrial ecosystems: a meta-analysis of 15N tracer field studies

    USGS Publications Warehouse

    Templer, P.H.; Mack, M.C.; Chapin, F. S.; Christenson, L.M.; Compton, J.E.; Crook, H.D.; Currie, W.S.; Curtis, C.J.; Dail, D.B.; D'Antonio, C. M.; Emmett, B.A.; Epstein, H.E.; Goodale, C.L.; Gundersen, P.; Hobbie, S.E.; Holland, K.; Hooper, D.U.; Hungate, B.A.; Lamontagne, S.; Nadelhoffer, K.J.; Osenberg, C.W.; Perakis, S.S.; Schleppi, P.; Schimel, J.; Schmidt, I.K.; Sommerkorn, M.; Spoelstra, J.; Tietema, A.; Wessel, W.W.; Zak, D.R.

    2012-01-01

    Effects of anthropogenic nitrogen (N) deposition and the ability of terrestrial ecosystems to store carbon (C) depend in part on the amount of N retained in the system and its partitioning among plant and soil pools. We conducted a meta-analysis of studies at 48 sites across four continents that used enriched 15N isotope tracers in order to synthesize information about total ecosystem N retention (i.e., total ecosystem 15N recovery in plant and soil pools) across natural systems and N partitioning among ecosystem pools. The greatest recoveries of ecosystem 15N tracer occurred in shrublands (mean, 89.5%) and wetlands (84.8%) followed by forests (74.9%) and grasslands (51.8%). In the short term (15N tracer application), total ecosystem 15N recovery was negatively correlated with fine-root and soil 15N natural abundance, and organic soil C and N concentration but was positively correlated with mean annual temperature and mineral soil C:N. In the longer term (3–18 months after 15N tracer application), total ecosystem 15N retention was negatively correlated with foliar natural-abundance 15N but was positively correlated with mineral soil C and N concentration and C: N, showing that plant and soil natural-abundance 15N and soil C:N are good indicators of total ecosystem N retention. Foliar N concentration was not significantly related to ecosystem 15N tracer recovery, suggesting that plant N status is not a good predictor of total ecosystem N retention. Because the largest ecosystem sinks for 15N tracer were below ground in forests, shrublands, and grasslands, we conclude that growth enhancement and potential for increased C storage in aboveground biomass from atmospheric N deposition is likely to be modest in these ecosystems. Total ecosystem 15N recovery decreased with N fertilization, with an apparent threshold fertilization rate of 46 kg N·ha-1·yr-1 above which most ecosystems showed net losses of applied 15N tracer in response to N fertilizer addition.

  18. Sinks for nitrogen inputs in terrestrial ecosystems: a meta-analysis of 15N tracer field studies.

    PubMed

    Templer, P H; Mack, M C; Chapin, F S; Christenson, L M; Compton, J E; Crook, H D; Currie, W S; Curtis, C J; Dail, D B; D'Antonio, C M; Emmett, B A; Epstein, H E; Goodale, C L; Gundersen, P; Hobbie, S E; Holland, K; Hooper, D U; Hungate, B A; Lamontagne, S; Nadelhoffer, K J; Osenberg, C W; Perakis, S S; Schleppi, P; Schimel, J; Schmidt, I K; Sommerkorn, M; Spoelstra, J; Tietema, A; Wessel, W W; Zak, D R

    2012-08-01

    Effects of anthropogenic nitrogen (N) deposition and the ability of terrestrial ecosystems to store carbon (C) depend in part on the amount of N retained in the system and its partitioning among plant and soil pools. We conducted a meta-analysis of studies at 48 sites across four continents that used enriched 15N isotope tracers in order to synthesize information about total ecosystem N retention (i.e., total ecosystem 15N recovery in plant and soil pools) across natural systems and N partitioning among ecosystem pools. The greatest recoveries of ecosystem 15N tracer occurred in shrublands (mean, 89.5%) and wetlands (84.8%) followed by forests (74.9%) and grasslands (51.8%). In the short term (< 1 week after 15N tracer application), total ecosystem 15N recovery was negatively correlated with fine-root and soil 15N natural abundance, and organic soil C and N concentration but was positively correlated with mean annual temperature and mineral soil C:N. In the longer term (3-18 months after 15N tracer application), total ecosystem 15N retention was negatively correlated with foliar natural-abundance 15N but was positively correlated with mineral soil C and N concentration and C:N, showing that plant and soil natural-abundance 15N and soil C:N are good indicators of total ecosystem N retention. Foliar N concentration was not significantly related to ecosystem 15N tracer recovery, suggesting that plant N status is not a good predictor of total ecosystem N retention. Because the largest ecosystem sinks for 15N tracer were below ground in forests, shrublands, and grasslands, we conclude that growth enhancement and potential for increased C storage in aboveground biomass from atmospheric N deposition is likely to be modest in these ecosystems. Total ecosystem 15N recovery decreased with N fertilization, with an apparent threshold fertilization rate of 46 kg N x ha(-1) x yr(-1) above which most ecosystems showed net losses of applied 15N tracer in response to N fertilizer

  19. Enrichment of natural (15)N abundance during soil N losses under 20years of continuous cereal cropping.

    PubMed

    Jones, Andrew R; Dalal, Ram C

    2017-01-01

    It is generally accepted that the enrichment of natural (15)N abundance in soil over time is reflective of historic N cycling and loss, but this process in cropping soils is not yet clear. In this study, we identified an enrichment gradient of natural (15)N abundance during 20-year chronosequence of cereal cropping on Alfisols in southwest Queensland, Australia, that have no history of fertilisation. We demonstrate that the increase in soil (15)N abundance is explained by isotopic fractionation of (15)N during organic N mineralisation and nitrification, which lead to isotopically heavier ammonium retained in the soil and isotopically lighter soil nitrate taken up and removed by seasonal crops during harvest. Here we present a framework for natural (15)N isotopic fractionation co-occurring with N losses during long-term cultivation.

  20. A 15N-poor isotopic composition for the solar system as shown by Genesis solar wind samples.

    PubMed

    Marty, B; Chaussidon, M; Wiens, R C; Jurewicz, A J G; Burnett, D S

    2011-06-24

    The Genesis mission sampled solar wind ions to document the elemental and isotopic compositions of the Sun and, by inference, of the protosolar nebula. Nitrogen was a key target element because the extent and origin of its isotopic variations in solar system materials remain unknown. Isotopic analysis of a Genesis Solar Wind Concentrator target material shows that implanted solar wind nitrogen has a (15)N/(14)N ratio of 2.18 ± 0.02 × 10(-3) (that is, ≈40% poorer in (15)N relative to terrestrial atmosphere). The (15)N/(14)N ratio of the protosolar nebula was 2.27 ± 0.03 × 10(-3), which is the lowest (15)N/(14)N ratio known for solar system objects. This result demonstrates the extreme nitrogen isotopic heterogeneity of the nascent solar system and accounts for the (15)N-depleted components observed in solar system reservoirs.

  1. Localization of 15N uptake in a Tibetan alpine Kobresia pasture

    NASA Astrophysics Data System (ADS)

    Schleuß, Per-Marten; Kuzyakov, Yakov

    2014-05-01

    The Kobresia Pygmea ecotone covers approximately 450.000 km2 and is of large global and regional importance due several socio-ecological aspects. For instance Kobresia pastures store high amounts of carbon, nitrogen and other nutrients, represent large grazing areas for herbivores, provide a fast regrowth after grazing events and protect against mechanical degradation and soil erosion. However, Kobresia pastures are assumed to be a grazing induced and are accompanied with distinct root mats varying in thickness between 5-30 cm. Yet, less is known about the morphology and the functions of this root mats, especially in the background of a progressing degradation due to changes of climate and management. Thus we aimed to identify the importance of single soil layers for plant nutrition. Accordingly, nitrogen uptake from different soil depths and its remain in above-ground biomass (AGB), belowground biomass (BGB) and soil were determined by using a 15N pulse labeling approach during the vegetation period in summer 2012. 15N urea was injected into six different soil depths (0.5 cm, 2.5 cm, 7.5 cm, 12.5 cm, 17.5 cm, 22.5 cm / for each 4 replicates) and plots were sampled 45 days after the labeling. For soil and BGB samples were taken in strict sample intervals of 0-1 cm, 1-5 cm, 5-10 cm, 10-15 cm, 15-20 cm, 20-25 cm. Results indicate that total recovery (including AGB, BGB and soil) was highest, if tracer was injected into the top 5 cm and subsequently decreased with decreasing injection depth. This is especially the case for the 15N recovery of BGB, which is clearly attributed to the root density and strongly decreased with soil depth. In contrast, the root activity derived from the 15N content of roots increased with soil depth, which is primary associated to a proportionate increase of living roots related to dead roots. However, most 15N was captured in plant biomass (67.5-85.3 % of total recovery), indicating high 15N uptake efficiency possibly due to N limitation

  2. Tracking the flow of bacterially derived 13C and 15N through soil faunal feeding channels.

    PubMed

    Crotty, F V; Blackshaw, R P; Murray, P J

    2011-06-15

    The soil food web has been referred to as a 'black box', a 'poor man's tropical rainforest' and an 'enigma', due to its opacity, diversity and the limited insight into feeding specificity. Here we investigate the flow of C and N through the soil food web as a way to gain understanding of the feeding interactions occurring. A bacterium, Pseudomonas lurida, was introduced to soil cores from two different habitats, a grassland and a woodland with the same soil type, enriched to 99 atom% in (13)C and (15)N, to trace the flow of bacterial C and N through the soil food web. Throughout the experiment the soil remained enriched in (13)C and (15)N. Almost all the invertebrates tested gained C and N enrichment indicative of the labelled bacteria, implying that bacterial feeding is a common mechanism within the soil. Only three groups were significantly enriched in both (13)C and (15)N in both habitats. These were Collembola (Entomobryomorpha), Acari (Oribatida), and Nematoda, indicating that these organisms are consuming the most bacteria within both systems. When the invertebrates were grouped into hypothesised trophic levels, those considered secondary decomposers were gaining the most enrichment across all invertebrates tested. This enrichment was also high in the micro-predators within the soil, implying that their main food source was the secondary decomposers, particularly the Collembola. Using an enriched bacterium to track the trophic transfer between organisms within the soil food web is a novel way of empirically showing that interactions are occurring, which normally cannot be seen.

  3. Provenancing Archaeological Wool Textiles from Medieval Northern Europe by Light Stable Isotope Analysis (δ13C, δ15N, δ2H)

    PubMed Central

    von Holstein, Isabella C. C.; Walton Rogers, Penelope; Craig, Oliver E.; Penkman, Kirsty E. H.; Newton, Jason; Collins, Matthew J.

    2016-01-01

    We investigate the origin of archaeological wool textiles preserved by anoxic waterlogging from seven medieval archaeological deposits in north-western Europe (c. 700–1600 AD), using geospatial patterning in carbon (δ13C), nitrogen (δ15N) and non-exchangeable hydrogen (δ2H) composition of modern and ancient sheep proteins. δ13C, δ15N and δ2H values from archaeological wool keratin (n = 83) and bone collagen (n = 59) from four sites were interpreted with reference to the composition of modern sheep wool from the same regions. The isotopic composition of wool and bone collagen samples clustered strongly by settlement; inter-regional relationships were largely parallel in modern and ancient samples, though landscape change was also significant. Degradation in archaeological wool samples, examined by elemental and amino acid composition, was greater in samples from Iceland (Reykholt) than in samples from north-east England (York, Newcastle) or northern Germany (Hessens). A nominal assignment approach was used to classify textiles into local/non-local at each site, based on maximal estimates of isotopic variability in modern sheep wool. Light element stable isotope analysis provided new insights into the origins of wool textiles, and demonstrates that isotopic provenancing of keratin preserved in anoxic waterlogged contexts is feasible. We also demonstrate the utility of δ2H analysis to understand the location of origin of archaeological protein samples. PMID:27764106

  4. Provenancing Archaeological Wool Textiles from Medieval Northern Europe by Light Stable Isotope Analysis (δ13C, δ15N, δ2H).

    PubMed

    von Holstein, Isabella C C; Walton Rogers, Penelope; Craig, Oliver E; Penkman, Kirsty E H; Newton, Jason; Collins, Matthew J

    2016-01-01

    We investigate the origin of archaeological wool textiles preserved by anoxic waterlogging from seven medieval archaeological deposits in north-western Europe (c. 700-1600 AD), using geospatial patterning in carbon (δ13C), nitrogen (δ15N) and non-exchangeable hydrogen (δ2H) composition of modern and ancient sheep proteins. δ13C, δ15N and δ2H values from archaeological wool keratin (n = 83) and bone collagen (n = 59) from four sites were interpreted with reference to the composition of modern sheep wool from the same regions. The isotopic composition of wool and bone collagen samples clustered strongly by settlement; inter-regional relationships were largely parallel in modern and ancient samples, though landscape change was also significant. Degradation in archaeological wool samples, examined by elemental and amino acid composition, was greater in samples from Iceland (Reykholt) than in samples from north-east England (York, Newcastle) or northern Germany (Hessens). A nominal assignment approach was used to classify textiles into local/non-local at each site, based on maximal estimates of isotopic variability in modern sheep wool. Light element stable isotope analysis provided new insights into the origins of wool textiles, and demonstrates that isotopic provenancing of keratin preserved in anoxic waterlogged contexts is feasible. We also demonstrate the utility of δ2H analysis to understand the location of origin of archaeological protein samples.

  5. Capture cross sections of 15N(n, γ)16N at astrophysical energies

    NASA Astrophysics Data System (ADS)

    Fan, Guang-Wei; Ma, Jun-Bing; Sheng, Zong-Qiang; Shi, Guo-Zhu; Tian, Feng; Wang, Jun; Zhang, Chao

    2016-12-01

    We have reanalyzed reaction cross sections of 16N on a 12C target. The nucleon density distribution of 16N, especially surface density distribution, was extracted using the modified Glauber model. On the basis of dilute surface densities, the 15N(n, γ)16N reaction is discussed within the framework of the direct capture reaction mechanism. The calculations agree quite well with the experimental data. Support given by National Natural Science Foundation of China (11447236, 11505002, 11247001) and Foundation of Anhui University of Science and Technology (11130, 12608)

  6. VizieR Online Data Catalog: 14N/15N isotopic ratio in L1544 (Bi

    NASA Astrophysics Data System (ADS)

    Bizzocchi, L.; Caselli, P.; Leonardo, E.; Dore, L.

    2013-06-01

    The observations towards L1544 were carried out with the IRAM 30m antenna, located at Pico Veleta (Spain) during observing sessions in June 2009 and July 2010. The J=1-0 transition of was observed with the EMIR receiver in the E090 configuration tuned at 90263.8360MHz and using the lower-inner sideband. The hyperfine-free rest frequencies were taken from the most recent laboratory investigation of 15N-dyazenilium species (Dore et al. 2009A&A...496..275D). (6 data files).

  7. Influenza A (H15N4) virus isolation in Western Siberia, Russia.

    PubMed

    Sivay, Mariya V; Baranovich, Tatiana; Marchenko, Vasiliy Y; Sharshov, Kirill A; Govorkova, Elena A; Shestopalov, Aleksander M; Webby, Richard J

    2013-03-01

    The rarely identified influenza A viruses of the H15 hemagglutinin subtype have been isolated exclusively in Australia. Here we report the isolation of an H15N4 influenza A virus (A/teal/Chany/7119/2008) in Western Siberia, Russia. Phylogenetic analysis demonstrated that the internal genes of the A/teal/Chany/7119/2008 strain belong to the Eurasian clade and that the H15 and N4 genes were introduced into the gene pool of circulating endemic avian influenza viruses through reassortment events.

  8. Optical Microscopy Characterization for Borehole U-15n#12 in Support of NCNS Source Physics Experiment

    SciTech Connect

    Wilson, Jennifer E.; Sussman, Aviva Joy

    2015-05-22

    Optical microscopy characterization of thin sections from corehole U-15n#12 is part of a larger material characterization effort for the Source Physics Experiment (SPE). The SPE program was conducted in Nevada with a series of explosive tests designed to study the generation and propagation of seismic waves inside Stock quartz monzonite. Optical microscopy analysis includes the following: 1) imaging of full thin sections (scans and mosaic maps); 2) high magnification imaging of petrographic texture (grain size, foliations, fractures, etc.); and 3) measurement of microfracture density.

  9. Uptake of stormwater nitrogen in bioretention systems demonstrated from 15N tracer techniques

    NASA Astrophysics Data System (ADS)

    Houdeshel, D.; Hultine, K. R.; Pomeroy, C. A.

    2012-12-01

    Bioretention stormwater management systems are engineered ecosystems that capture urban stormwater in order to reduce the harmful effects of stormwater pollution on receiving waters. Bioretention systems have been shown to be effective at reducing the volume of runoff, and thereby reduce the nutrient loading to receiving waters from urban areas. However, little work has been done to evaluate the treatment processes that are responsible for reductions in effluent nitrogen (N). We hypothesize that the pulses of inorganic nitrogen associated with urban runoff events are captured in the plat tissues within these systems and not adsorbed to the soil media, thus creating a long-term, sustainable treatment approach to reducing the total nutrient loading to receiving waters. Nitrogen treatment performance was tested on two bioretention systems in Salt Lake City, UT: 1) an upland native community that does not require irrigation in semi-arid climates, and 2) a wetland community that requires 250 l of daily irrigation to offset the relatively high evaporative demand in the region. Each cell is sized to treat a 2.5 cm storm from a 140 m2 impervious surface: the area of the bioretention system is 10 m2. To test the N removal performance of each system, runoff events were simulated to represent an average precipitation regime using a synthetic stormwater blend starting in January, 2012. Effluent was collected from an underdrain and analyzed for total nitrogen (TN); mass removal was calculated for each month by subtracting the TN mass added to the garden minus the TN mass that flowed out of the garden. To test the hypothesis that plants assimilate stormwater N, 4 g of 100 atom% 15N NH4NO3 tracer was used as the N source in the synthetic stormwater during the first 2,000 l synthetic storm event in May. This isotopic label was calculated to enrich the total N pool of each garden to 100‰ 15N/14Nair. New growth was harvested from each plant in both cells and analyzed for 15N

  10. Determination of the δ15N of nitrate in water; RSIL lab code 2899

    USGS Publications Warehouse

    Coplen, Tyler B.; Qi, Haiping; Revesz, Kinga; Casciotti, Karen; Hannon, Janet E.

    2007-01-01

    The purpose of the Reston Stable Isotope Laboratory (RSIL) lab code 2899 is to determine the δ15N of nitrate (NO3-) in water. The δ15N of the dissolved NO3- is analyzed by conversion of the NO3- to nitrous oxide (N2O), which serves as the analyte for mass spectrometry. A culture of denitrifying bacteria is used in the enzymatic conversion of the NO3- to N2O, which follows the pathway shown in equation 1: NO3- → NO2- → NO → 1/2 N2O (1) Because the bacteria Pseudomonas aureofaciens lack N2O reductive activity, the reaction stops at N2O, unlike the typical denitrification reaction that goes to N2. After several hours, the conversion is complete, and the N2O is extracted from the vial, separated from volatile organic vapor and water vapor by an automated -65 °C isopropanol-slush trap, a Nafion drier, a CO2 and water removal unit (Costech #021020 carbon dioxide absorbent with Mg(ClO4)2), and trapped in a small-volume trap immersed in liquid nitrogen with a modified Finnigan MAT (now Thermo Scientific) GasBench 2 introduction system. After the N2O is released, it is further purified by gas chromatography before introduction to the isotope-ratio mass spectrometer (IRMS). The IRMS is a Thermo Scientific Delta V Plus continuous flow IRMS (CF-IRMS). It has a universal triple collector, consisting of two wide cups with a narrow cup in the middle; it is capable of simultaneously measuring mass/charge (m/z) of the N2O molecule 44, 45, and 46. The ion beams from these m/z values are as follows: m/z = 44 = N2O = 14N14N16O; m/z = 45 = N2O = 14N15N16O or 14N14N17O; m/z = 46 = N2O = 14N14N18O. The 17O contributions to the m/z 44 and m/z 45 ion beams are accounted for before δ15N values are reported.

  11. Determination of the δ15N of nitrate in solids; RSIL lab code 2894

    USGS Publications Warehouse

    Coplen, Tyler B.; Qi, Haiping; Revesz, Kinga; Casciotti, Karen; Hannon, Janet E.

    2007-01-01

    The purpose of the Reston Stable Isotope Laboratory (RSIL) lab code 2894 is to determine the δ15N of nitrate (NO3-) in solids. The nitrate fraction of the nitrogen species is dissolved by water (called leaching) and can be analyzed by the bacterial method covered in RSIL lab code 2899. After leaching, the δ15N of the dissolved NO3- is analyzed by conversion of the NO3- to nitrous oxide (N2O), which serves as the analyte for mass spectrometry. A culture of denitrifying bacteria is used in the enzymatic conversion of NO3- to N2O, which follows the pathway shown in equation 1: NO3- → NO2- → NO → 1/2 N2O (1) Because the bacteria Pseudomonas aureofaciens lack N2O reductive activity, the reaction stops at N2O, unlike the typical denitrification reaction that goes to N2. After several hours, the conversion is complete, and the N2O is extracted from the vial, separated from volatile organic vapor and water vapor by an automated -65 °C isopropanol-slush trap, a Nafion drier, a CO2 and water removal unit (Costech #021020 carbon dioxide absorbent with Mg(ClO4)2), and trapped in a small-volume trap immersed in liquid nitrogen with a modified Finnigan MAT (now Thermo Scientific) GasBench 2 introduction system. After the N2O is released, it is further purified by gas chromatography before introduction to the isotope-ratio mass spectrometer (IRMS). The IRMS is a Thermo Scientific Delta V Plus continuous flow IRMS (CF-IRMS). It has a universal triple collector, consisting of two wide cups with a narrow cup in the middle; it is capable of simultaneously measuring mass/charge (m/z) of the N2O molecule 44, 45, and 46. The ion beams from these m/z values are as follows: m/z = 44 = N2O = 14N14N16O; m/z = 45 = N2O = 14N15N16O or 14N14N17O; m/z = 46 = N2O = 14N14N18O. The 17O contributions to the m/z 44 and m/z 45 ion beams are accounted for before δ15N values are reported.

  12. Rooting depth distribution and nitrogen acquisition using 15N tracer, Barrow, Alaska, 2013

    SciTech Connect

    Colleen Iversen

    2016-12-01

    Permafrost thaw and degradation may lead to altered thickness of the active soil layer and a changing distribution of plant-available nutrients throughout the soil, but little is known about the nutrient acquisition strategies of dominant tundra plant species. We conducted an 15N isotope tracer experiment to assess the vertical distribution of nutrient acquisition among three dominant species representing important plant functional types (PFTs) on the Barrow Environmental Observatory (BEO) in Barrow, Alaska. We found that vertical patterns of root distribution and nutrient acquisition varied among plant species, and that root density may not entirely explain patterns of nutrient acquisition for all species.

  13. Rapid, storm-induced changes in the natural abundance of sup 15 N in a planktonic ecosystem, Chesapeake Bay, USA

    SciTech Connect

    Montoya, J.P.; McCarthy, J.J. ); Horrigan, S.G. )

    1991-12-01

    Samples of dissolved inorganic nitrogen (DIN), particulate nitrogen (PN), and two species of zooplankton were collected during two north-south transects of the Chesapeake Bay in the autumn of 1984 (27-28 September and 3-5 October). During the first transect, the natural abundance of {sup 15}N ({delta} {sup 15}N) in the major dissolved and planktonic pools of nitrogen suggested that the {delta}{sup 15}N of PN was largely determined by isotopic fractionation during uptake of NH{sub 4}{sup +} by phytoplankton. Averaged over the transect as a whole, the {delta}{sup 15}N of the herbivorous calanoid copepod Acartia tonsa was 4.1% higher than that of the PN, while the {delta}{sup 15}N of the carnivorous ctenophore Mnemiopsis leidyi was 6.4% higher than that of the PN. In the interval between the two transects, storm-induced mixing of the water column resulted in the injection of NH{sub 4}{sup +} into the surface layer of the bay. In combination with ancillary physical, chemical, and biological data, these changes in {delta}{sup 15}N provided estimates of the isotopic fractionation factor for NH{sub 4}{sup +} uptake by phytoplankton ({alpha} = 1.0065-1.0080) as well as the turnover time of nitrogen in Acartia tonsa (6.0-9.6 days). Despite the changes in {delta}{sup 15}N observed during this cruise, the relative distribution of {sup 15}N between trophic levels was preserved: during the second transect, the difference in {delta}{sup 15}N between Acartia tonsa and PN was 3.6%, and the difference in {delta}{sup 15}N between Mnemiopsis leidyi and PN was 7.3%. These results demonstrate that the natural abundance of {sup 15}N can change dramatically on a time scale of days, and that time-series studies of the natural abundance of {sup 15}N can be a useful complement to studies using tracer additions of {sup 15}N to document nitrogen transformations in planktonic ecosystems.

  14. Structure and reactivity of lithium amides. /sup 6/Li, /sup 13/C, and /sup 15/N NMR spectroscopic studies and colligative measurements of lithium diphenylamide and lithium diphenylamide-lithium bromide complex solvated by tetrahydrofuran

    SciTech Connect

    DePue, J.S.; Collum, D.B.

    1988-08-03

    /sup 6/Li, /sup 13/C, and /sup 15/N NMR spectroscopic studies of lithium diphenylamide in THF/hydrocarbon solutions (THF = tetrahydrofuran) detected two different species. /sup 6/Li and /sup 15/N NMR spectroscopic studies of (/sup 6/Li, /sup 15/N)lithium diphenylamide showed the species observed at low THF concentrations to be a cyclic oligomer. Structural analogies provided strong support for a dimer while colligative measurements at 0/degrees/C indicated the dimer to be di- or trisolvated. On the basis of the observed mass action effects, the species appearing at intermediate THF concentrations is assigned as a contact or solvent-separated ion-paired monomer. Lithium diphenylamide forms a 1:1 adduct with lithium bromide at low THF concentrations. A combination of /sup 6/Li-/sup 15/N double labeling studies and colligative measurements supports a trisolvated cyclic mixed dimer structure. Although detailed spectroscopic studies at elevated THF concentrations were precluded by high fluctionality, the similarity of the /sup 13/C chemical shifts of lithium diphenylamide in the presence and absence of lithium bromide provide indirect evidence that the mixed dimer undergoes a THF concentration dependent dissociation to the monomeric amide and free lithium bromide. 24 references, 9 figures, 2 tables.

  15. Nitrogen removal in maturation ponds: tracer experiments with 15N-labelled ammonia.

    PubMed

    Camargo Valero, M A; Mara, D D

    2007-01-01

    A primary maturation pond (M1) was spiked with labelled ammonium chloride (15NH4Cl) to track ammonium transformations associated with algal uptake and subsequent sedimentation. Conventional sampling based on grab samples collected from M1 influent, water column and effluent, and processed for unfiltered and filtered TKN, ammonium, nitrite and nitrate, found low total nitrogen removal (8%) and high ammonium nitrogen removal (90%). Stable isotope analysis of 15N from suspended organic and ammonium nitrogen fractions in M1 effluent revealed that labelled ammonium was mainly found in the organic fraction (69% of the 15N recovered), rather than the inorganic fraction (5%). Algal uptake was the predominant pathway for ammonia removal, even though conditions were favourable for ammonia volatilization (8.9 < pH <10.1 units, 15.2 < temperature <18.8 degrees C). Total nitrogen was removed by ammonia volatilization at 15 g N/ha d (3%), organic nitrogen sedimentation at 105 g N/ha d (20%), and in-pond accumulation due to algal uptake at 377 g N/ha d (71%). Algal uptake of ammonium and subsequent sedimentation and retention in the benthic sludge, after partial ammonification of the algal organic nitrogen, is thus likely to be the dominant mechanism for permanent nitrogen removal in maturation ponds during warm summer months in England.

  16. The coral δ15N record of terrestrial nitrate loading varies with river catchment land use

    NASA Astrophysics Data System (ADS)

    Yamazaki, A.; Watanabe, T.; Tsunogai, U.; Hasegawa, H.; Yamano, H.

    2015-03-01

    We analysed the nitrogen isotopes in two coral cores (δ15Ncoral) from the mouth of the Todoroki River, Ishigaki Island, Japan, to examine whether the δ15Ncoral reflects the run-off of nitrate related to the land use in the river catchment. The two coral cores were used to examine the seasonal variation in δ15Ncoral for 14 years (CORE1; 1993-2007) and the annual variation of δ15Ncoral for 52 years (CORE2; 1958-2010). In CORE1, the 5-month running mean of δ15Ncoral was positively correlated with that of monthly precipitation, excluding all strong precipitation events (>150 mm d-1). In CORE2, the δ15Ncoral mean in the earlier period (1958-1980) was 1.0 ‰ greater than that in the later period (1981-2010). The annual averages of δ15Ncoral are positively correlated with the total precipitation in the rainy season (May-June) for both time periods. The difference in the δ15Ncoral between the earlier and later periods is probably caused by the land use changed from paddy fields with 15N-rich manure to sugar cane fields in the early 1980s. Although some uncertainties still remain regarding the precision of δ15N coral proxy records, this study emphasises the clear potential for their use in reconstructing terrestrial nitrate discharge records from corals.

  17. Covalent binding of aniline to humic substances. 2. 15N NMR studies of nucleophilic addition reactions

    USGS Publications Warehouse

    Thorn, K.A.; Pettigrew, P.J.; Goldenberg, W.S.; Weber, E.J.

    1996-01-01

    Aromatic amines are known to undergo covalent binding with humic substances in the environment. Although previous studies have examined reaction conditions and proposed mechanisms, there has been no direct spectroscopic evidence for the covalent binding of the amines to the functional groups in humic substances. In order to further elucidate the reaction mechanisms, the Suwannee River and IHSS soil fulvic and humic acids were reacted with 15N-labeled aniline at pH 6 and analyzed using 15N NMR spectrometry. Aniline underwent nucleophilic addition reactions with the quinone and other carbonyl groups in the samples and became incorporated in the form of anilinohydroquinone, anilinoquinone, anilide, imine, and heterocyclic nitrogen, the latter comprising 50% or more of the bound amine. The anilide and anilinohydroquinone nitrogens were determined to be susceptible to chemical exchange by ammonia. In the case of Suwannee River fulvic acid, reaction under anoxic conditions and pretreatment with sodium borohydride or hydroxylamine prior to reaction under oxic conditions resulted in a decrease in the proportion of anilinohydroquinone nitrogen incorporated. The relative decrease in the incorporation of anilinohydroquinone nitrogen with respect to anilinoquinone nitrogen under anoxic conditions suggested that inter- or intramolecular redox reactions accompanied the nucleophilic addition reactions.

  18. UV-visible and (1)H-(15)N NMR spectroscopic studies of colorimetric thiosemicarbazide anion sensors.

    PubMed

    Farrugia, Kristina N; Makuc, Damjan; Podborska, Agnieszka; Szaciłowski, Konrad; Plavec, Janez; Magri, David C

    2015-02-14

    Four model thiosemicarbazide anion chemosensors containing three N-H bonds, substituted with phenyl and/or 4-nitrophenyl units, were synthesised and studied for their anion binding abilities with hydroxide, fluoride, acetate, dihydrogen phosphate and chloride. The anion binding properties were studied in DMSO and 9 : 1 DMSO-H2O by UV-visible absorption and (1)H/(13)C/(15)N NMR spectroscopic techniques and corroborated with DFT studies. Significant changes were observed in the UV-visible absorption spectra with all anions, except for chloride, accompanied by dramatic colour changes visible to the naked eye. These changes were determined to be due to the deprotonation of the central N-H proton and not due to hydrogen bonding based on (1)H/(15)N NMR titration studies with acetate in DMSO-d6-0.5% water. Direct evidence for deprotonation was confirmed by the disappearance of the central thiourea proton and the formation of acetic acid. DFT and charge distribution calculations suggest that for all four compounds the central N-H proton is the most acidic. Hence, the anion chemosensors operate by a deprotonation mechanism of the central N-H proton rather than by hydrogen bonding as is often reported.

  19. Oxygen determination in materials by 18O(p,αγ)15N nuclear reaction

    NASA Astrophysics Data System (ADS)

    Kumar, Sanjiv; Sunitha, Y.; Reddy, G. L. N.; Sukumar, A. A.; Ramana, J. V.; Sarkar, A.; Verma, Rakesh

    2016-07-01

    The paper presents a proton induced γ-ray emission method based on 18O(p,αγ)15N nuclear reaction to determine bulk oxygen in materials. The determination involves the measurement of 5.27 MeV γ-rays emitted following the de-excitation of 15N nuclei. A description of the energetics of the reaction is given to provide an insight into the origin of 5.27 MeV γ-rays. In addition, thick target γ-ray yields and the limits of detection are measured to ascertain the analytical potential of the reaction. The thick-target γ-ray yields are measured with a high purity germanium detector and a bismuth germanate detector at 0° as well as 90° angles in 3.0-4.2 MeV proton energy region. The best limit of detection of about 1.3 at.% is achieved at 4.2 MeV proton energy for measurements at 0° as well 90° angles with the bismuth germanate detector while the uncertainty in quantitative analysis is <8%. The reaction has a probing depth of several tens of microns. Interferences can arise from fluorine due to the occurrence of 19F(p,αγ)16O reaction that emits 6-7 MeV γ-rays. The analytical potential of the methodology is demonstrated by determining oxygen in several oxide as well as non-oxide materials.

  20. Oligomeric complexes of some heteroaromatic ligands and aromatic diamines with rhodium and molybdenum tetracarboxylates: 13C and 15N CPMAS NMR and density functional theory studies.

    PubMed

    Leniak, Arkadiusz; Kamieński, Bohdan; Jaźwiński, Jarosław

    2015-05-01

    Seven new oligomeric complexes of 4,4'-bipyridine; 3,3'-bipyridine; benzene-1,4-diamine; benzene-1,3-diamine; benzene-1,2-diamine; and benzidine with rhodium tetraacetate, as well as 4,4'-bipyridine with molybdenum tetraacetate, have been obtained and investigated by elemental analysis and solid-state nuclear magnetic resonance spectroscopy, (13)C and (15)N CPMAS NMR. The known complexes of pyrazine with rhodium tetrabenzoate, benzoquinone with rhodium tetrapivalate, 4,4'-bipyridine with molybdenum tetrakistrifluoroacetate and the 1 : 1 complex of 2,2'-bipyridine with rhodium tetraacetate exhibiting axial-equatorial ligation mode have been obtained as well for comparison purposes. Elemental analysis revealed 1 : 1 complex stoichiometry of all complexes. The (15)N CPMAS NMR spectra of all new complexes consist of one narrow signal, indicating regular uniform structures. Benzidine forms a heterogeneous material, probably containing linear oligomers and products of further reactions. The complexes were characterized by the parameter complexation shift Δδ (Δδ = δcomplex  - δligand). This parameter ranged from around -40 to -90 ppm in the case of heteroaromatic ligands, from around -12 to -22 ppm for diamines and from -16 to -31 ppm for the complexes of molybdenum tetracarboxylates with 4,4'-bipyridine. The experimental results have been supported by a density functional theory computation of (15)N NMR chemical shifts and complexation shifts at the non-relativistic Becke, three-parameter, Perdew-Wang 91/[6-311++G(2d,p), Stuttgart] and GGA-PBE/QZ4P levels of theory and at the relativistic scalar and spin-orbit zeroth order regular approximation/GGA-PBE/QZ4P level of theory. Nucleus-independent chemical shifts have been calculated for the selected compounds.

  1. Alkaline Hydrolysis/Polymerization of 2,4,6-Trinitrotoluene:  Characterization of Products by 13C and 15N NMR

    USGS Publications Warehouse

    Thorn, Kevin A.; Thorne, Philip G.; Cox, Larry G.

    2004-01-01

    Alkaline hydrolysis has been investigated as a nonbiological procedure for the destruction of 2,4,6-trinitrotoluene (TNT) in explosives contaminated soils and munitions scrap. Nucleophilic substitutions of the nitro and methyl groups of TNT by hydroxide ion are the initial steps in the alkaline degradation of TNT. Potential applications of the technique include both in situ surface liming and ex situ alkaline treatment of contaminated soils. A number of laboratory studies have reported the formation of an uncharacterized polymeric material upon prolonged treatment of TNT in base. As part of an overall assessment of alkaline hydrolysis as a remediation technique, and to gain a better understanding of the chemical reactions underlying the hydrolysis/polymerization process, the soluble and precipitate fractions of polymeric material produced from the calcium hydroxide hydrolysis of unlabeled and 15N-labeled TNT were analyzed by elemental analysis and 13C and 15N nuclear magnetic resonance spectroscopy. Spectra indicated that reactions leading to polymerization included nucleophilic displacement of nitro groups by hydroxide ion, formation of ketone, carboxyl, alcohol, ether, and other aliphatic carbons, conversion of methyl groups to diphenyl methylene carbons, and recondensation of aromatic amines and reduced forms of nitrite, including ammonia and possibly hydroxylamine, into the polymer. Compared to the distribution of carbons in TNT as 14% sp3- and 86% sp2-hybridized, the precipitate fraction from hydrolysis of unlabeled TNT contained 33% sp3- and 67% sp2-hybridized carbons. The concentration of nitrogen in the precipitate was 64% of that in TNT. The 15N NMR spectra showed that, in addition to residual nitro groups, forms of nitrogen present in the filtrate and precipitate fractions include aminohydroquinone, primary amide, indole, imine, and azoxy, among others. Unreacted nitrite was recovered in the filtrate fraction. The toxicities and susceptibilities to

  2. Isotopic analysis of bulk, LMW, and HMW DON d15N indicates recycled nitrogen release from marine DON

    NASA Astrophysics Data System (ADS)

    Knapp, A. N.; Sigman, D. M.; Lipschultz, F.; Kustka, A.; Capone, D. G.

    2010-12-01

    Nitrogen (N) concentration and stable isotope ratio (d15N) measurements were made on bulk and size fractionated surface ocean dissolved organic nitrogen (DON) samples collected in the oligotrophic North Atlantic and Pacific Oceans. The bulk DON concentration in the upper 100 m is similar between the North Atlantic and North Pacific, between 4.5 and 5.0 uM, but the average d15N of bulk DON is significantly different, 3.9 per mil vs. air in the North Atlantic and 4.7 per mil in the North Pacific. The d15N of both bulk and HMW DON from the western tropical North Atlantic are similar to previous measurements, ~4.0 to 4.5 per mil. We report the first measurements of LMW DON d15N, which is consistently lower than HMW DON d15N. Neither the concentration nor d15N of bulk or size-fractionated DON varied with in situ N2 fixation rate, although significant variation in bulk and LMW DON d15N was observed between January and July of the same year in the western tropical North Atlantic. We propose a conceptual model to explain 1) the elevated d15N of bulk DON relative to other surface ocean N pools and fluxes, 2) the elevation of HMW DON d15N relative to LMW DON d15N, and 3) the inter-basin difference in the d15N of bulk DON. In this model, DON is produced from suspended particulate organic nitrogen (PON) without isotope fractionation because the conversion from PON to DON largely does not involve N-bearing bonds. In contrast, deamination and amide hydrolysis, with N isotope effects of 3 to 10 per mil, are major mechanisms by which DON is converted to ammonia and/or to other simple N compounds (e.g., amino acids). Thus these N-specific DON loss reactions result in an elevated d15N of residual DON relative to the parent DON and therefore also to the PON source. Moreover, the ammonium and simple organic N compounds released by microbial DON degradation are efficiently reassimilated back into the PON pool, as an integral part of the regenerated N cycle that further lowers the d15N

  3. Heterotrophic 15N2 Fixation and Distribution of Newly Fixed Nitrogen in a Rice-Flooded Soil System 1

    PubMed Central

    Eskew, David L.; Eaglesham, Allan R. J.; App, A. A.

    1981-01-01

    Rice (Oryza sativa L.) plants growing in pots of flooded soil were exposed to a 15N2-enriched atmosphere for 3 to 13 days in a gas-tight chamber. The floodwater and soil surface were shaded with a black cloth to reduce the activity of phototrophic N2-fixing micro-organisms. The highest 15N enrichments were consistently observed in the roots, although the total quantity of 15N incorporated into the soil was much greater. The rate of 15N incorporation into roots was much higher at the heading than at the tillering stage of growth. Definite enrichments were also found in the basal node and in the lower outer leaf sheath fractions after 3 days of exposure at the heading stage. Thirteen days was the shortest time period in which definite 15N enrichment was observed in the leaves and panicle. When plants were exposed to 15N2 for 13 days just before heading and then allowed to mature in a normal atmosphere, 11.3% of the total 15N in the system was found in the panicles, 2.3% in the roots, and 80.7% in the subsurface soil. These results provide direct evidence of heterotrophic N2 fixation associated with rice roots and the flooded soil and demonstrate that part of the newly fixed N is available to the plant. PMID:16661887

  4. δ15N-Size Relationships in River Invertebrate Communities: An Integrated Measure of Food web Structure?

    NASA Astrophysics Data System (ADS)

    Anderson, C.; Cabana, G.

    2005-05-01

    Anthropogenic perturbations can alter the composition and structure of benthic communities in rivers. Here, we examine the use of estimates of fish trophic position measured with stable nitrogen isotope ratios (δ15N), and δ15N-size relationships for the invertebrate community, as indicators of food web structure in rivers affected by various anthropogenic disturbances. Results revealed that δ15N-size relationships of invertebrate communities showed a variety of responses, some sites (50%) showing significant increases in δ15N with invertebrate size, whereas others did not. Similarly, the number of trophic levels between invertebrate-eating fish and primary consumers varied greatly among study sites, ranging from 0.6 to 2.2 levels. To investigate whether these estimates of fish trophic position integrated changes in invertebrate community structure, we compared slopes of invertebrate δ15N versus size with slopes of baseline-corrected fish δ15N versus size. Both slopes tended to be correlated, although the relationship was not significant (r2 = 0.26, p = 0.07). This trend suggests, however, that invertebrate-feeding fish trophic position measured with δ15N might reflect overall community structure, and could possibly be used as an easily measured indicator of food chain collapse following anthropogenic perturbations.

  5. Cereal grain, rachis and pulse seed amino acid δ15N values as indicators of plant nitrogen metabolism.

    PubMed

    Styring, Amy K; Fraser, Rebecca A; Bogaard, Amy; Evershed, Richard P

    2014-01-01

    Natural abundance δ(15)N values of plant tissue amino acids (AAs) reflect the cycling of N into and within plants, providing an opportunity to better understand environmental and anthropogenic effects on plant metabolism. In this study, the AA δ(15)N values of barley (Hordeum vulgare) and bread wheat (Triticum aestivum) grains and rachis and broad bean (Vicia faba) and pea (Pisum sativum) seeds, grown at the experimental farm stations of Rothamsted, UK and Bad Lauchstädt, Germany, were determined by GC-C-IRMS. It was found that the δ(15)N values of cereal grain and rachis AAs could be largely attributed to metabolic pathways involved in their biosynthesis and catabolism. The relative (15)N-enrichment of phenylalanine can be attributed to its involvement in the phenylpropanoid pathway and glutamate has a δ(15)N value which is an average of the other AAs due to its central role in AA-N cycling. The relative AA δ(15)N values of broad bean and pea seeds were very different from one another, providing evidence for differences in the metabolic routing of AAs to the developing seeds in these leguminous plants. This study has shown that AA δ(15)N values relate to known AA biosynthetic pathways in plants and thus have the potential to aid understanding of how various external factors, such as source of assimilated N, influence metabolic cycling of N within plants.

  6. Limitations in detection of 15N incorporation by mass spectrometry in protein-based stable isotope probing (protein-SIP).

    PubMed

    Taubert, Martin; von Bergen, Martin; Seifert, Jana

    2013-05-01

    The method of protein-based stable isotope probing (protein-SIP) has previously been shown to allow the modeling of carbon fluxes in microbial communities, thus tackling one of the key questions in microbial ecology. The method allows the analysis of stable isotope distribution in peptides, revealing metabolic activities of the species present in an ecosystem. Besides carbon, an application of protein-SIP with nitrogen is of interest for resolving the nitrogen fluxes in microbial communities. Thus, the sensitivity and reliability of a protein-SIP approach employing (15)N was analyzed. For this, cultivations of Pseudomonas fluorescens ATCC 17483 with different ratios of (14)N/(15)N were performed, from 10 % down to 0.1 % (15)N. After incubation leading to complete labeling of biomass, proteins were extracted and separated by one-dimensional gel electrophoresis, followed by tryptic digest and UPLC Orbitrap MS/MS analysis. (15)N relative isotope abundance (RIA) was calculated based on isotopic patterns from identified peptides in mass spectra. Proteomics data have been deposited to ProteomeXchange with identifier PXD000127. The distribution of (15)N RIA values among peptides was analyzed in samples with different (15)N amount, and potential causes for variations within individual samples of either technical or biological origin were investigated. Using a number of 50 peptides, significant differences (p ≤ 0.05) in (15)N incorporation were found between samples of different (15)N RIA down to 0.1 %. The study demonstrates that protein-SIP using (15)N is sufficiently sensitive for quantitative investigation of microbial activity in nitrogen cycling processes.

  7. Foliar δ15N is affected by foliar nitrogen uptake, soil nitrogen, and mycorrhizae along a nitrogen deposition gradient.

    PubMed

    Vallano, Dena M; Sparks, Jed P

    2013-05-01

    Foliar nitrogen isotope (δ(15)N) composition patterns have been linked to soil N, mycorrhizal fractionation, and within-plant fractionations. However, few studies have examined the potential importance of the direct foliar uptake of gaseous reactive N on foliar δ(15)N. Using an experimental set-up in which the rate of mycorrhizal infection was reduced using a fungicide, we examined the influence of mycorrhizae on foliar δ(15)N in potted red maple (Acer rubrum) seedlings along a regional N deposition gradient in New York State. Mycorrhizal associations altered foliar δ(15)N values in red maple seedlings from 0.06 to 0.74 ‰ across sites. At the same sites, we explored the predictive roles of direct foliar N uptake, soil δ(15)N, and mycorrhizae on foliar δ(15)N in adult stands of A. rubrum, American beech (Fagus grandifolia), black birch (Betula lenta), and red oak (Quercus rubra). Multiple regression analysis indicated that ambient atmospheric nitrogen dioxide (NO2) concentration explained 0, 69, 23, and 45 % of the variation in foliar δ(15)N in American beech, red maple, red oak, and black birch, respectively, after accounting for the influence of soil δ(15)N. There was no correlation between foliar δ(13)C and foliar %N with increasing atmospheric NO2 concentration in most species. Our findings suggest that total canopy uptake, and likely direct foliar N uptake, of pollution-derived atmospheric N deposition may significantly impact foliar δ(15)N in several dominant species occurring in temperate forest ecosystems.

  8. Plant community change mediates the response of foliar δ(15)N to CO 2 enrichment in mesic grasslands.

    PubMed

    Polley, H Wayne; Derner, Justin D; Jackson, Robert B; Gill, Richard A; Procter, Andrew C; Fay, Philip A

    2015-06-01

    Rising atmospheric CO2 concentration may change the isotopic signature of plant N by altering plant and microbial processes involved in the N cycle. CO2 may increase leaf δ(15)N by increasing plant community productivity, C input to soil, and, ultimately, microbial mineralization of old, (15)N-enriched organic matter. We predicted that CO2 would increase aboveground productivity (ANPP; g biomass m(-2)) and foliar δ(15)N values of two grassland communities in Texas, USA: (1) a pasture dominated by a C4 exotic grass, and (2) assemblages of tallgrass prairie species, the latter grown on clay, sandy loam, and silty clay soils. Grasslands were exposed in separate experiments to a pre-industrial to elevated CO2 gradient for 4 years. CO2 stimulated ANPP of pasture and of prairie assemblages on each of the three soils, but increased leaf δ(15)N only for prairie plants on a silty clay. δ(15)N increased linearly as mineral-associated soil C declined on the silty clay. Mineral-associated C declined as ANPP increased. Structural equation modeling indicted that CO2 increased ANPP partly by favoring a tallgrass (Sorghastrum nutans) over a mid-grass species (Bouteloua curtipendula). CO2 may have increased foliar δ(15)N on the silty clay by reducing fractionation during N uptake and assimilation. However, we interpret the soil-specific, δ(15)N-CO2 response as resulting from increased ANPP that stimulated mineralization from recalcitrant organic matter. By contrast, CO2 favored a forb species (Solanum dimidiatum) with higher δ(15)N than the dominant grass (Bothriochloa ischaemum) in pasture. CO2 enrichment changed grassland δ(15)N by shifting species relative abundances.

  9. Discrimination against 15N among recombinant inbred lines of Phaseolus vulgaris L. contrasting in phosphorus use efficiency for nitrogen fixation.

    PubMed

    Lazali, Mohamed; Bargaz, Adnane; Carlsson, Georg; Ounane, Sidi Mohamed; Drevon, Jean Jacques

    2014-02-15

    Although isotopic discrimination processes during nitrogen (N) transformations influence the outcome of (15)N based quantification of N2 fixation in legumes, little attention has been given to the effects of genotypic variability and environmental constraints such as phosphorus (P) deficiency, on discrimination against (15)N during N2 fixation. In this study, six Phaseolus vulgaris recombinant inbred lines (RILs), i.e. RILs 115, 104, 34 (P deficiency tolerant) and 147, 83, 70 (P deficiency sensitive), were inoculated with Rhizobium tropici CIAT899, and hydroaeroponically grown with P-sufficient (250 μmol P plant(-1) week(-1)) versus P-deficient (75 μmol P plant(-1) week(-1)) supply. Two harvests were done at 15 (before nodule functioning) and 42 (flowering stage) days after transplanting. Nodulation, plant biomass, P and N contents, and the ratios of (15)N over total N content ((15)N/Nt) for shoots, roots and nodules were determined. The results showed lower (15)N/Nt in shoots than in roots, both being much lower than in nodules. P deficiency caused a larger decrease in (15)N/Nt in shoots (-0.18%) than in nodules (-0.11%) for all of the genotypes, and the decrease in shoots was greatest for RILs 34 (-0.33%) and 104 (-0.25%). Nodule (15)N/Nt was significantly related to both the quantity of N2 fixed (R(2)=0.96***) and the P content of nodules (R(2)=0.66*). We conclude that the discrimination against (15)N in the legume N2-fixing symbiosis of common bean with R. tropici CIAT899 is affected by P nutrition and plant genotype, and that the (15)N/Nt in nodules may be used to screen for genotypic variation in P use efficiency for N2 fixation.

  10. Diffusion technique for 15N and inorganic N analysis of low-N aqueous solutions and Kjeldahl digests.

    PubMed

    Chen, Rui Rui; Dittert, Klaus

    2008-06-01

    Diffusion of ammonia is a common sample preparation method for the stable isotope analysis of inorganic nitrogen in aqueous solution. Classical diffusion methods usually require 6-12 days of diffusion and often focus on (15)N/(14)N analysis only. More recent studies have discussed whether complete N recovery was necessary for the precise analysis of stable N isotope ratios. In this paper we present a newly revised diffusion technique that allows correct and simultaneous determination of total N and (15)N at% from aqueous solutions and Kjeldahl digests, with N concentrations down to sub-0.5-mg N L(-1) levels, and it is tested under different conditions of (15)N isotope labelling. With the modification described, the diffusion time was reduced to 72 h, while the ratios of measured and expected (15)N at% were greater than 99% and the simultaneous recovery of total N was >95%. Analysis of soil microbial biomass N and its (15)N/(14)N ratio is one of the most important applications of this diffusion technique. An experiment with soil extracts spiked with (15)N-labelled yeast showed that predigestion was necessary to prevent serious N loss during Kjeldahl digestion of aqueous samples (i.e. soil extracts). The whole method of soil microbial biomass N preparation for (15)N/(14)N analysis included chloroform fumigation, predigestion, Kjeldahl digestion and diffusion. An experiment with soil spiked with (15)N-labelled yeast was carried out to evaluate the method. Results showed a highly significant correlation of recovered and added N, with the same recovery rate (0.21) of both total N and (15)N. A k(N) value of 0.25 was obtained based on the data. In conclusion, the diffusion method works for soil extracts and microbial biomass N determination and hence could be useful in many types of soil/water studies.

  11. Optimizing Marine Security Guard Assignments

    DTIC Science & Technology

    2011-06-01

    Bangkok , Thailand East Asia and Pacific 18 4 Fort Lauderdale, Florida Western Hemisphere - South 13 5 Frankfurt, Germany Western Europe and Scandinavia 15...2008). Each 7 stationing plan satisfies a myriad of unit requirements, such as building and land availability. Similarly, each assignment solution...optimize the assignment of enlisted Marines to billets. EAM-GLOBAL seeks to assign the best Marine-billet fit while balancing staffing shortages

  12. Mammalian DNA δ15N exhibits 40‰ intramolecular variation and is unresponsive to dietary protein level

    PubMed Central

    Strable, Maggie S.; Tschanz, Carolyn L.; Varamini, Behzad; Chikaraishi, Yoshito; Ohkouchi, Naohiko; Brenna, J. Thomas

    2014-01-01

    We report the first high precision characterization of molecular and intramolecular δ15N of nucleosides derived from mammalian DNA. The influence of dietary protein level on brain amino acids and deoxyribonucleosides was determined to investigate whether high protein turnover would alter amino acid 15N or 13C. Pregnant guinea pig dams were fed control diets, or high or low levels of dietary protein throughout gestation, and all pups were fed control diets. Cerebellar DNA of offspring was extracted at 2 and 120 days of life, nucleosides isolated and δ15N and δ13C characterized. Mean diet δ15N = 0.45±0.33‰, compared to cerebellar whole tissue and DNA δ15N = +4.1±0.7‰ and −4.5±0.4‰, respectively. Cerebellar deoxythymidine (dT), deoxycytidine (dC), deoxyadenosine (dA), and deoxyguanosine (dG) δ15N were +1.4±0.4, −2.1±0.9, −7.2±0.3, and −10.4±0.5‰, respectively. There were no changes in amino acid or deoxyribonucleoside δ15N due to dietary protein level. Using known metabolic relationships, we developed equations to calculate the intramolecular δ15N originating from aspartate (asp) in purines (pur) or pyrimidines (pyr), glutamine (glu), and glycine (gly) to be δ15NASP-PUR, δ15NASP-PYR, δ15NGLN, and δ15NGLY +11.9±2.3‰, +7.0±2.0‰, −9.1±2.4‰, and −31.8±8.9‰, respectively. A subset of twelve amino acids from food and brain had mean δ15N of 4.3±3.2‰ and 13.8±3.1‰, respectively, and δ15N for gly and asp were 12.6±2.2‰ and 15.2±0.8‰, respectively. A separate isotope tracer study detected no significant turnover of cerebellar DNA in the first six months of life. The large negative δ15N difference between gly and cerebellar purine N at the gly (7) position implies either that there is a major isotope effect during DNA synthesis, or that in utero gly has a different isotope ratio during rapid growth and metabolism than in adult life. Our data show that cerebellar nucleoside intramolecular δ15N vary over more than

  13. Simulating the density of HC15N in the Titan atmosphere with a coupled ion-neutral photochemical model

    NASA Astrophysics Data System (ADS)

    Vuitton, V.; Yelle, R. V.; Klippenstein, S. J.; Lavvas, P.; Hörst, S. M.

    2015-10-01

    The 14 N/ 15 N ratio for HCN in the atmosphere of Titan has been measured to be 2 to 3 times as less as the corresponding ratio for N2. Using a coupled ionneutral photochemical model incorporating state-of-the-art chemistry and cross-sections for N2, we show that the difference in the ratio of 14 N/15 N between HCN and N2 can be explained exclusively by the photo-induced isotopic fractionation of 14 N14 N and 14 N 15 N,without any further putative nitrogen input.

  14. Novel labeling technique illustrates transfer of 15N2 from Sphagnum moss to vascular plants via diazotrophic nitrogen fixation

    NASA Astrophysics Data System (ADS)

    Thorp, N. R.; Vile, M. A.; Wieder, R.

    2013-12-01

    We used 15N2 gas to trace nitrogen (N) from biological N2-fixation to vascular plant uptake in an Alberta bog in order to determine if neighboring bog plants acquire recently fixed N from diazotrophs associating with Sphagnum mosses. Recent evidence indicates high rates of N2-fixation in Sphagnum mosses of Alberta bogs (Vile et al. 2013). Our previous work has shown that mosses can assimilate fixed N from associated diazotrophs as evidenced by the high N content of mosses despite minimal inputs from atmospheric deposition, retranslocation, and N mineralization. Therefore, the potential exists for vascular plants to obtain N from ';leaky' tissues of live mosses, however, this phenomenon has not been tested previously. Here we document the potential for relatively rapid transfer to vascular plants of N fixed by Sphagnum moss-associated diazotrophs. We utilized the novel approach of incubating mosses in 15N2 to allow the process of diazotrophic N2-fixation to mechanistically provide the 15N label, which is subsequently transferred to Sphagnum mosses. The potential for vascular bog natives to tap this N was assessed by planting the vascular plants in the labeled moss. Sphagnum mosses (upper 3 cm of live plants) were incubated in the presence of 98 atom % 15N2 gas for 48 hours. Two vascular plants common to Alberta bogs; Picea mariana and Vaccinium oxycoccus were then placed in the labeled mosses, where the mosses served as the substrate. Tissue samples from these plants were collected at three time points during the incubation; prior to 15N2 exposure (to determine natural abundance 15N), and at one and two months after 15N2 exposure. Roots and leaves were separated and run separately on a mass spectrometer to determine 15N concentrations. Sphagnum moss capitula obtained N from N2-fixation (δ15N of -2.43 × 0.40, 122.76 × 23.78, 224.92 × 68.37, 143.74 × 54.38 prior to, immediately after, and at 1 and 2 months after exposure to 15N2, respectively). Nitrogen was

  15. Paleoenvironmental implications of taxonomic variation among δ15N values of chloropigments

    USGS Publications Warehouse

    Higgins, Meytal B.; Wolfe-Simon, Felisa; Robinson, Rebecca S.; Qin, Yelun; Saito, Mark A.; Pearson, Ann

    2011-01-01

    Natural variations in the ratios of nitrogen isotopes in biomass reflect variations in nutrient sources utilized for growth. In order to use δ15N values of chloropigments of photosynthetic organisms to determine the corresponding δ15N values of biomass – and by extension, surface waters – the isotopic offset between chlorophyll and biomass must be constrained. Here we examine this offset in various geologically-relevant taxa, grown using nutrient sources that may approximate ocean conditions at different times in Earth’s history. Phytoplankton in this study include cyanobacteria (diazotrophic and non-diazotrophic), eukaryotic algae (red and green), and anoxygenic photosynthetic bacteria (Proteobacteria), as well as environmental samples from sulfidic lake water. Cultures were grown using N2, NO3−, and NH4+ as nitrogen sources, and were examined under different light regimes and growth conditions. We find surprisingly high variability in the isotopic difference (δ15Nbiomass − δ15Nchloropigment) for prokaryotes, with average values for species ranging from −12.2‰ to +11.7‰. We define this difference as εpor, a term that encompasses diagenetic porphyrins and chlorins, as well as chlorophyll. Negative values of εpor reflect chloropigments that are 15N-enriched relative to biomass. Notably, this enrichment appears to occur only in cyanobacteria. The average value of εpor for freshwater cyanobacterial species is −9.8 ± 1.8‰, while for marine cyanobacteria it is −0.9 ± 1.3‰. These isotopic effects group environmentally but not phylogenetically, e.g., εpor values for freshwater Chroococcales resemble those of freshwater Nostocales but differ from those of marine Chroococcales. Our measured values of εpor for eukaryotic algae (range = 4.7–8.7‰) are similar to previous reports for pure cultures. For all taxa studied, values of εpor do not depend on the type of nitrogen substrate used for growth. The observed

  16. Ureide Assay for Measuring Nitrogen Fixation by Nodulated Soybean Calibrated by 15N Methods 1

    PubMed Central

    Herridge, David F.; Peoples, Mark B.

    1990-01-01

    We report experiments to quantify the relationships between the relative abundance of ureide-N in root-bleeding sap, vacuum-extracted sap, and hot water extracts of stems and petioles of nodulated soybean (Glycine max [L.] Merrill cv Bragg) and the proportion of plant N derived from nitrogen fixation. Additional experiments examined the effects of plant genotype and strain of rhizobia on these relationships. In each of the five experiments reported, plants of cv Bragg (experiment 1), cv Lincoln (experiments 3, 4, 5), or six cultivars/genotypes (experiment 2) were grown in a sand:vermiculite mixture in large pots in a naturally lit, temperature-controlled glasshouse during summer. Pots were inoculated at sowing with effective Bradyrhizobium japonicum CB1809 (USDA 136) or with one of 21 different strains of rhizobia. The proportions of plant N derived from nitrogen fixation were determined using 15N dilution. In one experiment with CB1809, plants were supplied throughout growth with either N-free nutrients or with nutrients supplemented with 1, 2, 4, or 8 millimolar 15N-nitrate and harvested on eight occasions between V6 and R7 for root-bleeding sap, vacuum-extracted sap, stems (including petioles), and whole plant dry matter. Analyses of the saps and stem extracts for ureides (allantoin plus allantoic acid), α-amino-N, and nitrate, and of dry matter for N and 15N, indicated a positive effect of nitrate supply on concentrations of nitrate in saps and extracts and a negative effect on ureides and on the proportion of plant N derived from nitrogen fixation. The relative abundance of ureide-N in root-bleeding sap, vacuum-extracted sap (100 [ureide-N]/[ureide-N+ α-amino-N + nitrate-N]) and stem extracts (100 [ureide-N]/[ureide-N + nitrate-N]) and the proportion of plant N, derived from nitrogen fixation between successive samplings were highly correlated (r = 0.97-1.00). For each variable, two standard curves were prepared to account for the shifts in the compositions

  17. Effects of climate on deer bone δ15N and δ13C: Lack of precipitation effects on δ15N for animals consuming low amounts of C 4 plants

    NASA Astrophysics Data System (ADS)

    Cormie, A. B.; Schwarcz, H. P.

    1996-11-01

    We have examined the relationship of bone collagen δ15N and δ13C to climatic variables, humidity, temperature, and amount of precipitation using fifty-nine specimens of North American white-tailed deer ( Odocoileus virginianus) from forty-six different locations. In previous studies of African mammals there was a significant correlation between bone collagen δ15N and local amount of precipitation. Results presented here similarly show an increase in δ15N with decreasing amount of precipitation but only for 25% of the animals, namely those consuming more than 10% C 4 plants. These animals also exhibited a significant correlation between δ13C and temperature which mirrors previous observations for grasses suggesting that these deer consume grasses during times of population and nutrient stress. In contrast, even in dry areas containing high proportions of C 4 grasses, the majority of the deer had consumed low amounts of C 4 plants and these deer did not have δ15N which correlate with amount of precipitation. Only when deer deviated from their normal feeding pattern by consuming C 4 plants or grasses did their δ15N correlate with amount of rainfall. For these animals, consumption of C 4 plants or grasses may signal conditions of water and nutrient stress. An increase in δ15N of bone collagen may result from combined effects from excretion of concentrated urine (to conserve water) and increased internal recycling of nitrogen (to conserve nitrogen).

  18. The Cyanide Ligands of [FeFe] Hydrogenase: Pulse EPR Studies of 13C and 15N-Labeled H-Cluster

    PubMed Central

    2015-01-01

    The two cyanide ligands in the assembled cluster of [FeFe] hydrogenase originate from exogenous l-tyrosine. Using selectively labeled tyrosine substrates, the cyanides were isotopically labeled via a recently developed in vitro maturation procedure allowing advanced electron paramagnetic resonance techniques to probe the electronic structure of the catalytic core of the enzyme. The ratio of the isotropic 13C hyperfine interactions for the two CN– ligands—a reporter of spin density on their respective coordinating iron ions—collapses from ≈5.8 for the Hox form of hydrogenase to <2 for the CO-inhibited form. Additionally, when the maturation was carried out using [15N]-tyrosine, no features previously ascribed to the nitrogen of the bridging dithiolate ligand were observed suggesting that this bridge is not sourced from tyrosine. PMID:25133957

  19. Determination of 15N/14N and 13C/12C in Solid and Aqueous Cyanides

    USGS Publications Warehouse

    Johnson, C.A.

    1996-01-01

    The stable isotopic compositions of nitrogen and carbon in cyanide compounds can be determined by combusting aliquots in sealed tubes to form N2 gas and CO2 gas and analyzing the gases by mass spectrometry. Free cyanide (CN-aq + HCNaq) in simple solutions can also be analyzed by first precipitating the cyanide as copper(II) ferrocyanide and then combusting the precipitate. Reproducibility is ??0.5??? or better for both ??15N and ??13C. If empirical corrections are made on the basis of carbon yields, the reproducibility of ??13C can be improved to ??0.2???. The analytical methods described herein are sufficiently accurate and precise to apply stable isotope techniques to problems of cyanide degradation in natural waters and industrial process solutions.

  20. Creating 13C- and 15N-enriched tree leaf litter for decomposition experiments

    NASA Astrophysics Data System (ADS)

    Szlavecz, K. A.; Pitz, S.; Chang, C.; Bernard, M.

    2013-12-01

    Labeling plant material with heavy isotopes of carbon and nitrogen can produce a traceable nutrient signal that can be followed into the different trophic levels and decomposer food web. We treated 60 tree saplings with 13C-enriched CO2 gas and 15N-enriched ammonium nitrate over a three-month period to create dually-labeled plant material for future decomposition experiments. The trees included both early (Red maple, Sweetgum, Tulip poplar) and late (American beech, White oak) successional deciduous tree species, and a conifer, White pine. We constructed a 2.4 m × 2.4 m × 2.4 m environmental chamber that was climate-controlled using an air conditioning system. An Arduino microcontroller interfaced with a Vaisala GMP343 CO2 probe maintained a CO2 concentration between 500-520 ppm by controlling a solenoid valve on the CO2 tank regulator. The trees were placed into the chamber in August 2012 and remained until senescence unless they were lost to death or disease. Ammonium nitrate was added twice, in September and October. Leaf samples were collected prior to the start of the experiment and after senescence, whereas root samples were collected only in December. Samples were dried, ground and analyzed using an isotope ratio mass spectrometer. American beech and White oak had 40% mortality, and 34% of tulip poplar trees were removed because of powdery mildew overgrowth or death. Most tulip poplar trees exhibited a second leaf out following senescence in late September. Nearly 1 kg of litter was produced with tulip poplar representing over half of the total mass. Levels of enrichment varied greatly by species. Beech (-14.2‰) and White oak (-4.8‰) had low levels of enrichment in comparison to early successional species such as Sweetgum (41.7‰) and Tulip poplar (30.7‰ [first leaf fall] and 238.0‰ [second leaf fall]). Leaf enrichment with 15N followed a similar pattern, though it was achieved at a higher level with δ15N values varying from 271.6‰ to 1354.2

  1. A 115-year δ15N record of cumulative nitrogen pollution in California serpentine grasslands

    NASA Astrophysics Data System (ADS)

    Vallano, D.; Zavaleta, E. S.

    2010-12-01

    Until the 1980s, California’s biodiverse serpentine grasslands were threatened primarily by development and protected by reserve creation. However, nitrogen (N) fertilization due to increasing fossil fuel emissions in the expanding Bay Area is thought to be contributing to rapid, recent invasion of these ecosystems by exotic annual grasses that are displacing rare and endemic serpentine species. Documenting the cumulative effects of N deposition in this ecosystem can direct policy and management actions to mitigate the role of N deposition in its transformation. Natural abundance stable isotopes of N in vegetation have been increasingly used as bio-indicators of N deposition patterns and subsequent changes to plant N cycling and assimilation. However, the long-term record of atmospheric reactive N enrichment and the resulting changes in ecosystem N dynamics have yet to be adequately reconstructed in many ecosystems. Museum archives of vascular plant tissue are valuable sources of materials to reconstruct temporal and spatial isotopic patterns of N inputs to ecosystems. Here, we present N stable isotope data from archived and current specimens of an endemic California serpentine grassland species, leather oak (Quercus durata), since 1895 across the greater San Francisco Bay region. We measured spatial and temporal trends in stable isotope composition (δ15N and δ13C) and concentration (%N and %C) of historical and current samples of leather oak leaves from sites within the Bay Area, impacted by increasing development, and sites northeast of the Bay Area, with significantly lower rates of urbanization and industrialization. Specifically, we sampled dry museum and fresh leaf specimens from serpentine sites within Lake (n=27) and Santa Clara (n=30) counties dating from 1895 to 2010. Leaf δ15N values were stable from 1895 to the 1950s and then decreased strongly throughout the last 50 years as fossil fuel emissions rapidly increased in the Bay Area, indicating that

  2. Nitrogen use efficiency evaluation of aerobic rice under field capacity water potential using {sup 15}N isotopic tracer technique

    SciTech Connect

    Wahid, Ahmad Nazrul Abd; Rahim, Sahibin Abd; Rahim, Khairuddin Abdul; Harun, Abdul Rahim

    2015-09-25

    This study was carried out to evaluate the efficiency use of the nitrogen fertilizer on aerobic rice varieties MR219-4 and MR219-9 which were grown aerobically under field capacity water potential at the controlled environment area or shield house. Direct {sup 15}N isotope tracer method was used in this study, whereby the {sup 15}N isotope was utilized as a tracer for nitrogen nutrient uptake. {sup 15}N isotope presence in the samples is determined by using emission spectrometer analysis and percentage of total nitrogen is determined by using Kjeldahl method. {sup 15}N atom access value contained in the sample will be used in determining the effectiveness of the use of nitrogen in fertilizers through the specific calculation formulas. In this work, the data several data of nitrogen derived from fertilizer (Ndff), total nitrogen, nitrogen uptake and nitrogen use efficiency was obtained.

  3. δ15N constraints on long-term nitrogen balances in temperate forests.

    PubMed

    Perakis, Steven S; Sinkhorn, Emily R; Compton, Jana E

    2011-11-01

    Biogeochemical theory emphasizes nitrogen (N) limitation and the many factors that can restrict N accumulation in temperate forests, yet lacks a working model of conditions that can promote naturally high N accumulation. We used a dynamic simulation model of ecosystem N and δ(15)N to evaluate which combination of N input and loss pathways could produce a range of high ecosystem N contents characteristic of forests in the Oregon Coast Range. Total ecosystem N at nine study sites ranged from 8,788 to 22,667 kg ha(-1) and carbon (C) ranged from 188 to 460 Mg ha(-1), with highest values near the coast. Ecosystem δ(15)N displayed a curvilinear relationship with ecosystem N content, and largely reflected mineral soil, which accounted for 96-98% of total ecosystem N. Model simulations of ecosystem N balances parameterized with field rates of N leaching required long-term average N inputs that exceed atmospheric deposition and asymbiotic and epiphytic N(2)-fixation, and that were consistent with cycles of post-fire N(2)-fixation by early-successional red alder. Soil water δ(15)NO(3)(-) patterns suggested a shift in relative N losses from denitrification to nitrate leaching as N accumulated, and simulations identified nitrate leaching as the primary N loss pathway that constrains maximum N accumulation. Whereas current theory emphasizes constraints on biological N(2)-fixation and disturbance-mediated N losses as factors that limit N accumulation in temperate forests, our results suggest that wildfire can foster substantial long-term N accumulation in ecosystems that are colonized by symbiotic N(2)-fixing vegetation.

  4. Regional, seasonal and interspecific variation in 15N and 13C in sympatric mouse lemurs

    NASA Astrophysics Data System (ADS)

    Rakotondranary, S. Jacques; Struck, Ulrich; Knoblauch, Christian; Ganzhorn, Jörg U.

    2011-11-01

    Madagascar provides some of the rare examples where two or more primate species of the same genus and with seemingly identical niche requirements occur in sympatry. If congeneric primate species co-occur in other parts of the world, they differ in size in a way that is consistent with Hutchinson's rule for coexisting species, or they occupy different ecological niches. In some areas of Madagascar, mouse lemurs do not follow these "rules" and thus seem to violate one of the principles of community ecology. In order to understand the mechanisms that allow coexistence of sympatric congeneric species without obvious niche differentiation, we studied food composition of two identical sized omnivorous mouse lemur species, Microcebus griseorufus and M. murinus with the help of stable isotope analyses ( δ 15N and δ 13C). The two species are closely related sister species. During the rich season, when food seems abundant, the two species do not differ in their nitrogen isotope composition, indicating that the two species occupy the same trophic level. But they differ in their δ 13C values, indicating that M. griseorufus feeds more on C4 and CAM (Crassulacean-acid-metabolism) plants than M. murinus. During the lean season, M. murinus has lower δ 15N values, indicating that the two species feed at different trophic levels during times of food shortage. Hybrids between the two species showed intermediate food composition. The results reflect subtle differences in foraging or metabolic adaptations that are difficult to quantify by traditional observations but that represent possibilities to allow coexistence of species.

  5. Search for d3/2 single particle strength in 15N in Unbound Levels

    NASA Astrophysics Data System (ADS)

    Mertin, C. E.; Caussyn, D. D.; Crisp, A. M.; Keeley, N.; Kemper, K. W.; Momotyuk, O.; Roeder, B. T.; Volya, A.

    2013-10-01

    The population of states in the nucleus 15N provides the opportunity to investigate both single particle and cluster structures in the 1p and 2s1d shells. Single, two, three and four particle transfer reactions selectively excite states in 15N thus providing a way to explore current nuclear structure models. Narrow structures are observed in the various transfer reactions up to at least 20 MeV in excitation well above the neutron (10.8 MeV) and proton (10.2 MeV) separation energies. In the present work new results for the reaction 14N(d,p) are presented that explore possible single particle strengths up to 18 MeV in excitation. The beam energies used in the present work were between 10.5 and 16 MeV. An early work with a beam energy of 8 MeV clearly populated strong sharp levels at 10.07 and 11.23 MeV and the present work confirms their existence. In addition, very weak broader levels are populated at 12.13 and 12.5 MeV but no other structures are found experimentally at higher excitation energies. The results of shell model calculations that include the 1p and 2s1d shells will be presented. The centroid energies for the 1d5/2 and 2s1/2 single particle strength have been obtained through comparison with FRESCO calculations. This work was supported by the NSF, DOE and Florida State University.

  6. δ15N constraints on long-term nitrogen balances in temperate forests

    USGS Publications Warehouse

    Perakis, S.S.; Sinkhorn, E.R.; Compton, J.E.

    2011-01-01

    Biogeochemical theory emphasizes nitrogen (N) limitation and the many factors that can restrict N accumulation in temperate forests, yet lacks a working model of conditions that can promote naturally high N accumulation. We used a dynamic simulation model of ecosystem N and δ15N to evaluate which combination of N input and loss pathways could produce a range of high ecosystem N contents characteristic of forests in the Oregon Coast Range. Total ecosystem N at nine study sites ranged from 8,788 to 22,667 kg ha−1 and carbon (C) ranged from 188 to 460 Mg ha−1, with highest values near the coast. Ecosystem δ15N displayed a curvilinear relationship with ecosystem N content, and largely reflected mineral soil, which accounted for 96–98% of total ecosystem N. Model simulations of ecosystem N balances parameterized with field rates of N leaching required long-term average N inputs that exceed atmospheric deposition and asymbiotic and epiphytic N2-fixation, and that were consistent with cycles of post-fire N2-fixation by early-successional red alder. Soil water δ15NO3 − patterns suggested a shift in relative N losses from denitrification to nitrate leaching as N accumulated, and simulations identified nitrate leaching as the primary N loss pathway that constrains maximum N accumulation. Whereas current theory emphasizes constraints on biological N2-fixation and disturbance-mediated N losses as factors that limit N accumulation in temperate forests, our results suggest that wildfire can foster substantial long-term N accumulation in ecosystems that are colonized by symbiotic N2-fixing vegetation.

  7. Heteronuclear NMR assignments and secondary structure of the coiled coil trimerization domain from cartilage matrix protein in oxidized and reduced forms.

    PubMed Central

    Wiltscheck, R.; Kammerer, R. A.; Dames, S. A.; Schulthess, T.; Blommers, M. J.; Engel, J.; Alexandrescu, A. T.

    1997-01-01

    The C-terminal oligomerization domain of chicken cartilage matrix protein is a trimeric coiled coil comprised of three identical 43-residue chains. NMR spectra of the protein show equivalent magnetic environments for each monomer, indicating a parallel coiled coil structure with complete threefold symmetry. Sequence-specific assignments for 1H-, 15N-, and 13C-NMR resonances have been obtained from 2D 1H NOESY and TOCSY spectra, and from 3D HNCA, 15N NOESY-HSQC, and HCCH-TOCSY spectra. A stretch of alpha-helix encompassing five heptad repeats (35 residues) has been identified from intra-chain HN-HN and HN-H alpha NOE connectivities. 3JHNH alpha coupling constants, and chemical shift indices. The alpha-helix begins immediately downstream of inter-chain disulfide bonds between residues Cys 5 and Cys 7, and extends to near the C-terminus of the molecule. The threefold symmetry of the molecule is maintained when the inter-chain disulfide bonds that flank the N-terminus of the coiled coil are reduced. Residues Ile 21 through Glu 36 show conserved chemical shifts and NOE connectivities, as well as strong protection from solvent exchange in the oxidized and reduced forms of the protein. By contrast, residues Ile 10 through Val 17 show pronounced chemical shift differences between the oxidized and reduced protein. Strong chemical exchange NOEs between HN resonances and water indicate solvent exchange on time scales faster than 10 s, and suggests a dynamic fraying of the N-terminus of the coiled coil upon reduction of the disulfide bonds. Possible roles for the disulfide crosslinks of the oligomerization domain in the function of cartilage matrix protein are proposed. PMID:9260286

  8. Species specific and environment induced variation of δ13C and δ15N in alpine plants

    PubMed Central

    Yang, Yang; Siegwolf, Rolf T. W.; Körner, Christian

    2015-01-01

    Stable carbon and nitrogen isotope signals in plant tissues integrate plant-environment interactions over long periods. In this study, we hypothesized that humid alpine life conditions are narrowing the scope for significant deviations from common carbon, water and nitrogen relations as captured by stable isotope signals. We explored the variation in δ13C and δ15N in 32 plant species from tissue type to ecosystem scale across a suite of locations at c. Two thousand five hundred meter elevation in the Swiss Alps. Foliar δ13C and δ15N varied among species by about 3–4‰ and 7–8‰ respectively. However, there was no overall difference in means of δ13C and δ15N for species sampled in different plant communities or when bulk plant dry matter harvests of different plant communities were compared. δ13C was found to be highly species specific, so that the ranking among species was mostly maintained across 11 habitats. However, δ15N varied significantly from place to place in all species (a range of 2.7‰) except in Fabaceae (Trifolium alpinum) and Juncaceae (Luzula lutea). There was also a substantial variation among individuals of the same species collected next to each other. No difference was found in foliar δ15N of non-legumes, which were either collected next to or away from the most common legume, T. alpinum. δ15N data place Cyperaceae and Juncaceae, just like Fabaceae, in a low discrimination category, well separated from other families. Soil δ15N was higher than in plants and increased with soil depth. The results indicate a high functional diversity in alpine plants that is similar to that reported for low elevation plants. We conclude that the surprisingly high variation in δ13C and δ15N signals in the studied high elevation plants is largely species specific (genetic) and insensitive to obvious environmental cues. PMID:26097487

  9. Detection of 2,4,6-Trinitrotoluene-Utilizing Anaerobic Bacteria by 15N and 13C Incorporation ▿

    PubMed Central

    Gallagher, Erin M.; Young, Lily Y.; McGuinness, Lora M.; Kerkhof, Lee J.

    2010-01-01

    2,4,6-Trinitrotoluene (15N or 13C labeled) was added to Norfolk Harbor sediments to test whether anaerobic bacteria use TNT for growth. Stable-isotope probing (SIP)-terminal restriction fragment length polymorphism (TRFLP) detected peaks in the [15N]TNT cultures (60, 163, and 168 bp). The 60-bp peak was also present in the [13C]TNT cultures and was related to Lysobacter taiwanensis. PMID:20081008

  10. Investigating patterns of symbiotic nitrogen fixation during vegetation change from grassland to woodland using fine scale δ(15) N measurements.

    PubMed

    Soper, Fiona M; Boutton, Thomas W; Sparks, Jed P

    2015-01-01

    Biological nitrogen fixation (BNF) in woody plants is often investigated using foliar measurements of δ(15) N and is of particular interest in ecosystems experiencing increases in BNF due to woody plant encroachment. We sampled δ(15) N along the entire N uptake pathway including soil solution, xylem sap and foliage to (1) test assumptions inherent to the use of foliar δ(15) N as a proxy for BNF; (2) determine whether seasonal divergences occur between δ(15) Nxylem sap and δ(15) Nsoil inorganic N that could be used to infer variation in BNF; and (3) assess patterns of δ(15) N with tree age as indicators of shifting BNF or N cycling. Measurements of woody N-fixing Prosopis glandulosa and paired reference non-fixing Zanthoxylum fagara at three seasonal time points showed that δ(15) Nsoil inorganic N varied temporally and spatially between species. Fractionation between xylem and foliar δ(15) N was consistently opposite in direction between species and varied on average by 2.4‰. Accounting for these sources of variation caused percent nitrogen derived from fixation values for Prosopis to vary by up to ∼70%. Soil-xylem δ(15) N separation varied temporally and increased with Prosopis age, suggesting seasonal variation in N cycling and BNF and potential long-term increases in BNF not apparent through foliar sampling alone.

  11. 15N/14N variations in Cretaceous Atlantic sedimentary sequences: implication for past changes in marine nitrogen biogeochemistry

    USGS Publications Warehouse

    Rau, G.H.; Arthur, M.A.; Dean, W.E.

    1987-01-01

    At two locations in the Atlantic Ocean (DSDP Sites 367 and 530) early to middle Cretaceous organic-carbon-rich beds ("black shales") were found to have significantly lower ??15N values (lower 15N/14N ratios) than adjacent organic-carbon-poor beds (white limestones or green claystones). While these lithologies are of marine origin, the black strata in particular have ??15N values that are significantly lower than those previously found in the marine sediment record and most contemporary marine nitrogen pools. In contrast, black, organic-carbon-rich beds at a third site (DSDP Site 603) contain predominantly terrestrial organic matter and have C- and N-isotopic compositions similar to organic matter of modern terrestrial origin. The recurring 15N depletion in the marine-derived Cretaceous sequences prove that the nitrogen they contain is the end result of an episodic and atypical biogeochemistry. Existing isotopic and other data indicate that the low 15N relative abundance is the consequence of pelagic rather than post-depositional processes. Reduced ocean circulation, increased denitrification, and, hence, reduced euphotic zone nitrate availability may have led to Cretaceous phytoplankton assemblages that were periodically dominated by N2-fixing blue-green algae, a possible source of this sediment 15N-depletion. Lack of parallel isotopic shifts in Cretaceous terrestrially-derived nitrogen (Site 603) argues that the above change in nitrogen cycling during this period did not extend beyond the marine environment. ?? 1987.

  12. Cryptic or day-to-day parts of the riverbed N cycle - new challenges for 15N

    NASA Astrophysics Data System (ADS)

    Trimmer, Mark; Ouyang, Liao; Lansdown, Katrina

    2016-04-01

    The discovery of anaerobic ammonium oxidation (anammox) not only changed our understanding of the nitrogen cycle in aquatic ecosystems but it also undermined some of the key 15N techniques used to study it. Reformulations of principle equations and the development of new 15N2 and 15N2O techniques enabled the simultaneous quantification of N2 production by anammox and denitrification in mainly soft, cohesive sediments where redox gradients are clearly defined and solute exchanged governed by diffusion. At the heart of the application of 15N, for the quantification of natural 14N cycling, is the key assumption that the respective pools of 15N and 14N are evenly mixed and that both are cycled without bias towards each other. Recent evidence, however, from a variety of aquatic ecosystems, suggests that this may not be the case. For example, organic N may be oxidised directly to N2 gas without ever mixing with the inorganic pool or inorganic intermediates (e.g. nitrite) are 'shunted' internally and also fail to mix evenly with the applied tracer pool. Our most recent work in permeable, oxic gravel riverbeds presents some particular challenges to the application of 15N. In these systems, a tight coupling between aerobic nitrification and anaerobic N2 production - in the presence of 100

  13. δ15N Values in Crassostrea virginica Shells Provides Early Direct Evidence for Nitrogen Loading to Chesapeake Bay

    PubMed Central

    Black, H. D.; Andrus, C. F. T.; Lambert, W. J.; Rick, T. C.; Gillikin, D. P.

    2017-01-01

    Crassostrea virginica is one of the most common estuarine bivalves in the United States’ east coast and is frequently found in archaeological sites and sub-fossil deposits. Although there have been several sclerochronological studies on stable carbon and oxygen isotopes in the shells of this species, less is known about δ15N values within their shells, which could be a useful paleoenvironmental proxy to assess estuarine nitrogen dynamics. Modern C. virginica samples were collected in Chesapeake Bay for comparison with archaeological shells from nearby sites ranging in age from ~100 to 3,200 years old. Left valves were sampled by milling the hinge area and the resulting powder was analyzed for %N and δ15N values. Comparison of δ15N values between C. virginica shells shows relatively constant values from ~1250 BC to ~1800 AD. After ~1800 AD, there are rapid increases in 15N enrichment in the shells, which continue to increase in value up to the modern shell values. The increase in δ15N values is evidence of early anthropogenic impact in Chesapeake Bay. These results corroborate the observation that coastal nitrogen pollution occurred earlier than the 19th century and support the use of oyster shell δ15N values as a useful environmental proxy. PMID:28281649

  14. δ15N Values in Crassostrea virginica Shells Provides Early Direct Evidence for Nitrogen Loading to Chesapeake Bay

    NASA Astrophysics Data System (ADS)

    Black, H. D.; Andrus, C. F. T.; Lambert, W. J.; Rick, T. C.; Gillikin, D. P.

    2017-03-01

    Crassostrea virginica is one of the most common estuarine bivalves in the United States’ east coast and is frequently found in archaeological sites and sub-fossil deposits. Although there have been several sclerochronological studies on stable carbon and oxygen isotopes in the shells of this species, less is known about δ15N values within their shells, which could be a useful paleoenvironmental proxy to assess estuarine nitrogen dynamics. Modern C. virginica samples were collected in Chesapeake Bay for comparison with archaeological shells from nearby sites ranging in age from ~100 to 3,200 years old. Left valves were sampled by milling the hinge area and the resulting powder was analyzed for %N and δ15N values. Comparison of δ15N values between C. virginica shells shows relatively constant values from ~1250 BC to ~1800 AD. After ~1800 AD, there are rapid increases in 15N enrichment in the shells, which continue to increase in value up to the modern shell values. The increase in δ15N values is evidence of early anthropogenic impact in Chesapeake Bay. These results corroborate the observation that coastal nitrogen pollution occurred earlier than the 19th century and support the use of oyster shell δ15N values as a useful environmental proxy.

  15. 15N Abundance of Nodules as an Indicator of N Metabolism in N2-Fixing Plants 1

    PubMed Central

    Shearer, Georgia; Feldman, Lori; Bryan, Barbara A.; Skeeters, Jerri L.; Kohl, Daniel H.; Amarger, Nöelle; Mariotti, Françoise; Mariotti, André

    1982-01-01

    This paper expands upon previous reports of 15N elevation in nodules (compared to other tissues) of N2-fixing plants. N2-Fixing nodules of Glycine max (soybeans), Vigna unguiculata (cowpea), Phaseolus vulgaris (common bean), Phaseolus coccineus (scarlet runner bean), Prosopis glandulosa (mesquite), and Olneya tesota (desert ironwood) were enriched in 15N. Nodules of Vicia faba (fava beans), Arachis hypogaea (peanut), Trifolium pratense (red clover), Pisum sativum (pea), Lathyrus sativus (grass pea), Medicago sativa (alfalfa), and Lupinus mutabilis (South American lupine) were not; nor were the nodules of nine species of N2-fixing nonlegumes. The nitrogen of ineffective nodules of soybeans and cowpeas was not enriched in 15N. Thus, 15N elevation in nodules of these plants depends on active N2-fixation. Results obtained so far on the generality of 15N enrichment in N2-fixing nodules suggest that only the nodules of plants which actively fix N2 and which transport allantoin or allantoic acid exhibit 15N enrichment. PMID:16662517

  16. Using dual-bacterial denitrification to improve δ15N determinations of nitrates containing mass-independent 17O

    USGS Publications Warehouse

    Coplen, T.B.; Böhlke, J.K.; Casciotti, K.L.

    2004-01-01

    The bacterial denitrification method for isotopic analysis of nitrate using N2O generated from Pseudomonas aureofaciens may overestimate ??15N values by as much as 1-2??? for samples containing atmospheric nitrate because of mass-independent 17O variations in such samples. By analyzing such samples for ??15N and ??18O using the denitrifier Pseudomonas chlororaphis, one obtains nearly correct ??15N values because oxygen in N 2O generated by P. chlororaphis is primarily derived from H 2O. The difference between the apparent ??15N value determined with P. aureofaciens and that determined with P. chlororaphis, assuming mass-dependent oxygen isotopic fractionation, reflects the amount of mass-independent 17O in a nitrate sample. By interspersing nitrate isotopic reference materials having substantially different ?? 18O values with samples, one can normalize oxygen isotope ratios and determine the fractions of oxygen in N2O derived from the nitrate and from water with each denitrifier. This information can be used to improve ??15N values of nitrates having excess 17O. The same analyses also yield estimates of the magnitude of 17O excess in the nitrate (expressed as ??17O) that may be useful in some environmental studies. The 1-?? uncertainties of ??15N, ??18O and ??17O measurements are ??0.2, ??0.3 and ??5???, respectively. Copyright ?? 2004 John Wiley & Sons, Ltd.

  17. Mycorrhizal fungi supply nitrogen to host plants in Arctic tundra and boreal forests: 15N is the key signal.

    PubMed

    Hobbie, John E; Hobbie, Erik A; Drossman, Howard; Conte, Maureen; Weber, J C; Shamhart, Julee; Weinrobe, Melissa

    2009-01-01

    Symbiotic fungi's role in providing nitrogen to host plants is well-studied in tundra at Toolik Lake, Alaska, but little-studied in the adjoining boreal forest ecosystem. Along a 570 km north-south transect from the Yukon River to the North Slope of Alaska, the 15N content was strongly reduced in ectomycorrhizal and ericoid mycorrhizal plants including Betula, Salix, Picea mariana (P. Mill.) B.S.P., Picea glauca Moench (Voss), and ericaceous plants. Compared with the 15N content of soil, the foliage of nonmycorrhizal plants (Carex and Eriophorum) was unchanged, whereas content of the ectomycorrhizal fungi was very much higher (e.g., Boletaceae, Leccinum and Cortinarius). It is hypothesized that similar processes operate in tundra and boreal forest, both nitrogen-limited ecosystems: (i) mycorrhizal fungi break down soil polymers and take up amino acids or other nitrogen compounds; (ii) mycorrhizal fungi fractionate against 15N during production of transfer compounds; (iii) host plants are accordingly depleted in 15N; and (iv) mycorrhizal fungi are enriched in 15N. Increased N availability for plant roots or decreased light availability to understory plants may have decreased N allocation to mycorrhizal partners and increased delta15N by 3-4 parts per million for southern populations of Vaccinium vitis-idaea L. and Salix. Fungal biomass, measured as ergosterol, correlated strongly with soil organic matter and attained amounts similar to those in temperate forest soils.

  18. Correlation between the synthetic origin of methamphetamine samples and their 15N and 13C stable isotope ratios.

    PubMed

    Billault, Isabelle; Courant, Frédérique; Pasquereau, Léo; Derrien, Solène; Robins, Richard J; Naulet, Norbert

    2007-06-12

    The active ingredient of ecstasy, N-methyl-3,4-methyldioxyphenylisopropylamine (MDMA) can be manufactured by a number of easy routes from simple precursors. We have synthesised 45 samples of MDMA following the five most common routes using N-precursors from 12 different origins and three different precursors for the aromatic moiety. The 13C and 15N contents of both the precursors and the MDMA samples derived therefrom were measured by isotope ratio mass spectrometry coupled to an elemental analyser (EA-IRMS). We show that within-pathway correlation between the 15N content of the precursor and that of the derived MDMA can be strong but that no general pattern of correlation can be defined. Rather, it is evident that the delta15N values of MDMA are strongly influenced by a combination of the delta15N values of the source of nitrogen used, the route by which the MDMA is synthesised, and the experimental conditions employed. Multivariate analysis (PCA) based on the delta15N values of the synthetic MDMA and of the delta15N and delta13C values of the N-precursors leads to good discrimination between the majority of the reaction conditions tested.

  19. Elemental formula annotation of polar and lipophilic metabolites using (13) C, (15) N and (34) S isotope labelling, in combination with high-resolution mass spectrometry.

    PubMed

    Giavalisco, Patrick; Li, Yan; Matthes, Annemarie; Eckhardt, Aenne; Hubberten, Hans-Michael; Hesse, Holger; Segu, Shruthi; Hummel, Jan; Köhl, Karin; Willmitzer, Lothar

    2011-10-01

    The unbiased and comprehensive analysis of metabolites in any organism presents a major challenge if proper peak annotation and unambiguous assignment of the biological origin of the peaks are required. Here we provide a comprehensive multi-isotope labelling-based strategy using fully labelled (13) C, (15) N and (34) S plant tissues, in combination with a fractionated metabolite extraction protocol. The extraction procedure allows for the simultaneous extraction of polar, semi-polar and hydrophobic metabolites, as well as for the extraction of proteins and starch. After labelling and extraction, the metabolites and lipids were analysed using a high-resolution mass spectrometer providing accurate MS and all-ion fragmentation data, providing an unambiguous readout for every detectable isotope-labelled peak. The isotope labelling assisted peak annotation process employed can be applied in either an automated database-dependent or a database-independent analysis of the plant polar metabolome and lipidome. As a proof of concept, the developed methods and technologies were applied and validated using Arabidopsis thaliana leaf and root extracts. Along with a large repository of assigned elemental compositions, which is provided, we show, using selected examples, the accuracy and reliability of the developed workflow.

  20. An Assignment Sequence for Underprepared Writers.

    ERIC Educational Resources Information Center

    Nimmo, Kristi

    2000-01-01

    Presents a sequenced writing assignment on shopping to aid basic writers. Describes a writing assignment focused around online and mail-order shopping. Notes steps in preparing for the assignment, the sequence, and discusses responses to the assignments. (SC)

  1. MILLIMETER-WAVE OBSERVATIONS OF CN AND HNC AND THEIR {sup 15}N ISOTOPOLOGUES: A NEW EVALUATION OF THE {sup 14}N/{sup 15}N RATIO ACROSS THE GALAXY

    SciTech Connect

    Adande, G. R.; Ziurys, L. M.

    2012-01-10

    The N = 1 {yields} 0 transitions of CN and C{sup 15}N (X{sup 2}{Sigma}{sup +}), as well as the J = 1 {yields} 0 lines of HN{sup 13}C and H{sup 15}NC, have been observed toward 11 molecular clouds using the new 3 mm ALMA-type receiver of the 12 m telescope of the Arizona Radio Observatory. These sources span a wide range of distances from the Galactic center and are all regions of star formation. From these observations, {sup 14}N/{sup 15}N ratios have been determined using two independent methods. First, the measurements of C{sup 14}N and C{sup 15}N were directly compared to establish this ratio, correcting for high opacities when needed, as indicated by the nitrogen hyperfine intensities. Second, the ratio was calculated from the quantity [HN{sup 13}C]/[H{sup 15}NC], determined from the HNC data, and then scaled by {sup 12}C/{sup 13}C ratios previously established, i.e., the so-called double isotope method. Values from both methods are in reasonable agreement, and fall in the range {approx}120-400, somewhat lower than previous {sup 14}N/{sup 15}N ratios derived from HCN. The ratios exhibit a distinct positive gradient with distance from the Galactic center, following the relationship{sup 14}N/{sup 15}N = 21.1 (5.2) kpc{sup -1} D{sub GC} + 123.8 (37.1). This gradient is consistent with predictions of Galactic chemical evolution models in which {sup 15}N has a secondary origin in novae, while primary and secondary sources exist for {sup 14}N. The local interstellar medium value was found to be {sup 4}N/{sup 15}N = 290 {+-} 40, in agreement with the ratio found in nearby diffuse clouds and close to the value of 272 found in Earth's atmosphere.

  2. Effect of body size and body mass on δ 13 C and δ 15 N in coastal fishes and cephalopods

    NASA Astrophysics Data System (ADS)

    Vinagre, C.; Máguas, C.; Cabral, H. N.; Costa, M. J.

    2011-11-01

    Carbon and nitrogen isotopes have been widely used in the investigation of trophic relations, energy pathways, trophic levels and migrations, under the assumption that δ 13C is independent of body size and that variation in δ 15N occurs exclusively due to ontogenetic changes in diet and not body size increase per se. However, several studies have shown that these assumptions are uncertain. Data from food-webs containing an important number of species lack theoretical support on these assumptions because very few species have been tested for δ 13C and δ 15N variation in captivity. However, if sampling comprises a wide range of body sizes from various species, the variation of δ 13C and δ 15N with body size can be investigated. While correlation between body size and δ 13C and δ 15N can be due to ontogenetic diet shifts, stability in such values throughout the size spectrum can be considered an indication that δ 13C and δ 15N in muscle tissues of such species is independent of body size within that size range, and thus the basic assumptions can be applied in the interpretation of such food webs. The present study investigated the variation in muscle δ 13C and δ 15N with body size and body mass of coastal fishes and cephalopods. It was concluded that muscle δ 13C and δ 15N did not vary with body size or mass for all bony fishes with only one exception, the dragonet Callionymus lyra. Muscle δ 13C and δ 15N also did not vary with body size or mass in cartilaginous fishes and cephalopods, meaning that body size/mass per se have no effect on δ 13C or δ 15N, for most species analysed and within the size ranges sampled. The assumption that δ 13C is independent of body size and that variation in δ 15N is not affected by body size increase per se was upheld for most organisms and can be applied to the coastal food web studied taking into account that C. lyra is an exception.

  3. NMR assignments of SPOC domain of the human transcriptional corepressor SHARP in complex with a C-terminal SMRT peptide.

    PubMed

    Mikami, Suzuka; Kanaba, Teppei; Ito, Yutaka; Mishima, Masaki

    2013-10-01

    The transcriptional corepressor SMRT/HDAC1-associated repressor protein (SHARP) recruits histone deacetylases. Human SHARP protein is thought to function in processes involving steroid hormone responses and the Notch signaling pathway. SHARP consists of RNA recognition motifs (RRMs) in the N-terminal region and the spen paralog and ortholog C-terminal (SPOC) domain in the C-terminal region. It is known that the SPOC domain binds the LSD motif in the C-terminal tail of corepressors silencing mediator for retinoid and thyroid receptor (SMRT)/nuclear receptor corepressor (NcoR). We are interested in delineating the mechanism by which the SPOC domain recognizes the LSD motif of the C-terminal tail of SMRT/NcoR. To this end, we are investigating the tertiary structure of the SPOC/SMRT peptide using NMR. Herein, we report on the (1)H, (13)C and (15)N resonance assignments of the SPOC domain in complex with a SMRT peptide, which contributes towards a structural understanding of the SPOC/SMRT peptide and its molecular recognition.

  4. Nuclear polarization of /sup 15/N via ion-beam-foil interaction

    SciTech Connect

    Deutch, B.I.; Liu, C.H. II; Lu, F.; Sun, C.; Tan, J.; Tang, G.; Xu, K.; Yang, F.; Ye, H.

    1981-10-01

    The ion beam surface interaction at grazing incidence (IBSIGI) generates highly oriented atomic states, and nuclear spin polarized ions are produced via hf-interactions. Both single and multiple IBSIGI were reported./sup 1/ By single reflection, nuclear polarizations of P/sub I/ = 14% in /sup 14/N(I = 1), and P/sub I/ = 6.8% in /sup 7/Li(I = 3/2) were produced. In this paper, the transmission rather than reflection technique is used. A 600 keV /sup 15/N/sup +/(I = 1/2) beam passed through a foil tilted 60 /sup 0/ with respect to the beam axis, and a perpendicular foil (both made of 20 ..mu..g/cm/sup 2/ thick carbon). After the first foil, highly oriented atomic states are produced, which result in large circular polarization fractions in the fluorescent radiation. By hf-interaction, the orientation can be transferred from the electronic shell to the nucleus, or vice versa. In the second foil, which is perpendicular, and therefore does not produce any polarization, the interaction does not affect the nuclear spin, but attaches a new unoriented electronic shell to the nucleus. Thus the circular polarization in the fluorescence after the second foil must stem from the transfer of orientation from the nucleus to the electronic shell and is therefore a direct measure of the nuclear spin orientation. To determine the degree of circular polarization, the Stokes parameter S/I is measured. For the multiplet exclamation/sup 5/N II 2s/sup 2/2p3s /sup 3/P--2s/sup 2/2p3p /sup 3/D after a tilted foil the S/I is equal to 8.5 +- 0.8%; after double foils (60 /sup 0/ tilted foil+perpendicular foil), S/I = 1.6 +- 0.4%. From the latter values, the nuclear polarization of /sup 15/N is calculated: P/sub I/ = 10.2%.

  5. [Platanus orientalis foliar N% and delta15 N responses to nitrogen of atmospheric wet deposition in urban area].

    PubMed

    Wang, Yan-Li; Xiao, Hua-Yun; Xiao, Hong-Wei

    2012-04-01

    Leaves of Platanus orientalis were collected since Mar. 2009 till Apr. 2010, in an urban area at Guiyang. After mass of experiments and analysis, we carried out constructing the temporal variation of foliar N% and delta15 N: both higher in Spring/Summer, lower in Autumn, no data of Winter because of leaf abscission. Results showed that foliar N% varied from 1.48% to 5.27%, with an annual average of 3.36%, while the average concentration of total N in rhizospheric soil was 0.29%. The foliar N% rose and fell relative to DIN in rainwater (range from 0.57 mg x L(-1) to 6.74 mg x L(-1)), indicating that the N% content in foliar tissue of plant was approximately proportional to atmospheric N inputs. The range of foliar delta15N were from 4.48 per thousand to 8.39 per thousand, with the average of 6.33 per thousand, much higher than the delta15N-NH4+ of rain water (-19.76 per thousand(-) -10.41 per thousand) and delta15TN of rhizospheric soil (3.19 per thousand +/- 1.04 per thousand). Besides, a good uniform correlation between foliar delta15N and delta15N-NH4+ of rain water were found. As synthesis of two main N sources, the more positive delta15N values of Platanus orientalis can be explained by isotopic fractionation during N uptake and basipetal translocation. These responses of both foliar N% and delta15N to atmospheric nitrogen deposition, revealed the potential value in using vascular leaves as bio-monitors for assessment of N deposition, furthermore, for prevention and control of air pollution in urban ecosystem.

  6. Qualitative study of substituent effects on NMR (15)N and (17)O chemical shifts.

    PubMed

    Contreras, Rubén H; Llorente, Tomás; Pagola, Gabriel I; Bustamante, Manuel G; Pasqualini, Enrique E; Melo, Juan I; Tormena, Cláudio F

    2009-09-10

    A qualitative approach to analyze the electronic origin of substituent effects on the paramagnetic part of chemical shifts is described and applied to few model systems, where its potentiality can be appreciated. The formulation of this approach is based on the following grounds. The influence of different inter- or intramolecular interactions on a second-order property can be qualitatively predicted if it can be known how they affect the main virtual excitations entering into that second-order property. A set of consistent approximations are introduced in order to analyze the behavior of occupied and virtual orbitals that define some experimental trends of magnetic shielding constants. This approach is applied first to study the electronic origin of methyl-beta substituent effects on both (15)N and (17)O chemical shifts, and afterward it is applied to a couple of examples of long-range substituent effects originated in charge transfer interactions such as the conjugative effect in aromatic compounds and sigma-hyperconjugative interactions in saturated multicyclic compounds.

  7. Tracking the incorporation of 15N from labeled beech litter into mineral-organic associations

    NASA Astrophysics Data System (ADS)

    Kleber, M.; Hatton, P.; Derrien, D.; Lajtha, K.; Zeller, B.

    2008-12-01

    Nitrogen containing organic compounds are thought to have a role in the complex web of processes that control the turnover time of soil organic matter. The sequential density fractionation technique is increasingly used for the purpose of investigating the association of organic materials with the mineral matrix. Organic materials in the denser fractions (>2.0 kg L-1) typically show 13C NMR signals indicative of carbohydrate and aliphatic structures, an absence of lignin and tannin structures and a narrow C:N ratio, suggesting a microbial origin of organic matter in these fractions. Here we take advantage of a labeling experiment conducted at two different sites in Germany and in France to investigate the incorporation of organic nitrogen into physical fractions of increasing density, representing a proximity gradient to mineral surfaces. 15N labeled beech litter was applied to two acidic forest topsoils 8 and 12 years ago. Although there are differences in the distribution patterns between the two soils, and the majority of the organic nitrogen was recovered in fractions representing organic matter of plant origin and not bound to the mineral matrix, our data clearly show that after a decade, significant amounts of the nitrogen had been incorporated in mineral-organic fractions of supposedly slow turnover. It remains to be shown to which extent the N in the densest fractions was incorporated by soil microbiota and associated with mineral surfaces in organic form or adsorbed to mineral surfaces in inorganic form (NH4+).

  8. 15N Solid-State NMR as a Probe of Flavin H-bonding

    PubMed Central

    Cui, Dongtao; Koder, Ronald L.; Dutton, P. Leslie; Miller, Anne-Frances

    2011-01-01

    Flavins mediate a wide variety of different chemical reactions in biology. To learn how one cofactor can be made to execute different reactions in different enzymes, we are developing solid-state NMR (SSNMR) to probe the flavin electronic structure, via the 15N chemical shift tensor principal values (δii). We find that SSNMR has superior responsiveness to H-bonds, compared to solution NMR. H-bonding to a model of the flavodoxin active site produced an increase of 10 ppm in the δ11 of N5 although none of the H-bonds directly engage N5, and solution NMR detected only a 4 ppm increase in the isotropic chemical shift (δiso). Moreover SSNMR responded differently to different H-bonding environments as H-bonding with water caused δ11 to decrease by 6 ppm whereas δiso increased by less than 1 ppm. Our density functional theoretical (DFT) calculations reproduce the observations, validating the use of computed electronic structures to understand how H-bonds modulate the flavin’s reactivity. PMID:21619002

  9. 13C and 15N natural isotope abundance reflects breast cancer cell metabolism

    NASA Astrophysics Data System (ADS)

    Tea, Illa; Martineau, Estelle; Antheaume, Ingrid; Lalande, Julie; Mauve, Caroline; Gilard, Francoise; Barillé-Nion, Sophie; Blackburn, Anneke C.; Tcherkez, Guillaume

    2016-09-01

    Breast cancer is the most common cancer in women worldwide. Despite the information provided by anatomopathological assessment and molecular markers (such as receptor expression ER, PR, HER2), breast cancer therapies and prognostics depend on the metabolic properties of tumor cells. However, metabolomics have not provided a robust and congruent biomarker yet, likely because individual metabolite contents are insufficient to encapsulate all of the alterations in metabolic fluxes. Here, we took advantage of natural 13C and 15N isotope abundance to show there are isotopic differences between healthy and cancer biopsy tissues or between healthy and malignant cultured cell lines. Isotope mass balance further suggests that these differences are mostly related to lipid metabolism, anaplerosis and urea cycle, three pathways known to be impacted in malignant cells. Our results demonstrate that the isotope signature is a good descriptor of metabolism since it integrates modifications in C partitioning and N excretion altogether. Our present study is thus a starting point to possible clinical applications such as patient screening and biopsy characterization in every cancer that is associated with metabolic changes.

  10. Effects of 15N application frequency on nitrogen uptake efficiency in citrus trees.

    PubMed

    Quiñones, Ana; Bañuls, Josefina; Millo, Eduardo Primo; Legaz, Francisco

    2003-12-01

    Two irrigation systems were used to compare nitrogen uptake efficiency in citrus trees and to evaluate the NO3- runoff in "Navelina" orange trees [Citrus sinensis (L.) Osbeck] on Carrizo citrange rootstock (Citrus sinensis x Poncirus trifoliata Raf.). These were fertilized with 125 g N as labelled K15NO3 and grown outdoors in containers filled with a sand-loamy soil. Two groups of 3 trees received this N dose either in five equally split applications by a flooding irrigation system or in 66 applications by drip. Trees were harvested at the end of the vegetative cycle (December) and the isotopic ratios of 15N/14N were measured in the soil-plant system. The N uptake efficiency of the whole tree was higher with drip irrigation (75%) than with flooding system (64%). In the 0-90 cm soil profile, the N immobilized in the organic fraction was similar for both irrigation methods (around 13 %), whereas the N retained as NO3- was 1% of the N applied under drip and 10% under flooding. In the last case, most of NO3- remained under root system and it could be lost to leaching either by heavy rainfalls or excessive water applications. These results showed that a drip irrigation system was more efficient for improving water use and N uptake from fertilizer, in addition to potentially reduced leaching losses.

  11. Simultaneous quantification of depolymerization and mineralization rates by a novel 15N tracing model

    NASA Astrophysics Data System (ADS)

    Andresen, Louise C.; Björsne, Anna-Karin; Bodé, Samuel; Klemedtsson, Leif; Boeckx, Pascal; Rütting, Tobias

    2016-09-01

    The depolymerization of soil organic matter, such as proteins and (oligo-)peptides, into monomers (e.g. amino acids) is currently considered to be the rate-limiting step for nitrogen (N) availability in terrestrial ecosystems. The mineralization of free amino acids (FAAs), liberated by the depolymerization of peptides, is an important fraction of the total mineralization of organic N. Hence, the accurate assessment of peptide depolymerization and FAA mineralization rates is important in order to gain a better process-based understanding of the soil N cycle. In this paper, we present an extended numerical 15N tracing model Ntrace, which incorporates the FAA pool and related N processes in order to provide a more robust and simultaneous quantification of depolymerization and gross mineralization rates of FAAs and soil organic N. We discuss analytical and numerical approaches for two forest soils, suggest improvements of the experimental work for future studies, and conclude that (i) when about half of all depolymerized peptide N is directly mineralized, FAA mineralization can be as important a rate-limiting step for total gross N mineralization as peptide depolymerization rate; (ii) gross FAA mineralization and FAA immobilization rates can be used to develop FAA use efficiency (NUEFAA), which can reveal microbial N or carbon (C) limitation.

  12. Ammonia 15N/14N Isotope Ratio in the Jovian Atmosphere

    NASA Technical Reports Server (NTRS)

    Mahaffy, P.R.; Niemann, H. B.; Atreya, S. K.; Wong, M. H.; Owen, T. C; Einaudi, Franco (Technical Monitor)

    2000-01-01

    Data from the Galileo Probe Mass Spectrometer has been used to derive the N-15/N-14 isotope ratio in ammonia at Jupiter. Although the mass spectral interference from the water contribution to 18 amu makes an accurate derivation of the (N-15)H3/(N-14)H3 ratio difficult from measurements of the singly ionized signals at 18 and 17 amu, this interference is not present in the doubly charged 8.5 and 9.0 amu signals from (N-14)H3++ and (N-15)H3++ respectively. Although the count rate from the 9 amu signal is low during the direct sampling of the atmosphere, the ammonia signal was considerably enhanced during the first enrichment cell (EC1) experiment that measured gas sampled between 0.8 and 2.8 bar. Count rates at 9 amu in the EC1 experiment reach 60/second and measure ammonia sampled from 0.88 to 2.8 bar. In the EC1 measurements the 8.5 amu signal is not measured directly, but can be calculated from the ammonia contribution to 17 amu and the ratio of NH3 ions of a double to single charged observed during a high resolution mass scan taken near the end of the descent. The high resolution scan gives this ratio from ammonia sampled much deeper in the atmosphere. These results are described and compared with Infrared Space Observatory-Short Wavelength Spectrometer (ISO-SWS) observations that give this ratio at 400 mbar.

  13. 13C and 15N natural isotope abundance reflects breast cancer cell metabolism

    PubMed Central

    Tea, Illa; Martineau, Estelle; Antheaume, Ingrid; Lalande, Julie; Mauve, Caroline; Gilard, Francoise; Barillé-Nion, Sophie; Blackburn, Anneke C.; Tcherkez, Guillaume

    2016-01-01

    Breast cancer is the most common cancer in women worldwide. Despite the information provided by anatomopathological assessment and molecular markers (such as receptor expression ER, PR, HER2), breast cancer therapies and prognostics depend on the metabolic properties of tumor cells. However, metabolomics have not provided a robust and congruent biomarker yet, likely because individual metabolite contents are insufficient to encapsulate all of the alterations in metabolic fluxes. Here, we took advantage of natural 13C and 15N isotope abundance to show there are isotopic differences between healthy and cancer biopsy tissues or between healthy and malignant cultured cell lines. Isotope mass balance further suggests that these differences are mostly related to lipid metabolism, anaplerosis and urea cycle, three pathways known to be impacted in malignant cells. Our results demonstrate that the isotope signature is a good descriptor of metabolism since it integrates modifications in C partitioning and N excretion altogether. Our present study is thus a starting point to possible clinical applications such as patient screening and biopsy characterization in every cancer that is associated with metabolic changes. PMID:27678172

  14. STELLAR ORIGINS OF EXTREMELY {sup 13}C- AND {sup 15}N-ENRICHED PRESOLAR SIC GRAINS: NOVAE OR SUPERNOVAE?

    SciTech Connect

    Liu, Nan; Nittler, Larry R.; Alexander, Conel M. O’D.; Wang, Jianhua; Pignatari, Marco; José, Jordi; Nguyen, Ann

    2016-04-01

    Extreme excesses of {sup 13}C ({sup 12}C/{sup 13}C < 10) and {sup 15}N ({sup 14}N/{sup 15}N < 20) in rare presolar SiC grains have been considered diagnostic of an origin in classical novae, though an origin in core collapse supernovae (CCSNe) has also been proposed. We report C, N, and Si isotope data for 14 submicron- to micron-sized {sup 13}C- and {sup 15}N-enriched presolar SiC grains ({sup 12}C/{sup 13}C < 16 and {sup 14}N/{sup 15}N < ∼100) from Murchison, and their correlated Mg–Al, S, and Ca–Ti isotope data when available. These grains are enriched in {sup 13}C and {sup 15}N, but with quite diverse Si isotopic signatures. Four grains with {sup 29,30}Si excesses similar to those of type C SiC grains likely came from CCSNe, which experienced explosive H burning occurred during explosions. The independent coexistence of proton- and neutron-capture isotopic signatures in these grains strongly supports heterogeneous H ingestion into the He shell in pre-supernovae. Two of the seven putative nova grains with {sup 30}Si excesses and {sup 29}Si depletions show lower-than-solar {sup 34}S/{sup 32}S ratios that cannot be explained by classical nova nucleosynthetic models. We discuss these signatures within the CCSN scenario. For the remaining five putative nova grains, both nova and supernova origins are viable because explosive H burning in the two stellar sites could result in quite similar proton-capture isotopic signatures. Three of the grains are sub-type AB grains that are also {sup 13}C enriched, but have a range of higher {sup 14}N/{sup 15}N. We found that {sup 15}N-enriched AB grains (∼50 < {sup 14}N/{sup 15}N < ∼100) have distinctive isotopic signatures compared to putative nova grains, such as higher {sup 14}N/{sup 15}N, lower {sup 26}Al/{sup 27}Al, and lack of {sup 30}Si excess, indicating weaker proton-capture nucleosynthetic environments.

  15. Spatial Patterns of Plant δ13C and δ15N Along a Topoedaphic Gradient in a Subtropical Savanna Landscape

    NASA Astrophysics Data System (ADS)

    Bai, E.; Boutton, T. W.; Liu, F.; Wu, B.; Archer, S. R.

    2005-12-01

    δ13C and δ15N values of plants are powerful tools in physiological ecology, ecosystem science, and global biogeochemistry, yet we know relatively little about their variation and controls at the landscape scale. In this study, we investigated landscape-scale spatial variations in the foliar isotopic composition of 3 woody plant species across a 308 m topoedaphic gradient, along which soil texture and plant resources (water and nitrogen availability) varied from upland (86 m) to lowland (84 m) portions of the landscape. The study was conducted in a subtropical savanna at the La Copita Research Area, approximately 60 km west of Corpus Christi, TX. Foliar δ13C, δ15N, leaf nitrogen concentration ([N]), and specific leaf area (SLA) were measured on all individuals of Prosopis glandulosa, Condalia hookeri, and Zanthoxylum fagara present within a belt transect 308 m long x 12 m wide. Soil texture, available soil moisture, and total N were measured at 1 m intervals along the center-line of the belt transect. Clay content, available soil moisture, and soil total N were all negatively correlated with elevation along the transect. Leaf δ13C and δ15N values for all 3 species increased by 1-4 o/oo with decreasing elevation along the transect. Contrary to theory and previous studies, δ13C values were highest where soil water was most available, suggesting that some other variable could be overriding or interacting with water availability. Foliar [N] appeared to exert the strongest control over landscape-level variation, and was positively correlated with δ13C of all species (R 2 = 0.58, p<0.0001). Since leaf [N] is positively related to photosynthetic capacity, plants with high [N] are likely to have low Ci/Ca ratios and therefore higher δ13C values. δ15N values of Zanthoxylum and Condalia were positively correlated with leaf [N] and soil water availability; however, these relationships were absent for Prosopis, an N-fixing tree legume. We speculate that the

  16. delta15N and delta13C diet-tissue discrimination factors for large sharks under semi-controlled conditions.

    PubMed

    Hussey, Nigel E; Brush, Jaclyn; McCarthy, Ian D; Fisk, Aaron T

    2010-04-01

    Stable isotopes (delta(15)N and delta(13)C) are being widely applied in ecological research but there has been a call for ecologists to determine species- and tissue-specific diet discrimination factors ((13)C and (15)N) for their study animals. For large sharks stable isotopes may provide an important tool to elucidate aspects of their ecological roles in marine systems, but laboratory based controlled feeding experiments are impractical. By utilizing commercial aquaria, we estimated (15)N and (13)C of muscle, liver, vertebral cartilage and a number of organs of three large sand tiger (Carcharias taurus) and one large lemon shark (Negaprion brevirostris) under a controlled feeding regime. For all sharks mean+/-SD for (15)N and (13)C in lipid extracted muscle using lipid extracted prey data were 2.29 per thousand+/-0.22 and 0.90 per thousand+/-0.33, respectively. The use of non-lipid extracted muscle and prey resulted in very similar (15)N and (13)C values but mixing of lipid and non-lipid extracted data produced variable estimates. Values of (15)N and (13)C in lipid extracted liver and prey were 1.50 per thousand+/-0.54 and 0.22 per thousand+/-1.18, respectively. Non-lipid extracted diet discrimination factors in liver were highly influenced by lipid content and studies that examine stable isotopes in shark liver, and likely any high lipid tissue, should strive to remove lipid effects through standardising C:N ratios, prior to isotope analysis. Mean vertebral cartilage (15)N and (13)C values were 1.45 per thousand+/-0.61 and 3.75 per thousand+/-0.44, respectively. Organ (15)N and (13)C values were more variable among individual sharks but heart tissue was consistently enriched by approximately 1-2.5 per thousand. Minimal variability in muscle and liver delta(15)N and delta(13)C sampled at different intervals along the length of individual sharks and between liver lobes suggests that stable isotope values are consistent within tissues of individual animals. To our

  17. (15)N natural abundance in plants of the Amazon River floodplain and potential atmospheric N2 fixation.

    PubMed

    Martinelli, L A; Victoria, R L; Trivelin, P C O; Devol, A H; Richey, J E

    1992-07-01

    The(15)N natural abundance values of various Amazon floodplain (várzea) plants was investigated. Samples of young leaf tissues were collected during three different periods of the river hydrography (low water, mid rising water and high water) and during one period in the Madeira River (high water). A large variation of(15)N abundance was observed, both among the different plant types and between the different flood stages. This variation probably, reflected, in part, the highly variable nature of the floodplain, sometimes dry and oxygenated and at other times inundated and anaerobic and, in part, changes in plant nitrogen metabolism. Comparison of the nitrogen isotopic composition of leguminous plants with that of non-leguminous plants showed that, on average, the(15)N abundance was lower in the legumes than non-legumes, suggesting active N-fixation. Also, the(15)N natural abundance in aquatic grasses of the generaPaspalum, was in general, lower than the(15)N abundance of aquatic grasses of the generaEchinochloa. As both of these grasses grow in the same general habitat, it appears thatPaspalum grasses may also be nitrogen fixers.

  18. Exploring the Nitrogen Ingestion of Aphids — A New Method Using Electrical Penetration Graph and 15N Labelling

    PubMed Central

    Kuhlmann, Franziska; Opitz, Sebastian E. W.; Inselsbacher, Erich; Ganeteg, Ulrika; Näsholm, Torgny; Ninkovic, Velemir

    2013-01-01

    Studying plant-aphid interactions is challenging as aphid feeding is a complex process hidden in the plant tissue. Here we propose a combination of two well established methods to study nutrient acquisition by aphids focusing on the uptake of isotopically labelled nitrogen (15N). We combined the Electrical Penetration Graph (EPG) technique that allows detailed recording of aphid feeding behaviour and stable isotope ratio mass spectrometry (IRMS) to precisely measure the uptake of nitrogen. Bird cherry-oat aphids Rhopalosiphum padi L. (Hemiptera, Aphididae) fed for 24 h on barley plants (Hordeum vulgare L., cultivar Lina, Poaceae) that were cultivated with a 15N enriched nutrient solution. The time aphids fed in the phloem was strongly positive correlated with their 15N uptake. All other single behavioural phases were not correlated with 15N enrichment in the aphids, which corroborates their classification as non-feeding EPG phases. In addition, phloem-feeding and 15N enrichment of aphids was divided into two groups. One group spent only short time in the phloem phase and was unsuccessful in nitrogen acquisition, while the other group displayed longer phloem-feeding phases and was successful in nitrogen acquisition. This suggests that several factors such as the right feeding site, time span of feeding and individual conditions play a role for the aphids to acquire nutrients successfully. The power of this combination of methods for studying plant-aphid interactions is discussed. PMID:24376642

  19. δ 15N Studies of Nitrogen Use by the Red Mangrove, Rhizophora mangle L. in South Florida

    NASA Astrophysics Data System (ADS)

    Fry, B.; Bern, A. L.; Ross, M. S.; Meeder, J. F.

    2000-02-01

    To help define nitrogen (N) sources and patterns of N processing in mangrove ecosystems, mangrove leaf nitrogen contents and δ 15N values were assayed in three marshes along the south Florida coast. In each marsh, leaf samples were collected from dwarf mangroves at interior locations and taller mangroves at the ocean fringe. Leaf % N and δ 15N values did not differ consistently between dwarf and tall mangroves, even though there were large variations in δ 15N (18‰ range, -5 to +13‰) and % N (1·2% range, 0·9-2·1%). Highest % N and δ 15N values occurred along the western margin of Biscayne Bay where canals draining agricultural lands deliver high-nitrate waters to fringing mangrove marshes. High mangrove δ 15N values may be good biomonitors of anthropogenic N loading to south Florida estuaries. Lower values likely reflect less anthropogenic N entering the mangrove marshes, as well as differences in plant physiology that occur along the fringe-dwarf gradient.

  20. Exploring the nitrogen ingestion of aphids--a new method using electrical penetration graph and (15)N labelling.

    PubMed

    Kuhlmann, Franziska; Opitz, Sebastian E W; Inselsbacher, Erich; Ganeteg, Ulrika; Näsholm, Torgny; Ninkovic, Velemir

    2013-01-01

    Studying plant-aphid interactions is challenging as aphid feeding is a complex process hidden in the plant tissue. Here we propose a combination of two well established methods to study nutrient acquisition by aphids focusing on the uptake of isotopically labelled nitrogen ((15)N). We combined the Electrical Penetration Graph (EPG) technique that allows detailed recording of aphid feeding behaviour and stable isotope ratio mass spectrometry (IRMS) to precisely measure the uptake of nitrogen. Bird cherry-oat aphids Rhopalosiphum padi L. (Hemiptera, Aphididae) fed for 24 h on barley plants (Hordeum vulgare L., cultivar Lina, Poaceae) that were cultivated with a (15)N enriched nutrient solution. The time aphids fed in the phloem was strongly positive correlated with their (15)N uptake. All other single behavioural phases were not correlated with (15)N enrichment in the aphids, which corroborates their classification as non-feeding EPG phases. In addition, phloem-feeding and (15)N enrichment of aphids was divided into two groups. One group spent only short time in the phloem phase and was unsuccessful in nitrogen acquisition, while the other group displayed longer phloem-feeding phases and was successful in nitrogen acquisition. This suggests that several factors such as the right feeding site, time span of feeding and individual conditions play a role for the aphids to acquire nutrients successfully. The power of this combination of methods for studying plant-aphid interactions is discussed.

  1. Range expansion of the jumbo squid in the NE Pacific: δ15N decrypts multiple origins, migration and habitat use.

    PubMed

    Ruiz-Cooley, Rocio I; Ballance, Lisa T; McCarthy, Matthew D

    2013-01-01

    Coincident with climate shifts and anthropogenic perturbations, the highly voracious jumbo squid Dosidicus gigas reached unprecedented northern latitudes along the NE Pacific margin post 1997-98. The physical or biological drivers of this expansion, as well as its ecological consequences remain unknown. Here, novel analysis from both bulk tissues and individual amino acids (Phenylalanine; Phe and Glutamic acid; Glu) in both gladii and muscle of D. gigas captured in the Northern California Current System (NCCS) documents for the first time multiple geographic origins and migration. Phe δ(15)N values, a proxy for habitat baseline δ(15)N values, confirm at least three different geographic origins that were initially detected by highly variable bulk δ(15)N values in gladii for squid at small sizes (<30 cm gladii length). In contrast, bulk δ(15)N values from gladii of large squid (>60 cm) converged, indicating feeding in a common ecosystem. The strong latitudinal gradient in Phe δ(15)N values from composite muscle samples further confirmed residency at a point in time for large squid in the NCCS. These results contrast with previous ideas, and indicate that small squid are highly migratory, move into the NCCS from two or more distinct geographic origins, and use this ecosystem mainly for feeding. These results represent the first direct information on the origins, immigration and habitat use of this key "invasive" predator in the NCCS, with wide implications for understanding both the mechanisms of periodic D. gigas population range expansions, and effects on ecosystem trophic structure.

  2. Sources of nitrate in the Arno River waters: Constraints from d15N and d18O

    USGS Publications Warehouse

    Nisi, Barbara; Vaselli, Orlando; Buccianti, Antonella; Silva, Steven R.

    2005-01-01

    Running waters in anthropogenically affected areas are susceptible to nitrate contamination. Source identification is a fundamental step for the development of effective remediation. Previous studies pointed to pollution by nitrogen-bearing contaminants in the Arno Basin. In this paper, eleven surface water samples have been analysed for main and trace components and 15N/14N and 18O/16O ratios, with the aim of identifying for the first time the origin of nitrate in the Arno River Basin so that further investigations can appropriately be designed. d18O(NO3)and d15N(NO3) values have allowed to hypothesise the main sources of nitrate, as follows: i) mineralized fertilizer, ii) soil-organic nitrogen, iii) manure and septic waste. The anomalously high d15N and d18O values in the Chiana (d15N=24.9‰ and d18O=15.5‰) and Usciana tributaries (d15N=30.1‰ and d18O=7.2‰) show a low probability of belonging to the same population as that of the other samples and can be related to denitrification process of nitrate from animal waste/sewage and/or an industrial process (e.g. tanneries).

  3. Resonance capture and Saturn's rings

    SciTech Connect

    Patterson, C.W.

    1986-05-01

    We have assigned the resonances apparently responsible for the stabilization of the Saturn's shepherd satellites and for the substructure seen in the F-ring and the ringlets in the C-ring. We show that Saturn's narrow ringlets have a substructure determined by three-body resonances with Saturn's ringmoons and the sun. We believe such resonances have important implications to satellite formation. 17 refs., 1 fig., 1 tab.

  4. Resonances and resonance widths

    SciTech Connect

    Collins, T.

    1986-05-01

    Two-dimensional betatron resonances are much more important than their simple one-dimensional counterparts and exhibit a strong dependence on the betatron phase advance per cell. A practical definition of ''width'' is expanded upon in order to display these relations in tables. A primarily pedagogical introduction is given to explain the tables, and also to encourage a wider capability for deriving resonance behavior and wider use of ''designer'' resonances.

  5. Choice of dietary protein of vegetarians and omnivores is reflected in their hair protein 13C and 15N abundance.

    PubMed

    Petzke, Klaus J; Boeing, Heiner; Metges, Cornelia C

    2005-01-01

    Stable isotopic (15N, 13C) composition of tissues depends on isotopic pattern of food sources. We investigated whether the isotopic compositions of human hair protein and amino acids reflect the habitual dietary protein intake. Hair samples were analyzed from 100 omnivores (selected randomly out of the 1987-1988 German nutrition survey VERA), and from 15 ovo-lacto-vegetarians (OLV), and from 6 vegans recruited separately. Hair bulk and amino acid specific isotopic compositions were analyzed by isotope-ratio mass spectrometry (EA/IRMS and GC/C/IRMS, respectively) and the results were correlated with data of the 7 day dietary records. Hair bulk 15N and 13C abundances clearly reflect the particular eating habits. Vegans can be distinguished from OLV and both are significantly distinct from omnivores in both 15N and 13C abundances. 15N and 13C abundances rose with a higher proportion of animal to total protein intake (PAPI). Individual proportions of animal protein consumption (IPAP) were calculated using isotopic abundances and a linear regression model using animal protein consumption data of vegans (PAPI = 0) and omnivores (mean PAPI = 0.639). IPAP values positively correlated with the intake of protein, meat, meat products, and animal protein. Distinct patterns for hair amino acid specific 15N and 13C abundances were measured but with lower resolution between food preference groups compared with bulk values. In conclusion, hair 13C and 15N values both reflected the extent of animal protein consumption. Bulk isotopic abundance of hair can be tested for future use in the validation of dietary assessment methods.

  6. Organic vs. conventional grassland management: do (15)N and (13)C isotopic signatures of hay and soil samples differ?

    PubMed

    Klaus, Valentin H; Hölzel, Norbert; Prati, Daniel; Schmitt, Barbara; Schöning, Ingo; Schrumpf, Marion; Fischer, Markus; Kleinebecker, Till

    2013-01-01

    Distinguishing organic and conventional products is a major issue of food security and authenticity. Previous studies successfully used stable isotopes to separate organic and conventional products, but up to now, this approach was not tested for organic grassland hay and soil. Moreover, isotopic abundances could be a powerful tool to elucidate differences in ecosystem functioning and driving mechanisms of element cycling in organic and conventional management systems. Here, we studied the δ(15)N and δ(13)C isotopic composition of soil and hay samples of 21 organic and 34 conventional grasslands in two German regions. We also used Δδ(15)N (δ(15)N plant - δ(15)N soil) to characterize nitrogen dynamics. In order to detect temporal trends, isotopic abundances in organic grasslands were related to the time since certification. Furthermore, discriminant analysis was used to test whether the respective management type can be deduced from observed isotopic abundances. Isotopic analyses revealed no significant differences in δ(13)C in hay and δ(15)N in both soil and hay between management types, but showed that δ(13)C abundances were significantly lower in soil of organic compared to conventional grasslands. Δδ(15)N values implied that management types did not substantially differ in nitrogen cycling. Only δ(13)C in soil and hay showed significant negative relationships with the time since certification. Thus, our result suggest that organic grasslands suffered less from drought stress compared to conventional grasslands most likely due to a benefit of higher plant species richness, as previously shown by manipulative biodiversity experiments. Finally, it was possible to correctly classify about two third of the samples according to their management using isotopic abundances in soil and hay. However, as more than half of the organic samples were incorrectly classified, we infer that more research is needed to improve this approach before it can be efficiently

  7. Towards interpreting nitrate-δ15N records in ice cores in terms of nitrogen oxide sources

    NASA Astrophysics Data System (ADS)

    Hastings, M. G.; Buffen, A. M.

    2011-12-01

    The isotopic composition of nitrate preserved in ice cores offers unique potential for reconstructing past contributions of nitrogen oxides (NOx = NO and NO2) to the atmosphere. Sources of NOx imprint a nitrogen stable isotopic (δ15N) signature, which can be conserved during subsequent oxidation to form nitrate. Major sources of NOx include fossil fuels combustion, biomass burning, microbial processes in soils, and lightning, and thus a quantitative tracer of emissions would help detail connections between the atmosphere, the biosphere, and climate. Unfortunately, the δ15N signatures of most NOx sources are not yet well enough constrained to allow for quantitative partitioning, though new methodology for directly collecting NOx for isotopic analysis is promising (Fibiger and Hastings, A43D-0265, AGU 2010). Still, a growing network of ice core δ15N records may offer insight into source signatures, as different sources are important to different regions of the world. For example, a 300-year ice core record of nitrate-δ15N from Summit, Greenland shows a clear and significant 12% (vs. N2) decrease since the Preindustrial that reflects emissions from fossil fuel combustion and/or soils related to changing agricultural practices in North America and Europe. Over the same time period, Antarctic ice cores show no such trend in δ15N. This would be consistent with previous work suggesting that biomass burning and/or stratospheric intrusion of NOx produced from N2O oxidation are dominant sources for nitrate formation at high southern latitudes. In comparison to the polar records, nitrate in tropical ice cores should represent more significant inputs from lightning, microbial processes in soils, and biomass burning. This may be reflected in new results from a high-elevation site in the Peruvian Andes that shows strong seasonal δ15N cycles of up to 15% (vs. N2). We compare and contrast these records in an effort to evaluate the contribution of NOx sources to nitrate over

  8. [Dynamics of 15N-isobutylidene diurea (IBDU) in sheep. 1. IBDU conversion in the digestive tract].

    PubMed

    Görsch, R; Bergner, H; Adam, K

    1978-07-01

    Four Merino Landrace wethers averaging 47.6 kg body weight were adapted to a semi-synthetic diet containing as the only N-source 60 g of IBDU per day. After the adaptation phase, on the 1 st experimental day the IBDU of the morning feed was given in 15N-labelled form (701 mg 15N-excess). After 2 1/2, 7 1/4, 12 and 24 hours the experimental animals were killed without having been fed again. The comparison of the IBDU-concentrations in the content of the rumen bottom with the residual rumen content did not allow to draw conclusions regarding IBDU-sedimentation at the bottom of the rumen. For the 15N-decline in the rumen content, a relationship was established following y = 76.3 - 2.62 (r = 0.96) (see fig. 2). In the order of killing times the following 15N-IBDU amounts were retrieved (% of intake): I = 15.6%, II = 24.1%, III = 3.3% and IV = 3.6%. 7 1/4 hours after starting the experiment, 40% of the 15N-labelled material were found in the rumen in the form IBDU; after 12 hours it came to 10%. Except for sheep I, 15N-urea was not found but in small amounts. Only sheep I and III revealed IBDU-traces in the abomasum, but in the small intestine of all sheep 2 to 6% of the amount taken in. This fact is explained with the endogenous influx of IBDU from the blood. An additional experimental sheep provided with a ligature at the abomasum entry, revealed that IBDU is absorbed from the rumen and allowed to enter the individual segments of the intestine in small amounts.

  9. Organic vs. Conventional Grassland Management: Do 15N and 13C Isotopic Signatures of Hay and Soil Samples Differ?

    PubMed Central

    Klaus, Valentin H.; Hölzel, Norbert; Prati, Daniel; Schmitt, Barbara; Schöning, Ingo; Schrumpf, Marion; Fischer, Markus; Kleinebecker, Till

    2013-01-01

    Distinguishing organic and conventional products is a major issue of food security and authenticity. Previous studies successfully used stable isotopes to separate organic and conventional products, but up to now, this approach was not tested for organic grassland hay and soil. Moreover, isotopic abundances could be a powerful tool to elucidate differences in ecosystem functioning and driving mechanisms of element cycling in organic and conventional management systems. Here, we studied the δ15N and δ13C isotopic composition of soil and hay samples of 21 organic and 34 conventional grasslands in two German regions. We also used Δδ15N15N plant - δ15N soil) to characterize nitrogen dynamics. In order to detect temporal trends, isotopic abundances in organic grasslands were related to the time since certification. Furthermore, discriminant analysis was used to test whether the respective management type can be deduced from observed isotopic abundances. Isotopic analyses revealed no significant differences in δ13C in hay and δ15N in both soil and hay between management types, but showed that δ13C abundances were significantly lower in soil of organic compared to conventional grasslands. Δδ15N values implied that management types did not substantially differ in nitrogen cycling. Only δ13C in soil and hay showed significant negative relationships with the time since certification. Thus, our result suggest that organic grasslands suffered less from drought stress compared to conventional grasslands most likely due to a benefit of higher plant species richness, as previously shown by manipulative biodiversity experiments. Finally, it was possible to correctly classify about two third of the samples according to their management using isotopic abundances in soil and hay. However, as more than half of the organic samples were incorrectly classified, we infer that more research is needed to improve this approach before it can be efficiently used in practice

  10. Nitrate removal in stream ecosystems measured by 15N addition experiments: 2. Denitrification

    SciTech Connect

    Mulholland, Patrick J; Hall, Robert; Sobota, Daniel; Dodds, Walter; Findlay, Stuart; Grimm, Nancy; Hamilton, Stephen; McDowell, William; O'Brien, Jon; Tank, Jennifer; Ashkenas, Linda; Cooper, Lee W; Dahm, Cliff; Gregory, Stanley; Johnson, Sherri; Meyer, Judy; Peterson, Bruce; Poole, Geoff; Valett, H. Maurice; Webster, Jackson; Arango, Clay; Beaulieu, Jake; Bernot, Melody; Burgin, Amy; Crenshaw, Chelsea; Helton, Ashley; Johnson, Laura; Niederlehner, Bobbie; Potter, Jody; Sheibley, Rich; Thomas, Suzanne

    2009-01-01

    We measured denitrification rates using a field {sup 15}N-NO{sub 3}{sup -} tracer-addition approach in a large, cross-site study of nitrate uptake in reference, agricultural, and suburban-urban streams. We measured denitrification rates in 49 of 72 streams studied. Uptake length due to denitrification (S{sub Wden}) ranged from 89 m to 184 km (median of 9050 m) and there were no significant differences among regions or land-use categories, likely because of the wide range of conditions within each region and land use. N{sub 2} production rates far exceeded N{sub 2}O production rates in all streams. The fraction of total NO{sub 3}{sup -} removal from water due to denitrification ranged from 0.5% to 100% among streams (median of 16%), and was related to NH{sub 4}{sup +} concentration and ecosystem respiration rate (ER). Multivariate approaches showed that the most important factors controlling S{sub Wden} were specific discharge (discharge/width) and NO{sub 3}{sup -} concentration (positive effects), and ER and transient storage zones (negative effects). The relationship between areal denitrification rate (U{sub den}) and NO{sub 3}{sup -} concentration indicated a partial saturation effect. A power function with an exponent of 0.5 described this relationship better than a Michaelis-Menten equation. Although U{sub den} increased with increasing NO{sub 3}{sup -} concentration, the efficiency of NO{sub 3}{sup -} removal from water via denitrification declined, resulting in a smaller proportion of streamwater NO{sub 3}{sup -} load removed over a given length of stream. Regional differences in stream denitrification rates were small relative to the proximate factors of NO{sub 3}{sup -} concentration and ecosystem respiration rate, and land use was an important but indirect control on denitrification in streams, primarily via its effect on NO{sub 3}{sup -} concentration.

  11. Determination of δ18O and δ15N in Nitrate

    USGS Publications Warehouse

    Revesz, K.; Böhlke, J.K.; Yoshinari, T.

    1997-01-01

    The analyses of both O and N isotopic compositions of nitrate have many potential applications in studies of nitrate sources and reactions in hydrology, oceanography, and atmospheric chemistry, but simple and precise methods for these analyses have yet to be developed. Testing of a new method involving reaction of potassium nitrate with catalyzed graphite (C + Pd + Au) at 520 °C resulted in quantitative recovery of N and O from nitrate as free CO2, K2CO3, and N2. The δ18O values of nitrate reference materials were obtained by analyzing both the CO2 and K2CO3 from catalyzed graphite combustion. Provisional values of δ18OVSMOW for the internationally distributed KNO3 reference materials IAEA-N3 and USGS-32 were both equal to +22.7 ± 0.5‰. Because the fraction of free CO2 and the isotopic fractionation factor between CO2 and K2CO3 were constant in the combustion products, the δ18O value of KNO3 could be calculated from measurements of the δ18O of free CO2. Thus, δ18OKNO3 = aδ18Ofree CO2 − b, where a and b were equal to 0.9967 and 3.3, respectively, for the specific conditions of the experiments. The catalyzed graphite combustion method can be used to determine δ18O of KNO3 from measurements of δ18O of free CO2 with reproducibility on the order of ±0.2‰ or better if local reference materials are prepared and analyzed with the samples. Reproducibility of δ15N was ±0.1‰ after trace amounts of CO were removed.

  12. Nitrate removal in stream ecosystems measured by 15N addition experiments: Denitrification

    USGS Publications Warehouse

    Mulholland, P.J.; Hall, R.O.; Sobota, D.J.; Dodds, W.K.; Findlay, S.E.G.; Grimm, N. B.; Hamilton, S.K.; McDowell, W.H.; O'Brien, J. M.; Tank, J.L.; Ashkenas, L.R.; Cooper, L.W.; Dahm, Clifford N.; Gregory, S.V.; Johnson, S.L.; Meyer, J.L.; Peterson, B.J.; Poole, G.C.; Valett, H.M.; Webster, J.R.; Arango, C.P.; Beaulieu, J.J.; Bernot, M.J.; Burgin, A.J.; Crenshaw, C.L.; Helton, A.M.; Johnson, L.T.; Niederlehner, B.R.; Potter, J.D.; Sheibley, R.W.; Thomasn, S.M.

    2009-01-01

    We measured denitrification rates using a field 15N-NO- 3 tracer-addition approach in a large, cross-site study of nitrate uptake in reference, agricultural, and suburban-urban streams. We measured denitrification rates in 49 of 72 streams studied. Uptake length due to denitrification (SWden) ranged from 89 m to 184 km (median of 9050 m) and there were no significant differences among regions or land-use categories, likely because of the wide range of conditions within each region and land use. N2 production rates far exceeded N2O production rates in all streams. The fraction of total NO-3 removal from water due to denitrification ranged from 0.5% to 100% among streams (median of 16%), and was related to NHz 4 concentration and ecosystem respiration rate (ER). Multivariate approaches showed that the most important factors controlling SWden were specific discharge (discharge / width) and NO-3 concentration (positive effects), and ER and transient storage zones (negative effects). The relationship between areal denitrification rate (Uden) and NO- 3 concentration indicated a partial saturation effect. A power function with an exponent of 0.5 described this relationship better than a Michaelis-Menten equation. Although Uden increased with increasing NO- 3 concentration, the efficiency of NO-3 removal from water via denitrification declined, resulting in a smaller proportion of streamwater NO-3 load removed over a given length of stream. Regional differences in stream denitrification rates were small relative to the proximate factors of NO-3 concentration and ecosystem respiration rate, and land use was an important but indirect control on denitrification in streams, primarily via its effect on NO-3 concentration. ?? 2009.

  13. Multi-Isotopic (δ2H, δ13C, δ15N) Tracing of Molt Origin for Red-Winged Blackbirds Associated with Agro-Ecosystems

    PubMed Central

    Werner, Scott J.; Hobson, Keith A.; Van Wilgenburg, Steven L.; Fischer, Justin W.

    2016-01-01

    We analyzed stable-hydrogen (δ2H), carbon (δ13C) and nitrogen (δ 15N) isotope ratios in feathers to better understand the molt origin and food habits of Red-winged Blackbirds (Agelaius phoeniceus) near sunflower production in the Upper Midwest and rice production in the Mid-South of the United States. Outer primary feathers were used from 661 after-second-year (ASY) male blackbirds collected in Minnesota, Montana, North Dakota and South Dakota (spring collection), and Arkansas, Louisiana, Mississippi, Missouri and Texas (winter collection). The best-fit model indicated that the combination of feather δ2H, δ13C and δ15N best predicted the state of sample collections and thus supported the use of this approach for tracing molt origins in Red-winged Blackbirds. When considering only birds collected in spring, 56% of birds were classified to their collection state on the basis of δ2H and δ13C alone. We then developed feather isoscapes for δ13C based upon these data and for δ2H based upon continental patterns of δ2H in precipitation. We used 81 birds collected at the ten independent sites for model validation. The spatially-explicit assignment of these 81 birds to the δ2H isoscape resulted in relatively high rates (~77%) of accurate assignment to collection states. We also modeled the spatial extent of C3 (e.g. rice, sunflower) and C4 (corn, millet, sorghum) agricultural crops grown throughout the Upper Midwest and Mid-South United States to predict the relative use of C3- versus C4-based foodwebs among sampled blackbirds. Estimates of C3 inputs to diet ranged from 50% in Arkansas to 27% in Minnesota. As a novel contribution to blackbird conservation and management, we demonstrate how such feather isoscapes can be used to predict the molt origin and interstate movements of migratory blackbirds for subsequent investigations of breeding biology (e.g. sex-specific philopatry), agricultural depredation, feeding ecology, physiology of migration and sensitivity to

  14. Multi-Isotopic (δ2H, δ13C, δ15N) Tracing of Molt Origin for Red-Winged Blackbirds Associated with Agro-Ecosystems.

    PubMed

    Werner, Scott J; Hobson, Keith A; Van Wilgenburg, Steven L; Fischer, Justin W

    2016-01-01

    We analyzed stable-hydrogen (δ2H), carbon (δ13C) and nitrogen (δ 15N) isotope ratios in feathers to better understand the molt origin and food habits of Red-winged Blackbirds (Agelaius phoeniceus) near sunflower production in the Upper Midwest and rice production in the Mid-South of the United States. Outer primary feathers were used from 661 after-second-year (ASY) male blackbirds collected in Minnesota, Montana, North Dakota and South Dakota (spring collection), and Arkansas, Louisiana, Mississippi, Missouri and Texas (winter collection). The best-fit model indicated that the combination of feather δ2H, δ13C and δ15N best predicted the state of sample collections and thus supported the use of this approach for tracing molt origins in Red-winged Blackbirds. When considering only birds collected in spring, 56% of birds were classified to their collection state on the basis of δ2H and δ13C alone. We then developed feather isoscapes for δ13C based upon these data and for δ2H based upon continental patterns of δ2H in precipitation. We used 81 birds collected at the ten independent sites for model validation. The spatially-explicit assignment of these 81 birds to the δ2H isoscape resulted in relatively high rates (~77%) of accurate assignment to collection states. We also modeled the spatial extent of C3 (e.g. rice, sunflower) and C4 (corn, millet, sorghum) agricultural crops grown throughout the Upper Midwest and Mid-South United States to predict the relative use of C3- versus C4-based foodwebs among sampled blackbirds. Estimates of C3 inputs to diet ranged from 50% in Arkansas to 27% in Minnesota. As a novel contribution to blackbird conservation and management, we demonstrate how such feather isoscapes can be used to predict the molt origin and interstate movements of migratory blackbirds for subsequent investigations of breeding biology (e.g. sex-specific philopatry), agricultural depredation, feeding ecology, physiology of migration and sensitivity to

  15. Potential for assessing long-term dynamics in soil nitrogen availability from variations in delta15N of tree rings.

    PubMed

    Hart, S C; Classen, A T

    2003-03-01

    Numerous researchers have used the isotopic signatures of C, H, and O in tree rings to provide a long-term record of changes in the physiological status, climate, or water-source use of trees. The frequently limiting element N is also found in tree rings, and variation in its isotopic signature may provide insight into long-term changes in soil N availability of a site. However, research has suggested that N is readily translocated among tree ring of different years; such infidelity between the isotopic compositions of the N taken up from the soil and the N contained in the ring of that growth year would obscure the long-term N isotopic record. We used a 15-year 15N-tracer study to assess the degree of N translocation among tree rings in ponderosa pine (Pinus ponderosa) trees growing in a young, mixed-conifer plantation. We also measured delta13C and delta15N values in unlabeled trees to assess the degree of their covariance in wood tissue, and to explore the potential for a biological linkage between them. We found that the maximum delta15N values in rings from the labeled trees occurred in the ring formed one-year after the 15N was applied to the roots. The delta15N value of rings from labeled trees declined exponentially and bidirectionally from this maximum peak, toward younger and older rings. The unlabeled trees showed considerable interannual variation in the delta15N values of their rings (up to 3 and 5 per thousand), but these values correlated poorly between trees over time and differed by as much as 6 per thousand. Removal of extractives from the wood reduced their delta15N value, but the change was fairly small and consistent among unlabeled trees. The delta13C and delta15N values of tree rings were correlated over time in only one of the unlabeled trees. Across all trees, both delta13C values of tree rings and annual stem wood production were well correlated with annual precipitation, suggesting that soil water balance is an important environmental

  16. Compound specific amino acid δ15N in marine sediments: A new approach for studies of the marine nitrogen cycle

    NASA Astrophysics Data System (ADS)

    Batista, Fabian C.; Ravelo, A. Christina; Crusius, John; Casso, Michael A.; McCarthy, Matthew D.

    2014-10-01

    The nitrogen (N) isotopic composition (δ15N) of bulk sedimentary N (δ15Nbulk) is a common tool for studying past biogeochemical cycling in the paleoceanographic record. Empirical evidence suggests that natural fluctuations in the δ15N of surface nutrient N are reflected in the δ15N of exported planktonic biomass and in sedimentary δ15Nbulk. However, δ15Nbulk is an analysis of total combustible sedimentary N, and therefore also includes mixtures of N sources and/or selective removal or preservation of N-containing compounds. Compound-specific nitrogen isotope analyses of individual amino acids (δ15NAA) are novel measurements with the potential to decouple δ15N changes in nutrient N from trophic effects, two main processes that can influence δ15Nbulk records. As a proof of concept study to examine how δ15NAA can be applied in marine sedimentary systems, we compare the δ15NAA signatures of surface and sinking POM sources with shallow surface sediments from the Santa Barbara Basin, a sub-oxic depositional environmental that exhibits excellent preservation of sedimentary organic matter. Our results demonstrate that δ15NAA signatures of both planktonic biomass and sinking POM are well preserved in such surface sediments. However, we also observed an unexpected inverse correlation between δ15N value of phenylalanine (δ15NPhe; the best AA proxy for N isotopic value at the base of the food web) and calculated trophic position. We used a simple N isotope mass balance model to confirm that over long time scales, δ15NPhe values should in fact be directly dependent on shifts in ecosystem trophic position. While this result may appear incongruent with current applications of δ15NAA in food webs, it is consistent with expectations that paleoarchives will integrate N dynamics over much longer timescales. We therefore propose that for paleoceanographic applications, key δ15NAA parameters are ecosystem trophic position, which determines relative partitioning of 15N

  17. Nitrogen mineralization from selected /sup 15/N-labelled crop residues and humus as affected by inorganic nitrogen

    SciTech Connect

    Santos, J.A.

    1987-01-01

    The use of cover crops or crop residues as a source of N to succeeding crops has become a matter of increasing importance for economic and environmental reason. Greenhouse and field studies were conducted to determine the N contribution of four /sup 15/N labelled crop residues, rye (Secale cereale L.), wheat (Triticum aestivum L.), crimson clover (Trifolium encarnatum L.), and hairy vetch (Vicia sativa L.), to successive crops and to evaluate the effect of different organic (ON) and inorganic N (IN) combinations on mineralization of the above residues. Total /sup 15/N recovery from the residues ranged from 51% to 85% and 4% to 74% for the greenhouse and field studies, respectively.

  18. Expert systems for personnel assignment

    SciTech Connect

    Hardee, J.L.; Liepins, G.

    1986-01-01

    In order to reduce stress on assignment personnel (detailers) and ensure maximum fairness and consistency in the Navy's personnel assignment process, The Navy Military Personnel Command (NMPC) has begun to explore the potential use of expert systems to supplement current manual and computerized distribution methods. The Detailer's Assistant expert system is being developed to improve the detailers' ability to satisfy the needs of their constituents and Navy management. An initial prototype of the Detailer's Assistant is now being evaluated. Numerous upgrades and extensions should lead to an operational system in the near future. Further development to a production system will involve additional research in machine learning, intelligent database methods, and cooperating expert systems.

  19. Using natural 15N abundances to trace the fate of waste-derived nitrogen in forest ecosystems: New Zealand case studies.

    PubMed

    Wang, H; Magesan, G N; Clinton, P W; Lavery, J M

    2005-03-01

    Treatment of wastewater generally results in elevated natural 15N abundance (delta15N) in the effluent and sludges. For example, high delta15N values are found in treated sewage effluent, biosolids, and other wastes that are commonly applied to land. In contrast, N deficient coniferous forest soils usually have a low delta15N. When wastes with high delta15N values are applied to land, their distinctive delta15N signature can potentially be used to trace the fate of waste-derived N in the ecosystem. In this paper, we provide an overview of the use of delta15N in land application of wastes, including New Zealand case studies on tracing nitrogen in forest ecosystems.

  20. CARBON-RICH PRESOLAR GRAINS FROM MASSIVE STARS: SUBSOLAR {sup 12}C/{sup 13}C AND {sup 14}N/{sup 15}N RATIOS AND THE MYSTERY OF {sup 15}N

    SciTech Connect

    Pignatari, M.; Zinner, E.; Hoppe, P.; Jordan, C. J.; Gibson, B. K.; Trappitsch, R.; Herwig, F.; Fryer, C.; Hirschi, R.; Timmes, F. X.

    2015-08-01

    Carbon-rich grains with isotopic anomalies compared to the Sun are found in primitive meteorites. They were made by stars, and carry the original stellar nucleosynthesis signature. Silicon carbide grains of Type X and C and low-density (LD) graphites condensed in the ejecta of core-collapse supernovae. We present a new set of models for the explosive He shell and compare them with the grains showing {sup 12}C/{sup 13}C and {sup 14}N/{sup 15}N ratios lower than solar. In the stellar progenitor H was ingested into the He shell and not fully destroyed before the explosion. Different explosion energies and H concentrations are considered. If the supernova shock hits the He-shell region with some H still present, the models can reproduce the C and N isotopic signatures in C-rich grains. Hot-CNO cycle isotopic signatures are obtained, including a large production of {sup 13}C and {sup 15}N. The short-lived radionuclides {sup 22}Na and {sup 26}Al are increased by orders of magnitude. The production of radiogenic {sup 22}Ne from the decay of {sup 22}Na in the He shell might solve the puzzle of the Ne-E(L) component in LD graphite grains. This scenario is attractive for the SiC grains of type AB with {sup 14}N/{sup 15}N ratios lower than solar, and provides an alternative solution for SiC grains originally classified as nova grains. Finally, this process may contribute to the production of {sup 14}N and {sup 15}N in the Galaxy, helping to produce the {sup 14}N/{sup 15}N ratio in the solar system.

  1. Assessing Contributions to Group Assignments

    ERIC Educational Resources Information Center

    Johnston, Lucy; Miles, Lynden

    2004-01-01

    We report the use of a combination of self- and peer-assessment in an undergraduate social psychology laboratory course. Students worked in small groups on a self-directed empirical project that they each wrote up independently as a laboratory report. Marks for the written assignment were moderated by a contribution index measure based on the…

  2. The Year of Secret Assignments

    ERIC Educational Resources Information Center

    Moriarty, Jaclyn

    2004-01-01

    The path to "novelist" was a convoluted one for Moriarty, who began writing fiction as doctoral student at Cambridge University. Her interest in young adults stems from an appreciation for the "troubles, strengths, and surprises of that age group." Now, in a uniquely formatted book titled "The Year of Secret Assignments," we peek inside the mind…

  3. Radiocarbon, 13C and 15N analysis of fossil bone: Removal of humates with XAD-2 resin

    NASA Astrophysics Data System (ADS)

    Stafford, Thomas W., Jr.; Brendel, Klaus; Duhamel, Raymond C.

    1988-09-01

    Humic acids are the predominant source of error in the 14C and stable isotope analysis of fossil bone organic matter. XAD-2 resin will quantitatively remove humates and give the highest yields of protein from bones with variable types of preservation. Decalcified bone, gelatin and base-leached residues can vary up to 5%. for δ 13C and by 1%. on δ 15N relative to XAD-treated fractions. Simultaneous analysis of 14C age, δ 13C and δ 15N is recommended because each isotope value can be independently affected by the bone's diagenetic history. Radiocarbon analysis is the most sensitive and δ 15N is least sensitive for detecting exogenous organic matter. The uncertainty of analyses on the best pretreated protein is ±0.5%. for both δ 13C and δ 15N and is larger than previous estimates. The accuracy for all isotope analyses will be better assessed by using individual amino acids instead of total collagenous residues. Inaccurate 14C dates on severely degraded bone are an indication that this class of fossils may be unsuitable for any isotopic analysis.

  4. Determination of the natural abundance δ15N of taurine by gas chromatography-isotope ratio measurement mass spectrometry.

    PubMed

    Tea, Illa; Antheaume, Ingrid; Besnard, Jorick; Robins, Richard J

    2010-12-15

    The measurement of the nitrogen isotope ratio of taurine (2-aminoethanesulphonic acid) in biological samples has a large number of potential applications. Taurine is a small water-soluble molecule which is notoriously difficult to analyze due to its polarity and functionality. A method is described which allows the determination of the natural abundance δ(15)N values of taurine and structural analogues, such as 3-amino-1-propanesulphonic acid (APSA), by isotope ratio mass spectrometry interfaced to gas chromatography (GC-irm-MS). The one-step protocol exploits the simultaneous derivatization of both functionalities of these aminosulphonic acids by reaction with triethylorthoacetate (TEOA). Conditions have been established which ensure quantitative reaction thus avoiding any nitrogen isotope fractionation during derivatization and workup. The differences in the δ(15)N values of derivatized and non-derivatized taurine and APSA all fall within the working range of 0.4‰ (-0.02 to 0.39‰). When applied to four sources of taurine with various δ(15)N values, the method achieved excellent reproducibility and accuracy. The optimized method enables the determination of the natural abundance δ(15)N values of taurine over the concentration range 1.5-7.84 µmol.mL(-1) in samples of biological origin.

  5. Using macroalgal δ15N bioassay to detect cruise ship waste water effluent inputs in Skagway, AK

    EPA Science Inventory

    Nitrogen stable isotopes are a powerful tool for tracking sources of N to marine ecosystems. I used green macroalgae as a bioassay organism to evaluate if the δ15N signature of cruise ship waste water effluent (CSWWE) could be detected in Skagway Harbor, AK. Opportunistic green...

  6. STATISTICAL ESTIMATES OF VARIANCE FOR 15N ISOTOPE DILUTION MEASUREMENTS OF GROSS RATES OF NITROGEN CYCLE PROCESSES

    EPA Science Inventory

    It has been fifty years since Kirkham and Bartholmew (1954) presented the conceptual framework and derived the mathematical equations that formed the basis of the now commonly employed method of 15N isotope dilution. Although many advances in methodology and analysis have been ma...

  7. Contrasting food web linkages for the grazing pathway in 3 temperate forested streams using {sup 15}N as a tracer

    SciTech Connect

    Tank, J.L.; Mulholland, P.J.; Meyer, J.L.; Bowden, W.B.; Webster, J.R.; Peterson, B.J.

    1998-11-01

    Nitrogen is a critical element controlling the productivity and dynamics of stream ecosystems and many streams are limited by the supply of biologically available nitrogen. The authors are learning more about the fate of inorganic nitrogen entering streams through {sup 15}N tracer additions. The Lotic Intersite Nitrogen Experiment (LINX) is studying the uptake, cycling, and fate of {sup 15}N-NH{sub 4} in the stream food web of 10 streams draining different biomes. Using the {sup 15}N tracer method and data from 3 sites in the study, the authors can differentiate patterns in the cycling of nitrogen through the grazing pathway (N from the epilithon to grazing macroinvertebrates) for 3 temperate forested streams. Here, they quantify the relationship between the dominant grazer and its proposed food resource, the epilithon, by comparing {sup 15}N levels of grazers with those of the epilithon, as well as the biomass, nitrogen content, and chlorophyll a standing stocks of the epilithon in 3 streams.

  8. Plant community change mediates the response of foliar delta15N to CO2 enrichment in mesic grasslands

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Rising atmospheric CO2 concentration may change the isotopic signature of plant N by altering plant and microbial processes involved in the N cycle. Isotope fractionation theory and limited experimental evidence indicate that CO2 may increase leaf delta15N by increasing plant community productivity,...

  9. The influence of fish cage culture on δ13C and δ15N of filter-feeding Bivalvia (Mollusca).

    PubMed

    Benedito, E; Figueroa, L; Takeda, A M; Manetta, G I

    2013-11-01

    The objective of this study was to evaluate the effect of Oreochromis niloticus cage culture promoted variations in the δ13C and δ15N in Corbicula fluminea (Mollusca; Bivalvia) and in the sediment of an aquatic food web. Samples were taken before and after net cage installation in the Rosana Reservoir (Paranapanema River, PR-SP). Samples of specimens of the bivalve filterer C. fluminea and samples of sediment were collected using a modified Petersen grab. All samples were dried in an oven (60 °C) for 72 hours, macerated to obtain homogenous fine powders and sent for carbon (δ13C) and nitrogen (δ15N) isotopic value analysis in a mass spectrometer. There were significant differences in the δ13C and δ15N values of the invertebrate C. fluminea between the beginning and the end of the experiment. There were no differences between the δ13C and δ15N values of sediment. These results indicate that the installation of fish cage culture promoted impacts in the isotopic composition of the aquatic food web organisms, which could exert influence over the native species and the ecosystem.

  10. Variation in hair δ13C and δ15N values in long-tailed macaques (Macaca fascicularis) from Singapore

    USGS Publications Warehouse

    Schillaci, Michael A.; Castellini, J. Margaret; Stricker, Craig A.; Jones-Engel, Lisa; Lee, Benjamin P.Y.-H.

    2014-01-01

    Much of the primatology literature on stable isotope ratios of carbon (δ13C) and nitrogen (δ15N) has focused on African and New World species, with comparatively little research published on Asian primates. Here we present hair δ13C and δ15N isotope values for a sample of 33 long-tailed macaques from Singapore. We evaluate the suggestion by a previous researcher that forest degradation and biodiversity loss in Singapore have led to a decline in macaque trophic level. The results of our analysis indicated significant spatial variability in δ13C but not δ15N. The range of variation in δ13C was consistent with a diet based on C3 resources, with one group exhibiting low values consistent with a closed canopy environment. Relative to other macaque species from Europe and Asia, the macaques from Singapore exhibited a low mean δ13C value but mid-range mean δ15N value. Previous research suggesting a decline in macaque trophic level is not supported by the results of our study.

  11. Plant δ15N Correlates with the Transpiration Efficiency of Nitrogen Acquisition in Tropical Trees1[OA

    PubMed Central

    Cernusak, Lucas A.; Winter, Klaus; Turner, Benjamin L.

    2009-01-01

    Based upon considerations of a theoretical model of 15N/14N fractionation during steady-state nitrate uptake from soil, we hypothesized that, for plants grown in a common soil environment, whole-plant δ15N (δP) should vary as a function of the transpiration efficiency of nitrogen acquisition (FN/v) and the difference between δP and root δ15N (δP − δR). We tested these hypotheses with measurements of several tropical tree and liana species. Consistent with theoretical expectations, both FN/v and δP − δR were significant sources of variation in δP, and the relationship between δP and FN/v differed between non-N2-fixing and N2-fixing species. We interpret the correlation between δP and FN/v as resulting from variation in mineral nitrogen efflux-to-influx ratios across plasma membranes of root cells. These results provide a simple explanation of variation in δ15N of terrestrial plants and have implications for understanding nitrogen cycling in ecosystems. PMID:19726571

  12. Proton-decoupled CPMG: a better experiment for measuring (15)N R2 relaxation in disordered proteins.

    PubMed

    Yuwen, Tairan; Skrynnikov, Nikolai R

    2014-04-01

    (15)N R2 relaxation is one of the most informative experiments for characterization of intrinsically disordered proteins (IDPs). Small changes in nitrogen R2 rates are often used to determine how IDPs respond to various biologically relevant perturbations such as point mutations, posttranslational modifications and weak ligand interactions. However collecting high-quality (15)N relaxation data can be difficult. Of necessity, the samples of IDPs are often prepared with low protein concentration and the measurement time can be limited because of rapid sample degradation. Furthermore, due to hardware limitations standard experiments such as (15)N spin-lock and CPMG can sample the relaxation decay only to ca. 150ms. This is much shorter than (15)N T2 times in disordered proteins at or near physiological temperature. As a result, the sampling of relaxation decay profiles in these experiments is suboptimal, which further lowers the precision of the measurements. Here we report a new implementation of the proton-decoupled (PD) CPMG experiment which allows one to sample (15)N R2 relaxation decay up to ca. 0.5-1s. The new experiment has been validated through comparison with the well-established spin-lock measurement. Using dilute samples of denatured ubiquitin, we have demonstrated that PD-CPMG produces up to 3-fold improvement in the precision of the data. It is expected that for intrinsically disordered proteins the gains may be even more substantial. We have also shown that this sequence has a number of favorable properties: (i) the spectra are recorded with narrow linewidth in nitrogen dimension; (ii) (15)N offset correction is small and easy to calculate; (iii) the experiment is immune to various spurious effects arising from solvent exchange; (iv) the results are stable with respect to pulse miscalibration and rf field inhomogeneity; (v) with minimal change, the pulse sequence can also be used to measure R2 relaxation of (15)N(ε) spins in arginine side chains. We

  13. The Titan 14N/ 15N and 12C/ 13C isotopic ratios in HCN from Cassini/CIRS

    NASA Astrophysics Data System (ADS)

    Vinatier, Sandrine; Bézard, Bruno; Nixon, Conor A.

    2007-11-01

    We report the detection of H 13CN and HC 15N in mid-infrared spectra recorded by the Composite Infrared Spectrometer (CIRS) aboard Cassini, along with the determination of the 12C/ 13C and 14N/ 15N isotopic ratios. We analyzed two sets of limb spectra recorded near 13-15° S (Tb flyby) and 83° N (T4 flyby) at 0.5 cm -1 resolution. The spectral range 1210-1310 cm -1 was used to retrieve the temperature profile in the range 145-490 km at 13° S and 165-300 km at 83° N. These two temperature profiles were then incorporated in the atmospheric model to retrieve the abundance profile of H 12C 14N, H 13CN and HC 15N from their bands at 713, 706 and 711 cm -1, respectively. The HCN abundance profile was retrieved in the range 90-460 km at 15° S and 165-305 km at 83° N. There is no evidence for vertical variations of the isotopic ratios. Constraining the isotopic abundance profiles to be proportional to the HCN one, we find C12/C13=89-18+22 at 15° S, and 68-12+16 at 83° N, two values that are statistically consistent. A combination of these results yields a 12C/ 13C value equal to 75±12. This global result, as well as the 15° S one, envelop the value in Titan's methane ( 82.3±1) [Niemann, H.B., and 17 colleagues, 2005. Nature 438, 779-784] measured at 10° S and is slightly lower than the terrestrial inorganic standard value (89). The 14N/ 15N isotopic ratio is found equal to 56-13+16 at 15° S and 56-9+10 at 83° N. Combining the two values yields 14N/ 15N = 56 ± 8, which corresponds to an enrichment in 15N of about 4.9 compared with the terrestrial ratio. These results agree with the values obtained from previous ground-based millimeter observations [Hidayat, T., Marten, A., Bézard, B., Gautier, D., Owen, T., Matthews, H.E., Paubert, G., 1997. Icarus 126, 170-182; Marten, A., Hidayat, T., Biraud, Y., Moreno, R., 2002. Icarus 158, 532-544]. The 15N/ 14N ratio found in HCN is ˜3 times higher than in N 2 [Niemann, H.B., and 17 colleagues, 2005. Nature 438, 779

  14. Measuring denitrification after grassland renewal and grassland conversion to cropland by using the 15N gas-flux method

    NASA Astrophysics Data System (ADS)

    Buchen, Caroline; Eschenbach, Wolfram; Flessa, Heinz; Giesemann, Anette; Lewicka-Szczebak, Dominika; Well, Reinhard

    2015-04-01

    Denitrification, the reduction of oxidized forms of inorganic N to N2O and N2 is an important pathway of gaseous nitrogen losses. Measuring denitrification, especially the reduction of N2O to N2, expressed in the product ratio (N2O/(N2O + N2)), is rather difficult and hence rarely performed under field conditions. But using the 15N gas-flux method allows determining N transformation processes in their natural environment. In order to develop effective climate mitigation strategies understanding the N2O source is essential. We used the 15N gas-flux method to determine N2O and N2 emissions following grassland renewal and conversion techniques. Therefore we selected three different treatments: control (C), mechanical grassland renovation (GR) (autumn 2013) and grassland conversion to maize (GM) (spring 2014) from field plot trials on two different sites (Histic Gleysoil and Plaggic Anthrosol) near Oldenburg, Lower Saxony, Germany. We applied 15N labeled KNO3- (60 atom. % 15N) at a rate equivalent to common farming practices (150 kg N*ha-1) using needle injection of fertilizer solution in three different depths (10 cm, 15 cm, 20 cm) for homogeneous soil labeling up to 30 cm in microplots. During the first 10 days after application (May 2014) gas flux measurements from closed chambers were performed every second day and then weekly following a period of 8 weeks. Gas samples were analyzed for δ15N of N2 and N2O by IRMS according to Lewicka-Szczebak et al. (2013). Concentration and 15N enrichment of NO3- in soil water was determined on weekly samples using the SPIN-MAS technique (Stange et al. 2007). Fluxes of N2 and N2O evolved from the 15N labeled soil nitrogen pool were calculated using the equations of Spott et al. (2006). Peak events of N2 and N2O emissions occurred during the first 10 days of measurement, showing differences in soil types, as well as treatment variations. N2 fluxes up to 178 g*ha-1*day-1 and N2O fluxes up to 280 g*ha-1*day-1 were measured on the

  15. Oceanic δ15N biogeography: a novel top-down approach to examine nutrient dynamics in the equatorial Pacific Ocean

    NASA Astrophysics Data System (ADS)

    Graham, B. S.; Fry, B.; Popp, B. N.; Allain, V.; Olson, R.; Galvan, F.

    2010-12-01

    By mapping the δ15N and δ13C values of three top-level pelagic predators, yellowfin (Thunnus albacares), bigeye (T. obesus), and skipjack (Katsuwonus pelamis) tuna throughout the equatorial Pacific Ocean, we demonstrated systematic geographic isotopic variation (up to ~12‰ for the δ15N values) that reflect nutrient dynamics that occur at the base of the food web. Remarkably the variation observed in the δ15N values of the tunas is geographically similar to δ15N values previously reported in surface particulate organic matter and deep-sea sediments in the tropical Pacific. We discuss the mechanisms occurring at the base of the food web that could produce the spatial variability observed in tropical tuna δ15N values. We present a simple Rayleigh fractionation model that can explain much of the spatial structure. We also discuss the temporal stability in the isotopic compositions at the base and top of the food web. Overall, this nitrogen isotope cartography or “isoscapes” suggests nitrogen is tightly retained in the marine food web, up to the top predators, and that the uptake of nitrate from the equatorial upwelling zone, denitrification in the oxygen minimum zones, and nitrogen fixation at the base of the food web play major roles in the observed geographical variation. In addition to providing insight into the nutrient dynamics of the open ocean, these predator isoscapes can begin to be used to characterize regional residency in tropical tunas, which is important for the successful management of tuna fisheries.

  16. Variability and directionality of temporal changes in δ(13)C and δ (15)N of aquatic invertebrate primary consumers.

    PubMed

    Woodland, Ryan J; Magnan, Pierre; Glémet, Hélène; Rodríguez, Marco A; Cabana, Gilbert

    2012-05-01

    Seasonal oscillations in the carbon (δ(13)C) and nitrogen (δ(15)N) isotope signatures of aquatic algae can cause seasonal enrichment-depletion cycles in the isotopic composition of planktonic invertebrates (e.g., copepods). Yet, there is growing evidence that seasonal enrichment-depletion cycles also occur in the isotope signatures of larger invertebrate consumers, taxa used to define reference points in isotope-based trophic models (e.g., trophic baselines). To evaluate the general assumption of temporal stability in non-zooplankton aquatic invertebrates, δ(13)C and δ(15)N time series data from the literature were analyzed for seasonality and the influence of biotic (feeding group) and abiotic (trophic state, climate regime) factors on isotope temporal patterns. The amplitude of δ(13)C and δ(15)N enrichment-depletion cycles was negatively related to body size, although all size-classes of invertebrates displayed a winter-to-summer enrichment in δ(13)C and depletion in δ(15)N. Among feeding groups, periphytic grazers were more variable and displayed larger temporal changes in δ(13)C than detritivores. For nitrogen, temporal variability and magnitude of directional change of δ(15)N was most strongly related to ecosystem trophic state (eutrophic > mesotrophic, oligotrophic). This study provides evidence of seasonality in the isotopic composition of aquatic invertebrates across very broad geographical and ecological gradients as well as identifying factors that are likely to modulate the strength and variability of seasonality. These results emphasize the need for researchers to recognize the likelihood of temporal changes in non-zooplankton aquatic invertebrate consumers at time scales relevant to seasonal studies and, if present, to account for temporal dynamics in isotope trophic models.

  17. Range Expansion of the Jumbo Squid in the NE Pacific: δ15N Decrypts Multiple Origins, Migration and Habitat Use

    PubMed Central

    Ruiz-Cooley, Rocio I.; Ballance, Lisa T.; McCarthy, Matthew D.

    2013-01-01

    Coincident with climate shifts and anthropogenic perturbations, the highly voracious jumbo squid Dosidicus gigas reached unprecedented northern latitudes along the NE Pacific margin post 1997–98. The physical or biological drivers of this expansion, as well as its ecological consequences remain unknown. Here, novel analysis from both bulk tissues and individual amino acids (Phenylalanine; Phe and Glutamic acid; Glu) in both gladii and muscle of D. gigas captured in the Northern California Current System (NCCS) documents for the first time multiple geographic origins and migration. Phe δ15N values, a proxy for habitat baseline δ15N values, confirm at least three different geographic origins that were initially detected by highly variable bulk δ15N values in gladii for squid at small sizes (<30 cm gladii length). In contrast, bulk δ15N values from gladii of large squid (>60 cm) converged, indicating feeding in a common ecosystem. The strong latitudinal gradient in Phe δ15N values from composite muscle samples further confirmed residency at a point in time for large squid in the NCCS. These results contrast with previous ideas, and indicate that small squid are highly migratory, move into the NCCS from two or more distinct geographic origins, and use this ecosystem mainly for feeding. These results represent the first direct information on the origins, immigration and habitat use of this key “invasive” predator in the NCCS, with wide implications for understanding both the mechanisms of periodic D. gigas population range expansions, and effects on ecosystem trophic structure. PMID:23527242

  18. Production of 15N-Labelled Liquid Organic Fertilisers Based on Manure and Crop Residue for Use in Fertigation Studies.

    PubMed

    Martínez-Alcántara, Belén; Martínez-Cuenca, Mary-Rus; Fernández, Carlos; Legaz, Francisco; Quiñones, Ana

    2016-01-01

    Large quantities of crop residue and animal manure from agricultural and livestock activities are annually produced worldwide. With proper management, these residues are potentially valuable sources of plant nutrients, mainly N. Recycling such subproducts in sustainably-based agricultural systems can minimise the use of mineral fertilisers, and hence reduce the potential risk of surface and groundwater pollution. Therefore, the purpose of this study was to obtain (small scale) two liquid labelled-organic fertilisers, an animal- and a vegetal-based organic (AO and VO, respectively) fertiliser, to be used as organic N sources in subsequent fertigation studies. Forage maize (Zea mays L.) grown under 15N-labelled fertiliser supply was used as raw material for VO fertiliser production, and also as 15N-labelled sheep feed to obtain 15N-labelled manure. The labelled faeces fraction was used as raw material for the AO fertiliser. The VO fertiliser was obtained after an acidic and an enzyme-driven hydrolysis. The AO fertiliser was obtained after acidic hydrolysis. The VO liquid fertiliser presented an N concentration of 330 mg·L-1, 85% of total N was organic, while ammonium and nitrate N accounted for 55% and 45% of the mineral nitrogen fraction, respectively. This fertiliser also exhibited high K, Ca and S concentrations and notable values for the remaining macro- and micronutrients. The AO liquid fertiliser had a similar total N concentration (496 mg·L-1, 82% of total N in an organic form) to that of VO, but its mineral N fraction significantly differed, which came in a predominantly (95%) ammonia form. It also had a high content of N, P, K and other macronutrients, and sufficient Fe, Zn, Mn, Cu and B levels, which suggests its suitability as a potential fertiliser. The percentage of 15N enrichment in both VO and AO liquid fertilisers exceeded 2% 15N atom excess, which enabled their use in subsequent assays run to assess nitrogen uptake efficiency.

  19. Retrospective characterization of ontogenetic shifts in killer whale diets via δ13C and δ15N analysis of teeth

    USGS Publications Warehouse

    Newsome, Seth D.; Etnier, Michael A.; Monson, Daniel H.; Fogel, Marilyn L.

    2009-01-01

    Metabolically inert, accretionary structures such as the dentin growth layers in teeth provide a life history record of individual diet with near-annual resolution. We constructed ontogenetic δ13C and δ15N profiles by analyzing tooth dentin growth layers from 13 individual killer whales Orcinus orca collected in the eastern northeast Pacific Ocean between 1961 and 2003. The individuals sampled were 6 to 52 yr old, representing 2 ecotypes—resident and transient—collected across ~25° of latitude. The average isotopic values of transient individuals (n = 10) are consistent with a reliance on mammalian prey, while the average isotopic values of residents (n = 3) are consistent with piscivory. Regardless of ecotype, most individuals show a decrease in δ15N values of ~2.5‰ through the first 3 yr of life, roughly equivalent to a decrease of one trophic level. We interpret this as evidence of gradual weaning, after which, ontogenetic shifts in isotopic values are highly variable. A few individuals (n = 2) maintained relatively stable δ15N and δ13C values throughout the remainder of their lives, whereas δ15N values of most (n = 11) increased by ~1.5‰, suggestive of an ontogenetic increase in trophic level. Significant differences in mean δ13C and δ15N values among transients collected off California suggest that individuality in prey preferences may be prevalent within this ecotype. Our approach provides retrospective individual life history and dietary information that cannot be obtained through traditional field observations of free-ranging and elusive species such as killer whales, including unique historic ecological information that pre-dates modern studies. By providing insights into individual diet composition, stable isotope analysis of teeth and/or bones may be the only means of evaluating a number of hypothesized historical dietary shifts in killer whales of the northeast Pacific Ocean

  20. Retrospective characterization of ontogenetic shifts in killer whale diets via δ13C and δ15N analysis of teeth

    USGS Publications Warehouse

    Newsome, Seth D.; Etnier, Michael A.; Monson, Daniel H.; Fogel, Marilyn L.

    2009-01-01

    Metabolically inert, accretionary structures such as the dentin growth layers in teeth provide a life history record of individual diet with near-annual resolution. We constructed ontogenetic ??13C and ??15N profiles by analyzing tooth dentin growth layers from 13 individual killer whales Orcinus orca collected in the eastern northeast Pacific Ocean between 1961 and 2003. The individuals sampled were 6 to 52 yr old, representing 2 ecotypes-resident and transient - collected across ???25?? of latitude. The average isotopic values of transient individuals (n = 10) are consistent with a reliance on mammalian prey, while the average isotopic values of residents (n = 3) are consistent with piscivory. Regardless of ecotype, most individuals show a decrease in ??15N values of ???2.5% through the first 3 yr of life, roughly equivalent to a decrease of one trophic level. We interpret this as evidence of gradual weaning, after which, ontogenetic shifts in isotopic values are highly variable. A few individuals (n = 2) maintained relatively stable ??15N and ??13C values throughout the remainder of their lives, whereas ??15N values of most (n = 11) increased by ???1.5%, suggestive of an ontogenetic increase in trophic level. Significant differences in mean ??13C and ??15N values among transients collected off California suggest that individuality in prey preferences may be prevalent within this ecotype. Our approach provides retrospective individual life history and dietary information that cannot be obtained through traditional field observations of free-ranging and elusive species such as killer whales, including unique historic ecological information that pre-dates modern studies. By providing insights into individual diet composition, stable isotope analysis of teeth and/or bones may be the only means of evaluating a number of hypothesized historical dietary shifts in killer whales of the northeast Pacific Ocean. ?? Inter-Research 2009.

  1. Coordination Environment of a Site-Bound Metal Ion in the Hammerhead Ribozyme Determined by 15N and 2H ESEEM Spectroscopy

    PubMed Central

    Vogt, Matthew; Lahiri, Simanti; Hoogstraten, Charles G.; Britt, R. David; DeRose, Victoria J.

    2010-01-01

    Although site-bound Mg2+ ions have been proposed to influence RNA structure and function, establishing the molecular properties of such sites has been challenging due largely to the unique electrostatic properties of the RNA biopolymer. We have previously determined that, in solution, the hammerhead ribozyme (a self-cleaving RNA) has a high-affinity metal ion binding site characterized by a Kd,app < 10 µM for Mn2+ in 1 M NaCl and speculated that this site has functional importance in the ribozyme cleavage reaction. Here we determine both the precise location and the hydration level of Mn2+ in this site using ESEEM (electron spin–echo envelope modulation) spectroscopy. Definitive assignment of the high-affinity site to the activity-sensitive A9/G10.1 region is achieved by site-specific labeling of G10.1 with 15N guanine. The coordinated metal ion retains four water ligands as measured by 2H ESEEM spectroscopy. The results presented here show that a functionally important, specific metal binding site is uniquely populated in the hammerhead ribozyme even in a background of high ionic strength. Although it has a relatively high thermodynamic affinity, this ion remains partially hydrated and is chelated to the RNA by just two ligands. PMID:17177426

  2. 15N, 17O NMR and X-ray diffraction study of mesoionic 1,2,3,4-thiatriazolium-5-olate and its ethylated derivative

    NASA Astrophysics Data System (ADS)

    Jaźiwińsk, J.; Staszewska, O.; Staszewski, P.; Stefaniak, L.; Wiench, J. W.; Webb, G. A.

    1999-02-01

    Two mesoionic compounds with oxygenous exocyclic groups: 3-phenyl-1,2,3,4-thiatriazolium-5-olate 1 and its ethylated derivative 2 were investigated by means of 15N, 17O NMR and X-ray diffraction techniques. The exocyclic C5-O6 bond of thiatriazole 1 [1.224(3) Å] has a strong double bond character. Bond lengths in the thiatriazole ring are intermediate between single and double bond values except for S1-C5 [1.800(2) Å] which is close to a single Csp 3-S bond. The C5-O6 bond is significantly longer for the ethylated derivative 2 [1.314(4) Å]. The ethyl group attached to O6 is located in the trans position in relation to the ring S1 atom. The experimental data are compared with the results of ab initio molecular orbital calculations. The calculated absolute nuclear shieldings, chemical shifts and charge densities, in spite of some limitations, can be useful as an aid to signal assignments and for an understanding of the NMR parameters.

  3. Automatic NOESY assignment in CS-RASREC-Rosetta.

    PubMed

    Lange, Oliver F

    2014-07-01

    We have developed an approach for simultaneous structure calculation and automatic Nuclear Overhauser Effect (NOE) assignment to solve nuclear magnetic resonance (NMR) structures from unassigned NOESY data. The approach, autoNOE-Rosetta, integrates Resolution Adapted Structural RECombination (RASREC) Rosetta NMR calculations with algorithms for automatic NOE assignment. The method was applied to two proteins in the 15-20 kDa size range for which both, NMR and X-ray data, is available. The autoNOE-Rosetta calculations converge for both proteins and yield accurate structures with an RMSD of 1.9 Å to the X-ray reference structures. The method greatly expands the radius of convergence for automatic NOE assignment, and should be broadly useful for NMR structure determination.

  4. Selective excitation for spectral editing and assignment in separated local field experiments of oriented membrane proteins

    NASA Astrophysics Data System (ADS)

    Koroloff, Sophie N.; Nevzorov, Alexander A.

    2017-01-01

    Spectroscopic assignment of NMR spectra for oriented uniformly labeled membrane proteins embedded in their native-like bilayer environment is essential for their structure determination. However, sequence-specific assignment in oriented-sample (OS) NMR is often complicated by insufficient resolution and spectral crowding. Therefore, the assignment process is usually done by a laborious and expensive "shotgun" method involving multiple selective labeling of amino acid residues. Presented here is a strategy to overcome poor spectral resolution in crowded regions of 2D spectra by selecting resolved "seed" residues via soft Gaussian pulses inserted into spin-exchange separated local-field experiments. The Gaussian pulse places the selected polarization along the z-axis while dephasing the other signals before the evolution of the 1H-15N dipolar couplings. The transfer of magnetization is accomplished via mismatched Hartmann-Hahn conditions to the nearest-neighbor peaks via the proton bath. By optimizing the length and amplitude of the Gaussian pulse, one can also achieve a phase inversion of the closest peaks, thus providing an additional phase contrast. From the superposition of the selective spin-exchanged SAMPI4 onto the fully excited SAMPI4 spectrum, the 15N sites that are directly adjacent to the selectively excited residues can be easily identified, thereby providing a straightforward method for initiating the assignment process in oriented membrane proteins.

  5. Fleet Assignment Using Collective Intelligence

    NASA Technical Reports Server (NTRS)

    Antoine, Nicolas E.; Bieniawski, Stefan R.; Kroo, Ilan M.; Wolpert, David H.

    2004-01-01

    Product distribution theory is a new collective intelligence-based framework for analyzing and controlling distributed systems. Its usefulness in distributed stochastic optimization is illustrated here through an airline fleet assignment problem. This problem involves the allocation of aircraft to a set of flights legs in order to meet passenger demand, while satisfying a variety of linear and non-linear constraints. Over the course of the day, the routing of each aircraft is determined in order to minimize the number of required flights for a given fleet. The associated flow continuity and aircraft count constraints have led researchers to focus on obtaining quasi-optimal solutions, especially at larger scales. In this paper, the authors propose the application of this new stochastic optimization algorithm to a non-linear objective cold start fleet assignment problem. Results show that the optimizer can successfully solve such highly-constrained problems (130 variables, 184 constraints).

  6. Fleet Assignment Using Collective Intelligence

    NASA Technical Reports Server (NTRS)

    Antoine, Nicolas E.; Bieniawski, Stefan R.; Kroo, Ilan M.; Wolpert, David H.

    2004-01-01

    Airline fleet assignment involves the allocation of aircraft to a set of flights legs in order to meet passenger demand, while satisfying a variety of constraints. Over the course of the day, the routing of each aircraft is determined in order to minimize the number of required flights for a given fleet. The associated flow continuity and aircraft count constraints have led researchers to focus on obtaining quasi-optimal solutions, especially at larger scales. In this paper, the authors propose the application of an agent-based integer optimization algorithm to a "cold start" fleet assignment problem. Results show that the optimizer can successfully solve such highly- constrained problems (129 variables, 184 constraints).

  7. Model Refinement Using Eigensystem Assignment

    NASA Technical Reports Server (NTRS)

    Maghami, Peiman G.

    2000-01-01

    IA novel approach for the refinement of finite-element-based analytical models of flexible structures is presented. The proposed approach models the possible refinements in the mass, damping, and stiffness matrices of the finite element model in the form of a constant gain feedback with acceleration, velocity, and displacement measurements, respectively. Once the free elements of the structural matrices have been defined, the problem of model refinement reduces to obtaining position, velocity, and acceleration gain matrices with appropriate sparsity that reassign a desired subset of the eigenvalues of the model, along with partial mode shapes, from their baseline values to those obtained from system identification test data. A sequential procedure is used to assign one conjugate pair of eigenvalues at each step using symmetric output feedback gain matrices, and the eigenvectors are partially assigned, while ensuring that the eigenvalues assigned in the previous steps are not disturbed. The procedure can also impose that gain matrices be dissipative to guarantee the stability of the refined model. A numerical example, involving finite element model refinement for a structural testbed at NASA Langley Research Center (Controls-Structures-Interaction Evolutionary model) is presented to demonstrate the feasibility of the proposed approach.

  8. Proton-decoupled CPMG: A better experiment for measuring 15N R2 relaxation in disordered proteins

    NASA Astrophysics Data System (ADS)

    Yuwen, Tairan; Skrynnikov, Nikolai R.

    2014-04-01

    15N R2 relaxation is one of the most informative experiments for characterization of intrinsically disordered proteins (IDPs). Small changes in nitrogen R2 rates are often used to determine how IDPs respond to various biologically relevant perturbations such as point mutations, posttranslational modifications and weak ligand interactions. However collecting high-quality 15N relaxation data can be difficult. Of necessity, the samples of IDPs are often prepared with low protein concentration and the measurement time can be limited because of rapid sample degradation. Furthermore, due to hardware limitations standard experiments such as 15N spin-lock and CPMG can sample the relaxation decay only to ca. 150 ms. This is much shorter than 15N T2 times in disordered proteins at or near physiological temperature. As a result, the sampling of relaxation decay profiles in these experiments is suboptimal, which further lowers the precision of the measurements. Here we report a new implementation of the proton-decoupled (PD) CPMG experiment which allows one to sample 15N R2 relaxation decay up to ca. 0.5-1 s. The new experiment has been validated through comparison with the well-established spin-lock measurement. Using dilute samples of denatured ubiquitin, we have demonstrated that PD-CPMG produces up to 3-fold improvement in the precision of the data. It is expected that for intrinsically disordered proteins the gains may be even more substantial. We have also shown that this sequence has a number of favorable properties: (i) the spectra are recorded with narrow linewidth in nitrogen dimension; (ii) 15N offset correction is small and easy to calculate; (iii) the experiment is immune to various spurious effects arising from solvent exchange; (iv) the results are stable with respect to pulse miscalibration and rf field inhomogeneity; (v) with minimal change, the pulse sequence can also be used to measure R2 relaxation of 15Nε spins in arginine side chains. We anticipate that

  9. Simple approach for the preparation of 15−15N2-enriched water for nitrogen fixation assessments: evaluation, application and recommendations

    PubMed Central

    Klawonn, Isabell; Lavik, Gaute; Böning, Philipp; Marchant, Hannah K.; Dekaezemacker, Julien; Mohr, Wiebke; Ploug, Helle

    2015-01-01

    Recent findings revealed that the commonly used 15N2 tracer assay for the determination of dinitrogen (N2) fixation can underestimate the activity of aquatic N2-fixing organisms. Therefore, a modification to the method using pre-prepared 15−15N2-enriched water was proposed. Here, we present a rigorous assessment and outline a simple procedure for the preparation of 15−15N2-enriched water. We recommend to fill sterile-filtered water into serum bottles and to add 15−15N2 gas to the water in amounts exceeding the standard N2 solubility, followed by vigorous agitation (vortex mixing ≥ 5 min). Optionally, water can be degassed at low-pressure (≥950 mbar) for 10 min prior to the 15−15N2 gas addition to indirectly enhance the 15−15N2 concentration. This preparation of 15−15N2-enriched water can be done within 1 h using standard laboratory equipment. The final 15N-atom% excess was 5% after replacing 2–5% of the incubation volume with 15−15N2-enriched water. Notably, the addition of 15−15N2-enriched water can alter levels of trace elements in the incubation water due to the contact of 15−15N2-enriched water with glass, plastic and rubber ware. In our tests, levels of trace elements (Fe, P, Mn, Mo, Cu, Zn) increased by up to 0.1 nmol L−1 in the final incubation volume, which may bias rate measurements in regions where N2 fixation is limited by trace elements. For these regions, we tested an alternative way to enrich water with 15−15N2. The 15−15N2 was injected as a bubble directly to the incubation water, followed by gentle shaking. Immediately thereafter, the bubble was replaced with water to stop the 15−15N2 equilibration. This approach achieved a 15N-atom% excess of 6.6 ± 1.7% when adding 2 mL 15−15N2 per liter of incubation water. The herein presented methodological tests offer guidelines for the 15N2 tracer assay and thus, are crucial to circumvent methodological draw-backs for future N2 fixation assessments. PMID:26300853

  10. Semantic Gender Assignment Regularities in German

    ERIC Educational Resources Information Center

    Schwichtenberg, Beate; Schiller, Niels O.

    2004-01-01

    Gender assignment relates to a native speaker's knowledge of the structure of the gender system of his/her language, allowing the speaker to select the appropriate gender for each noun. Whereas categorical assignment rules and exceptional gender assignment are well investigated, assignment regularities, i.e., tendencies in the gender distribution…

  11. Stereospecific assignment of the asparagine and glutamine sidechain amide protons in proteins from chemical shift analysis.

    PubMed

    Harsch, Tobias; Schneider, Philipp; Kieninger, Bärbel; Donaubauer, Harald; Kalbitzer, Hans Robert

    2017-02-01

    Side chain amide protons of asparagine and glutamine residues in random-coil peptides are characterized by large chemical shift differences and can be stereospecifically assigned on the basis of their chemical shift values only. The bimodal chemical shift distributions stored in the biological magnetic resonance data bank (BMRB) do not allow such an assignment. However, an analysis of the BMRB shows, that a substantial part of all stored stereospecific assignments is not correct. We show here that in most cases stereospecific assignment can also be done for folded proteins using an unbiased artificial chemical shift data base (UACSB). For a separation of the chemical shifts of the two amide resonance lines with differences ≥0.40 ppm for asparagine and differences ≥0.42 ppm for glutamine, the downfield shifted resonance lines can be assigned to H(δ21) and H(ε21), respectively, at a confidence level >95%. A classifier derived from UASCB can also be used to correct the BMRB data. The program tool AssignmentChecker implemented in AUREMOL calculates the Bayesian probability for a given stereospecific assignment and automatically corrects the assignments for a given list of chemical shifts.

  12. High-resolution laser spectroscopy and magnetic effect of the B̃(2)E(')←X̃(2)A2(') transition of the (15)N substituted nitrate radical.

    PubMed

    Tada, Kohei; Teramoto, Kanon; Ishiwata, Takashi; Hirota, Eizi; Kasahara, Shunji

    2015-03-21

    Rotationally resolved high-resolution fluorescence excitation spectra of the 0-0 band of the B̃(2)E(')←X̃(2)A2(') transition of the (15)N substituted nitrate radical were observed for the first time, by crossing a jet-cooled molecular beam and a single-mode dye laser beam at right angles. Several thousand rotational lines were detected in the 15 080-15 103 cm(-1) region. We observed the Zeeman splitting of intense lines up to 360 G in order to obtain secure rotational assignment. Two, nine, and seven rotational line pairs with 0.0248 cm(-1) spacing were assigned to the transitions from the X̃(2)A2(') (υ″ = 0, k″ = 0, N″ = 1, J″ = 0.5 and 1.5) to the (2)E3/2(') (J' = 1.5), (2)E1/2(') (J' = 0.5), and (2)E1/2(') (J' = 1.5) levels, respectively, based on the ground state combination differences and the Zeeman splitting patterns. The observed spectrum was complicated due to the vibronic coupling between the bright B̃(2)E(') (υ = 0) state and surrounding dark vibronic states. Some series of rotational lines other than those from the X̃(2)A2(') (J = 0.5 and 1.5) levels were also assigned by the ground state combination differences and the observed Zeeman splitting. The rotational branch structures were identified, and the molecular constants of the B̃(2)E1/2(') (υ = 0) state were estimated by a deperturbed analysis to be T0 = 15 098.20(4) cm(-1), B = 0.4282(7) cm(-1), and DJ = 4 × 10(-4) cm(-1). In the observed region, both the (2)E1/2(') and (2)E3/2(') spin-orbit components were identified, and the spin-orbit interaction constant of the B̃(2)E(') (υ = 0) state was estimated to be -12 cm(-1) as the lower limit.

  13. Target-specific NMR detection of protein-ligand interactions with antibody-relayed (15)N-group selective STD.

    PubMed

    Hetényi, Anasztázia; Hegedűs, Zsófia; Fajka-Boja, Roberta; Monostori, Éva; Kövér, Katalin E; Martinek, Tamás A

    2016-12-01

    Fragment-based drug design has been successfully applied to challenging targets where the detection of the weak protein-ligand interactions is a key element. (1)H saturation transfer difference (STD) NMR spectroscopy is a powerful technique for this work but it requires pure homogeneous proteins as targets. Monoclonal antibody (mAb)-relayed (15)N-GS STD spectroscopy has been developed to resolve the problem of protein mixtures and impure proteins. A (15)N-labelled target-specific mAb is selectively irradiated and the saturation is relayed through the target to the ligand. Tests on the anti-Gal-1 mAb/Gal-1/lactose system showed that the approach is experimentally feasible in a reasonable time frame. This method allows detection and identification of binding molecules directly from a protein mixture in a multicomponent system.

  14. Solving the woolly mammoth conundrum: amino acid 15N-enrichment suggests a distinct forage or habitat

    NASA Astrophysics Data System (ADS)

    Schwartz-Narbonne, Rachel; Longstaffe, Fred J.; Metcalfe, Jessica Z.; Zazula, Grant

    2015-06-01

    Understanding woolly mammoth ecology is key to understanding Pleistocene community dynamics and evaluating the roles of human hunting and climate change in late Quaternary megafaunal extinctions. Previous isotopic studies of mammoths’ diet and physiology have been hampered by the ‘mammoth conundrum’: woolly mammoths have anomalously high collagen δ15N values, which are more similar to coeval carnivores than herbivores, and which could imply a distinct diet and (or) habitat, or a physiological adaptation. We analyzed individual amino acids from collagen of adult woolly mammoths and coeval species, and discovered greater  15N enrichment in source amino acids of woolly mammoths than in most other herbivores or carnivores. Woolly mammoths consumed an isotopically distinct food source, reflective of extreme aridity, dung fertilization, and (or) plant selection. This dietary signal suggests that woolly mammoths occupied a distinct habitat or forage niche relative to other Pleistocene herbivores.

  15. Solving the woolly mammoth conundrum: amino acid 15N-enrichment suggests a distinct forage or habitat

    PubMed Central

    Schwartz-Narbonne, Rachel; Longstaffe, Fred J.; Metcalfe, Jessica Z.; Zazula, Grant

    2015-01-01

    Understanding woolly mammoth ecology is key to understanding Pleistocene community dynamics and evaluating the roles of human hunting and climate change in late Quaternary megafaunal extinctions. Previous isotopic studies of mammoths’ diet and physiology have been hampered by the ‘mammoth conundrum’: woolly mammoths have anomalously high collagen δ15N values, which are more similar to coeval carnivores than herbivores, and which could imply a distinct diet and (or) habitat, or a physiological adaptation. We analyzed individual amino acids from collagen of adult woolly mammoths and coeval species, and discovered greater  15N enrichment in source amino acids of woolly mammoths than in most other herbivores or carnivores. Woolly mammoths consumed an isotopically distinct food source, reflective of extreme aridity, dung fertilization, and (or) plant selection. This dietary signal suggests that woolly mammoths occupied a distinct habitat or forage niche relative to other Pleistocene herbivores. PMID:26056037

  16. Quantification of the separate matrix constituents of spheroidal graphite cast iron implanted with 15N by nuclear reaction analysis using an ion muprobe

    NASA Astrophysics Data System (ADS)

    Matthews, A. P.; Jeynes, C.; Reeson, K. J.; Thornton, J.; Spyrou, N. M.

    1992-02-01

    The retained dose of nitrogen in a spheroidal graphite (SG) cast iron (4% carbon) implanted with 2 × 10 1715{solN}/{cm 2} at 200 keV has been determined separately in the iron matrix and in the graphite inclusions of 30 μm average diameter randomly dispersed in the matrix, using the 15N(p, αγ) 12C resonance at 898 keV and a proton muprobe focussed to less than 20 μm spot diameter. In normalised and tempered SG cast iron the retained doses were 1.09 × 10 17 and 1.74 × 10 17{N}/{cm}2 in the pearlitic matrix and graphite nodules, respectively, and in induction hardened SG cast iron the retained doses were 1.18 × 10 15 and 0.97 × 10 17{N}/{cm 2} in the martensitic matrix and nodules, respectively. The profile shapes are also quite different in both types of samples, and in both matrix and nodule.

  17. Fate of nitrogen deposition and decomposed nitrogen from litter in a 15N-tracer mesocosm experiment

    NASA Astrophysics Data System (ADS)

    Nair, R.; Perks, M.; Mencuccini, M.

    2013-12-01

    Atmospheric deposition of anthropogenic-derived nitrogen may be a major driver of the 0.6-0.7 Pg y-1 increase in the carbon sink in historically N-limited northern and boreal forests, but the magnitude of its effect is still uncertain. A strong effect depends on the allocation of N to trees, because of their high C:N ratio in woody tissues, and isotope tracer experiments have shown that the majority of 15N tracers applied directly to the soil are lost via leeching or retained in soil pools rather than being acquired by tree root systems. However, ambient anthropogenic inputs of N to these systems are transported in the atmosphere and intercepted by foliage before they reach the soil system, while labelled fertilization experiments also can only explicitly trace the fate of the 15N-tracer from deposition, as opposed to changes in the fate of N from litter, where decomposition rates may be enhanced at low ambient levels of deposition, affecting the availability of N from this pool for tree nutrition. We present initial results from a potted Sitka Spruce mesocosm 15N-tracer experiment where ambient nitrogen deposition was supplemented with a minor (0.4 kg ha-1 y-1) input of additional N, applied to either the soil or the foliage. Either this deposition, or litter in the pots, was enriched in 15N, allowing the fate of the isotope from two different methods of deposition to be compared with that of nitrogen released from the litter under the deposition treatment.

  18. Use of a 15N tracer to determine linkages between a mangrove and an upland freshwater swamp

    NASA Astrophysics Data System (ADS)

    MacKenzie, R. A.; Cormier, N.

    2005-05-01

    Mangrove forests and adjacent upland freshwater swamps are important components of subsistence-based economies of Pacific islands. Mangroves provide valuable firewood (Rhizophora apiculata) and mangrove crabs (Scylla serrata); intact freshwater swamps are often used for agroforestry (e.g., taro cultivation). While these two systems are connected hydrologically via groundwater and surface flows, little information is available on how they may be biogeochemically or ecologically linked. For example, mangrove leaf litter was once thought to be an important food source for resident and transient nekton and invertebrates, but this value may have been overestimated. Instead, nutrients or allochthonous material (e.g., phytoplankton, detritus) delivered via groundwater or surface water from upland freshwater swamps may play a larger role in mangrove food webs. Understanding the linkages between these two ecologically and culturally important ecosystems will help us to understand the potential impacts of hydrological alterations that occur when roads or bridges are constructed through them. We conducted a 15N tracer study in the Yela watershed on the island of Kosrae, Federated States of Micronesia. K15NO3 was continually added at trace levels for 4 weeks to the Yela River in an upland freshwater swamp adjacent to a mangrove forest. Nitrate and ammonium pools, major primary producers, macroinvertebrates, and fish were sampled from stations 5 m upstream (freshwater swamp) and 138, 188, 213, and 313 m downstream (mangrove) from the tracer addition. Samples were collected once a week prior to, during, and after the 15N addition for a total of 6 weeks. Preliminary results revealed no significant enrichment (< 1 ‰) in the 15N isotope composition of either resident shrimp (Macrobrachium sp.) or mudskipper fish (Periophthalmus sp.). However, the 15N signature of ammonium pools was enriched 10-60 ‰ by the end of the third week. These results suggest that the tracer was present

  19. 15N natural abundance during early and late succession in a middle-European dry acidic grassland.

    PubMed

    Beyschlag, W; Hanisch, S; Friedrich, S; Jentsch, A; Werner, C

    2009-09-01

    delta(15)N and total nitrogen content of above- and belowground tissues of 13 plant species from two successional stages (open pioneer community and ruderal grass stage) of a dry acidic grassland in Southern Germany were analysed, in order to evaluate whether resource use partitioning by niche separation and N input by N(2)-fixing legumes are potential determinants for species coexistence and successional changes. Within each stage, plants from plots with different legume cover were compared. Soil inorganic N content, total plant biomass and delta(15)N values of bulk plant material were significantly lower in the pioneer stage than in the ruderal grass community. The observed delta(15)N differences were rather species- than site-specific. Within both stages, there were also species-specific differences in isotopic composition between above- and belowground plant dry matter. Species-specific delta(15)N signatures may theoretically be explained by (i) isotopic fractionation during microbial-mediated soil N transformations; (ii) isotopic fractionation during plant N uptake or fractionation during plant-mycorrhiza transfer processes; (iii) differences in metabolic pathways and isotopic fractionation within the plant; or (iv) partitioning of available N resources (or pools) among plant groups or differential use of the same resources by different species, which seems to be the most probable route in the present case. A significant influence of N(2)-fixing legumes on the N balance of the surrounding plant community was not detectable. This was confirmed by the results of an independent in situ removal experiment, showing that after 3 years there were no measurable differences in the frequency distribution between plots with and without N(2)-fixing legumes.

  20. Determining the source of nitrate pollution in the Niger discontinuous aquifers using the natural {15N }/{14N } ratios

    NASA Astrophysics Data System (ADS)

    Girard, Pierre; Hillaire-Marcel, Claude

    1997-12-01

    In the semi-arid Niamey area (Niger), more than 10% of the deep wells exploiting the fracture network of the Precambrian aquifer are contaminated by nitrates, with concentrations as high as 10 meq l -1. In order to identify the source(s) of this pollution, nitrate and 15N contents in the polluted wells were monitored over a 20-month period. Potential sources of nitrate contamination were also analyzed for their 15N content. The isotopic compositions of nitrate in polluted waters were > + 12‰ and in rare cases exceeded +17‰. Latrines (˜ + 15‰) may be the major nitrate source for wells showing δ15N values above +15‰. Below this value, waters may be polluted by a combination of nitrates from both latrine and soil sources (˜ + 10‰). In some cases, the soil may account for up to 85% of the groundwater nitrate load. This mode of groundwater pollution is thought to be a consequence of deforestation. Despite their reputation as polluting agents, fertilizers ( +0.5 < δ 15N < + 3.6‰ ) which are used in rice paddies close to the contaminated areas, do not appear to be a significant source of nitrate contamination. Denitrification is probably not a significant process in the study area. Results suggest that nitrate contamination of the aquifer is a consequence of unregulated urbanization (home-made latrines) and deforestation. While latrines are limited to the urban zones, intensive cutting of the forest to meet the city dwellers' wood demand occurs in an ever increasing area around the capital, threatening the local water supply.

  1. Ntrace a 15N tracing model to analyse gross N transformations and sources of gaseous N emissions

    NASA Astrophysics Data System (ADS)

    Muller, Christoph

    2013-04-01

    Anthropogenically generated reactive nitrogen (N) cascades throughout the global environment ...(Galloway and Cowling, 2002). This reactive N may be lost from ecosystems via leaching, as nitrate (NO3-), or in gaseous forms such as ammonia and nitrous oxide (N2O) and the loss is governed by the N dynamics of the system. Thus, to be rendered environmentally benign NO3- must be reduced to a non-reactive form, dinitrogen (N2) which requires the evaluation of three major biological pathways of NO3- reduction: i) assimilatory NO3- reduction into biomass, ii) dissimilatory NO3- reduction to NH4+ (DNRA) and iii) dissimilatory NO3- reduction to N2 (denitrification) ...(Burger and Jackson, 2004). Advanced techniques based on 15N tracing in combination with suitable model analyses are the method of choice to analyse complex N interactions and simultaneous N transformation process. Techniques are based on dilution - enrichment principles and usually rely on the simultaneous labelling of various N pools. The data sets are then analysed by suitable 15N tracing models which allow the individual N transformation rates to be calculated based on realistic kinetic settings. The 15N tracing model Ntrace has been developed to analyse the simultaneously occurring N transformation rates in soil-plant systems and includes submodels for the evaluation of the processes associated with gaseous N emissions. The 15N tracing model and some typical model results will be presented. Literature cited .Burger, M., and Jackson, L. E. (2004). Plant and microbial use and turnover: rapid conversion of nitrate to ammonium in soil with roots. Plant and Soil 266, 289-301. Galloway, J. N., and Cowling, E. B. (2002). Reactive nitrogen and the world: 200 years of change. Ambio 31, 64-71. .

  2. Quantitative study on the fate of residual soil nitrate in winter wheat based on a 15N-labeling method

    PubMed Central

    Zhang, Jing-Ting; Wang, Zhi-Min; Liang, Shuang-Bo; Zhang, Ying-Hua; Lu, Lai-Qing; Wang, Run-Zheng

    2017-01-01

    A considerable amount of surplus nitrogen (N), which primarily takes the form of nitrate, accumulates in the soil profile after harvesting crops from an intensive production system in the North China Plain. The residual soil nitrate (RSN) is a key factor that is included in the N recommendation algorithm. Quantifying the utilization and losses of RSN is a fundamental necessity for optimizing crop N management, improving N use efficiency, and reducing the impact derived from farmland N losses on the environment. In this study, a 15N-labeling method was introduced to study the fate of the RSN quantitatively during the winter wheat growing season by 15N tracer technique combined with a soil column study. A soil column with a 2 m height was vertically divided into 10 20-cm layers, and the RSN in each layer was individually labeled with a 15N tracer before the wheat was sown. The results indicated that approximately 17.68% of the crop N derived from RSN was located in the 0–2 m soil profile prior to wheat sowing. The wheat recovery proportions of RSN at various layers ranged from 0.21% to 33.46%. The percentages that still remained in the soil profile after the wheat harvest ranged from 47.08% to 75.44%, and 19.46–32.64% of the RSN was unaccounted for. Upward and downward movements in the RSN were observed, and the maximum upward and downward distances were 40 cm and 100 cm, respectively. In general, the 15N-labeling method contributes to a deeper understanding of the fates of the RSN. Considering the low crop recovery of the RSN from deep soil layers, water and N saving practices should be adopted during crop production. PMID:28170440

  3. Plant δ(15) N reflects the high landscape-scale heterogeneity of soil fertility and vegetation productivity in a Mediterranean semiarid ecosystem.

    PubMed

    Ruiz-Navarro, Antonio; Barberá, Gonzalo G; Albaladejo, Juan; Querejeta, José I

    2016-12-01

    We investigated the magnitude and drivers of spatial variability in soil and plant δ(15) N across the landscape in a topographically complex semiarid ecosystem. We hypothesized that large spatial heterogeneity in water availability, soil fertility and vegetation cover would be positively linked to high local-scale variability in δ(15) N. We measured foliar δ(15) N in three dominant plant species representing contrasting plant functional types (tree, shrub, grass) and mycorrhizal association types (ectomycorrhizal or arbuscular mycorrhizal). This allowed us to investigate whether δ(15) N responds to landscape-scale environmental heterogeneity in a consistent way across species. Leaf δ(15) N varied greatly within species across the landscape and was strongly spatially correlated among co-occurring individuals of the three species. Plant δ(15) N correlated tightly with soil δ(15) N and key measures of soil fertility, water availability and vegetation productivity, including soil nitrogen (N), organic carbon (C), plant-available phosphorus (P), water-holding capacity, topographic moisture indices and normalized difference vegetation index. Multiple regression models accounted for 62-83% of within-species variation in δ(15) N across the landscape. The tight spatial coupling and interdependence of the water, N and C cycles in drylands may allow the use of leaf δ(15) N as an integrative measure of variations in moisture availability, biogeochemical activity, soil fertility and vegetation productivity (or 'site quality') across the landscape.

  4. [Detection of microbial protein synthesis in the small intestine of sheep using an intraduodenal 15N-urea infusion].

    PubMed

    Bergner, H; Bartelt, J; Kijora, C; Götz, K P

    1991-10-01

    Sheep (3 animals, 50 kg LW) with reentrant cannulas in duodenum and at the end of the ileum received 700 g hay and 800 g alfalfa pellets per animal and day. In a previous 1st period of three days duodenal digesta and in a 2nd period of four days ileal digesta were collected and stored deep frozen. In the main period the digesta flow was interrupted for 28 hours. The duodenal and ileal digesta were collected quantitatively. The previously collected duodenal and ileal digesta portions were introduced hourly. The duodenal digesta was supplemented with 15N-labelled urea for a 24 hour period. 4.5% of the introduced 15N-excess were detected at the end of the ileum in the 24 hour period. 5.6% of the 15N-excess at the end of the ileum were incorporated in bacterial protein. It was measured that the ileal digesta contained 4.62 g N in the TCE precipitable fraction and 24.4% of the TCE precipitable N-fraction was bacterial nitrogen.

  5. Symbiotic nitrogen fixation in a tropical rainforest: 15N natural abundance measurements supported by experimental isotopic enrichment.

    PubMed

    Pons, Thijs L; Perreijn, Kristel; van Kessel, Chris; Werger, Marinus J A

    2007-01-01

    * Leguminous trees are very common in the tropical rainforests of Guyana. Here, species-specific differences in N(2) fixation capability among nodulating legumes growing on different soils and a possible limitation of N(2) fixation by a relatively high nitrogen (N) and low phosphorus (P) availability in the forest were investigated. * Leaves of 17 nodulating species and 17 non-nodulating reference trees were sampled and their delta(15)N values measured. Estimates of N(2) fixation rates were calculated using the (15)N natural abundance method. Pot experiments were conducted on the effect of N and P availability on N(2) fixation using the (15)N-enriched isotope dilution method. * Nine species showed estimates of > 33% leaf N derived from N(2) fixation, while the others had low or undetectable N(2) fixation rates. High N and low P availability reduced N(2) fixation substantially. * The results suggest that a high N and low P availability in the forest limit N(2) fixation. At the forest ecosystem level, N(2) fixation was estimated at c. 6% of total N uptake by the tree community. We conclude that symbiotic N(2) fixation plays an important role in maintaining high amounts of soil available N in undisturbed forest.

  6. Tracing the diet of the monitor lizard Varanus mabitang by stable isotope analyses (δ15N, δ13C)

    NASA Astrophysics Data System (ADS)

    Struck, Ulrich; Altenbach, Alexander; Gaulke, Maren; Glaw, Frank

    2002-09-01

    In this study, we used analyses of stable isotopes of nitrogen (δ15N) and carbon (δ13C) to determine the trophic ecology of the monitor lizard Varanus mabitang. Stable isotopes from claws, gut contents, and soft tissues were measured from the type specimen. Samples from Varanus olivaceus, Varanus prasinus, Varanus salvator, the herbivorous agamid lizard Hydrosaurus pustulatus, and some plant matter were included for comparison. Our data show a rapid decrease in δ13C (about10‰) from food plants towards gut contents and soft tissues of herbivorous species. For the varanids, we found a significant linear correlation of decreasing δ13C and increasing δ15N from herbivorous towards carnivorous species. In terms of trophic isotope ecology, the type specimen of V. mabitang is a strict herbivore. Thus it differs significantly in its isotopic composition from the morphologically next closest related species V. olivaceus. The most highly carnivorous species is V. salvator, while δ15N values for V. prasinus and V. olivaceus are intermediate. Claws provide very valuable samples for such measurements, because they can be sampled from living animals without harm. Additionally, their range of variability is relatively small in comparison with measurements from soft tissues.

  7. [Absorption and distribution of nitrogen from 15N labelled urea applied at core-hardening stage in winter jujube].

    PubMed

    Zhao, Dengchao; Jiang, Yuanmao; Peng, Futian; Zhang, Jin; Zhang, Xu; Ju, Xiaotang; Zhang, Fusuo

    2006-01-01

    The study with pot experiment showed that at the rapid-swelling stage of winter jujube fruit, the percent of nitrogen derived from fertilizer (Ndff%) was the highest (10.64%) in fine roots, followed by new-growth nutritive organs. The absorbed urea-15N decreased in leaves and deciduous supers, and accumulated preferentially in root systems after harvest. The Ndff% in coarse roots was the highest (3.69%) before budding stage, while that in new-growth organs (new branches, deciduous supers, leaves and flowers) was the highest at full-blooming stage. The urea-15N applied at core-hardening stage mainly allocated in nutritive organs (leaves, deciduous supers, roots) in the first year, with the distribution rate 54.01% in root systems in winter, which was higher than that in branches (45.99%). The 15N stored in main branches changed drastically from post-harvest to budding stage. Main branches could be regarded as the 'target organs' of N storage, while coarse roots were the 'long-term sink' of N storage. The N reserve distributed preferentially in contiguity organs, and the distribution center changed with the growth and development of winter jujube in next spring.

  8. Seasonal δ13C and δ15N isoscapes of fish populations along a continental shelf trophic gradient

    NASA Astrophysics Data System (ADS)

    Radabaugh, Kara R.; Hollander, David J.; Peebles, Ernst B.

    2013-10-01

    The West Florida Shelf, located in the eastern Gulf of Mexico, transitions from a eutrophic ecosystem dominated by the Mississippi River plume to mesotrophic and oligotrophic ecosystems off the coast of peninsular Florida. Three extensive trawl surveys in this region were used to acquire samples of fish muscle, benthic algae from sea urchin stomach contents, and filtered particulate organic matter (POM) to create δ13C and δ15N isoscapes. Muscle δ15N from three widely distributed fish species, Synodus foetens (inshore lizardfish), Calamus proridens (littlehead porgy), and Syacium papillosum (dusky flounder), exhibited strong longitudinal correlations (Pearson‧s r=-0.67 to -0.90, p<0.001) that coincided with the principal trophic gradient, whereas δ13C values of fish muscle and benthic algae were correlated with depth (Pearson‧s r=-0.34 to -0.73, p<0.05). Correlations between isotopic values and surface concentrations of chlorophyll and particulate organic carbon (POC) imply linkages between the isotopic baseline and transitions from eutrophic to oligotrophic waters. The δ13C depth gradient and the δ15N longitudinal gradient were consistent between seasons and years, providing a foundation for future stable isotope studies of animal migration in the Gulf of Mexico.

  9. Bioconcentration of (15)N-tamoxifen at environmental concentration in liver, gonad and muscle of Danio rerio.

    PubMed

    Orias, Frédéric; Simon, Laurent; Mialdea, Gladys; Clair, Angéline; Brosselin, Vanessa; Perrodin, Yves

    2015-10-01

    Pharmaceutical compounds (PCs) are ubiquitous in aquatic ecosystems. In addition to the direct ecotoxicological risk presented by certain PCs, others can accumulate inside organisms and along trophic webs, subsequently contaminating whole ecosystems. We studied the bioconcentration of a bioaccumulative PC already found several times in the environment: tamoxifen. To this end, we exposed Danio rerio for 21d to (15)N-tamoxifen concentrations ranging from 0.1 to 10µg/L and used an analytic method based on stable isotopes to evaluate the tamoxifen content in these organisms. The evolution of the (15)N/(14)N ratio was thus measured in liver, muscle and gonads of exposed fish compared to control fish. We succeeded in quantifying (15)N-tamoxifen bioconcentrations at all the exposure concentrations tested. The highest bioconcentration factors of tamoxifen measured were 14,920 in muscle, 73,800 in liver and 85,600 in gonads of fish after 21d exposure at a nominal concentration of 10µg/L. However, these bioconcentration factors have to be considered as maximal values (BCFMAX). Indeed, despite its proven stability, tamoxifen can be potentially partially degraded during experiments. We now need to refine these results by using a direct analytic method (i.e. LC-MS/MS).

  10. 15N discrimination and the sensitivity of nitrogen fixation to changes in dietary nitrogen in Reticulitermes flavipes (Isoptera: Rhinotermitidae).