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Sample records for initiator protein dnaa

  1. The escherichia coli chromosome replication initiator protein, DnaA

    DEFF Research Database (Denmark)

    Nyborg, Malene

    The experimental work presented in this thesis involve mutational analysis of the DNA binding domain of the DnaA protein and analysis of the A184V substitution in the ATP area of domain III and other amino acid substitutions found in the DnaA5 and DnaA4G proteins....

  2. Multiple conformational states of DnaA protein regulate its interaction with DnaA boxes in the initiation of DNA replication.

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    Patel, Meera J; Bhatia, Lavesh; Yilmaz, Gulden; Biswas-Fiss, Esther E; Biswas, Subhasis B

    2017-09-01

    DnaA protein is the initiator of genomic DNA replication in prokaryotes. It binds to specific DNA sequences in the origin of DNA replication and unwinds small AT-rich sequences downstream for the assembly of the replisome. The mechanism of activation of DnaA that enables it to bind and organize the origin DNA and leads to replication initiation remains unclear. In this study, we have developed double-labeled fluorescent DnaA probes to analyze conformational states of DnaA protein upon binding DNA, nucleotide, and Soj sporulation protein using Fluorescence Resonance Energy Transfer (FRET). Our studies demonstrate that DnaA protein undergoes large conformational changes upon binding to substrates and there are multiple distinct conformational states that enable it to initiate DNA replication. DnaA protein adopted a relaxed conformation by expanding ~15Å upon binding ATP and DNA to form the ATP·DnaA·DNA complex. Hydrolysis of bound ATP to ADP led to a contraction of DnaA within the complex. The relaxed conformation of DnaA is likely required for the formation of the multi-protein ATP·DnaA·DNA complex. In the initiation of sporulation, Soj binding to DnaA prevented relaxation of its conformation. Soj·ADP appeared to block the activation of DnaA, suggesting a mechanism for Soj·ADP in switching initiation of DNA replication to sporulation. Our studies demonstrate that multiple conformational states of DnaA protein regulate its binding to DNA in the initiation of DNA replication. Copyright © 2017 Elsevier B.V. All rights reserved.

  3. The DnaA Cycle in Escherichia coli: Activation, Function and Inactivation of the Initiator Protein

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    Tsutomu Katayama

    2017-12-01

    Full Text Available This review summarizes the mechanisms of the initiator protein DnaA in replication initiation and its regulation in Escherichia coli. The chromosomal origin (oriC DNA is unwound by the replication initiation complex to allow loading of DnaB helicases and replisome formation. The initiation complex consists of the DnaA protein, DnaA-initiator-associating protein DiaA, integration host factor (IHF, and oriC, which contains a duplex-unwinding element (DUE and a DnaA-oligomerization region (DOR containing DnaA-binding sites (DnaA boxes and a single IHF-binding site that induces sharp DNA bending. DiaA binds to DnaA and stimulates DnaA assembly at the DOR. DnaA binds tightly to ATP and ADP. ATP-DnaA constructs functionally different sub-complexes at DOR, and the DUE-proximal DnaA sub-complex contains IHF and promotes DUE unwinding. The first part of this review presents the structures and mechanisms of oriC-DnaA complexes involved in the regulation of replication initiation. During the cell cycle, the level of ATP-DnaA level, the active form for initiation, is strictly regulated by multiple systems, resulting in timely replication initiation. After initiation, regulatory inactivation of DnaA (RIDA intervenes to reduce ATP-DnaA level by hydrolyzing the DnaA-bound ATP to ADP to yield ADP-DnaA, the inactive form. RIDA involves the binding of the DNA polymerase clamp on newly synthesized DNA to the DnaA-inactivator Hda protein. In datA-dependent DnaA-ATP hydrolysis (DDAH, binding of IHF at the chromosomal locus datA, which contains a cluster of DnaA boxes, results in further hydrolysis of DnaA-bound ATP. SeqA protein inhibits untimely initiation at oriC by binding to newly synthesized oriC DNA and represses dnaA transcription in a cell cycle dependent manner. To reinitiate DNA replication, ADP-DnaA forms oligomers at DnaA-reactivating sequences (DARS1 and DARS2, resulting in the dissociation of ADP and the release of nucleotide-free apo-DnaA, which then

  4. NMR structure of the N-terminal domain of the replication initiator protein DnaA

    Energy Technology Data Exchange (ETDEWEB)

    Wemmer, David E.; Lowery, Thomas J.; Pelton, Jeffrey G.; Chandonia, John-Marc; Kim, Rosalind; Yokota, Hisao; Wemmer, David E.

    2007-08-07

    DnaA is an essential component in the initiation of bacterial chromosomal replication. DnaA binds to a series of 9 base pair repeats leading to oligomerization, recruitment of the DnaBC helicase, and the assembly of the replication fork machinery. The structure of the N-terminal domain (residues 1-100) of DnaA from Mycoplasma genitalium was determined by NMR spectroscopy. The backbone r.m.s.d. for the first 86 residues was 0.6 +/- 0.2 Angstrom based on 742 NOE, 50 hydrogen bond, 46 backbone angle, and 88 residual dipolar coupling restraints. Ultracentrifugation studies revealed that the domain is monomeric in solution. Features on the protein surface include a hydrophobic cleft flanked by several negative residues on one side, and positive residues on the other. A negatively charged ridge is present on the opposite face of the protein. These surfaces may be important sites of interaction with other proteins involved in the replication process. Together, the structure and NMR assignments should facilitate the design of new experiments to probe the protein-protein interactions essential for the initiation of DNA replication.

  5. Regulation of the activity of the dual-function DnaA protein in Caulobacter crescentus.

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    Carmen Fernandez-Fernandez

    Full Text Available DnaA is a conserved essential bacterial protein that acts as the initiator of chromosomal replication as well as a master transcriptional regulator in Caulobacter crescentus. Thus, the intracellular levels of active DnaA need to be tightly regulated during the cell cycle. Our previous work suggested that DnaA may be regulated at the level of its activity by the replisome-associated protein HdaA. Here, we describe the construction of a mutant DnaA protein [DnaA(R357A]. The R357 residue in the AAA+ domain of the C. crescentus DnaA protein is equivalent to the R334 residue of the E. coli DnaA protein, which is required for the Regulatory Inactivation of DnaA (RIDA. We found that the expression of the DnaA(R357A mutant protein in C. crescentus, but not the expression of the wild-type DnaA protein at similar levels, causes a severe phenotype of over-initiation of chromosomal replication and that it blocks cell division. Thus, the mutant DnaA(R357A protein is hyper-active to promote the initiation of DNA replication, compared to the wild-type DnaA protein. DnaA(R357A could not replace DnaA in vivo, indicating that the switch in DnaA activity once chromosomal replication has started may be an essential process in C. crescentus. We propose that the inactivation of DnaA is the main mechanism ensuring that chromosomal replication starts only once per cell cycle. We further observed that the R357A substitution in DnaA does not promote the activity of DnaA as a direct transcriptional activator of four important genes, encoding HdaA, the GcrA master cell cycle regulator, the FtsZ cell division protein and the MipZ spatial regulator of cell division. Thus, the AAA+ domain of DnaA may play a role in temporally regulating the bifunctionality of DnaA by reallocating DnaA molecules from initiating DNA replication to transcribing genes within the unique DnaA regulon of C. crescentus.

  6. Hda-mediated inactivation of the DnaA protein and dnaA gene autoregulation act in concert to ensure homeostatic maintenance of the Escherichia coli chromosome.

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    Riber, Leise; Olsson, Jan A; Jensen, Rasmus B; Skovgaard, Ole; Dasgupta, Santanu; Marinus, Martin G; Løbner-Olesen, Anders

    2006-08-01

    Initiation of DNA replication in Eschericia coli requires the ATP-bound form of the DnaA protein. The conversion of DnaA-ATP to DnaA-ADP is facilitated by a complex of DnaA, Hda (homologous to DnaA), and DNA-loaded beta-clamp proteins in a process termed RIDA (regulatory inactivation of DnaA). Hda-deficient cells initiate replication at each origin mainly once per cell cycle, and the rare reinitiation events never coincide with the end of the origin sequestration period. Therefore, RIDA is not the predominant mechanism to prevent immediate reinitiation from oriC. The cellular level of Hda correlated directly with dnaA gene expression such that Hda deficiency led to reduced dnaA gene expression, and overproduction of Hda led to DnaA overproduction. Hda-deficient cells were very sensitive to variations in the cellular level of DnaA, and DnaA overproduction led to uncontrolled initiation of replication from oriC, causing severe growth retardation or cell death. Based on these observations, we propose that both RIDA and dnaA gene autoregulation are required as homeostatic mechanisms to ensure that initiation of replication occurs at the same time relative to cell mass in each cell cycle.

  7. Sequence characteristics required for cooperative binding and efficient in vivo titration of the replication initiator protein DnaA in E. coli

    DEFF Research Database (Denmark)

    Hansen, Flemming G.; Christensen, Bjarke Bak; Atlung, Tove

    2007-01-01

    Plasmids carrying the mioC promoter region, which contains two DnaA boxes, R5 and R6 with one misfit to the consensus TT(A)/(T)TNCACA, are as efficient in in vivo titration of the DnaA protein as plasmids carrying a replication-inactivated oriC region with its eight DnaA boxes. Three additional Dna......A boxes around the promoter proximal R5 DnaA box were identified and shown by mutational analysis to be necessary for the cooperative binding of DnaA required for titration. These four DnaA boxes are located in the same orientation and with a spacing of two or three base-pairs. The cooperative binding...... was eliminated by insertion of half a helical turn between any of the DnaA boxes. Titration strongly depends on the presence and orientation of the promoter distal R6 DnaA box located 104 bp upstream of the R5 box as well as neighbouring sequences downstream of R6. Titration depends on the integrity of a 43 bp...

  8. The Escherichia coli cryptic prophage protein YfdR binds to DnaA and initiation of chromosomal replication is inhibited by overexpression of the gene cluster yfdQ-yfdR-yfdS-yfdT

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    Yaunori eNoguchi

    2016-03-01

    Full Text Available The initiation of bacterial chromosomal replication is regulated by multiple pathways. To explore novel regulators, we isolated multicopy suppressors for the cold-sensitive hda-185 ΔsfiA(sulA mutant. Hda is crucial for the negative regulation of the initiator DnaA and the hda-185 mutation causes severe replication overinitiation at the replication origin oriC. The SOS-associated division inhibitor SfiA inhibits FtsZ ring formation, an essential step for cell division during the SOS response, and ΔsfiA enhances the cold sensitivity of hda-185 cells in colony formation. One of the suppressors comprised the yfdQ-yfdR-yfdS-yfdT gene cluster carried on a cryptic prophage. Increased copy numbers of yfdQRT or yfdQRS inhibited not only hda-185-dependent overinitiation, but also replication overinitiation in a hyperactive dnaA mutant, and in a mutant lacking an oriC-binding initiation-inhibitor SeqA. In addition, increasing the copy number of the gene set inhibited the growth of cells bearing specific, initiation-impairing dnaA mutations. In wild-type cells, multicopy supply of yfdQRT or yfdQRS also inhibited replication initiation and increased hydroxyurea (HU-resistance, as seen in cells lacking DiaA, a stimulator of DnaA assembly on oriC. Deletion of the yfdQ-yfdR-yfdS-yfdT genes did not affect either HU resistance or initiation regulation. Furthermore, we found that DnaA bound specifically to YfdR in soluble protein extracts oversupplied with YfdQRST. Purified YfdR also bound to DnaA, and DnaA Phe46, an amino acid residue crucial for DnaA interactions with DiaA and DnaB replicative helicase was important for this interaction. Consistently, YfdR moderately inhibited DiaA-DnaA and DnaB-DnaA interactions. In addition, protein extracts oversupplied with YfdQRST inhibited replication initiation in vitro. Given the roles of yfdQ and yfdS in cell tolerance to specific environmental stresses, the yfdQ-yfdR-yfdS-yfdT genes might downregulate the initiator

  9. The Escherichia coli Cryptic Prophage Protein YfdR Binds to DnaA and Initiation of Chromosomal Replication Is Inhibited by Overexpression of the Gene Cluster yfdQ-yfdR-yfdS-yfdT

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    Noguchi, Yasunori; Katayama, Tsutomu

    2016-01-01

    The initiation of bacterial chromosomal replication is regulated by multiple pathways. To explore novel regulators, we isolated multicopy suppressors for the cold-sensitive hda-185 ΔsfiA(sulA) mutant. Hda is crucial for the negative regulation of the initiator DnaA and the hda-185 mutation causes severe replication overinitiation at the replication origin oriC. The SOS-associated division inhibitor SfiA inhibits FtsZ ring formation, an essential step for cell division regulation during the SOS response, and ΔsfiA enhances the cold sensitivity of hda-185 cells in colony formation. One of the suppressors comprised the yfdQ-yfdR-yfdS-yfdT gene cluster carried on a cryptic prophage. Increased copy numbers of yfdQRT or yfdQRS inhibited not only hda-185-dependent overinitiation, but also replication overinitiation in a hyperactive dnaA mutant, and in a mutant lacking an oriC-binding initiation-inhibitor SeqA. In addition, increasing the copy number of the gene set inhibited the growth of cells bearing specific, initiation-impairing dnaA mutations. In wild-type cells, multicopy supply of yfdQRT or yfdQRS also inhibited replication initiation and increased hydroxyurea (HU)-resistance, as seen in cells lacking DiaA, a stimulator of DnaA assembly on oriC. Deletion of the yfdQ-yfdR-yfdS-yfdT genes did not affect either HU resistance or initiation regulation. Furthermore, we found that DnaA bound specifically to YfdR in soluble protein extracts oversupplied with YfdQRST. Purified YfdR also bound to DnaA, and DnaA Phe46, an amino acid residue crucial for DnaA interactions with DiaA and DnaB replicative helicase was important for this interaction. Consistently, YfdR moderately inhibited DiaA-DnaA and DnaB-DnaA interactions. In addition, protein extracts oversupplied with YfdQRST inhibited replication initiation in vitro. Given the roles of yfdQ and yfdS in cell tolerance to specific environmental stresses, the yfdQ-yfdR-yfdS-yfdT genes might downregulate the initiator Dna

  10. The Escherichia coli Cryptic Prophage Protein YfdR Binds to DnaA and Initiation of Chromosomal Replication Is Inhibited by Overexpression of the Gene Cluster yfdQ-yfdR-yfdS-yfdT.

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    Noguchi, Yasunori; Katayama, Tsutomu

    2016-01-01

    The initiation of bacterial chromosomal replication is regulated by multiple pathways. To explore novel regulators, we isolated multicopy suppressors for the cold-sensitive hda-185 ΔsfiA(sulA) mutant. Hda is crucial for the negative regulation of the initiator DnaA and the hda-185 mutation causes severe replication overinitiation at the replication origin oriC. The SOS-associated division inhibitor SfiA inhibits FtsZ ring formation, an essential step for cell division regulation during the SOS response, and ΔsfiA enhances the cold sensitivity of hda-185 cells in colony formation. One of the suppressors comprised the yfdQ-yfdR-yfdS-yfdT gene cluster carried on a cryptic prophage. Increased copy numbers of yfdQRT or yfdQRS inhibited not only hda-185-dependent overinitiation, but also replication overinitiation in a hyperactive dnaA mutant, and in a mutant lacking an oriC-binding initiation-inhibitor SeqA. In addition, increasing the copy number of the gene set inhibited the growth of cells bearing specific, initiation-impairing dnaA mutations. In wild-type cells, multicopy supply of yfdQRT or yfdQRS also inhibited replication initiation and increased hydroxyurea (HU)-resistance, as seen in cells lacking DiaA, a stimulator of DnaA assembly on oriC. Deletion of the yfdQ-yfdR-yfdS-yfdT genes did not affect either HU resistance or initiation regulation. Furthermore, we found that DnaA bound specifically to YfdR in soluble protein extracts oversupplied with YfdQRST. Purified YfdR also bound to DnaA, and DnaA Phe46, an amino acid residue crucial for DnaA interactions with DiaA and DnaB replicative helicase was important for this interaction. Consistently, YfdR moderately inhibited DiaA-DnaA and DnaB-DnaA interactions. In addition, protein extracts oversupplied with YfdQRST inhibited replication initiation in vitro. Given the roles of yfdQ and yfdS in cell tolerance to specific environmental stresses, the yfdQ-yfdR-yfdS-yfdT genes might downregulate the initiator Dna

  11. DnaA protein DNA-binding domain binds to Hda protein to promote inter-AAA+ domain interaction involved in regulatory inactivation of DnaA.

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    Keyamura, Kenji; Katayama, Tsutomu

    2011-08-19

    Chromosomal replication is initiated from the replication origin oriC in Escherichia coli by the active ATP-bound form of DnaA protein. The regulatory inactivation of DnaA (RIDA) system, a complex of the ADP-bound Hda and the DNA-loaded replicase clamp, represses extra initiations by facilitating DnaA-bound ATP hydrolysis, yielding the inactive ADP-bound form of DnaA. However, the mechanisms involved in promoting the DnaA-Hda interaction have not been determined except for the involvement of an interaction between the AAA+ domains of the two. This study revealed that DnaA Leu-422 and Pro-423 residues within DnaA domain IV, including a typical DNA-binding HTH motif, are specifically required for RIDA-dependent ATP hydrolysis in vitro and that these residues support efficient interaction with the DNA-loaded clamp·Hda complex and with Hda in vitro. Consistently, substitutions of these residues caused accumulation of ATP-bound DnaA in vivo and oriC-dependent inhibition of cell growth. Leu-422 plays a more important role in these activities than Pro-423. By contrast, neither of these residues is crucial for DNA replication from oriC, although they are highly conserved in DnaA orthologues. Structural analysis of a DnaA·Hda complex model suggested that these residues make contact with residues in the vicinity of the Hda AAA+ sensor I that participates in formation of a nucleotide-interacting surface. Together, the results show that functional DnaA-Hda interactions require a second interaction site within DnaA domain IV in addition to the AAA+ domain and suggest that these interactions are crucial for the formation of RIDA complexes that are active for DnaA-ATP hydrolysis.

  12. DnaA Protein DNA-binding Domain Binds to Hda Protein to Promote Inter-AAA+ Domain Interaction Involved in Regulatory Inactivation of DnaA*

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    Keyamura, Kenji; Katayama, Tsutomu

    2011-01-01

    Chromosomal replication is initiated from the replication origin oriC in Escherichia coli by the active ATP-bound form of DnaA protein. The regulatory inactivation of DnaA (RIDA) system, a complex of the ADP-bound Hda and the DNA-loaded replicase clamp, represses extra initiations by facilitating DnaA-bound ATP hydrolysis, yielding the inactive ADP-bound form of DnaA. However, the mechanisms involved in promoting the DnaA-Hda interaction have not been determined except for the involvement of an interaction between the AAA+ domains of the two. This study revealed that DnaA Leu-422 and Pro-423 residues within DnaA domain IV, including a typical DNA-binding HTH motif, are specifically required for RIDA-dependent ATP hydrolysis in vitro and that these residues support efficient interaction with the DNA-loaded clamp·Hda complex and with Hda in vitro. Consistently, substitutions of these residues caused accumulation of ATP-bound DnaA in vivo and oriC-dependent inhibition of cell growth. Leu-422 plays a more important role in these activities than Pro-423. By contrast, neither of these residues is crucial for DNA replication from oriC, although they are highly conserved in DnaA orthologues. Structural analysis of a DnaA·Hda complex model suggested that these residues make contact with residues in the vicinity of the Hda AAA+ sensor I that participates in formation of a nucleotide-interacting surface. Together, the results show that functional DnaA-Hda interactions require a second interaction site within DnaA domain IV in addition to the AAA+ domain and suggest that these interactions are crucial for the formation of RIDA complexes that are active for DnaA-ATP hydrolysis. PMID:21708944

  13. Evidence for roles of the Escherichia coli Hda protein beyond regulatory inactivation of DnaA.

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    Baxter, Jamie C; Sutton, Mark D

    2012-08-01

    The ATP-bound form of the Escherichia coli DnaA protein binds 'DnaA boxes' present in the origin of replication (oriC) and operator sites of several genes, including dnaA, to co-ordinate their transcription with initiation of replication. The Hda protein, together with the β sliding clamp, stimulates the ATPase activity of DnaA via a process termed regulatory inactivation of DnaA (RIDA), to regulate the activity of DnaA in DNA replication. Here, we used the mutant dnaN159 strain, which expresses the β159 clamp protein, to gain insight into how the actions of Hda are co-ordinated with replication. Elevated expression of Hda impeded growth of the dnaN159 strain in a Pol II- and Pol IV-dependent manner, suggesting a role for Hda managing the actions of these Pols. In a wild-type strain, elevated levels of Hda conferred sensitivity to nitrofurazone, and suppressed the frequency of -1 frameshift mutations characteristic of Pol IV, while loss of hda conferred cold sensitivity. Using the dnaN159 strain, we identified 24 novel hda alleles, four of which supported E. coli viability despite their RIDA defect. Taken together, these findings suggest that although one or more Hda functions are essential for cell viability, RIDA may be dispensable. © 2012 Blackwell Publishing Ltd.

  14. In Vitro Whole Genome DNA Binding Analysis of the Bacterial Replication Initiator and Transcription Factor DnaA.

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    Janet L Smith

    2015-05-01

    Full Text Available DnaA, the replication initiation protein in bacteria, is an AAA+ ATPase that binds and hydrolyzes ATP and exists in a heterogeneous population of ATP-DnaA and ADP-DnaA. DnaA binds cooperatively to the origin of replication and several other chromosomal regions, and functions as a transcription factor at some of these regions. We determined the binding properties of Bacillus subtilis DnaA to genomic DNA in vitro at single nucleotide resolution using in vitro DNA affinity purification and deep sequencing (IDAP-Seq. We used these data to identify 269 binding regions, refine the consensus sequence of the DnaA binding site, and compare the relative affinity of binding regions for ATP-DnaA and ADP-DnaA. Most sites had a slightly higher affinity for ATP-DnaA than ADP-DnaA, but a few had a strong preference for binding ATP-DnaA. Of the 269 sites, only the eight strongest binding ones have been observed to bind DnaA in vivo, suggesting that other cellular factors or the amount of available DnaA in vivo restricts DnaA binding to these additional sites. Conversely, we found several chromosomal regions that were bound by DnaA in vivo but not in vitro, and that the nucleoid-associated protein Rok was required for binding in vivo. Our in vitro characterization of the inherent ability of DnaA to bind the genome at single nucleotide resolution provides a backdrop for interpreting data on in vivo binding and regulation of DnaA, and is an approach that should be adaptable to many other DNA binding proteins.

  15. Molecular mechanism of DNA replication-coupled inactivation of the initiator protein in Escherichia coli: interaction of DnaA with the sliding clamp-loaded DNA and the sliding clamp-Hda complex.

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    Su'etsugu, Masayuki; Takata, Makoto; Kubota, Toshio; Matsuda, Yusaku; Katayama, Tsutomu

    2004-06-01

    In Escherichia coli, the ATP-DnaA protein initiates chromosomal replication. After the DNA polymerase III holoenzyme is loaded on to DNA, DnaA-bound ATP is hydrolysed in a manner depending on Hda protein and the DNA-loaded form of the DNA polymerase III sliding clamp subunit, which yields ADP-DnaA, an inactivated form for initiation. This regulatory DnaA-inactivation represses extra initiation events. In this study, in vitro replication intermediates and structured DNA mimicking replicational intermediates were first used to identify structural prerequisites in the process of DnaA-ATP hydrolysis. Unlike duplex DNA loaded with sliding clamps, primer RNA-DNA heteroduplexes loaded with clamps were not associated with DnaA-ATP hydrolysis, and duplex DNA provided in trans did not rescue this defect. At least 40-bp duplex DNA is competent for the DnaA-ATP hydrolysis when a single clamp was loaded. The DnaA-ATP hydrolysis was inhibited when ATP-DnaA was tightly bound to a DnaA box-bearing oligonucleotide. These results imply that the DnaA-ATP hydrolysis involves the direct interaction of ATP-DnaA with duplex DNA flanking the sliding clamp. Furthermore, Hda protein formed a stable complex with the sliding clamp. Based on these, we suggest a mechanical basis in the DnaA-inactivation that ATP-DnaA interacts with the Hda-clamp complex with the aid of DNA binding. Copyright Blackwell Publishing Limited

  16. The DnaA Tale

    DEFF Research Database (Denmark)

    Hansen, Flemming G.; Atlung, Tove

    2018-01-01

    , and translation efficiency, as well as, the DnaA protein, its concentration, its binding to DnaA-boxes, and its binding of ATP or ADP. Furthermore, we will discuss the different models for regulation of initiation which have been proposed over the years, with particular emphasis on the Initiator Titration Model....

  17. Isolation and characterization of the dnaA gene of Rickettsia prowazekii

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    Waite, R.T.; Shaw, E.I.; Winkler, H.H.; Wood, D.G.

    1998-01-01

    The dnaA gene encoding the initiator protein of DNA replication was isolated from the obligate intracellular bacterium, Rickettsia prowazekii. Comparison of the deduced amino acid sequence of R. prowazekii DnaA with other bacterial DnaA proteins revealed extensive similarity. However, the rickettsial sequence is unique in the number of basic lysine residues found within a highly conserved portion of the putative DNA binding region, suggesting that the rickettsial protein may recognize a DNA sequence that differs from the consensus DnaA box sequence identified in other bacteria. Consensus DnaA box sequences, found upstream of many bacterial dnaA genes, were not identified upstream of rickettsial dnaA gene. In addition, gene organization within this region differed from that of other bacteria. The putative start of transcription of the rickettsial dnaA gene was localized to a site 522 nucleotides upstream of the DnaA start codon. Key words: Rickettsia prowazekii; dnaA gene; initiator protein (authors)

  18. Independent control of replication initiation of the two Vibrio cholerae chromosomes by DnaA and RctB

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    Duigou, Stephane; Knudsen, Kristine Groth; Skovgaard, Ole

    2006-01-01

    Although the two Vibrio cholerae chromosomes initiate replication in a coordinated fashion, we show here that each chromosome appears to have a specific replication initiator. DnaA overproduction promoted overinitiation of chromosome I and not chromosome II. In contrast, overproduction of RctB, a...

  19. Escherichia coli DnaA forms helical structures along the longitudinal cell axis distinct from MreB filaments.

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    Boeneman, Kelly; Fossum, Solveig; Yang, Yanhua; Fingland, Nicholas; Skarstad, Kirsten; Crooke, Elliott

    2009-05-01

    DnaA initiates chromosomal replication in Escherichia coli at a well-regulated time in the cell cycle. To determine how the spatial distribution of DnaA is related to the location of chromosomal replication and other cell cycle events, the localization of DnaA in living cells was visualized by confocal fluorescence microscopy. The gfp gene was randomly inserted into a dnaA-bearing plasmid via in vitro transposition to create a library that included internally GFP-tagged DnaA proteins. The library was screened for the ability to rescue dnaA(ts) mutants, and a candidate gfp-dnaA was used to replace the dnaA gene of wild-type cells. The resulting cells produce close to physiological levels of GFP-DnaA from the endogenous promoter as their only source of DnaA and somewhat under-initiate replication with moderate asynchrony. Visualization of GFP-tagged DnaA in living cells revealed that DnaA adopts a helical pattern that spirals along the long axis of the cell, a pattern also seen in wild-type cells by immunofluorescence with affinity purified anti-DnaA antibody. Although the DnaA helices closely resemble the helices of the actin analogue MreB, co-visualization of GFP-tagged DnaA and RFP-tagged MreB demonstrates that DnaA and MreB adopt discrete helical structures along the length of the longitudinal cell axis.

  20. Insights into the quality of DnaA boxes and their cooperativity

    DEFF Research Database (Denmark)

    Hansen, Flemming G.; Christensen, Bjarke Bak; Nielsen, Christina Bang

    2006-01-01

    Plasmids carrying the mioC promoter region with its two DnaA boxes are as efficient in titration of DnaA protein as plasmids carrying a replicationinactivated oriC region with its five DnaA boxes. The two DnaA boxes upstream of the mioC promoter were mutated in various ways to study the cooperati......Plasmids carrying the mioC promoter region with its two DnaA boxes are as efficient in titration of DnaA protein as plasmids carrying a replicationinactivated oriC region with its five DnaA boxes. The two DnaA boxes upstream of the mioC promoter were mutated in various ways to study...... the cooperativity between the DnaA boxes, and to study in vivo the in vitrodefined 9mer DnaA box consensus sequence TTA/TTNCACA). The quality and cooperativity of the DnaA oxes were determined in two complementary ways: as titration of DnaA protein leading to derepression of the dnaA promoter, and as repression...... of the mioC promoter caused by the DnaA protein binding to the DnaA boxes. Titration of DnaA protein correlated with repression of the mioC promoter. The level of titration and repression with the normal promoter-proximal box (TTTTCCACA) depends strongly on the presence and the quality of a DnaA box...

  1. The DnaA N-terminal domain interacts with Hda to facilitate replicase clamp-mediated inactivation of DnaA.

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    Su'etsugu, Masayuki; Harada, Yuji; Keyamura, Kenji; Matsunaga, Chika; Kasho, Kazutoshi; Abe, Yoshito; Ueda, Tadashi; Katayama, Tsutomu

    2013-12-01

    DnaA activity for replication initiation of the Escherichia coli chromosome is negatively regulated by feedback from the DNA-loaded form of the replicase clamp. In this process, called RIDA (regulatory inactivation of DnaA), ATP-bound DnaA transiently assembles into a complex consisting of Hda and the DNA-clamp, which promotes inter-AAA+ domain association between Hda and DnaA and stimulates hydrolysis of DnaA-bound ATP, producing inactive ADP-DnaA. Using a truncated DnaA mutant, we previously demonstrated that the DnaA N-terminal domain is involved in RIDA. However, the precise role of the N-terminal domain in RIDA has remained largely unclear. Here, we used an in vitro reconstituted system to demonstrate that the Asn-44 residue in the N-terminal domain of DnaA is crucial for RIDA but not for replication initiation. Moreover, an assay termed PDAX (pull-down after cross-linking) revealed an unstable interaction between a DnaA-N44A mutant and Hda. In vivo, this mutant exhibited an increase in the cellular level of ATP-bound DnaA. These results establish a model in which interaction between DnaA Asn-44 and Hda stabilizes the association between the AAA+ domains of DnaA and Hda to facilitate DnaA-ATP hydrolysis during RIDA. © 2013 Society for Applied Microbiology and John Wiley & Sons Ltd.

  2. A structural basis for the regulatory inactivation of DnaA.

    Science.gov (United States)

    Xu, Qingping; McMullan, Daniel; Abdubek, Polat; Astakhova, Tamara; Carlton, Dennis; Chen, Connie; Chiu, Hsiu-Ju; Clayton, Thomas; Das, Debanu; Deller, Marc C; Duan, Lian; Elsliger, Marc-Andre; Feuerhelm, Julie; Hale, Joanna; Han, Gye Won; Jaroszewski, Lukasz; Jin, Kevin K; Johnson, Hope A; Klock, Heath E; Knuth, Mark W; Kozbial, Piotr; Sri Krishna, S; Kumar, Abhinav; Marciano, David; Miller, Mitchell D; Morse, Andrew T; Nigoghossian, Edward; Nopakun, Amanda; Okach, Linda; Oommachen, Silvya; Paulsen, Jessica; Puckett, Christina; Reyes, Ron; Rife, Christopher L; Sefcovic, Natasha; Trame, Christine; van den Bedem, Henry; Weekes, Dana; Hodgson, Keith O; Wooley, John; Deacon, Ashley M; Godzik, Adam; Lesley, Scott A; Wilson, Ian A

    2009-01-16

    Regulatory inactivation of DnaA is dependent on Hda (homologous to DnaA), a protein homologous to the AAA+ (ATPases associated with diverse cellular activities) ATPase region of the replication initiator DnaA. When bound to the sliding clamp loaded onto duplex DNA, Hda can stimulate the transformation of active DnaA-ATP into inactive DnaA-ADP. The crystal structure of Hda from Shewanella amazonensis SB2B at 1.75 A resolution reveals that Hda resembles typical AAA+ ATPases. The arrangement of the two subdomains in Hda (residues 1-174 and 175-241) differs dramatically from that of DnaA. A CDP molecule anchors the Hda domains in a conformation that promotes dimer formation. The Hda dimer adopts a novel oligomeric assembly for AAA+ proteins in which the arginine finger, crucial for ATP hydrolysis, is fully exposed and available to hydrolyze DnaA-ATP through a typical AAA+ type of mechanism. The sliding clamp binding motifs at the N-terminus of each Hda monomer are partially buried and combine to form an antiparallel beta-sheet at the dimer interface. The inaccessibility of the clamp binding motifs in the CDP-bound structure of Hda suggests that conformational changes are required for Hda to form a functional complex with the clamp. Thus, the CDP-bound Hda dimer likely represents an inactive form of Hda.

  3. Novel essential residues of Hda for interaction with DnaA in the regulatory inactivation of DnaA: unique roles for Hda AAA Box VI and VII motifs.

    Science.gov (United States)

    Nakamura, Kenta; Katayama, Tsutomu

    2010-04-01

    Escherichia coli ATP-DnaA initiates chromosomal replication. For preventing extra-initiations, a complex of ADP-Hda and the DNA-loaded replicase clamp promotes DnaA-ATP hydrolysis, yielding inactive ADP-DnaA. However, the Hda-DnaA interaction mode remains unclear except that the Hda Box VII Arg finger (Arg-153) and DnaA sensor II Arg-334 within each AAA(+) domain are crucial for the DnaA-ATP hydrolysis. Here, we demonstrate that direct and functional interaction of ADP-Hda with DnaA requires the Hda residues Ser-152, Phe-118 and Asn-122 as well as Hda Arg-153 and DnaA Arg-334. Structural analyses suggest intermolecular interactions between Hda Ser-152 and DnaA Arg-334 and between Hda Phe-118 and the DnaA Walker B motif region, in addition to an intramolecular interaction between Hda Asn-122 and Arg-153. These interactions likely sustain a specific association of ADP-Hda and DnaA, promoting DnaA-ATP hydrolysis. Consistently, ATP-DnaA and ADP-DnaA interact with the ADP-Hda-DNA-clamp complex with similar affinities. Hda Phe-118 and Asn-122 are contained in the Box VI region, and their hydrophobic and electrostatic features are basically conserved in the corresponding residues of other AAA(+) proteins, suggesting a conserved role for Box VI. These findings indicate novel interaction mechanisms for Hda-DnaA as well as a potentially fundamental mechanism in AAA(+) protein interactions.

  4. Hda inactivation of DnaA is the predominant mechanism preventing hyperinitiation of Escherichia coli DNA replication.

    Science.gov (United States)

    Camara, Johanna E; Breier, Adam M; Brendler, Therese; Austin, Stuart; Cozzarelli, Nicholas R; Crooke, Elliott

    2005-08-01

    Initiation of DNA replication from the Escherichia coli chromosomal origin is highly regulated, assuring that replication occurs precisely once per cell cycle. Three mechanisms for regulation of replication initiation have been proposed: titration of free DnaA initiator protein by the datA locus, sequestration of newly replicated origins by SeqA protein and regulatory inactivation of DnaA (RIDA), in which active ATP-DnaA is converted to the inactive ADP-bound form. DNA microarray analyses showed that the level of initiation in rapidly growing cells that lack datA was indistinguishable from that in wild-type cells, and that the absence of SeqA protein caused only a modest increase in initiation, in agreement with flow-cytometry data. In contrast, cells lacking Hda overinitiated replication twofold, implicating RIDA as the predominant mechanism preventing extra initiation events in a cell cycle.

  5. Characterization of Staphylococcus aureus Primosomal DnaD Protein: Highly Conserved C-Terminal Region Is Crucial for ssDNA and PriA Helicase Binding but Not for DnaA Protein-Binding and Self-Tetramerization.

    Directory of Open Access Journals (Sweden)

    Yen-Hua Huang

    Full Text Available The role of DnaD in the recruitment of replicative helicase has been identified. However, knowledge of the DNA, PriA, and DnaA binding mechanism of this protein for the DnaA- and PriA-directed replication primosome assemblies is limited. We characterized the DNA-binding properties of DnaD from Staphylococcus aureus (SaDnaD and analyzed its interactions with SaPriA and SaDnaA. The gel filtration chromatography analysis of purified SaDnaD and its deletion mutant proteins (SaDnaD1-195, SaDnaD1-200 and SaDnaD1-204 showed a stable tetramer in solution. This finding indicates that the C-terminal region aa 196-228 is not crucial for SaDnaD oligomerization. SaDnaD forms distinct complexes with ssDNA of different lengths. In fluorescence titrations, SaDnaD bound to ssDNA with a binding-site size of approximately 32 nt. A stable complex of SaDnaD1-195, SaDnaD1-200, and SaDnaD1-204 with ssDNA dT40 was undetectable, indicating that the C-terminal region of SaDnaD (particularly aa 205-228 is crucial for ssDNA binding. The SPR results revealed that SaDnaD1-195 can interact with SaDnaA but not with SaPriA, which may indicate that DnaD has different binding sites for PriA and DnaA. Both SaDnaD and SaDnaDY176A mutant proteins, but not SaDnaD1-195, can significantly stimulate the ATPase activity of SaPriA. Hence, the stimulation effect mainly resulted from direct contact within the protein-protein interaction, not via the DNA-protein interaction. Kinetic studies revealed that the SaDnaD-SaPriA interaction increases the Vmax of the SaPriA ATPase fivefold without significantly affecting the Km. These results indicate that the conserved C-terminal region is crucial for ssDNA and PriA helicase binding, but not for DnaA protein-binding and self-tetramerization.

  6. A role for the weak DnaA binding sites in bacterial replication origins

    DEFF Research Database (Denmark)

    Charbon, Godefroid; Løbner-Olesen, Anders

    2011-01-01

    DnaA initiates the chromosomal DNA replication in nearly all bacteria, and replication origins are characterized by binding sites for the DnaA protein (DnaA-boxes) along with an ‘AT-rich’ region. However, great variation in number, spatial organization and specificity of DnaA-boxes is observed...... between species. In the study by Taylor et al. (2011), new and unexpectedly weak DnaA-boxes were identified within the Caulobacter crescentus origin of replication (Cori). The position of weak and stronger DnaA-boxes follows a pattern seen in Escherichia coli oriC. This raises the possibility...... that bacterial origins might be more alike than previously thought....

  7. Diversification of DnaA dependency for DNA replication in cyanobacterial evolution.

    Science.gov (United States)

    Ohbayashi, Ryudo; Watanabe, Satoru; Ehira, Shigeki; Kanesaki, Yu; Chibazakura, Taku; Yoshikawa, Hirofumi

    2016-05-01

    Regulating DNA replication is essential for all living cells. The DNA replication initiation factor DnaA is highly conserved in prokaryotes and is required for accurate initiation of chromosomal replication at oriC. DnaA-independent free-living bacteria have not been identified. The dnaA gene is absent in plastids and some symbiotic bacteria, although it is not known when or how DnaA-independent mechanisms were acquired. Here, we show that the degree of dependency of DNA replication on DnaA varies among cyanobacterial species. Deletion of the dnaA gene in Synechococcus elongatus PCC 7942 shifted DNA replication from oriC to a different site as a result of the integration of an episomal plasmid. Moreover, viability during the stationary phase was higher in dnaA disruptants than in wild-type cells. Deletion of dnaA did not affect DNA replication or cell growth in Synechocystis sp. PCC 6803 or Anabaena sp. PCC 7120, indicating that functional dependency on DnaA was already lost in some nonsymbiotic cyanobacterial lineages during diversification. Therefore, we proposed that cyanobacteria acquired DnaA-independent replication mechanisms before symbiosis and such an ancestral cyanobacterium was the sole primary endosymbiont to form a plastid precursor.

  8. Mutant DnaAs of Escherichia coli that are refractory to negative control.

    Science.gov (United States)

    Chodavarapu, Sundari; Felczak, Magdalena M; Simmons, Lyle A; Murillo, Alec; Kaguni, Jon M

    2013-12-01

    DnaA is the initiator of DNA replication in bacteria. A mutant DnaA named DnaAcos is unusual because it is refractory to negative regulation. We developed a genetic method to isolate other mutant DnaAs that circumvent regulation to extend our understanding of mechanisms that control replication initiation. Like DnaAcos, one mutant bearing a tyrosine substitution for histidine 202 (H202Y) withstands the regulation exerted by datA, hda and dnaN (β clamp), and both DnaAcos and H202Y resist inhibition by the Hda-β clamp complex in vitro. Other mutant DnaAs carrying G79D, E244K, V303M or E445K substitutions are either only partially sensitive or refractory to inhibition by the Hda-β clamp complex in vitro but are responsive to hda expression in vivo. All mutant DnaAs remain able to interact directly with Hda. Of interest, both DnaAcos and DnaAE244K bind more avidly to Hda. These mutants, by sequestrating Hda, may limit its availability to regulate other DnaA molecules, which remain active to induce extra rounds of DNA replication. Other evidence suggests that a mutant bearing a V292M substitution hyperinitiates by escaping the effect of an unknown regulatory factor. Together, our results provide new insight into the mechanisms that regulate replication initiation in Escherichia coli.

  9. Multiple DNA binding proteins contribute to timing of chromosome replication in E. coli

    DEFF Research Database (Denmark)

    Riber, Leise; Frimodt-Møller, Jakob; Charbon, Godefroid

    2016-01-01

    Chromosome replication in Escherichia coli is initiated from a single origin, oriC. Initiation involves a number of DNA binding proteins, but only DnaA is essential and specific for the initiation process. DnaA is an AAA+ protein that binds both ATP and ADP with similar high affinities. Dna...... replication is initiated, or the time window in which all origins present in a single cell are initiated, i.e. initiation synchrony, or both. Overall, these DNA binding proteins modulate the initiation frequency from oriC by: (i) binding directly to oriC to affect DnaA binding, (ii) altering the DNA topology...... in or around oriC, (iii) altering the nucleotide bound status of DnaA by interacting with non-coding chromosomal sequences, distant from oriC, that are important for DnaA activity. Thus, although DnaA is the key protein for initiation of replication, other DNA-binding proteins act not only on ori...

  10. Controlled initiation of chromosomal replication in Escherichia coli requires functional Hda protein.

    Science.gov (United States)

    Camara, Johanna Eltz; Skarstad, Kirsten; Crooke, Elliott

    2003-05-01

    Regulatory inactivation of DnaA helps ensure that the Escherichia coli chromosome is replicated only once per cell cycle, through accelerated hydrolysis of active replication initiator ATP-DnaA to inactive ADP-DnaA. Analysis of deltahda strains revealed that the regulatory inactivation of DnaA component Hda is necessary for maintaining controlled initiation but not for cell growth or viability.

  11. Initiation of Replication in Escherichia coli

    DEFF Research Database (Denmark)

    Frimodt-Møller, Jakob

    The circular chromosome of Escherichia coli is replicated by two replisomes assembled at the unique origin and moving in the opposite direction until they meet in the less well defined terminus. The key protein in initiation of replication, DnaA, facilitates the unwinding of double-stranded DNA...... to single-stranded DNA in oriC. Although DnaA is able to bind both ADP and ATP, DnaA is only active in initiation when bound to ATP. Although initiation of replication, and the regulation of this, is thoroughly investigated it is still not fully understood. The overall aim of the thesis was to investigate...... the regulation of initiation, the effect on the cell when regulation fails, and if regulation was interlinked to chromosomal organization. This thesis uncovers that there exists a subtle balance between chromosome replication and reactive oxygen species (ROS) inflicted DNA damage. Thus, failure in regulation...

  12. Hda, a novel DnaA-related protein, regulates the replication cycle in Escherichia coli.

    Science.gov (United States)

    Kato , J; Katayama, T

    2001-08-01

    The bacterial DnaA protein binds to the chromosomal origin of replication to trigger a series of initiation reactions, which leads to the loading of DNA polymerase III. In Escherichia coli, once this polymerase initiates DNA synthesis, ATP bound to DnaA is efficiently hydrolyzed to yield the ADP-bound inactivated form. This negative regulation of DnaA, which occurs through interaction with the beta-subunit sliding clamp configuration of the polymerase, functions in the temporal blocking of re-initiation. Here we show that the novel DnaA-related protein, Hda, from E.coli is essential for this regulatory inactivation of DnaA in vitro and in vivo. Our results indicate that the hda gene is required to prevent over-initiation of chromosomal replication and for cell viability. Hda belongs to the chaperone-like ATPase family, AAA(+), as do DnaA and certain eukaryotic proteins essential for the initiation of DNA replication. We propose that the once-per-cell-cycle rule of replication depends on the timely interaction of AAA(+) proteins that comprise the apparatus regulating the activity of the initiator of replication.

  13. Interaction of the Sliding Clamp β-Subunit and Hda, a DnaA-Related Protein

    OpenAIRE

    Kurz, Mareike; Dalrymple, Brian; Wijffels, Gene; Kongsuwan, Kritaya

    2004-01-01

    In Escherichia coli, interactions between the replication initiation protein DnaA, the β subunit of DNA polymerase III (the sliding clamp protein), and Hda, the recently identified DnaA-related protein, are required to convert the active ATP-bound form of DnaA to an inactive ADP-bound form through the accelerated hydrolysis of ATP. This rapid hydrolysis of ATP is proposed to be the main mechanism that blocks multiple initiations during cell cycle and acts as a molecular switch from initiation...

  14. Initiation of chromosomal replication in predatory bacterium Bdellovibrio bacteriovorus

    Directory of Open Access Journals (Sweden)

    Lukasz Makowski

    2016-11-01

    Full Text Available Bdellovibrio bacteriovorus is a small Gram-negative predatory bacterium that attacks other Gram-negative bacteria, including many animal, human, and plant pathogens. This bacterium exhibits a peculiar biphasic life cycle during which two different types of cells are produced: non-replicating highly motile cells (the free-living phase and replicating cells (the intracellular-growth phase. The process of chromosomal replication in B. bacteriovorus must therefore be temporally and spatially regulated to ensure that it is coordinated with cell differentiation and cell cycle progression. Recently, B. bacteriovorus has received considerable research interest due to its intriguing life cycle and great potential as a prospective antimicrobial agent. Although we know that chromosomal replication in bacteria is mainly regulated at the initiation step, no data exists about this process in B. bacteriovorus. We report the first characterization of key elements of initiation of chromosomal replication – DnaA protein and oriC region from the predatory bacterium, B. bacteriovorus. In vitro studies using different approaches demonstrate that the B. bacteriovorus oriC (BdoriC is specifically bound and unwound by the DnaA protein. Sequence comparison of the DnaA-binding sites enabled us to propose a consensus sequence for the B. bacteriovorus DnaA box (5’-NN(A/TTCCACA-3’. Surprisingly, in vitro analysis revealed that BdoriC is also bound and unwound by the host DnaA proteins (relatively distantly related from B. bacteriovorus. We compared the architecture of the DnaA–oriC complexes (orisomes in homologous (oriC and DnaA from B. bacteriovorus and heterologous (BdoriC and DnaA from prey, E. coli or P. aeruginosa systems. This work provides important new entry points toward improving our understanding of the initiation of chromosomal replication in this predatory bacterium.

  15. Protein associations in DnaA-ATP hydrolysis mediated by the Hda-replicase clamp complex.

    Science.gov (United States)

    Su'etsugu, Masayuki; Shimuta, Toh-Ru; Ishida, Takuma; Kawakami, Hironori; Katayama, Tsutomu

    2005-02-25

    In Escherichia coli, the activity of ATP-bound DnaA protein in initiating chromosomal replication is negatively controlled in a replication-coordinated manner. The RIDA (regulatory inactivation of DnaA) system promotes DnaA-ATP hydrolysis to produce the inactivated form DnaA-ADP in a manner depending on the Hda protein and the DNA-loaded form of the beta-sliding clamp, a subunit of the replicase holoenzyme. A highly functional form of Hda was purified and shown to form a homodimer in solution, and two Hda dimers were found to associate with a single clamp molecule. Purified mutant Hda proteins were used in a staged in vitro RIDA system followed by a pull-down assay to show that Hda-clamp binding is a prerequisite for DnaA-ATP hydrolysis and that binding is mediated by an Hda N-terminal motif. Arg(168) in the AAA(+) Box VII motif of Hda plays a role in stable homodimer formation and in DnaA-ATP hydrolysis, but not in clamp binding. Furthermore, the DnaA N-terminal domain is required for the functional interaction of DnaA with the Hda-clamp complex. Single cells contain approximately 50 Hda dimers, consistent with the results of in vitro experiments. These findings and the features of AAA(+) proteins, including DnaA, suggest the following model. DnaA-ATP is hydrolyzed at a binding interface between the AAA(+) domains of DnaA and Hda; the DnaA N-terminal domain supports this interaction; and the interaction of DnaA-ATP with the Hda-clamp complex occurs in a catalytic mode.

  16. Evidence for Roles of the Escherichia coli Hda Protein Beyond RIDA

    Science.gov (United States)

    Baxter, Jamie C.; Sutton, Mark D.

    2012-01-01

    The ATP-bound form of the Escherichia coli DnaA protein binds ‘DnaA boxes’ present in the origin of replication (oriC) and operator sites of several genes, including dnaA, to coordinate their transcription with initiation of replication. The Hda protein, together with the β sliding clamp, stimulates the ATPase activity of DnaA via a process termed Regulatory Inactivation of DnaA (RIDA), to regulate the activity of DnaA in DNA replication. Here, we used the mutant dnaN159 strain, which expresses the β159 clamp protein, to gain insight into how the actions of Hda are coordinated with replication. Elevated expression of Hda impeded growth of the dnaN159 strain in a Pol II- and Pol IV-dependent manner, suggesting a role for Hda managing the actions of these Pols. In a wild type strain, elevated levels of Hda conferred sensitivity to nitrofurazone, and suppressed the frequency of −1 frameshift mutations characteristic of Pol IV, while loss of hda conferred cold sensitivity. Using the dnaN159 strain, we identified 24 novel hda alleles, four of which supported E. coli viability despite their RIDA defect. Taken together, these findings suggest that although one or more Hda functions are essential for cell viability, RIDA may be dispensable. PMID:22716942

  17. Data of evolutionary structure change: 1WEEA-2DNAA [Confc[Archive

    Lifescience Database Archive (English)

    Full Text Available 1WEEA-2DNAA 1WEE 2DNA A A GSSGSSGMERGVDNWKVDCKCGTKDDDGERMLACDGCGV...WHHT--RCIGINNADALPSKFLCFRCIELS----GPSSG GSSGSSGPSHSLQAPEVRFSKEMECLQAMGFV----------NYNANLQALIATDG-DTNAA... 2DNA A 2DNAA IATDG-DTNAA

  18. Modes of overinitiation, dnaA gene expression, and inhibition of cell division in a novel cold-sensitive hda mutant of Escherichia coli.

    Science.gov (United States)

    Fujimitsu, Kazuyuki; Su'etsugu, Masayuki; Yamaguchi, Yoko; Mazda, Kensaku; Fu, Nisi; Kawakami, Hironori; Katayama, Tsutomu

    2008-08-01

    The chromosomal replication cycle is strictly coordinated with cell cycle progression in Escherichia coli. ATP-DnaA initiates replication, leading to loading of the DNA polymerase III holoenzyme. The DNA-loaded form of the beta clamp subunit of the polymerase binds the Hda protein, which promotes ATP-DnaA hydrolysis, yielding inactive ADP-DnaA. This regulation is required to repress overinitiation. In this study, we have isolated a novel cold-sensitive hda mutant, the hda-185 mutant. The hda-185 mutant caused overinitiation of chromosomal replication at 25 degrees C, which most likely led to blockage of replication fork progress. Consistently, the inhibition of colony formation at 25 degrees C was suppressed by disruption of the diaA gene, an initiation stimulator. Disruption of the seqA gene, an initiation inhibitor, showed synthetic lethality with hda-185 even at 42 degrees C. The cellular ATP-DnaA level was increased in an hda-185-dependent manner. The cellular concentrations of DnaA protein and dnaA mRNA were comparable at 25 degrees C to those in a wild-type hda strain. We also found that multiple copies of the ribonucleotide reductase genes (nrdAB or nrdEF) or dnaB gene repressed overinitiation. The cellular levels of dATP and dCTP were elevated in cells bearing multiple copies of nrdAB. The catalytic site within NrdA was required for multicopy suppression, suggesting the importance of an active form of NrdA or elevated levels of deoxyribonucleotides in inhibition of overinitiation in the hda-185 cells. Cell division in the hda-185 mutant was inhibited at 25 degrees C in a LexA regulon-independent manner, suggesting that overinitiation in the hda-185 mutant induced a unique division inhibition pathway.

  19. Modes of Overinitiation, dnaA Gene Expression, and Inhibition of Cell Division in a Novel Cold-Sensitive hda Mutant of Escherichia coli▿

    Science.gov (United States)

    Fujimitsu, Kazuyuki; Su'etsugu, Masayuki; Yamaguchi, Yoko; Mazda, Kensaku; Fu, Nisi; Kawakami, Hironori; Katayama, Tsutomu

    2008-01-01

    The chromosomal replication cycle is strictly coordinated with cell cycle progression in Escherichia coli. ATP-DnaA initiates replication, leading to loading of the DNA polymerase III holoenzyme. The DNA-loaded form of the β clamp subunit of the polymerase binds the Hda protein, which promotes ATP-DnaA hydrolysis, yielding inactive ADP-DnaA. This regulation is required to repress overinitiation. In this study, we have isolated a novel cold-sensitive hda mutant, the hda-185 mutant. The hda-185 mutant caused overinitiation of chromosomal replication at 25°C, which most likely led to blockage of replication fork progress. Consistently, the inhibition of colony formation at 25°C was suppressed by disruption of the diaA gene, an initiation stimulator. Disruption of the seqA gene, an initiation inhibitor, showed synthetic lethality with hda-185 even at 42°C. The cellular ATP-DnaA level was increased in an hda-185-dependent manner. The cellular concentrations of DnaA protein and dnaA mRNA were comparable at 25°C to those in a wild-type hda strain. We also found that multiple copies of the ribonucleotide reductase genes (nrdAB or nrdEF) or dnaB gene repressed overinitiation. The cellular levels of dATP and dCTP were elevated in cells bearing multiple copies of nrdAB. The catalytic site within NrdA was required for multicopy suppression, suggesting the importance of an active form of NrdA or elevated levels of deoxyribonucleotides in inhibition of overinitiation in the hda-185 cells. Cell division in the hda-185 mutant was inhibited at 25°C in a LexA regulon-independent manner, suggesting that overinitiation in the hda-185 mutant induced a unique division inhibition pathway. PMID:18502852

  20. Loss of Hda activity stimulates replication initiation from I-box, but not R4 mutant origins in Escherichia coli.

    Science.gov (United States)

    Riber, Leise; Fujimitsu, Kazuyuki; Katayama, Tsutomu; Løbner-Olesen, Anders

    2009-01-01

    Initiation of chromosome replication in Escherichia coli is limited by the initiator protein DnaA associated with ATP. Within the replication origin, binding sites for DnaA associated with ATP or ADP (R boxes) and the DnaA(ATP) specific sites (I-boxes, tau-boxes and 6-mer sites) are found. We analysed chromosome replication of cells carrying mutations in conserved regions of oriC. Cells carrying mutations in DnaA-boxes I2, I3, R2, R3 and R5 as well as FIS and IHF binding sites resembled wild-type cells with respect to origin concentration. Initiation of replication in these mutants occurred in synchrony or with slight asynchrony only. Furthermore, lack of Hda stimulated initiation in all these mutants. The DnaA(ATP) containing complex that leads to initiation can therefore be formed in the absence of several of the origin DnaA binding sites including both DnaA(ATP) specific I-boxes. However, competition between I-box mutant and wild-type origins, revealed a positive role of I-boxes on initiation. On the other hand, mutations affecting DnaA-box R4 were found to be compromised for initiation and could not be augmented by an increase in cellular DnaA(ATP)/DnaA(ADP) ratio. Compared with the sites tested here, R4 therefore seems to contribute to initiation most critically.

  1. Suppression of the Escherichia coli dnaA46 mutation by changes in the activities of the pyruvate-acetate node links DNA replication regulation to central carbon metabolism.

    Science.gov (United States)

    Tymecka-Mulik, Joanna; Boss, Lidia; Maciąg-Dorszyńska, Monika; Matias Rodrigues, João F; Gaffke, Lidia; Wosinski, Anna; Cech, Grzegorz M; Szalewska-Pałasz, Agnieszka; Węgrzyn, Grzegorz; Glinkowska, Monika

    2017-01-01

    To ensure faithful transmission of genetic material to progeny cells, DNA replication is tightly regulated, mainly at the initiation step. Escherichia coli cells regulate the frequency of initiation according to growth conditions. Results of the classical, as well as the latest studies, suggest that the DNA replication in E. coli starts at a predefined, constant cell volume per chromosome but the mechanisms coordinating DNA replication with cell growth are still not fully understood. Results of recent investigations have revealed a role of metabolic pathway proteins in the control of cell division and a direct link between metabolism and DNA replication has also been suggested both in Bacillus subtilis and E. coli cells. In this work we show that defects in the acetate overflow pathway suppress the temperature-sensitivity of a defective replication initiator-DnaA under acetogenic growth conditions. Transcriptomic and metabolic analyses imply that this suppression is correlated with pyruvate accumulation, resulting from alterations in the pyruvate dehydrogenase (PDH) activity. Consequently, deletion of genes encoding the pyruvate dehydrogenase subunits likewise resulted in suppression of the thermal-sensitive growth of the dnaA46 strain. We propose that the suppressor effect may be directly related to the PDH complex activity, providing a link between an enzyme of the central carbon metabolism and DNA replication.

  2. Initiation of protein synthesis in bacteria

    DEFF Research Database (Denmark)

    Laursen, Brian Søgaard; Sørensen, Hans Peter; Mortensen, Kim Kusk

    2005-01-01

    Valuable information on translation initiation is available from biochemical data and recently solved structures. We present a detailed description of current knowledge about the structure, function, and interactions of the individual components involved in bacterial translation initiation. The f...

  3. Initial photoinduced dynamics of the photoactive yellow protein

    NARCIS (Netherlands)

    Larsen, D.S.; van Grondelle, R.

    2005-01-01

    The photoactive yellow protein (PYP) is the photoreceptor protein responsible for initiating the blue-light repellent response of the Halorhodospira halophila bacterium. Optical excitation of the intrinsic chromophore in PYP, p-coumaric acid, leads to the initiation of a photocycle that comprises

  4. Interaction of the Sliding Clamp β-Subunit and Hda, a DnaA-Related Protein

    Science.gov (United States)

    Kurz, Mareike; Dalrymple, Brian; Wijffels, Gene; Kongsuwan, Kritaya

    2004-01-01

    In Escherichia coli, interactions between the replication initiation protein DnaA, the β subunit of DNA polymerase III (the sliding clamp protein), and Hda, the recently identified DnaA-related protein, are required to convert the active ATP-bound form of DnaA to an inactive ADP-bound form through the accelerated hydrolysis of ATP. This rapid hydrolysis of ATP is proposed to be the main mechanism that blocks multiple initiations during cell cycle and acts as a molecular switch from initiation to replication. However, the biochemical mechanism for this crucial step in DNA synthesis has not been resolved. Using purified Hda and β proteins in a plate binding assay and Ni-nitrilotriacetic acid pulldown analysis, we show for the first time that Hda directly interacts with β in vitro. A new β-binding motif, a hexapeptide with the consensus sequence QL[SP]LPL, related to the previously identified β-binding pentapeptide motif (QL[SD]LF) was found in the amino terminus of the Hda protein. Mutants of Hda with amino acid changes in the hexapeptide motif are severely defective in their ability to bind β. A 10-amino-acid peptide containing the E. coli Hda β-binding motif was shown to compete with Hda for binding to β in an Hda-β interaction assay. These results establish that the interaction of Hda with β is mediated through the hexapeptide sequence. We propose that this interaction may be crucial to the events that lead to the inactivation of DnaA and the prevention of excess initiation of rounds of replication. PMID:15150238

  5. Interaction of the sliding clamp beta-subunit and Hda, a DnaA-related protein.

    Science.gov (United States)

    Kurz, Mareike; Dalrymple, Brian; Wijffels, Gene; Kongsuwan, Kritaya

    2004-06-01

    In Escherichia coli, interactions between the replication initiation protein DnaA, the beta subunit of DNA polymerase III (the sliding clamp protein), and Hda, the recently identified DnaA-related protein, are required to convert the active ATP-bound form of DnaA to an inactive ADP-bound form through the accelerated hydrolysis of ATP. This rapid hydrolysis of ATP is proposed to be the main mechanism that blocks multiple initiations during cell cycle and acts as a molecular switch from initiation to replication. However, the biochemical mechanism for this crucial step in DNA synthesis has not been resolved. Using purified Hda and beta proteins in a plate binding assay and Ni-nitrilotriacetic acid pulldown analysis, we show for the first time that Hda directly interacts with beta in vitro. A new beta-binding motif, a hexapeptide with the consensus sequence QL[SP]LPL, related to the previously identified beta-binding pentapeptide motif (QL[SD]LF) was found in the amino terminus of the Hda protein. Mutants of Hda with amino acid changes in the hexapeptide motif are severely defective in their ability to bind beta. A 10-amino-acid peptide containing the E. coli Hda beta-binding motif was shown to compete with Hda for binding to beta in an Hda-beta interaction assay. These results establish that the interaction of Hda with beta is mediated through the hexapeptide sequence. We propose that this interaction may be crucial to the events that lead to the inactivation of DnaA and the prevention of excess initiation of rounds of replication.

  6. Regulation of protein translation initiation in response to ionizing radiation

    International Nuclear Information System (INIS)

    Trivigno, Donatella; Bornes, Laura; Huber, Stephan M; Rudner, Justine

    2013-01-01

    Proliferating tumor cells require continuous protein synthesis. De novo synthesis of most proteins is regulated through cap-dependent translation. Cellular stress such as ionizing radiation (IR) blocks cap-dependent translation resulting in shut-down of global protein translation which saves resources and energy needed for the stress response. At the same time, levels of proteins required for stress response are maintained or even increased. The study aimed to analyze the regulation of signaling pathways controlling protein translation in response to IR and the impact on Mcl-1, an anti-apoptotic and radioprotective protein, which levels rapidly decline upon IR. Protein levels and processing were analyzed by Western blot. The assembly of the translational pre-initiation complex was examined by Immunoprecipitation and pull-down experiments with 7-methyl GTP agarose. To analyze IR-induced cell death, dissipation of the mitochondrial membrane potential and DNA fragmentation were determined by flow cytometry. Protein levels of the different initiation factors were down-regulated using RNA interference approach. IR induced caspase-dependent cleavage of the translational initiation factors eIF4G1, eIF3A, and eIF4B resulting in disassembly of the cap-dependent initiation complex. In addition, DAP5-dependent initiation complex that regulates IRES-dependent translation was disassembled in response to IR. Moreover, IR resulted in dephosphorylation of 4EBP1, an inhibitor of cap-dependent translation upstream of caspase activation. However, knock-down of eIF4G1, eIF4B, DAP5, or 4EBP1 did not affect IR-induced decline of the anti-apoptotic protein Mcl-1. Our data shows that cap-dependent translation is regulated at several levels in response to IR. However, the experiments indicate that IR-induced Mcl-1 decline is not a consequence of translational inhibition in Jurkat cells

  7. Regulation of protein translation initiation in response to ionizing radiation

    Directory of Open Access Journals (Sweden)

    Trivigno Donatella

    2013-02-01

    Full Text Available Abstract Background Proliferating tumor cells require continuous protein synthesis. De novo synthesis of most proteins is regulated through cap-dependent translation. Cellular stress such as ionizing radiation (IR blocks cap-dependent translation resulting in shut-down of global protein translation which saves resources and energy needed for the stress response. At the same time, levels of proteins required for stress response are maintained or even increased. The study aimed to analyze the regulation of signaling pathways controlling protein translation in response to IR and the impact on Mcl-1, an anti-apoptotic and radioprotective protein, which levels rapidly decline upon IR. Methods Protein levels and processing were analyzed by Western blot. The assembly of the translational pre-initiation complex was examined by Immunoprecipitation and pull-down experiments with 7-methyl GTP agarose. To analyze IR-induced cell death, dissipation of the mitochondrial membrane potential and DNA fragmentation were determined by flow cytometry. Protein levels of the different initiation factors were down-regulated using RNA interference approach. Results IR induced caspase-dependent cleavage of the translational initiation factors eIF4G1, eIF3A, and eIF4B resulting in disassembly of the cap-dependent initiation complex. In addition, DAP5-dependent initiation complex that regulates IRES-dependent translation was disassembled in response to IR. Moreover, IR resulted in dephosphorylation of 4EBP1, an inhibitor of cap-dependent translation upstream of caspase activation. However, knock-down of eIF4G1, eIF4B, DAP5, or 4EBP1 did not affect IR-induced decline of the anti-apoptotic protein Mcl-1. Conclusion Our data shows that cap-dependent translation is regulated at several levels in response to IR. However, the experiments indicate that IR-induced Mcl-1 decline is not a consequence of translational inhibition in Jurkat cells.

  8. Timely binding of IHF and Fis to DARS2 regulates ATP–DnaA production and replication initiation

    Science.gov (United States)

    Kasho, Kazutoshi; Fujimitsu, Kazuyuki; Matoba, Toshihiro; Oshima, Taku; Katayama, Tsutomu

    2014-01-01

    In Escherichia coli, the ATP-bound form of DnaA (ATP–DnaA) promotes replication initiation. During replication, the bound ATP is hydrolyzed to ADP to yield the ADP-bound form (ADP–DnaA), which is inactive for initiation. The chromosomal site DARS2 facilitates the regeneration of ATP–DnaA by catalyzing nucleotide exchange between free ATP and ADP bound to DnaA. However, the regulatory mechanisms governing this exchange reaction are unclear. Here, using in vitro reconstituted experiments, we show that two nucleoid-associated proteins, IHF and Fis, bind site-specifically to DARS2 to activate coordinately the exchange reaction. The regenerated ATP–DnaA was fully active in replication initiation and underwent DnaA–ATP hydrolysis. ADP–DnaA formed heteromultimeric complexes with IHF and Fis on DARS2, and underwent nucleotide dissociation more efficiently than ATP–DnaA. Consistently, mutant analyses demonstrated that specific binding of IHF and Fis to DARS2 stimulates the formation of ATP–DnaA production, thereby promoting timely initiation. Moreover, we show that IHF–DARS2 binding is temporally regulated during the cell cycle, whereas Fis only binds to DARS2 in exponentially growing cells. These results elucidate the regulation of ATP–DnaA and replication initiation in coordination with the cell cycle and growth phase. PMID:25378325

  9. Timely binding of IHF and Fis to DARS2 regulates ATP-DnaA production and replication initiation.

    Science.gov (United States)

    Kasho, Kazutoshi; Fujimitsu, Kazuyuki; Matoba, Toshihiro; Oshima, Taku; Katayama, Tsutomu

    2014-12-01

    In Escherichia coli, the ATP-bound form of DnaA (ATP-DnaA) promotes replication initiation. During replication, the bound ATP is hydrolyzed to ADP to yield the ADP-bound form (ADP-DnaA), which is inactive for initiation. The chromosomal site DARS2 facilitates the regeneration of ATP-DnaA by catalyzing nucleotide exchange between free ATP and ADP bound to DnaA. However, the regulatory mechanisms governing this exchange reaction are unclear. Here, using in vitro reconstituted experiments, we show that two nucleoid-associated proteins, IHF and Fis, bind site-specifically to DARS2 to activate coordinately the exchange reaction. The regenerated ATP-DnaA was fully active in replication initiation and underwent DnaA-ATP hydrolysis. ADP-DnaA formed heteromultimeric complexes with IHF and Fis on DARS2, and underwent nucleotide dissociation more efficiently than ATP-DnaA. Consistently, mutant analyses demonstrated that specific binding of IHF and Fis to DARS2 stimulates the formation of ATP-DnaA production, thereby promoting timely initiation. Moreover, we show that IHF-DARS2 binding is temporally regulated during the cell cycle, whereas Fis only binds to DARS2 in exponentially growing cells. These results elucidate the regulation of ATP-DnaA and replication initiation in coordination with the cell cycle and growth phase. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

  10. Influence of Translation Initiation on Organellar Protein Targeting in Arabidopsis

    Energy Technology Data Exchange (ETDEWEB)

    Sally A. Mackenzie

    2011-04-18

    A primary focus of the Mackenzie laboratory is the elucidation of processes and machinery for mitochondrial genome maintenance and transmission in higher plants. We have found that numerous organellar DNA maintenance components in plants appear to be dual targeted to mitochondria and plastids. Of particular interest was the observation that some twin (tandemly arrayed) dual targeting presequences appeared to utilize non-AUG alternative translation initiation, allowing for multiple translation starts at a single gene. Two aspects of this phenomenon were of particular interest: (1) Alternative translation initiation might provide a mechanism to regulate protein targeting temporally and spatially, a possibility that had not been demonstrated previously, and (2) alternative translation initiation might occur in genes involved in nuclear-controlled mitochondrial genome recombination, thought to be exclusively mitochondrial in their function. During the course of this research, we pursued three aims, with an emphasis on two specific genes of interest: POLgamma2, an organellar DNA polymerase, and MSH1, a MutS homolog thought to participate in mitochondrial, but not plastid, genome recombination surveillance. Our aims were to (1) Identify additional genes within Arabidopsis and other genomes that employ non-AUG alternative translation initiation, (2) Locate sequences upstream to the annotated AUG that confer alternative non-AUG translation initiation activity, and (3) Identify cis and trans factors that influence start site selection in genes with non-AUG starts. Toward these ends, we have shown that non-AUG initiation occurs in a number of genes, likely influencing targeting behavior of the protein. We have also shown that start site selection is strongly influenced by Kozak consensus sequence environment, indicating that alternative translation initiation in plants occurs by relaxation of ribosome scanning.

  11. Translation initiation mediated by nuclear cap-binding protein complex.

    Science.gov (United States)

    Ryu, Incheol; Kim, Yoon Ki

    2017-04-01

    In mammals, cap-dependent translation of mRNAs is initiated by two distinct mechanisms: cap-binding complex (CBC; a heterodimer of CBP80 and 20)-dependent translation (CT) and eIF4E-dependent translation (ET). Both translation initiation mechanisms share common features in driving cap- dependent translation; nevertheless, they can be distinguished from each other based on their molecular features and biological roles. CT is largely associated with mRNA surveillance such as nonsense-mediated mRNA decay (NMD), whereas ET is predominantly involved in the bulk of protein synthesis. However, several recent studies have demonstrated that CT and ET have similar roles in protein synthesis and mRNA surveillance. In a subset of mRNAs, CT preferentially drives the cap-dependent translation, as ET does, and ET is responsible for mRNA surveillance, as CT does. In this review, we summarize and compare the molecular features of CT and ET with a focus on the emerging roles of CT in translation. [BMB Reports 2017; 50(4): 186-193].

  12. DNA-A of a highly pathogenic Indian cassava mosaic virus isolated from Jatropha curcas causes symptoms in Nicotiana benthamiana.

    Science.gov (United States)

    Wang, Gang; Sun, Yanwei; Xu, Ruirui; Qu, Jing; Tee, Chuansia; Jiang, Xiyuan; Ye, Jian

    2014-04-01

    Jatropha curcas mosaic disease (JcMD) is a newly emerging disease that has been reported in Africa and India. Here, we report the complete nucleotide sequence of a new Indian cassava mosaic virus isolate (ICMV-SG) from Singapore. Infection of ICMV-SG showed more severe JcMD in Jatropha curcas and Nicotiana benthamiana than the other ICMV isolates reported previously, though ICMV-SG shares high sequence identity with the other ICMV isolates. Agroinfectious DNA-A alone sufficiently induced systemic symptoms in N. benthamiana, but not in J. curcas. Results from agroinfection assays showed that systemic infection of ICMV-SG in J. curcas required both DNA-A and DNA-B components.

  13. Induction of protein X in Escherichia coli

    International Nuclear Information System (INIS)

    Little, J.W.; Hanawalt, P.C.

    1977-01-01

    The authors have examined some of the treatments that might induce protein X and they have, in particular, tested the hypothesis that DNA degradation products play an essential role in the induction process. UV irradiation, nalidixic acid treatment, or thymine starvation result in protein X synthesis in wild type strains. However, UV irradiation, unlike nalidixic acid, also induced protein X in recB strains, in which little DNA degradation occurs. The presence of DNA fragments resulting from hostcontrolled restriction of phage lambda DNA did not affect protein X synthesis. It was concluded that no causal relationship exists bewteen the production of DNA fragments and induction of protein X. The presence of the plasmid R 46, which confers enhanced mutagenesis and UV resistance on its host, did not affect protein X synthesis. Growth in the presence of 5-bromouracil, which does not result in production of degradation fragments, resulted eventually in a low rate of protein X synthesis. In dnaA mutants, deficient in the initiation of new rounds of replication, UV irradiation induced protein X, again unlike nalidixic acid. Thus, the inhibition of active replication forks is not an essential requirement for protein X induction. (orig./MG) [de

  14. Loss of Hda activity stimulates replication initiation from I-box, but not R4 mutant origins in Escherichia coli

    DEFF Research Database (Denmark)

    Riber, Leise; Fujimitsu, K.; Katayama, T.

    2009-01-01

    in synchrony or with slight asynchrony only. Furthermore, lack of Hda stimulated initiation in all these mutants. The DnaA(ATP) containing complex that leads to initiation can therefore be formed in the absence of several of the origin DnaA binding sites including both DnaA(ATP) specific I-boxes. However...

  15. Isolation and sequence characterization of DNA-A genome of a new begomovirus strain associated with severe leaf curling symptoms of Jatropha curcas L.

    KAUST Repository

    Chauhan, Sushma

    2018-04-22

    Begomoviruses belong to the family Geminiviridae are associated with several disease symptoms, such as mosaic and leaf curling in Jatropha curcas. The molecular characterization of these viral strains will help in developing management strategies to control the disease. In this study, J. curcas that was infected with begomovirus and showed acute leaf curling symptoms were identified. DNA-A segment from pathogenic viral strain was isolated and sequenced. The sequenced genome was assembled and characterized in detail. The full-length DNA-A sequence was covered by primer walking. The genome sequence showed the general organization of DNA-A from begomovirus by the distribution of ORFs in both viral and anti-viral strands. The genome size ranged from 2844 bp–2852 bp. Three strains with minor nucleotide variations were identified, and a phylogenetic analysis was performed by comparing the DNA-A segments from other reported begomovirus isolates. The maximum sequence similarity was observed with Euphorbia yellow mosaic virus (FN435995). In the phylogenetic tree, no clustering was observed with previously reported begomovirus strains isolated from J. curcas host. The strains isolated in this study belong to new begomoviral strain that elicits symptoms of leaf curling in J. curcas. The results indicate that the probable origin of the strains is from Jatropha mosaic virus infecting J. gassypifolia. The strains isolated in this study are referred as Jatropha curcas leaf curl India virus (JCLCIV) based on the major symptoms exhibited by host J. curcas.

  16. Specificity and function of Archaeal DNA replication initiator proteins

    DEFF Research Database (Denmark)

    Samson, Rachel Y.; Xu, Yanqun; Gadelha, Catarina

    2013-01-01

    Chromosomes with multiple DNA replication origins are a hallmark of Eukaryotes and some Archaea. All eukaryal nuclear replication origins are defined by the origin recognition complex (ORC) that recruits the replicative helicase MCM(2-7) via Cdc6 and Cdt1. We find that the three origins...... to investigate the role of ATP binding and hydrolysis in initiator function in vivo and in vitro. We find that the ATP-bound form of Orc1-1 is proficient for replication and implicates hydrolysis of ATP in downregulation of origin activity. Finally, we reveal that ATP and DNA binding by Orc1-1 remodels...

  17. Specificity and Function of Archaeal DNA Replication Initiator Proteins

    Directory of Open Access Journals (Sweden)

    Rachel Y. Samson

    2013-02-01

    Full Text Available Chromosomes with multiple DNA replication origins are a hallmark of Eukaryotes and some Archaea. All eukaryal nuclear replication origins are defined by the origin recognition complex (ORC that recruits the replicative helicase MCM(2-7 via Cdc6 and Cdt1. We find that the three origins in the single chromosome of the archaeon Sulfolobus islandicus are specified by distinct initiation factors. While two origins are dependent on archaeal homologs of eukaryal Orc1 and Cdc6, the third origin is instead reliant on an archaeal Cdt1 homolog. We exploit the nonessential nature of the orc1-1 gene to investigate the role of ATP binding and hydrolysis in initiator function in vivo and in vitro. We find that the ATP-bound form of Orc1-1 is proficient for replication and implicates hydrolysis of ATP in downregulation of origin activity. Finally, we reveal that ATP and DNA binding by Orc1-1 remodels the protein’s structure rather than that of the DNA template.

  18. Effect of initial protein concentration and pH on in vitro gastric digestion of heated whey proteins.

    Science.gov (United States)

    Zhang, Sha; Vardhanabhuti, Bongkosh

    2014-02-15

    The in vitro digestion of heated whey protein aggregates having different structure and physicochemical properties was evaluated under simulated gastric conditions. Aggregates were formed by heating whey protein isolates (WPI) at 3-9% w/w initial protein concentration and pH 3.0-7.0. Results showed that high protein concentration led to formation of larger WPI aggregates with fewer remaining monomers. Aggregates formed at high protein concentrations showed slower degradation rate compared to those formed at low protein concentration. The effect of initial protein concentration on peptide release pattern was not apparent. Heating pH was a significant factor affecting digestion pattern. At pH above the isoelectric point, the majority of the proteins involved in the aggregation, and aggregates formed at pH 6.0 were more susceptible to pepsin digestion than at pH 7.0. At acidic conditions, only small amount of proteins was involved in the aggregation and heated aggregates were easily digested by pepsin, while the remaining unaggregated proteins were very resistant to gastric digestion. The potential physiological implication of these results on satiety was discussed. Copyright © 2013 Elsevier Ltd. All rights reserved.

  19. Physical interactions between bacteriophage and Escherichia coli proteins required for initiation of lambda DNA replication.

    Science.gov (United States)

    Liberek, K; Osipiuk, J; Zylicz, M; Ang, D; Skorko, J; Georgopoulos, C

    1990-02-25

    The process of initiation of lambda DNA replication requires the assembly of the proper nucleoprotein complex at the origin of replication, ori lambda. The complex is composed of both phage and host-coded proteins. The lambda O initiator protein binds specifically to ori lambda. The lambda P initiator protein binds to both lambda O and the host-coded dnaB helicase, giving rise to an ori lambda DNA.lambda O.lambda P.dnaB structure. The dnaK and dnaJ heat shock proteins have been shown capable of dissociating this complex. The thus freed dnaB helicase unwinds the duplex DNA template at the replication fork. In this report, through cross-linking, size chromatography, and protein affinity chromatography, we document some of the protein-protein interactions occurring at ori lambda. Our results show that the dnaK protein specifically interacts with both lambda O and lambda P, and that the dnaJ protein specifically interacts with the dnaB helicase.

  20. CRISPR-mediated control of the bacterial initiation of replication.

    Science.gov (United States)

    Wiktor, Jakub; Lesterlin, Christian; Sherratt, David J; Dekker, Cees

    2016-05-05

    Programmable control of the cell cycle has been shown to be a powerful tool in cell-biology studies. Here, we develop a novel system for controlling the bacterial cell cycle, based on binding of CRISPR/dCas9 to the origin-of-replication locus. Initiation of replication of bacterial chromosomes is accurately regulated by the DnaA protein, which promotes the unwinding of DNA at oriC We demonstrate that the binding of CRISPR/dCas9 to any position within origin or replication blocks the initiation of replication. Serial-dilution plating, single-cell fluorescence microscopy, and flow-cytometry experiments show that ongoing rounds of chromosome replication are finished upon CRISPR/dCas9 binding, but no new rounds are initiated. Upon arrest, cells stay metabolically active and accumulate cell mass. We find that elevating the temperature from 37 to 42°C releases the CRISR/dCas9 replication inhibition, and we use this feature to recover cells from the arrest. Our simple and robust method of controlling the bacterial cell cycle is a useful asset for synthetic biology and DNA-replication studies in particular. The inactivation of CRISPR/dCas9 binding at elevated temperatures may furthermore be of wide interest for CRISPR/Cas9 applications in genomic engineering. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  1. Enhanced protein retention on poly(caprolactone) via surface initiated polymerization of acrylamide

    International Nuclear Information System (INIS)

    Ma, Yuhao; Cai, Mengtan; He, Liu; Luo, Xianglin

    2016-01-01

    Graphical abstract: - Highlights: • Dense package of poly(acrylamide) on poly(caprolactone) surface was achieved by surface-initiated atom transfer radical polymerization. • Poly(acrylamide) grafted surface exhibited high protein retention ability. • Loaded protein was resistant to detachment and maintained its structure without denaturation. - Abstract: To enhance the biocompatibility or extend the biomedical application of poly(caprolactone) (PCL), protein retention on PCL surface is often required. In this study, poly(acrylamide) (PAAm) brushes were grown from PCL surface via surface-initiated atom transfer radical polymerization (SI-ATRP) and served as a protein-capturing platform. Grafted PAAm was densely packed on surface and exhibited superior protein retention ability. Captured protein was found to be resistant to washing under detergent environment. Furthermore, protein structure after being captured was investigated by circular dichroism (CD) spectroscopy, and the CD spectra verified that secondary structure of captured proteins was maintained, indicating no denaturation of protein happened for retention process.

  2. Isolation and sequence characterization of DNA-A genome of a new begomovirus strain associated with severe leaf curling symptoms of Jatropha curcas L.

    Science.gov (United States)

    Chauhan, Sushma; Rahman, Hifzur; Mastan, Shaik G; Pamidimarri, D V N Sudheer; Reddy, Muppala P

    2018-07-20

    Begomoviruses belong to the family Geminiviridae are associated with several disease symptoms, such as mosaic and leaf curling in Jatropha curcas. The molecular characterization of these viral strains will help in developing management strategies to control the disease. In this study, J. curcas that was infected with begomovirus and showed acute leaf curling symptoms were identified. DNA-A segment from pathogenic viral strain was isolated and sequenced. The sequenced genome was assembled and characterized in detail. The full-length DNA-A sequence was covered by primer walking. The genome sequence showed the general organization of DNA-A from begomovirus by the distribution of ORFs in both viral and anti-viral strands. The genome size ranged from 2844 bp-2852 bp. Three strains with minor nucleotide variations were identified, and a phylogenetic analysis was performed by comparing the DNA-A segments from other reported begomovirus isolates. The maximum sequence similarity was observed with Euphorbia yellow mosaic virus (FN435995). In the phylogenetic tree, no clustering was observed with previously reported begomovirus strains isolated from J. curcas host. The strains isolated in this study belong to new begomoviral strain that elicits symptoms of leaf curling in J. curcas. The results indicate that the probable origin of the strains is from Jatropha mosaic virus infecting J. gassypifolia. The strains isolated in this study are referred as Jatropha curcas leaf curl India virus (JCLCIV) based on the major symptoms exhibited by host J. curcas. Copyright © 2018 Elsevier B.V. All rights reserved.

  3. Heat-shock proteins in stromal joint tissues: innocent bystanders or disease-initiating proteins?

    Science.gov (United States)

    Lambrecht, Stijn; Juchtmans, Nele; Elewaut, Dirk

    2014-02-01

    Heat-shock proteins (HSPs) are molecular chaperones that are highly conserved between species. In recent decades it has become clear that these proteins play an important role in the pathogenesis of inflammatory and degenerative joint diseases by (dys)regulating the immune system and by direct effects on the stromal tissues of the joint. In this review we discuss current insights into the expression pattern of HSPs in connective tissues, the direct biological role of HSPs in stromal tissues and the potential clinical applications.

  4. Initial steps of signal generation in photoactive yellow protein revealed with femtosecond mid-infrared spectroscopy

    NARCIS (Netherlands)

    Groot, M.L.; van Wilderen, L.; Larsen, D.S.; Horst, M.A.; van Stokkum, I.H.M.; Hellingwerf, K.J.; van Grondelle, R.

    2003-01-01

    Photoactive yellow protein (PYP) is a bacterial blue light sensor that induces Halorhodospira halophila to swim away from intense blue light. Light absorption by PYP's intrinsic chromophore, p-coumaric acid, leads to the initiation of a photocycle that comprises several distinct intermediates. Here

  5. Initial crystallographic studies of a small heat-shock protein from Xylella fastidiosa

    International Nuclear Information System (INIS)

    Tada, Susely F. S.; Saraiva, Antonio Marcos; Lorite, Gabriela S.; Rosselli-Murai, Luciana K.; Pelloso, Alexandre César; Santos, Marcelo Leite dos; Trivella, Daniela B. B.; Cotta, Mônica A.; Souza, Anete Pereira de; Aparicio, Ricardo

    2012-01-01

    Initial crystallographic studies of the X. fastidiosa small heat-shock protein HSP17.9 are reported. The ORF XF2234 in the Xylella fastidiosa genome was identified as encoding a small heat-shock protein of 17.9 kDa (HSP17.9). HSP17.9 was found as one of the proteins that are induced during X. fastidiosa proliferation and infection in citrus culture. Recombinant HSP17.9 was crystallized and surface atomic force microscopy experiments were conducted with the aim of better characterizing the HSP17.9 crystals. X-ray diffraction data were collected at 2.7 Å resolution. The crystal belonged to space group P4 3 22, with unit-cell parameters a = 68.90, b = 68.90, c = 72.51 Å, and is the first small heat-shock protein to crystallize in this space group

  6. Protein synthesis during the initial phase of the temperature-induced bleaching response in Euglena gracilis

    International Nuclear Information System (INIS)

    Ortiz, W.

    1990-01-01

    Growing cultures of photoheterotrophic Euglena gracilis experience an increase in chlorophyll accumulation during the initial phase of the temperature-induced bleaching response suggesting an increase in the synthesis of plastid components at the bleaching temperature of 33 degree C. A primary goal of this work was to establish whether an increase in the synthesis of plastid proteins accompanies the observed increase in chlorophyll accumulation. In vivo pulse-labeling experiments with [ 35 S]sodium sulfate were carried out with cells grown at room temperature or at 33 degree C. The synthesis of a number of plastid polypeptides of nucleocytoplasmic origin, including some presumably novel polypeptides, increased in cultures treated for 15 hours at 33 degree C. In contrast, while synthesis of thylakoid proteins by the plastid protein synthesis machinery decreased modestly, synthesis of the large subunit of the enzyme ribulosebisphosphate carboxylase was strongly affected at the elevated temperature. Synthesis of novel plastid-encoded polypeptides was not induced at the bleaching temperature. It is concluded that protein synthesis in plastids declines during the initial phase of the temperature response in Euglena despite an overall increase in cellular protein synthesis and an increase in chlorophyll accumulation per cell

  7. RecO protein initiates DNA recombination and strand annealing through two alternative DNA binding mechanisms.

    Science.gov (United States)

    Ryzhikov, Mikhail; Gupta, Richa; Glickman, Michael; Korolev, Sergey

    2014-10-17

    Recombination mediator proteins (RMPs) are important for genome stability in all organisms. Several RMPs support two alternative reactions: initiation of homologous recombination and DNA annealing. We examined mechanisms of RMPs in both reactions with Mycobacterium smegmatis RecO (MsRecO) and demonstrated that MsRecO interacts with ssDNA by two distinct mechanisms. Zinc stimulates MsRecO binding to ssDNA during annealing, whereas the recombination function is zinc-independent and is regulated by interaction with MsRecR. Thus, different structural motifs or conformations of MsRecO are responsible for interaction with ssDNA during annealing and recombination. Neither annealing nor recombinase loading depends on MsRecO interaction with the conserved C-terminal tail of single-stranded (ss) DNA-binding protein (SSB), which is known to bind Escherichia coli RecO. However, similarly to E. coli proteins, MsRecO and MsRecOR do not dismiss SSB from ssDNA, suggesting that RMPs form a complex with SSB-ssDNA even in the absence of binding to the major protein interaction motif. We propose that alternative conformations of such complexes define the mechanism by which RMPs initiate the repair of stalled replication and support two different functions during recombinational repair of DNA breaks. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  8. Self-assembly of protein-based biomaterials initiated by titania nanotubes.

    Science.gov (United States)

    Forstater, Jacob H; Kleinhammes, Alfred; Wu, Yue

    2013-12-03

    Protein-based biomaterials are a promising strategy for creating robust highly selective biocatalysts. The assembled biomaterials must sufficiently retain the near-native structure of proteins and provide molecular access to catalytically active sites. These requirements often exclude the use of conventional assembly techniques, which rely on covalent cross-linking of proteins or entrapment within a scaffold. Here we demonstrate that titania nanotubes can initiate and template the self-assembly of enzymes, such as ribonuclease A, while maintaining their catalytic activity. Initially, the enzymes form multilayer thick ellipsoidal aggregates centered on the nanotube surface; subsequently, these nanosized entities assemble into a micrometer-sized enzyme material that has enhanced enzymatic activity and contains as little as 0.1 wt % TiO2 nanotubes. This phenomenon is uniquely associated with the active anatase (001)-like surface of titania nanotubes and does not occur on other anatase nanomaterials, which contain significantly fewer undercoordinated Ti surface sites. These findings present a nanotechnology-enabled mechanism of biomaterial growth and open a new route for creating stable protein-based biomaterials and biocatalysts without the need for chemical modification.

  9. Phosphorylation of protein synthesis initiation factor 2 (elF-2) in the yeast Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    Romero, D.P.

    1986-01-01

    Initiation Factor 2 (elF-2) in the yeast Saccharomyces cerevisiae is comprised of 3 subunits. The control of protein synthesis in mammalian cells have been shown to involve the phosphorylation of the small (alpha) subunit by a specific protein kinase. Phosphorylation results in an inhibition of protein synthesis. In order to determine whether or not an analogous system is operative in yeast, the phosphorylation state of the alpha subunit of elF-2 in Saccharomyces was determined during various growth and nongrowth conditions. Cells were radiolabelled with 32 P and 35 S, and the whole cell lysates were analyzed by two dimensional gel electrophoresis. These experiments revealed that the smallest subunit (alpha, M/sub r/ = 31,000) is a phosphoprotein in vivo under a variety of growth and nongrowth conditions. This is in direct contrast to the pattern exhibited in mammalian cells. The fact that the small subunit of elF-2 in yeast is phosphorylated under a variety of physiological conditions indicates that such a covalent modification is important for some aspects of elF-2 function. In order to investigate this problem further, a protein kinase that specifically labels the alpha subunit of elF-2 in vitro was isolated. The kinase is not autophosphorylating, utilizes ATP as a phosphate donor, phosphorylates an exogenous protein, casein, modifies serine residues in elF-2, is cyclic nucleotide-independent, and is strongly inhibited by heparin

  10. Laboratory information management system for membrane protein structure initiative--from gene to crystal.

    Science.gov (United States)

    Troshin, Petr V; Morris, Chris; Prince, Stephen M; Papiz, Miroslav Z

    2008-12-01

    Membrane Protein Structure Initiative (MPSI) exploits laboratory competencies to work collaboratively and distribute work among the different sites. This is possible as protein structure determination requires a series of steps, starting with target selection, through cloning, expression, purification, crystallization and finally structure determination. Distributed sites create a unique set of challenges for integrating and passing on information on the progress of targets. This role is played by the Protein Information Management System (PIMS), which is a laboratory information management system (LIMS), serving as a hub for MPSI, allowing collaborative structural proteomics to be carried out in a distributed fashion. It holds key information on the progress of cloning, expression, purification and crystallization of proteins. PIMS is employed to track the status of protein targets and to manage constructs, primers, experiments, protocols, sample locations and their detailed histories: thus playing a key role in MPSI data exchange. It also serves as the centre of a federation of interoperable information resources such as local laboratory information systems and international archival resources, like PDB or NCBI. During the challenging task of PIMS integration, within the MPSI, we discovered a number of prerequisites for successful PIMS integration. In this article we share our experiences and provide invaluable insights into the process of LIMS adaptation. This information should be of interest to partners who are thinking about using LIMS as a data centre for their collaborative efforts.

  11. Phosphorylation in vitro of eukaryotic initiation factors IF-E2 and IF-E3 by protein kinases

    DEFF Research Database (Denmark)

    Issinger, O G; Benne, R; Hershey, J W

    1976-01-01

    Purified protein synthesis initiation factors IF-E2 and IF-E3 from rabbit reticulocytes were phosphorylated in vitro with protein kinases isolated from the same source. The highest levels of phosphorylation resulted from incubation of the factors with a cyclic nucleotide-independent protein kinase...

  12. DNA structure modulates the oligomerization properties of the AAV initiator protein Rep68.

    Directory of Open Access Journals (Sweden)

    Jorge Mansilla-Soto

    2009-07-01

    Full Text Available Rep68 is a multifunctional protein of the adeno-associated virus (AAV, a parvovirus that is mostly known for its promise as a gene therapy vector. In addition to its role as initiator in viral DNA replication, Rep68 is essential for site-specific integration of the AAV genome into human chromosome 19. Rep68 is a member of the superfamily 3 (SF3 helicases, along with the well-studied initiator proteins simian virus 40 large T antigen (SV40-LTag and bovine papillomavirus (BPV E1. Structurally, SF3 helicases share two domains, a DNA origin interaction domain (OID and an AAA(+ motor domain. The AAA(+ motor domain is also a structural feature of cellular initiators and it functions as a platform for initiator oligomerization. Here, we studied Rep68 oligomerization in vitro in the presence of different DNA substrates using a variety of biophysical techniques and cryo-EM. We found that a dsDNA region of the AAV origin promotes the formation of a complex containing five Rep68 subunits. Interestingly, non-specific ssDNA promotes the formation of a double-ring Rep68, a known structure formed by the LTag and E1 initiator proteins. The Rep68 ring symmetry is 8-fold, thus differing from the hexameric rings formed by the other SF3 helicases. However, similiar to LTag and E1, Rep68 rings are oriented head-to-head, suggesting that DNA unwinding by the complex proceeds bidirectionally. This novel Rep68 quaternary structure requires both the DNA binding and AAA(+ domains, indicating cooperativity between these regions during oligomerization in vitro. Our study clearly demonstrates that Rep68 can oligomerize through two distinct oligomerization pathways, which depend on both the DNA structure and cooperativity of Rep68 domains. These findings provide insight into the dynamics and oligomeric adaptability of Rep68 and serve as a step towards understanding the role of this multifunctional protein during AAV DNA replication and site-specific integration.

  13. Crystallization and initial X-ray analysis of polyhydroxyalkanoate granule-associated protein from Aeromonas hydrophila

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Minglian; Li, Zhenguo; Zheng, Wei; Lou, Zhiyong [MOE Key Laboratory of Protein Science, Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084 (China); Chen, Guo-Qiang, E-mail: chengq@stu.edu.cn [MOE Key Laboratory of Protein Science, Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084 (China); Multidisciplinary Research Center, Shantou University, Shantou 515063, Guangdong (China)

    2006-08-01

    The phasin PhaP{sub Ah} from A. hydrophila strain 4AK4 was crystallized using the hanging-drop vapour-diffusion method. Polyhydroxyalkanoate (PHA) granule-associated proteins (phasins) were discovered in PHA-accumulating bacteria. They play a crucial role as a structural protein during initial PHA-granule formation and granule growth and also serve as interfaces for granule stabilization in vivo. The phasin PhaP{sub Ah} from Aeromonas hydrophila strain 4AK4 was crystallized using the hanging-drop vapour-diffusion method. Single crystals were cryocooled for X-ray diffraction analysis. The phasin crystals belonged to space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 80.8, b = 108.9, c = 134.4 Å.

  14. Photo-initiated dynamics and spectroscopy of the deprotonated Green Fluorescent Protein chromophore

    DEFF Research Database (Denmark)

    Bochenkova, Anastasia; Andersen, Lars Henrik

    2013-01-01

    . Knowledge of intrinsic properties of the GFP photoabsorbing molecular unit is a prerequisite in understanding the atomic-scale interactions that play a key role for the diverse functioning of these proteins. Here, we show how recent developments in action and photoelectron spectroscopy combined with state...... efficiently compete with each other in spite of their inherently different intrinsic timescales. The reason behind this is an efficient coupling between the nuclear and electronic motion in the photo-initiated dynamics, where the energy may be transferred from nuclei to electrons and from electrons to nuclei...

  15. In silico platform for predicting and initiating β-turns in a protein at desired locations.

    Science.gov (United States)

    Singh, Harinder; Singh, Sandeep; Raghava, Gajendra P S

    2015-05-01

    Numerous studies have been performed for analysis and prediction of β-turns in a protein. This study focuses on analyzing, predicting, and designing of β-turns to understand the preference of amino acids in β-turn formation. We analyzed around 20,000 PDB chains to understand the preference of residues or pair of residues at different positions in β-turns. Based on the results, a propensity-based method has been developed for predicting β-turns with an accuracy of 82%. We introduced a new approach entitled "Turn level prediction method," which predicts the complete β-turn rather than focusing on the residues in a β-turn. Finally, we developed BetaTPred3, a Random forest based method for predicting β-turns by utilizing various features of four residues present in β-turns. The BetaTPred3 achieved an accuracy of 79% with 0.51 MCC that is comparable or better than existing methods on BT426 dataset. Additionally, models were developed to predict β-turn types with better performance than other methods available in the literature. In order to improve the quality of prediction of turns, we developed prediction models on a large and latest dataset of 6376 nonredundant protein chains. Based on this study, a web server has been developed for prediction of β-turns and their types in proteins. This web server also predicts minimum number of mutations required to initiate or break a β-turn in a protein at specified location of a protein. © 2015 Wiley Periodicals, Inc.

  16. Phage Lambda P Protein: Trans-Activation, Inhibition Phenotypes and their Suppression

    Science.gov (United States)

    Hayes, Sidney; Erker, Craig; Horbay, Monique A.; Marciniuk, Kristen; Wang, Wen; Hayes, Connie

    2013-01-01

    The initiation of bacteriophage λ replication depends upon interactions between the oriλ DNA site, phage proteins O and P, and E. coli host replication proteins. P exhibits a high affinity for DnaB, the major replicative helicase for unwinding double stranded DNA. The concept of P-lethality relates to the hypothesis that P can sequester DnaB and in turn prevent cellular replication initiation from oriC. Alternatively, it was suggested that P-lethality does not involve an interaction between P and DnaB, but is targeted to DnaA. P-lethality is assessed by examining host cells for transformation by ColE1-type plasmids that can express P, and the absence of transformants is attributed to a lethal effect of P expression. The plasmid we employed enabled conditional expression of P, where under permissive conditions, cells were efficiently transformed. We observed that ColE1 replication and plasmid establishment upon transformation is extremely sensitive to P, and distinguish this effect from P-lethality directed to cells. We show that alleles of dnaB protect the variant cells from P expression. P-dependent cellular filamentation arose in ΔrecA or lexA[Ind-] cells, defective for SOS induction. Replication propagation and restart could represent additional targets for P interference of E. coli replication, beyond the oriC-dependent initiation step. PMID:23389467

  17. Zinc in the prevention of Fe2initiated lipid and protein oxidation

    Directory of Open Access Journals (Sweden)

    M. PAOLA ZAGO

    2000-01-01

    Full Text Available In the present study we characterized the capacity of zinc to protect lipids and proteins from Fe2+-initiated oxidative damage. The effects of zinc on lipid oxidation were investigated in liposomes composed of brain phosphatidylcholine (PC and phosphatidylserine (PS at a molar relationship of 60:40 (PC:PS, 60:40. Lipid oxidation was evaluated as the oxidation of cis-parinaric acid or as the formation of 2-thiobarbituric acid-reactive substances (TBARS. Zinc protected liposomes from Fe2+ (2.5-50 muM-supported lipid oxidation. However, zinc (50 muM did not prevent the oxidative inactivation of glutamine synthelase and glucose 6-phosphate dehydrogenase when rat brain superntants were oxidized in the presence of 5 muM Fe2+ and 0.5 mM H2O2 .We also studied the interactions of zinc with epicatechin in the prevention of liid oxidation in liposomes. The simulaneous addition of 0.5 muM epicatechin (EC and 50 muM zinc or EC separately. Zinc (50 muM also protecte liposomes from the stimulatory effect of aluminum on Fe2+-initiated lipid oxidation. Zinc could play an important role as an antioxidant in biological systems, replacing iron and other metals with pro-oxidant activity from binding sites and interacting with other components of the oxidant defense system.

  18. The Phosphotransfer Protein CD1492 Represses Sporulation Initiation in Clostridium difficile.

    Science.gov (United States)

    Childress, Kevin O; Edwards, Adrianne N; Nawrocki, Kathryn L; Anderson, Sarah E; Woods, Emily C; McBride, Shonna M

    2016-12-01

    The formation of spores is critical for the survival of Clostridium difficile outside the host gastrointestinal tract. Persistence of C. difficile spores greatly contributes to the spread of C. difficile infection (CDI), and the resistance of spores to antimicrobials facilitates the relapse of infection. Despite the importance of sporulation to C. difficile pathogenesis, the molecular mechanisms controlling spore formation are not well understood. The initiation of sporulation is known to be regulated through activation of the conserved transcription factor Spo0A. Multiple regulators influence Spo0A activation in other species; however, many of these factors are not conserved in C. difficile and few novel factors have been identified. Here, we investigated the function of a protein, CD1492, that is annotated as a kinase and was originally proposed to promote sporulation by directly phosphorylating Spo0A. We found that deletion of CD1492 resulted in increased sporulation, indicating that CD1492 is a negative regulator of sporulation. Accordingly, we observed increased transcription of Spo0A-dependent genes in the CD1492 mutant. Deletion of CD1492 also resulted in decreased toxin production in vitro and in decreased virulence in the hamster model of CDI. Further, the CD1492 mutant demonstrated effects on gene expression that are not associated with Spo0A activation, including lower sigD and rstA transcription, suggesting that this protein interacts with factors other than Spo0A. Altogether, the data indicate that CD1492 negatively affects sporulation and positively influences motility and virulence. These results provide further evidence that C. difficile sporulation is regulated differently from that of other endospore-forming species. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  19. Photo-initiated crosslinking extends mapping of the protein-protein interface to membrane-embedded portions of cytochromes P450 2B4 and b(5)

    Czech Academy of Sciences Publication Activity Database

    Ječmen, Tomáš; Ptáčková, Renata; Černá, V.; Dračínská, H.; Hodek, P.; Stiborová, M.; Hudeček, J.; Šulc, Miroslav

    2015-01-01

    Roč. 89, NOV 2015 (2015), s. 128-137 ISSN 1046-2023 R&D Projects: GA ČR(CZ) GAP207/12/0627 Grant - others:OPPC(XE) CZ.2.16/3.1.00/24023 Institutional support: RVO:61388971 Keywords : Protein-protein interaction * Trans-membrane segments * Photo-initiated crosslinking Subject RIV: CE - Biochemistry Impact factor: 3.503, year: 2015

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

    DEFF Research Database (Denmark)

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

    1996-01-01

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

  1. Development of human protein reference database as an initial platform for approaching systems biology in humans

    DEFF Research Database (Denmark)

    Peri, Suraj; Navarro, J Daniel; Amanchy, Ramars

    2003-01-01

    Human Protein Reference Database (HPRD) is an object database that integrates a wealth of information relevant to the function of human proteins in health and disease. Data pertaining to thousands of protein-protein interactions, posttranslational modifications, enzyme/substrate relationships...

  2. Tuning of Recombinant Protein Expression in Escherichia coli by Manipulating Transcription, Translation Initiation Rates, and Incorporation of Noncanonical Amino Acids.

    Science.gov (United States)

    Schlesinger, Orr; Chemla, Yonatan; Heltberg, Mathias; Ozer, Eden; Marshall, Ryan; Noireaux, Vincent; Jensen, Mogens Høgh; Alfonta, Lital

    2017-06-16

    Protein synthesis in cells has been thoroughly investigated and characterized over the past 60 years. However, some fundamental issues remain unresolved, including the reasons for genetic code redundancy and codon bias. In this study, we changed the kinetics of the Eschrichia coli transcription and translation processes by mutating the promoter and ribosome binding domains and by using genetic code expansion. The results expose a counterintuitive phenomenon, whereby an increase in the initiation rates of transcription and translation lead to a decrease in protein expression. This effect can be rescued by introducing slow translating codons into the beginning of the gene, by shortening gene length or by reducing initiation rates. On the basis of the results, we developed a biophysical model, which suggests that the density of co-transcriptional-translation plays a role in bacterial protein synthesis. These findings indicate how cells use codon bias to tune translation speed and protein synthesis.

  3. Sonic hedgehog initiates cochlear hair cell regeneration through downregulation of retinoblastoma protein

    Energy Technology Data Exchange (ETDEWEB)

    Lu, Na [Otology Skull Base Surgery Department, Hearing Research Institute, Eye and ENT Hospital of Shanghai Medical School, Fudan University, Shanghai 200031 (China); Department of Otolaryngology and Program in Neuroscience, Harvard Medical School and Eaton Peabody Laboratory, Massachusetts Eye and Ear Infirmary, Boston, MA 02114 (United States); Chen, Yan [Central Laboratory, Hearing Research Institute, Eye and ENT Hospital of Shanghai Medical School, Fudan University, Shanghai 200031 (China); Wang, Zhengmin [Otology Skull Base Surgery Department, Hearing Research Institute, Eye and ENT Hospital of Shanghai Medical School, Fudan University, Shanghai 200031 (China); Institute of Biomedical Sciences, Fudan University, Shanghai 200032 (China); Chen, Guoling [Otology Skull Base Surgery Department, Hearing Research Institute, Eye and ENT Hospital of Shanghai Medical School, Fudan University, Shanghai 200031 (China); Lin, Qin [Otology Skull Base Surgery Department, Hearing Research Institute, Eye and ENT Hospital of Shanghai Medical School, Fudan University, Shanghai 200031 (China); Department of Otolaryngology, First Affiliated Hospital of Fujian Medical University, Otolaryngology Institute of Fujian Province, Fuzhou (China); Chen, Zheng-Yi, E-mail: Zheng-yi_chen@meei.harvard.edu [Department of Otolaryngology and Program in Neuroscience, Harvard Medical School and Eaton Peabody Laboratory, Massachusetts Eye and Ear Infirmary, Boston, MA 02114 (United States); Li, Huawei, E-mail: hwli@shmu.edu.cn [Otology Skull Base Surgery Department, Hearing Research Institute, Eye and ENT Hospital of Shanghai Medical School, Fudan University, Shanghai 200031 (China); Institute of Biomedical Sciences, Fudan University, Shanghai 200032 (China)

    2013-01-11

    Highlights: Black-Right-Pointing-Pointer Shh activation in neonatal cochleae enhances sensory cell proliferation. Black-Right-Pointing-Pointer Proliferating supporting cells can transdifferentiate into hair cells. Black-Right-Pointing-Pointer Shh promotes proliferation by transiently modulating pRb activity. Black-Right-Pointing-Pointer Shh inhibits pRb by inhibiting transcription and increasing phosphorylation of pRb. -- Abstract: Cell cycle re-entry by cochlear supporting cells and/or hair cells is considered one of the best approaches for restoring hearing loss as a result of hair cell damage. To identify mechanisms that can be modulated to initiate cell cycle re-entry and hair cell regeneration, we studied the effect of activating the sonic hedgehog (Shh) pathway. We show that Shh signaling in postnatal rat cochleae damaged by neomycin leads to renewed proliferation of supporting cells and hair cells. Further, proliferating supporting cells are likely to transdifferentiate into hair cells. Shh treatment leads to inhibition of retinoblastoma protein (pRb) by increasing phosphorylated pRb and reducing retinoblastoma gene transcription. This results in upregulation of cyclins B1, D2, and D3, and CDK1. These results suggest that Shh signaling induces cell cycle re-entry in cochlear sensory epithelium and the production of new hair cells, in part by attenuating pRb function. This study provides an additional route to modulate pRb function with important implications in mammalian hair cell regeneration.

  4. Sonic Hedgehog Initiates Cochlear Hair Cell Regeneration through Downregulation of Retinoblastoma Protein

    Science.gov (United States)

    Lu, Na; Chen, Yan; Wang, Zhengmin; Chen, Guoling; Lin, Qin; Chen, Zheng-Yi; Li, Huawei

    2013-01-01

    Cell cycle re-entry by cochlear supporting cells and/or hair cells is considered one of the best approaches for restoring hearing loss as a result of hair cell damage. To identify mechanisms that can be modulated to initiate cell cycle re-entry and hair cell regeneration, we studied the effect of activating the sonic hedgehog (Shh) pathway. We show that Shh signaling in postnatal rat cochleae damaged by neomycin leads to renewed proliferation of supporting cells and hair cells. Further, proliferating supporting cells are likely to transdifferentiate into hair cells. Shh treatment leads to inhibition of retinoblastoma protein (pRb) by increasing phosphorylated pRb and reducing retinoblastoma gene transcription. This results in upregulation of cyclins B1, D2, and D3, and CDK1. These results suggest that Shh signaling induces cell cycle re-entry in cochlear sensory epithelium and the production of new hair cells, in part by attenuating pRb function. This study provides an additional route to modulate pRb function with important implications in mammalian hair cell regeneration. PMID:23211596

  5. Scaffold protein JLP mediates TCR-initiated CD4+T cell activation and CD154 expression.

    Science.gov (United States)

    Yan, Qi; Yang, Cheng; Fu, Qiang; Chen, Zhaowei; Liu, Shan; Fu, Dou; Rahman, Rahmat N; Nakazato, Ryota; Yoshioka, Katsuji; Kung, Sam K P; Ding, Guohua; Wang, Huiming

    2017-07-01

    CD4 + T-cell activation and its subsequent induction of CD154 (CD40 ligand, CD40L) expression are pivotal in shaping both the humoral and cellular immune responses. Scaffold protein JLP regulates signal transduction pathways and molecular trafficking inside cells, thus represents a critical component in maintaining cellular functions. Its role in regulating CD4 + T-cell activation and CD154 expression, however, is unclear. Here, we demonstrated expression of JLP in mouse tissues of lymph nodes, thymus, spleen, and also CD4 + T cells. Using CD4+ T cells from jlp-deficient and jlp-wild-type mice, we demonstrated that JLP-deficiency impaired T-cell proliferation, IL-2 production, and CD154 induction upon TCR stimulations, but had no impacts on the expression of other surface molecules such as CD25, CD69, and TCR. These observed impaired T-cell functions in the jlp-/- CD4 + T cells were associated with defective NF-AT activation and Ca 2 + influx, but not the MAPK, NF-κB, as well as AP-1 signaling pathways. Our findings indicated that, for the first time, JLP plays a critical role in regulating CD4 + T cells response to TCR stimulation partly by mediating the activation of TCR-initiated Ca 2+ /NF-AT. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. Sonic hedgehog initiates cochlear hair cell regeneration through downregulation of retinoblastoma protein

    International Nuclear Information System (INIS)

    Lu, Na; Chen, Yan; Wang, Zhengmin; Chen, Guoling; Lin, Qin; Chen, Zheng-Yi; Li, Huawei

    2013-01-01

    Highlights: ► Shh activation in neonatal cochleae enhances sensory cell proliferation. ► Proliferating supporting cells can transdifferentiate into hair cells. ► Shh promotes proliferation by transiently modulating pRb activity. ► Shh inhibits pRb by inhibiting transcription and increasing phosphorylation of pRb. -- Abstract: Cell cycle re-entry by cochlear supporting cells and/or hair cells is considered one of the best approaches for restoring hearing loss as a result of hair cell damage. To identify mechanisms that can be modulated to initiate cell cycle re-entry and hair cell regeneration, we studied the effect of activating the sonic hedgehog (Shh) pathway. We show that Shh signaling in postnatal rat cochleae damaged by neomycin leads to renewed proliferation of supporting cells and hair cells. Further, proliferating supporting cells are likely to transdifferentiate into hair cells. Shh treatment leads to inhibition of retinoblastoma protein (pRb) by increasing phosphorylated pRb and reducing retinoblastoma gene transcription. This results in upregulation of cyclins B1, D2, and D3, and CDK1. These results suggest that Shh signaling induces cell cycle re-entry in cochlear sensory epithelium and the production of new hair cells, in part by attenuating pRb function. This study provides an additional route to modulate pRb function with important implications in mammalian hair cell regeneration.

  7. Role of the Escherichia coli grpE heat shock protein in the initiation of bacteriophage lambda DNA replication.

    Science.gov (United States)

    Osipiuk, J; Zylicz, M

    1991-01-01

    Initiation of replication of lambda DNA requires assembly of the proper nucleoprotein complex consisting of the lambda origin of replication-lambda O-lambda P-dnaB proteins. The dnaJ, dnaK and grpE heat shock proteins destabilize the lambda P-dnaB interaction in this complex permitting dnaB helicase to unwind lambda DNA near ori lambda sequence. First step of this disassembling reaction is the binding of dnaK protein to lambda P protein. In this report we examined the influence of dnaJ and grpE proteins on stability of the lambda P-dnaK complex. Our results show that grpE alone dissociates this complex, but both grpE and dnaJ together do not. These results suggest that, in the presence of grpE protein, dnaK protein has a higher affinity for lambda P protein complexed with dnaJ protein than in the situation where grpE protein is not used.

  8. Protein C activation during the initial phase of experimental acute pancreatitis in the rabbit

    DEFF Research Database (Denmark)

    Ottesen, L H; Bladbjerg, E-M; Osman, M

    2000-01-01

    activity), anticoagulant proteins (protein C, antithrombin) and fibrinolytic factors (tissue plasminogen activator, plasminogen activator inhibitor-1) were performed for 5 h. RESULTS: ANP was confirmed by elevated serum amylase, development of ascites, and histological changes of the pancreas. A moderate...

  9. Immunodominant fragments of myelin basic protein initiate T cell-dependent pain

    Directory of Open Access Journals (Sweden)

    Liu Huaqing

    2012-06-01

    Full Text Available Abstract Background The myelin sheath provides electrical insulation of mechanosensory Aβ-afferent fibers. Myelin-degrading matrix metalloproteinases (MMPs damage the myelin sheath. The resulting electrical instability of Aβ-fibers is believed to activate the nociceptive circuitry in Aβ-fibers and initiate pain from innocuous tactile stimulation (mechanical allodynia. The precise molecular mechanisms, responsible for the development of this neuropathic pain state after nerve injury (for example, chronic constriction injury, CCI, are not well understood. Methods and results Using mass spectrometry of the whole sciatic nerve proteome followed by bioinformatics analyses, we determined that the pathways, which are classified as the Infectious Disease and T-helper cell signaling, are readily activated in the nerves post-CCI. Inhibition of MMP-9/MMP-2 suppressed CCI-induced mechanical allodynia and concomitant TNF-α and IL-17A expression in nerves. MMP-9 proteolysis of myelin basic protein (MBP generated the MBP84-104 and MBP68-86 digest peptides, which are prominent immunogenic epitopes. In agreement, the endogenous MBP69-86 epitope co-localized with MHCII and MMP-9 in Schwann cells and along the nodes of Ranvier. Administration of either the MBP84-104 or MBP68-86 peptides into the naïve nerve rapidly produced robust mechanical allodynia with a concomitant increase in T cells and MHCII-reactive cell populations at the injection site. As shown by the genome-wide expression profiling, a single intraneural MBP84-104 injection stimulated the inflammatory, immune cell trafficking, and antigen presentation pathways in the injected naïve nerves and the associated spinal cords. Both MBP84-104-induced mechanical allodynia and characteristic pathway activation were remarkably less prominent in the T cell-deficient athymic nude rats. Conclusions These data implicate MBP as a novel mediator of pain. Furthermore, the action of MMPs expressed within 1

  10. Immunodominant fragments of myelin basic protein initiate T cell-dependent pain.

    Science.gov (United States)

    Liu, Huaqing; Shiryaev, Sergey A; Chernov, Andrei V; Kim, Youngsoon; Shubayev, Igor; Remacle, Albert G; Baranovskaya, Svetlana; Golubkov, Vladislav S; Strongin, Alex Y; Shubayev, Veronica I

    2012-06-07

    The myelin sheath provides electrical insulation of mechanosensory Aβ-afferent fibers. Myelin-degrading matrix metalloproteinases (MMPs) damage the myelin sheath. The resulting electrical instability of Aβ-fibers is believed to activate the nociceptive circuitry in Aβ-fibers and initiate pain from innocuous tactile stimulation (mechanical allodynia). The precise molecular mechanisms, responsible for the development of this neuropathic pain state after nerve injury (for example, chronic constriction injury, CCI), are not well understood. Using mass spectrometry of the whole sciatic nerve proteome followed by bioinformatics analyses, we determined that the pathways, which are classified as the Infectious Disease and T-helper cell signaling, are readily activated in the nerves post-CCI. Inhibition of MMP-9/MMP-2 suppressed CCI-induced mechanical allodynia and concomitant TNF-α and IL-17A expression in nerves. MMP-9 proteolysis of myelin basic protein (MBP) generated the MBP84-104 and MBP68-86 digest peptides, which are prominent immunogenic epitopes. In agreement, the endogenous MBP69-86 epitope co-localized with MHCII and MMP-9 in Schwann cells and along the nodes of Ranvier. Administration of either the MBP84-104 or MBP68-86 peptides into the naïve nerve rapidly produced robust mechanical allodynia with a concomitant increase in T cells and MHCII-reactive cell populations at the injection site. As shown by the genome-wide expression profiling, a single intraneural MBP84-104 injection stimulated the inflammatory, immune cell trafficking, and antigen presentation pathways in the injected naïve nerves and the associated spinal cords. Both MBP84-104-induced mechanical allodynia and characteristic pathway activation were remarkably less prominent in the T cell-deficient athymic nude rats. These data implicate MBP as a novel mediator of pain. Furthermore, the action of MMPs expressed within 1 day post-injury is critical to the generation of tactile allodynia

  11. Targeting tumor-initiating cells: Eliminating anabolic cancer stem cells with inhibitors of protein synthesis or by mimicking caloric restriction

    Science.gov (United States)

    Lamb, Rebecca; Harrison, Hannah; Smith, Duncan L.; Townsend, Paul A.; Jackson, Thomas; Ozsvari, Bela; Martinez-Outschoorn, Ubaldo E.; Pestell, Richard G.; Howell, Anthony; Lisanti, Michael P.; Sotgia, Federica

    2015-01-01

    We have used an unbiased proteomic profiling strategy to identify new potential therapeutic targets in tumor-initiating cells (TICs), a.k.a., cancer stem cells (CSCs). Towards this end, the proteomes of mammospheres from two breast cancer cell lines were directly compared to attached monolayer cells. This allowed us to identify proteins that were highly over-expressed in CSCs and/or progenitor cells. We focused on ribosomal proteins and protein folding chaperones, since they were markedly over-expressed in mammospheres. Overall, we identified >80 molecules specifically associated with protein synthesis that were commonly upregulated in mammospheres. Most of these proteins were also transcriptionally upregulated in human breast cancer cells in vivo, providing evidence for their potential clinical relevance. As such, increased mRNA translation could provide a novel mechanism for enhancing the proliferative clonal expansion of TICs. The proteomic findings were functionally validated using known inhibitors of protein synthesis, via three independent approaches. For example, puromycin (which mimics the structure of tRNAs and competitively inhibits protein synthesis) preferentially targeted CSCs in both mammospheres and monolayer cultures, and was ~10-fold more potent for eradicating TICs, than “bulk” cancer cells. In addition, rapamycin, which inhibits mTOR and hence protein synthesis, was very effective at reducing mammosphere formation, at nanomolar concentrations. Finally, mammosphere formation was also markedly inhibited by methionine restriction, which mimics the positive effects of caloric restriction in cultured cells. Remarkably, mammosphere formation was >18-fold more sensitive to methionine restriction and replacement, as directly compared to monolayer cell proliferation. Methionine is absolutely required for protein synthesis, since every protein sequence starts with a methionine residue. Thus, the proliferation and survival of CSCs is very sensitive to

  12. Amino acids augment muscle protein synthesis in neonatal pigs during acute endotoxemia by stimulating mTOR-dependent translation initiation.

    Science.gov (United States)

    Orellana, Renán A; Jeyapalan, Asumthia; Escobar, Jeffery; Frank, Jason W; Nguyen, Hanh V; Suryawan, Agus; Davis, Teresa A

    2007-11-01

    In skeletal muscle of adults, sepsis reduces protein synthesis by depressing translation initiation and induces resistance to branched-chain amino acid stimulation. Normal neonates maintain a high basal muscle protein synthesis rate that is sensitive to amino acid stimulation. In the present study, we determined the effect of amino acids on protein synthesis in skeletal muscle and other tissues in septic neonates. Overnight-fasted neonatal pigs were infused with endotoxin (LPS, 0 and 10 microg.kg(-1).h(-1)), whereas glucose and insulin were maintained at fasting levels; amino acids were clamped at fasting or fed levels. In the presence of fasting insulin and amino acids, LPS reduced protein synthesis in longissimus dorsi (LD) and gastrocnemius muscles and increased protein synthesis in the diaphragm, but had no effect in masseter and heart muscles. Increasing amino acids to fed levels accelerated muscle protein synthesis in LD, gastrocnemius, masseter, and diaphragm. LPS stimulated protein synthesis in liver, lung, spleen, pancreas, and kidney in fasted animals. Raising amino acids to fed levels increased protein synthesis in liver of controls, but not LPS-treated animals. The increase in muscle protein synthesis in response to amino acids was associated with increased mTOR, 4E-BP1, and S6K1 phosphorylation and eIF4G-eIF4E association in control and LPS-infused animals. These findings suggest that amino acids stimulate skeletal muscle protein synthesis during acute endotoxemia via mTOR-dependent ribosomal assembly despite reduced basal protein synthesis rates in neonatal pigs. However, provision of amino acids does not further enhance the LPS-induced increase in liver protein synthesis.

  13. The potential role of ribosomal protein S5 on cell cycle arrest and initiation of murine erythroleukemia cell differentiation.

    Science.gov (United States)

    Matragkou, Christina N; Papachristou, Eleni T; Tezias, Sotirios S; Tsiftsoglou, Asterios S; Choli-Papadopoulou, Theodora; Vizirianakis, Ioannis S

    2008-07-01

    Evidence now exists to indicate that some ribosomal proteins besides being structural components of the ribosomal subunits are involved in the regulation of cell differentiation and apoptosis. As we have shown earlier, initiation of erythroid differentiation of murine erythroleukemia (MEL) cells is associated with transcriptional inactivation of genes encoding ribosomal RNAs and ribosomal proteins S5 (RPS5) and L35a. In this study, we extended these observations and investigated whether transfection of MEL cells with RPS5 cDNA affects the onset of initiation of erythroid maturation and their entrance in cell cycle arrest. Stably transfected MEL cloned cells (MEL-C14 and MEL-C56) were established and assessed for their capacity to produce RPS5 RNA transcript and its translated product. The impact of RPS5 cDNA transfection on the RPS5 gene expression patterns and the accumulation of RPS5 protein in inducible transfected MEL cells were correlated with their ability to: (a) initiate differentiation, (b) enter cell cycle arrest at G(1)/G(0) phase, and (c) modulate the level of cyclin-dependent kinases CDK2, CDK4, and CDK6. The data presented indicate that deregulation of RPS5 gene expression (constitutive expression) affects RPS5 protein level and delays both the onset of initiation of erythroid maturation and entrance in cell cycle arrest in inducer-treated MEL cells. 2008 Wiley-Liss, Inc.

  14. Initial investigation of dietitian perception of plant-based protein quality

    OpenAIRE

    Hughes, Glenna J; Kress, Kathleen S; Armbrecht, Eric S; Mukherjea, Ratna; Mattfeldt-Beman, Mildred

    2014-01-01

    Interest in plant-based diets is increasing, evidenced by scientific and regulatory recommendations, including Dietary Guidelines for Americans. Dietitians provide guidance in dietary protein selection but little is known about how familiar dietitians are with the quality of plant versus animal proteins or methods for measuring protein quality. Likewise, there is a need to explore their beliefs related to dietary recommendations. The aim of this study was to assess dietitians' perceptions of ...

  15. Teaching Protein Purification and Characterization Techniques: A Student-Initiated, Project-Oriented Biochemistry Laboratory Course

    Science.gov (United States)

    MacDonald, Gina

    2008-01-01

    This report describes a biochemistry laboratory that is completely project-oriented. Upper-level biology and chemistry majors work in teams to purify a protein of their choice. After the student groups have completed literature searches, ordered reagents, and made buffers they continue to learn basic protein purification and biochemical techniques…

  16. Purification and Initial Functions of Sex-Specific Storage Protein 2 in Bombyx mori.

    Science.gov (United States)

    Chen, Jianqing; Shu, Tejun; Chen, Jian; Ye, Man; Lv, Zhengbing; Nie, Zuoming; Gai, Qijing; Yu, Wei; Zhang, Yaozhou

    2015-08-01

    In this study, we identified a heat-resistant protein from the chrysalis stage of the silkworm which we named sex-specific storage protein 2 (SSP2). This protein was stable even at 80 °C, and has an amino acid sequence that is 90.65 % homologous to SP2. We utilized the heat-resistant characteristics of SSP2 to purify the protein and maintain its biological activity. In addition, using flow cytometry and the MTT assay, we found that SSP2 had anti-apoptotic effects on BmN cells, and that SSP2 could also inhibit cell apoptosis induced by chemical factors. These results suggest that SSP2 has a cell-protective function, and provides a basis for future work on the function of storage proteins in silkworm.

  17. Structure of the protein core of translation initiation factor 2 in apo, GTP-bound and GDP-bound forms

    International Nuclear Information System (INIS)

    Simonetti, Angelita; Marzi, Stefano; Fabbretti, Attilio; Hazemann, Isabelle; Jenner, Lasse; Urzhumtsev, Alexandre; Gualerzi, Claudio O.; Klaholz, Bruno P.

    2013-01-01

    The crystal structures of the eubacterial translation initiation factor 2 in apo form and with bound GDP and GTP reveal conformational changes upon nucleotide binding and hydrolysis, notably of the catalytically important histidine in the switch II region. Translation initiation factor 2 (IF2) is involved in the early steps of bacterial protein synthesis. It promotes the stabilization of the initiator tRNA on the 30S initiation complex (IC) and triggers GTP hydrolysis upon ribosomal subunit joining. While the structure of an archaeal homologue (a/eIF5B) is known, there are significant sequence and functional differences in eubacterial IF2, while the trimeric eukaryotic IF2 is completely unrelated. Here, the crystal structure of the apo IF2 protein core from Thermus thermophilus has been determined by MAD phasing and the structures of GTP and GDP complexes were also obtained. The IF2–GTP complex was trapped by soaking with GTP in the cryoprotectant. The structures revealed conformational changes of the protein upon nucleotide binding, in particular in the P-loop region, which extend to the functionally relevant switch II region. The latter carries a catalytically important and conserved histidine residue which is observed in different conformations in the GTP and GDP complexes. Overall, this work provides the first crystal structure of a eubacterial IF2 and suggests that activation of GTP hydrolysis may occur by a conformational repositioning of the histidine residue

  18. Structure of the protein core of translation initiation factor 2 in apo, GTP-bound and GDP-bound forms

    Energy Technology Data Exchange (ETDEWEB)

    Simonetti, Angelita [IGBMC (Institute of Genetics and of Molecular and Cellular Biology), Centre National de la Recherche Scientifique (CNRS) UMR 7104/Institut National de la Santé de la Recherche Médicale - INSERM U964/Université de Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch (France); Marzi, Stefano [Architecture et Réactivité de l’ARN, UPR 9002 CNRS, IBMC (Institute of Molecular and Cellular Biology), 15 Rue R. Descartes, 67084 Strasbourg, France, Université de Strasbourg, 67000 Strasbourg (France); Fabbretti, Attilio [University of Camerino, 62032 Camerino (Monaco) (Italy); Hazemann, Isabelle; Jenner, Lasse [IGBMC (Institute of Genetics and of Molecular and Cellular Biology), Centre National de la Recherche Scientifique (CNRS) UMR 7104/Institut National de la Santé de la Recherche Médicale -INSERM U964/Université de Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch (France); Urzhumtsev, Alexandre [IGBMC (Institute of Genetics and of Molecular and Cellular Biology), Centre National de la Recherche Scientifique (CNRS) UMR 7104/Institut National de la Santé de la Recherche Médicale - INSERM U964/Université de Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch (France); Université de Lorraine, 54506 Vandoeuvre-lès-Nancy (France); Gualerzi, Claudio O. [University of Camerino, 62032 Camerino (Monaco) (Italy); Klaholz, Bruno P., E-mail: klaholz@igbmc.fr [IGBMC (Institute of Genetics and of Molecular and Cellular Biology), Centre National de la Recherche Scientifique (CNRS) UMR 7104/Institut National de la Santé de la Recherche Médicale - INSERM U964/Université de Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch (France)

    2013-06-01

    The crystal structures of the eubacterial translation initiation factor 2 in apo form and with bound GDP and GTP reveal conformational changes upon nucleotide binding and hydrolysis, notably of the catalytically important histidine in the switch II region. Translation initiation factor 2 (IF2) is involved in the early steps of bacterial protein synthesis. It promotes the stabilization of the initiator tRNA on the 30S initiation complex (IC) and triggers GTP hydrolysis upon ribosomal subunit joining. While the structure of an archaeal homologue (a/eIF5B) is known, there are significant sequence and functional differences in eubacterial IF2, while the trimeric eukaryotic IF2 is completely unrelated. Here, the crystal structure of the apo IF2 protein core from Thermus thermophilus has been determined by MAD phasing and the structures of GTP and GDP complexes were also obtained. The IF2–GTP complex was trapped by soaking with GTP in the cryoprotectant. The structures revealed conformational changes of the protein upon nucleotide binding, in particular in the P-loop region, which extend to the functionally relevant switch II region. The latter carries a catalytically important and conserved histidine residue which is observed in different conformations in the GTP and GDP complexes. Overall, this work provides the first crystal structure of a eubacterial IF2 and suggests that activation of GTP hydrolysis may occur by a conformational repositioning of the histidine residue.

  19. Mammalian poly(A)-binding protein is a eukaryotic translation initiation factor, which acts via multiple mechanisms

    OpenAIRE

    Kahvejian, Avak; Svitkin, Yuri V.; Sukarieh, Rami; M'Boutchou, Marie-Noël; Sonenberg, Nahum

    2005-01-01

    Translation initiation is a multistep process involving several canonical translation factors, which assemble at the 5′-end of the mRNA to promote the recruitment of the ribosome. Although the 3′ poly(A) tail of eukaryotic mRNAs and its major bound protein, the poly(A)-binding protein (PABP), have been studied extensively, their mechanism of action in translation is not well understood and is confounded by differences between in vivo and in vitro systems. Here, we provide direct evidence for ...

  20. Initial investigation of dietitian perception of plant-based protein quality.

    Science.gov (United States)

    Hughes, Glenna J; Kress, Kathleen S; Armbrecht, Eric S; Mukherjea, Ratna; Mattfeldt-Beman, Mildred

    2014-07-01

    Interest in plant-based diets is increasing, evidenced by scientific and regulatory recommendations, including Dietary Guidelines for Americans. Dietitians provide guidance in dietary protein selection but little is known about how familiar dietitians are with the quality of plant versus animal proteins or methods for measuring protein quality. Likewise, there is a need to explore their beliefs related to dietary recommendations. The aim of this study was to assess dietitians' perceptions of plant-based protein quality and to determine if these are affected by demographic factors such as age and dietary practice group (DPG) membership. This was a cross-sectional design using an online survey. The survey was sent to all members of the Missouri Dietetic Association. All completed surveys (136) were analyzed. The main outcome measures were responses to belief and knowledge questions about the protein quality of plant-based diets, along with demographic information including age and DPG membership. Descriptive statistics and frequencies were determined, and chi-square analysis was used to determine the associations between belief and knowledge responses and demographic characteristics. Responses to belief statements suggested a high level of support for plant-based diets. No associations were found between any of the belief questions and demographic factors. A majority of respondents were not familiar with protein quality determination methods that are currently recognized by global regulatory and advisory agencies. Potential barriers identified in shifting to a more plant-based diet were lack of interest and perceived difficulty. Knowledge among dietitians of plant-based protein quality in general, and methods of protein quality measurement more specifically, needs to be addressed to enhance their knowledge base for making dietary protein recommendations. Two potential avenues for training are university curricula and continuing education opportunities provided to

  1. Mammalian poly(A)-binding protein is a eukaryotic translation initiation factor, which acts via multiple mechanisms.

    Science.gov (United States)

    Kahvejian, Avak; Svitkin, Yuri V; Sukarieh, Rami; M'Boutchou, Marie-Noël; Sonenberg, Nahum

    2005-01-01

    Translation initiation is a multistep process involving several canonical translation factors, which assemble at the 5'-end of the mRNA to promote the recruitment of the ribosome. Although the 3' poly(A) tail of eukaryotic mRNAs and its major bound protein, the poly(A)-binding protein (PABP), have been studied extensively, their mechanism of action in translation is not well understood and is confounded by differences between in vivo and in vitro systems. Here, we provide direct evidence for the involvement of PABP in key steps of the translation initiation pathway. Using a new technique to deplete PABP from mammalian cell extracts, we show that extracts lacking PABP exhibit dramatically reduced rates of translation, reduced efficiency of 48S and 80S ribosome initiation complex formation, and impaired interaction of eIF4E with the mRNA cap structure. Supplementing PABP-depleted extracts with wild-type PABP completely rectified these deficiencies, whereas a mutant of PABP, M161A, which is incapable of interacting with eIF4G, failed to restore translation. In addition, a stronger inhibition (approximately twofold) of 80S as compared to 48S ribosome complex formation (approximately 65% vs. approximately 35%, respectively) by PABP depletion suggests that PABP plays a direct role in 60S subunit joining. PABP can thus be considered a canonical translation initiation factor, integral to initiation complex formation at the 5'-end of mRNA.

  2. Molecular dynamics analysis of conformational change of paramyxovirus F protein during the initial steps of membrane fusion

    International Nuclear Information System (INIS)

    Martín-García, Fernando; Mendieta-Moreno, Jesús Ignacio; Mendieta, Jesús; Gómez-Puertas, Paulino

    2012-01-01

    Highlights: ► Initial conformational change of paramyxovirus F protein is caused only by mechanical forces. ► HRA region undergoes a structural change from a beta + alpha conformation to an extended coil and then to an all-alpha conformation. ► HRS domains of F protein form three single α-helices prior to generation of the coiled coil. -- Abstract: The fusion of paramyxovirus to the cell membrane is mediated by fusion protein (F protein) present in the virus envelope, which undergoes a dramatic conformational change during the process. Unlike hemagglutinin in orthomyxovirus, this change is not mediated by an alteration of environmental pH, and its cause remains unknown. Steered molecular dynamics analysis leads us to suggest that the conformational modification is mediated only by stretching mechanical forces once the transmembrane fusion peptide of the protein is anchored to the cell membrane. Such elongating forces will generate major secondary structure rearrangement in the heptad repeat A region of the F protein; from β-sheet conformation to an elongated coil and then spontaneously to an α-helix. In addition, it is proposed that the heptad repeat A region adopts a final three-helix coiled coil and that this structure appears after the formation of individual helices in each monomer.

  3. Effects of high-protein vs. high- fat snacks on appetite control, satiety, and eating initiation in healthy women.

    Science.gov (United States)

    Ortinau, Laura C; Hoertel, Heather A; Douglas, Steve M; Leidy, Heather J

    2014-09-29

    The purpose of this study was to determine whether a high-protein afternoon yogurt snack improves appetite control, satiety, and reduces subsequent food intake compared to other commonly-consumed, energy dense, high-fat snacks. Twenty, healthy women (age: 27 ± 2 y; BMI: 23.4 ± 0.7 kg/m2) completed the randomized crossover design study which included 3, 8-h testing days comparing the following 160 kcal afternoon snacks: high-protein yogurt (14 g protein/25 g CHO/0 g fat); high-fat crackers (0 g protein/19 g CHO/9 g fat); and high-fat chocolate (2 g protein/19 g CHO/9 g fat). Participants were acclimated to each snack for 3 consecutive days. On day 4, the participants consumed a standardized breakfast and lunch; the respective snack was consumed 3-h post-lunch. Perceived hunger and fullness were assessed throughout the afternoon until dinner was voluntarily requested. An ad libitum dinner was then provided. The consumption of the yogurt snack led to greater reductions in afternoon hunger vs. chocolate (p snack also delayed eating initiation by approximately 30 min compared to the chocolate snack (p snack led to approximately 100 fewer kcals consumed at dinner vs. the crackers (p = 0.08) and chocolate (p snacks, eating less energy dense, high-protein snacks like yogurt improves appetite control, satiety, and reduces subsequent food intake in healthy women.

  4. 2D proteome analysis initiates new Insights on the Salmonella Typhimurium LuxS protein

    Directory of Open Access Journals (Sweden)

    Vanderleyden Jos

    2009-09-01

    Full Text Available Abstract Background Quorum sensing is a term describing a bacterial communication system mediated by the production and recognition of small signaling molecules. The LuxS enzyme, catalyzing the synthesis of AI-2, is conserved in a wide diversity of bacteria. AI-2 has therefore been suggested as an interspecies quorum sensing signal. To investigate the role of endogenous AI-2 in protein expression of the Gram-negative pathogen Salmonella enterica serovar Typhimurium (S. Typhimurium, we performed a 2D-DIGE proteomics experiment comparing total protein extract of wildtype S. Typhimurium with that of a luxS mutant, unable to produce AI-2. Results Differential proteome analysis of wildtype S. Typhimurium versus a luxS mutant revealed relatively few changes beyond the known effect on phase 2 flagellin. However, two highly differentially expressed protein spots with similar molecular weight but differing isoelectric point, were identified as LuxS whereas the S. Typhimurium genome contains only one luxS gene. This observation was further explored and we show that the S. Typhimurium LuxS protein can undergo posttranslational modification at a catalytic cysteine residue. Additionally, by constructing LuxS-βla and LuxS-PhoA fusion proteins, we demonstrate that S. Typhimurium LuxS can substitute the cognate signal peptide sequences of β-lactamase and alkaline phosphatase for translocation across the cytoplasmic membrane in S. Typhimurium. This was further confirmed by fractionation of S. Typhimurium protein extracts, followed by Western blot analysis. Conclusion 2D-DIGE analysis of a luxS mutant vs. wildtype Salmonella Typhimurium did not reveal new insights into the role of AI-2/LuxS in Salmonella as only a small amount of proteins were differentially expressed. However, subsequent in depth analysis of the LuxS protein itself revealed two interesting features: posttranslational modification and potential translocation across the cytoplasmic membrane. As

  5. Rotavirus NSP2 interferes with the core lattice protein VP2 in initiation of minus-strand synthesis

    International Nuclear Information System (INIS)

    Vende, Patrice; Tortorici, M. Alejandra; Taraporewala, Zenobia F.; Patton, John T.

    2003-01-01

    The rotavirus nonstructural protein NSP2 self-assembles into stable octameric structures that possess nonspecific affinity for single-stranded (ss)RNA and RNA-RNA helix-destabilizing and NTPase activities. Furthermore, NSP2 is a component of replication intermediates with replicase activity and plays a critical role in the packaging and replication of the segmented dsRNA genome of rotavirus. To better understand the function of the protein in genome replication, we examined the effect that purified recombinant NSP2 had on the synthesis of dsRNA by the open core replication system. The results showed that NSP2 inhibited the synthesis of dsRNA from viral mRNA in vitro, in a concentration-dependent manner. The inhibition was overcome by adding increasing amounts of viral mRNA or nonviral ssRNA to the system, indicating that the inhibition was mediated by the nonspecific RNA-binding activity of NSP2. Further analysis revealed that NSP2 interfered with the ability of the open core proteins, GTP, and viral mRNA to form the initiation complex for (-) strand synthesis. Additional experiments indicated that NSP2 did not perturb recognition of viral mRNA by the viral RNA polymerase VP1, but rather interfered with the function of VP2, a protein that is essential for (-) strand initiation and dsRNA synthesis and that forms the T = 1 lattice of the virion core. In contrast to initiation, NSP2 did not inhibit (-) strand elongation. Collectively, the findings provide evidence that the temporal order of interaction of RNA-binding proteins with viral mRNA is a crucial factor impacting the formation of replication intermediates

  6. Effect of a high-protein diet on maintenance of blood pressure levels achieved after initial weight loss

    DEFF Research Database (Denmark)

    Engberink, M F; Geleijnse, J M; Bakker, S J L

    2015-01-01

    Randomized trials have shown significant blood pressure (BP) reductions after increased protein compared with carbohydrate intake, but the effect on BP maintenance after initial weight loss is unclear. We examined the effect of a high-protein diet on the maintenance of reduced BP after weight loss......(-2), BP was 118/73 mm Hg and 28 subjects (6.5%) used antihypertensive agents. Systolic BP during 26 weeks of weight maintenance dietary intervention increased in both treatment groups, but it was 2.2 mm Hg less (95% CI: -4.6 to 0.2 mm Hg, P=0.08) in the high-protein group than in the lower...

  7. Isolation and initial structural characterization of a 27 kDa protein from Zingiber officinale

    Science.gov (United States)

    Rasheed, Saima; Malik, Shoaib Ahmad; Falke, Sven; Arslan, Ali; Fazel, Ramin; Schlüter, Hartmut; Betzel, Christian; Choudhary, M. Iqbal

    2018-03-01

    Zingiber officinale Roscoe (Ginger) is a widely used traditional medicinal plant (for different ailments such as arthritis, constipation, and hypertension). This article describes the isolation and characterization of a so far unknown protein from ginger rhizomes applying ion exchange, affinity, size-exclusion chromatography, small angle X-ray scattering (SAXS), and mass spectrometry techniques. One-dimensional Coomassie-stained SDS-PAGE was performed under non-reducing conditions, showing one band corresponding to approx. 27 kDa. Dynamic light scattering (DLS) analysis of the protein solution revealed monodispersity and a monomeric state of the purified protein. Circular dichroism (CD) spectroscopy strongly indicated a β-sheet-rich protein, and disordered regions. MALDI-TOF-MS, and LC-MS/MS analysis resulted in the identification of 27.29 kDa protein, having 32.13% and 25.34% sequence coverage with Zingipain-1 and 2, respectively. The monomeric state and molecular weight were verified by small angle X-ray scattering (SAXS) studies. An elongated ab-initio model was calculated based on the scattering intensity distribution.

  8. Protein adsorption resistance of PVP-modified polyurethane film prepared by surface-initiated atom transfer radical polymerization

    Energy Technology Data Exchange (ETDEWEB)

    Yuan, Huihui; Qian, Bin; Zhang, Wei [Shanghai Key Laboratory of Functional Materials Chemistry and Research Center of Analysis and Test, East China University of Science and Technology, Shanghai 200237 (China); Lan, Minbo, E-mail: minbolan@ecust.edu.cn [Shanghai Key Laboratory of Functional Materials Chemistry and Research Center of Analysis and Test, East China University of Science and Technology, Shanghai 200237 (China); State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237 (China)

    2016-02-15

    Highlights: • Antifouling PVP brushes were successfully grafted on PU films by SI-ATRP. • The effect of polymerization time on surface property and topography was studied. • Hydrophilicity and protein fouling resistance of PVP–PU films were greatly promoted. • Competitive adsorption of three proteins on PVP–PU films was evaluated. - Abstract: An anti-fouling surface of polyurethane (PU) film grafted with Poly(N-vinylpyrrolidone) (PVP) was prepared through surface-initiated atom transfer radical polymerization (SI-ATRP). And the polymerization time was investigated to obtain PU films with PVP brushes of different lengths. The surface properties and protein adsorption of modified PU films were evaluated. The results showed that the hydrophilicity of PU–PVP films were improved with the increase of polymerization time, which was not positive correlation with the surface roughness due to the brush structure. Additionally, the protein resistance performance was promoted when prolonging the polymerization time. The best antifouling PU–PVP (6.0 h) film reduced the adsoption level of bovine serum albumin (BSA), lysozyme (LYS), and brovin serum fibrinogen (BFG) by 93.4%, 68.3%, 85.6%, respectively, compared to the unmodified PU film. The competitive adsorption of three proteins indicated that LYS preferentially adsorbed on the modified PU film, while BFG had the lowest adsorption selectivity. And the amount of BFG on PU–PVP (6.0 h) film reduced greatly to 0.08 μg/cm{sup 2}, which was almost one-tenth of its adsorption from the single-protein system. Presented results suggested that both hydrophilicity and surface roughness might be the important factors in all cases of protein adsorption, and the competitive or selective adsorption might be related to the size of the proteins, especially on the non-charged films.

  9. Protein adsorption resistance of PVP-modified polyurethane film prepared by surface-initiated atom transfer radical polymerization

    International Nuclear Information System (INIS)

    Yuan, Huihui; Qian, Bin; Zhang, Wei; Lan, Minbo

    2016-01-01

    Highlights: • Antifouling PVP brushes were successfully grafted on PU films by SI-ATRP. • The effect of polymerization time on surface property and topography was studied. • Hydrophilicity and protein fouling resistance of PVP–PU films were greatly promoted. • Competitive adsorption of three proteins on PVP–PU films was evaluated. - Abstract: An anti-fouling surface of polyurethane (PU) film grafted with Poly(N-vinylpyrrolidone) (PVP) was prepared through surface-initiated atom transfer radical polymerization (SI-ATRP). And the polymerization time was investigated to obtain PU films with PVP brushes of different lengths. The surface properties and protein adsorption of modified PU films were evaluated. The results showed that the hydrophilicity of PU–PVP films were improved with the increase of polymerization time, which was not positive correlation with the surface roughness due to the brush structure. Additionally, the protein resistance performance was promoted when prolonging the polymerization time. The best antifouling PU–PVP (6.0 h) film reduced the adsoption level of bovine serum albumin (BSA), lysozyme (LYS), and brovin serum fibrinogen (BFG) by 93.4%, 68.3%, 85.6%, respectively, compared to the unmodified PU film. The competitive adsorption of three proteins indicated that LYS preferentially adsorbed on the modified PU film, while BFG had the lowest adsorption selectivity. And the amount of BFG on PU–PVP (6.0 h) film reduced greatly to 0.08 μg/cm"2, which was almost one-tenth of its adsorption from the single-protein system. Presented results suggested that both hydrophilicity and surface roughness might be the important factors in all cases of protein adsorption, and the competitive or selective adsorption might be related to the size of the proteins, especially on the non-charged films.

  10. Functions of alternative Replication Protein A (aRPA) in initiation and elongation

    OpenAIRE

    Mason, Aaron C.; Roy, Rupa; Simmons, Daniel T.; Wold, Marc S.

    2010-01-01

    Replication protein A (RPA) is a single-stranded DNA-binding complex that is essential for DNA replication, repair and recombination in eukaryotic cells. In addition to this canonical complex, we have recently characterized an alternative Replication Protein A complex (aRPA) that is unique to primates. aRPA is composed of three subunits: RPA1 and RPA3, also present in canonical RPA, and a primate-specific subunit RPA4, homologous to canonical RPA2. aRPA has biochemical properties similar to t...

  11. Sputter deposited bioceramic coatings: surface characterisation and initial protein adsorption studies using surface-MALDI-MS

    DEFF Research Database (Denmark)

    Boyd, A. R.; Burke, G. A.; Duffy, H.

    2011-01-01

    Protein adsorption onto calcium phosphate (Ca–P) bioceramics utilised in hard tissue implant applications has been highlighted as one of the key events that influences the subsequent biological response, in vivo. This work reports on the use of surface-matrix assisted laser desorption ionisation...... to a combination of growth factors and lipoproteins present in serum. From the data obtained here it is evident that surface-MALDI-MS has significant utility as a tool for studying the dynamic nature of protein adsorption onto the surfaces of bioceramic coatings, which most likely plays a significant role...

  12. Protein repellent hydrophilic grafts prepared by surface-initiated atom transfer radical polymerization from polypropylene

    DEFF Research Database (Denmark)

    Fristrup, Charlotte Juel; Jankova Atanasova, Katja; Eskimergen, Rüya

    2012-01-01

    with Attenuated Total Reflectance (ATR) Fourier Transform Infrared (FTIR) spectroscopy and Water Contact Angle (WCA) measurements. Confocal fluorescence microscopy of modified and unmodified substrates immersed in labelled insulin aspart showed superior repulsion of this protein for the poly(PEGMA) grafts, due...

  13. Microgravity alters protein phosphorylation changes during initiation of sea urchin sperm motility

    Science.gov (United States)

    Tash, J. S.; Bracho, G. E.

    1999-01-01

    European Space Agency (ESA) studies demonstrated that bull sperm swim with higher velocity in microgravity (microG) than at 1 G. Coupling between protein phosphorylation and sperm motility during activation in microG and at 1 G was examined in the ESA Biorack on two space shuttle missions. Immotile sperm were activated to swim (86-90% motility) at launch +20 h by dilution into artificial seawater (ASW). Parallel ground controls were performed 2 h after the flight experiment. Activation after 0, 30, and 60 s was terminated with electrophoresis sample buffer and samples analyzed for phosphoamino acids by Western blotting. Phosphorylation of a 130-kDa phosphothreonine-containing protein (FP130) occurred three to four times faster in microG than at 1 G. A 32-kDa phosphoserine-containing protein was significantly stimulated at 30 s but returned to 1 G control levels at 60 s. The rate of FP130 phosphorylation in microG was attenuated by D2O, suggesting that changes in water properties participate in altering signal transduction. Changes in FP130 phosphorylation triggered by the egg peptide speract were delayed in microG. These results demonstrate that previously observed effects of microG on sperm motility are coupled to changes in phosphorylation of specific flagellar proteins and that early events of sperm activation and fertilization are altered in microG.

  14. The Ezrin Metastatic Phenotype Is Associated with the Initiation of Protein Translation

    Directory of Open Access Journals (Sweden)

    Joseph W. Briggs

    2012-04-01

    Full Text Available We previously associated the cytoskeleton linker protein, Ezrin, with the metastatic phenotype of pediatric sarcomas, including osteosarcoma and rhabdomyosarcoma. These studies have suggested that Ezrin contributes to the survival of cancer cells after their arrival at secondary metastatic locations. To better understand this role in metastasis, we undertook two noncandidate analyses of Ezrin function including a microarray subtraction of high-and low-Ezrin-expressing cells and a proteomic approach to identify proteins that bound the N-terminus of Ezrin in tumor lysates. Functional analyses of these data led to a novel and unifying hypothesis that Ezrin contributes to the efficiency of metastasis through regulation of protein translation. In support of this hypothesis, we found Ezrin to be part of the ribonucleoprotein complex to facilitate the expression of complex messenger RNA in cells and to bind with poly A binding protein 1 (PABP1; PABPC1. The relevance of these findings was supported by our identification of Ezrin and components of the translational machinery in pseudopodia of highly metastatic cells during the process of cell invasion. Finally, two small molecule inhibitors recently shown to inhibit the Ezrin metastatic phenotype disrupted the Ezrin/PABP1 association. Taken together, these results provide a novel mechanistic basis by which Ezrin may contribute to metastasis.

  15. The N-terminal region of eukaryotic translation initiation factor 5A signals to nuclear localization of the protein

    International Nuclear Information System (INIS)

    Parreiras-e-Silva, Lucas T.; Gomes, Marcelo D.; Oliveira, Eduardo B.; Costa-Neto, Claudio M.

    2007-01-01

    The eukaryotic translation initiation factor 5A (eIF5A) is a ubiquitous protein of eukaryotic and archaeal organisms which undergoes hypusination, a unique post-translational modification. We have generated a polyclonal antibody against murine eIF5A, which in immunocytochemical assays in B16-F10 cells revealed that the endogenous protein is preferentially localized to the nuclear region. We therefore analyzed possible structural features present in eIF5A proteins that could be responsible for that characteristic. Multiple sequence alignment analysis of eIF5A proteins from different eukaryotic and archaeal organisms showed that the former sequences have an extended N-terminal segment. We have then performed in silico prediction analyses and constructed different truncated forms of murine eIF5A to verify any possible role that the N-terminal extension might have in determining the subcellular localization of the eIF5A in eukaryotic organisms. Our results indicate that the N-terminal extension of the eukaryotic eIF5A contributes in signaling this protein to nuclear localization, despite of bearing no structural similarity with classical nuclear localization signals

  16. [The effects of TorR protein on initiation of DNA replication in Escherichia coli].

    Science.gov (United States)

    Yuan, Yao; Jiaxin, Qiao; Jing, Li; Hui, Li; Morigen, Morigen

    2015-03-01

    The two-component systems, which could sense and respond to environmental changes, widely exist in bacteria as a signal transduction pathway. The bacterial CckA/CtrA, ArcA/ArcB and PhoP/PhoQ two-component systems are associated with initiation of DNA replication and cell division, however, the effects of the TorS/TorR system on cell cycle and DNA replication remains unknown. The TorS/TorR system in Escherichia coli can sense changes in trimethylamine oxide (TMAO) concentration around the cells. However, it is unknown if it also affects initiation of DNA replication. We detected DNA replication patterns in ΔtorS and ΔtorR mutant strains by flow cytometry. We found that the average number of replication origins (oriCs) per cell and doubling time in ΔtorS mutants were the same while the average number of oriCs in ΔtorR mutants was increased compared with that in wild-type cells. These results indicated that absence of TorR led to an earlier initiation of DNA replication than that in wild-type cells. Strangely, neither overexpression of TorR nor co-expression of TorR and TorS could restore ΔtorR mutant phenotype to the wild type. However, overexpression of SufD in both wild type and ΔtorR mutants promoted initiation of DNA replication, while mutation of SufD delayed it in ΔtorR mutants. Thus, TorR may affect initiation of DNA replication indirectly through regulating gene expression of sufD.

  17. The influenza A virus matrix protein as a marker to monitor initial virus internalisation.

    Science.gov (United States)

    Eierhoff, Thorsten; Ludwig, Stephan; Ehrhardt, Christina

    2009-01-01

    The uptake of influenza A viruses (IAV) into cells represents an attractive antiviral drug target, e.g., by interfering with essential cellular or viral entry factors. So far, this process could only be studied by time-consuming microscopical methods. Thus, there is a lack of rapid and easy assay systems to monitor viral entry. Here, we describe a rapid procedure to analyse internalisation of IAV via Western blot detection of virion-associated matrix protein (M1), the most abundant protein within the viral particle. The assay is broadly applicable and detects different virus strains of various subtypes. As a proof of principle, treatment of cells with various known or presumed entry inhibitors resulted in reduced M1 levels. Removal of sialic acids, the receptors for IAV, led to a complete loss of the M1 signal, indicating that virus internalisation can be monitored already at the stage of attachment. Prevention of endosomal acidification resulted in a delayed degradation of M1 indicative of IAV particles trapped in endosomes. Thus, early detection of the virus-associated M1 protein is a rapid method to monitor different steps of influenza virus internalisation and has potential for application as a screening method for drugs that interfere with the uptake of IAV.

  18. A sporozoite asparagine-rich protein controls initiation of Plasmodium liver stage development.

    Directory of Open Access Journals (Sweden)

    Olivier Silvie

    2008-06-01

    Full Text Available Plasmodium sporozoites invade host hepatocytes and develop as liver stages (LS before the onset of erythrocytic infection and malaria symptoms. LS are clinically silent, and constitute ideal targets for causal prophylactic drugs and vaccines. The molecular and cellular mechanisms underlying LS development remain poorly characterized. Here we describe a conserved Plasmodium asparagine-rich protein that is specifically expressed in sporozoites and liver stages. Gene disruption in Plasmodium berghei results in complete loss of sporozoite infectivity to rodents, due to early developmental arrest after invasion of hepatocytes. Mutant sporozoites productively invade host cells by forming a parasitophorous vacuole (PV, but subsequent remodelling of the membrane of the PV (PVM is impaired as a consequence of dramatic down-regulation of genes encoding PVM-resident proteins. These early arrested mutants confer only limited protective immunity in immunized animals. Our results demonstrate the role of an asparagine-rich protein as a key regulator of Plasmodium sporozoite gene expression and LS development, and suggest a requirement of partial LS maturation to induce optimal protective immune responses against malaria pre-erythrocytic stages. These findings have important implications for the development of genetically attenuated parasites as a vaccine approach.

  19. Rapid changes in plasma membrane protein phosphorylation during initiation of cell wall digestion

    International Nuclear Information System (INIS)

    Blowers, D.P.; Boss, W.F.; Trewavas, A.J.

    1988-01-01

    Plasma membrane vesicles from wild carrot cells grown in suspension culture were isolated by aqueous two-phase partitioning, and ATP-dependent phosphorylation was measured with [γ- 32 P]ATP in the presence and absence of calcium. Treatment of the carrot cells with the cell wall digestion enzymes, driselase, in a sorbitol osmoticum for 1.5 min altered the protein phosphorylation pattern compared to that of cells treated with sorbitol alone. Driselase treatment resulted in decreased phosphorylation of a band of M r 80,000 which showed almost complete calcium dependence in the osmoticum treated cells; decreased phosphorylation of a band of M r 15,000 which showed little calcium activation, and appearance of a new band of calcium-dependent phosphorylation at M r 22,000. However, protein phosphorylation was decreased. Adding driselase to the in vitro reaction mixture caused a general decrease in the membrane protein phosphorylation either in the presence or absence of calcium which did not mimic the in vivo response. Cells labeled in vivo with inorganic 32 P also showed a response to the Driselase treatment. An enzymically active driselas preparation was required for the observed responses

  20. Structures of minute virus of mice replication initiator protein N-terminal domain: Insights into DNA nicking and origin binding

    International Nuclear Information System (INIS)

    Tewary, Sunil K.; Liang, Lingfei; Lin, Zihan; Lynn, Annie; Cotmore, Susan F.; Tattersall, Peter; Zhao, Haiyan; Tang, Liang

    2015-01-01

    Members of the Parvoviridae family all encode a non-structural protein 1 (NS1) that directs replication of single-stranded viral DNA, packages viral DNA into capsid, and serves as a potent transcriptional activator. Here we report the X-ray structure of the minute virus of mice (MVM) NS1 N-terminal domain at 1.45 Å resolution, showing that sites for dsDNA binding, ssDNA binding and cleavage, nuclear localization, and other functions are integrated on a canonical fold of the histidine-hydrophobic-histidine superfamily of nucleases, including elements specific for this Protoparvovirus but distinct from its Bocaparvovirus or Dependoparvovirus orthologs. High resolution structural analysis reveals a nickase active site with an architecture that allows highly versatile metal ligand binding. The structures support a unified mechanism of replication origin recognition for homotelomeric and heterotelomeric parvoviruses, mediated by a basic-residue-rich hairpin and an adjacent helix in the initiator proteins and by tandem tetranucleotide motifs in the replication origins. - Highlights: • The structure of a parvovirus replication initiator protein has been determined; • The structure sheds light on mechanisms of ssDNA binding and cleavage; • The nickase active site is preconfigured for versatile metal ligand binding; • The binding site for the double-stranded replication origin DNA is identified; • A single domain integrates multiple functions in virus replication

  1. Structures of minute virus of mice replication initiator protein N-terminal domain: Insights into DNA nicking and origin binding

    Energy Technology Data Exchange (ETDEWEB)

    Tewary, Sunil K.; Liang, Lingfei; Lin, Zihan; Lynn, Annie [Department of Molecular Biosciences, University of Kansas, Lawrence, KS 66045 (United States); Cotmore, Susan F. [Departments of Laboratory Medicine, Yale University Medical School, New Haven, CT 06510 (United States); Tattersall, Peter [Departments of Laboratory Medicine, Yale University Medical School, New Haven, CT 06510 (United States); Departments of Genetics, Yale University Medical School, New Haven, CT 06510 (United States); Zhao, Haiyan, E-mail: zhaohy@ku.edu [Department of Molecular Biosciences, University of Kansas, Lawrence, KS 66045 (United States); Tang, Liang, E-mail: tangl@ku.edu [Department of Molecular Biosciences, University of Kansas, Lawrence, KS 66045 (United States)

    2015-02-15

    Members of the Parvoviridae family all encode a non-structural protein 1 (NS1) that directs replication of single-stranded viral DNA, packages viral DNA into capsid, and serves as a potent transcriptional activator. Here we report the X-ray structure of the minute virus of mice (MVM) NS1 N-terminal domain at 1.45 Å resolution, showing that sites for dsDNA binding, ssDNA binding and cleavage, nuclear localization, and other functions are integrated on a canonical fold of the histidine-hydrophobic-histidine superfamily of nucleases, including elements specific for this Protoparvovirus but distinct from its Bocaparvovirus or Dependoparvovirus orthologs. High resolution structural analysis reveals a nickase active site with an architecture that allows highly versatile metal ligand binding. The structures support a unified mechanism of replication origin recognition for homotelomeric and heterotelomeric parvoviruses, mediated by a basic-residue-rich hairpin and an adjacent helix in the initiator proteins and by tandem tetranucleotide motifs in the replication origins. - Highlights: • The structure of a parvovirus replication initiator protein has been determined; • The structure sheds light on mechanisms of ssDNA binding and cleavage; • The nickase active site is preconfigured for versatile metal ligand binding; • The binding site for the double-stranded replication origin DNA is identified; • A single domain integrates multiple functions in virus replication.

  2. The application of an emerging technique for protein-protein interaction interface mapping: the combination of photo-initiated cross-linking protein nanoprobes with mass spectrometry

    Czech Academy of Sciences Publication Activity Database

    Ptáčková, Renata; Ječmen, Tomáš; Novák, Petr; Šulc, Miroslav; Hudeček, J.; Stiborová, M.

    2014-01-01

    Roč. 15, č. 6 (2014), s. 9224-9241 E-ISSN 1422-0067 R&D Projects: GA ČR(CZ) GAP207/12/0627 Grant - others:Universita Karlova(CZ) 903413; Magistrát hlavního města Prahy(CZ) CZ.2.16/3.1.00/24023; UNCE(BE) 204025/2012 Institutional support: RVO:61388971 Keywords : nanoprobes * mass spectrometry * protein-protein interactions Subject RIV: CE - Biochemistry Impact factor: 2.862, year: 2014

  3. Protein Structure Initiative Material Repository: an open shared public resource of structural genomics plasmids for the biological community

    Science.gov (United States)

    Cormier, Catherine Y.; Mohr, Stephanie E.; Zuo, Dongmei; Hu, Yanhui; Rolfs, Andreas; Kramer, Jason; Taycher, Elena; Kelley, Fontina; Fiacco, Michael; Turnbull, Greggory; LaBaer, Joshua

    2010-01-01

    The Protein Structure Initiative Material Repository (PSI-MR; http://psimr.asu.edu) provides centralized storage and distribution for the protein expression plasmids created by PSI researchers. These plasmids are a resource that allows the research community to dissect the biological function of proteins whose structures have been identified by the PSI. The plasmid annotation, which includes the full length sequence, vector information and associated publications, is stored in a freely available, searchable database called DNASU (http://dnasu.asu.edu). Each PSI plasmid is also linked to a variety of additional resources, which facilitates cross-referencing of a particular plasmid to protein annotations and experimental data. Plasmid samples can be requested directly through the website. We have also developed a novel strategy to avoid the most common concern encountered when distributing plasmids namely, the complexity of material transfer agreement (MTA) processing and the resulting delays this causes. The Expedited Process MTA, in which we created a network of institutions that agree to the terms of transfer in advance of a material request, eliminates these delays. Our hope is that by creating a repository of expression-ready plasmids and expediting the process for receiving these plasmids, we will help accelerate the accessibility and pace of scientific discovery. PMID:19906724

  4. Cooperative Interactions between 480 kDa Ankyrin-G and EB Proteins Assemble the Axon Initial Segment.

    Science.gov (United States)

    Fréal, Amélie; Fassier, Coralie; Le Bras, Barbara; Bullier, Erika; De Gois, Stéphanie; Hazan, Jamilé; Hoogenraad, Casper C; Couraud, François

    2016-04-20

    The axon initial segment (AIS) is required for generating action potentials and maintaining neuronal polarity. Significant progress has been made in deciphering the basic building blocks composing the AIS, but the underlying mechanisms required for AIS formation remains unclear. The scaffolding protein ankyrin-G is the master-organizer of the AIS. Microtubules and their interactors, particularly end-binding proteins (EBs), have emerged as potential key players in AIS formation. Here, we show that the longest isoform of ankyrin-G (480AnkG) selectively associates with EBs via its specific tail domain and that this interaction is crucial for AIS formation and neuronal polarity in cultured rodent hippocampal neurons. EBs are essential for 480AnkG localization and stabilization at the AIS, whereas 480AnkG is required for the specific accumulation of EBs in the proximal axon. Our findings thus provide a conceptual framework for understanding how the cooperative relationship between 480AnkG and EBs induces the assembly of microtubule-AIS structures in the proximal axon. Neuronal polarity is crucial for the proper function of neurons. The assembly of the axon initial segment (AIS), which is the hallmark of early neuronal polarization, relies on the longest 480 kDa ankyrin-G isoform. The microtubule cytoskeleton and its interacting proteins were suggested to be early key players in the process of AIS formation. In this study, we show that the crosstalk between 480 kDa ankyrin-G and the microtubule plus-end tracking proteins, EBs, at the proximal axon is decisive for AIS assembly and neuronal polarity. Our work thus provides insight into the functional mechanisms used by 480 kDa ankyrin-G to drive the AIS formation and thereby to establish neuronal polarity. Copyright © 2016 the authors 0270-6474/16/364421-13$15.00/0.

  5. Translation initiation on mRNAs bound by nuclear cap-binding protein complex CBP80/20 requires interaction between CBP80/20-dependent translation initiation factor and eukaryotic translation initiation factor 3g.

    Science.gov (United States)

    Choe, Junho; Oh, Nara; Park, Sungjin; Lee, Ye Kyung; Song, Ok-Kyu; Locker, Nicolas; Chi, Sung-Gil; Kim, Yoon Ki

    2012-05-25

    In the cytoplasm of mammalian cells, either cap-binding proteins 80 and 20 (CBP80/20) or eukaryotic translation initiation factor (eIF) 4E can direct the initiation of translation. Although the recruitment of ribosomes to mRNAs during eIF4E-dependent translation (ET) is well characterized, the molecular mechanism for CBP80/20-dependent translation (CT) remains obscure. Here, we show that CBP80/20-dependent translation initiation factor (CTIF), which has been shown to be preferentially involved in CT but not ET, specifically interacts with eIF3g, a component of the eIF3 complex involved in ribosome recruitment. By interacting with eIF3g, CTIF serves as an adaptor protein to bridge the CBP80/20 and the eIF3 complex, leading to efficient ribosome recruitment during CT. Accordingly, down-regulation of CTIF using a small interfering RNA causes a redistribution of CBP80 from polysome fractions to subpolysome fractions, without significant consequence to eIF4E distribution. In addition, down-regulation of eIF3g inhibits the efficiency of nonsense-mediated mRNA decay, which is tightly coupled to CT but not to ET. Moreover, the artificial tethering of CTIF to an intercistronic region of dicistronic mRNA results in translation of the downstream cistron in an eIF3-dependent manner. These findings support the idea that CT mechanistically differs from ET.

  6. The effects of increased dietary protein yogurt snack in the afternoon on appetite control and eating initiation in healthy women.

    Science.gov (United States)

    Ortinau, Laura C; Culp, Julie M; Hoertel, Heather A; Douglas, Steve M; Leidy, Heather J

    2013-06-06

    A large portion of daily intake comes from snacking. One of the increasingly common, healthier snacks includes Greek-style yogurt, which is typically higher in protein than regular yogurt. This study evaluated whether a 160 kcal higher-protein (HP) Greek-style yogurt snack improves appetite control, satiety, and delays subsequent eating compared to an isocaloric normal protein (NP) regular yogurt in healthy women. This study also identified the factors that predict the onset of eating. Thirty-two healthy women (age: 27 ± 2y; BMI: 23.0 ± 0.4 kg/m2) completed the acute, randomized crossover-design study. On separate days, participants came to our facility to consume a standardized lunch followed by the consumption of the NP (5.0 g protein) or HP (14.0 g protein) yogurt at 3 h post-lunch. Perceived hunger and fullness were assessed throughout the afternoon until dinner was voluntarily requested; ad libitum dinner was then provided. Snacking led to reductions in hunger and increases in fullness. No differences in post-snack perceived hunger or fullness were observed between the NP and HP yogurt snacks. Dinner was voluntarily requested at approximately 2:40 ± 0:05 h post-snack with no differences between the HP vs. NP yogurts. Ad libitum dinner intake was not different between the snacks (NP: 686 ± 33 kcal vs. HP: 709 ± 34 kcal; p = 0.324). In identifying key factors that predict eating initiation, perceived hunger, fullness, and habitual dinner time accounted for 30% of the variability of time to dinner request (r = 0.55; p < 0.001). The additional 9 g of protein contained in the high protein Greek yogurt was insufficient to elicit protein-related improvements in markers of energy intake regulation.

  7. The effects of increased dietary protein yogurt snack in the afternoon on appetite control and eating initiation in healthy women

    Science.gov (United States)

    2013-01-01

    Background A large portion of daily intake comes from snacking. One of the increasingly common, healthier snacks includes Greek-style yogurt, which is typically higher in protein than regular yogurt. This study evaluated whether a 160 kcal higher-protein (HP) Greek-style yogurt snack improves appetite control, satiety, and delays subsequent eating compared to an isocaloric normal protein (NP) regular yogurt in healthy women. This study also identified the factors that predict the onset of eating. Findings Thirty-two healthy women (age: 27 ± 2y; BMI: 23.0 ± 0.4 kg/m2) completed the acute, randomized crossover-design study. On separate days, participants came to our facility to consume a standardized lunch followed by the consumption of the NP (5.0 g protein) or HP (14.0 g protein) yogurt at 3 h post-lunch. Perceived hunger and fullness were assessed throughout the afternoon until dinner was voluntarily requested; ad libitum dinner was then provided. Snacking led to reductions in hunger and increases in fullness. No differences in post-snack perceived hunger or fullness were observed between the NP and HP yogurt snacks. Dinner was voluntarily requested at approximately 2:40 ± 0:05 h post-snack with no differences between the HP vs. NP yogurts. Ad libitum dinner intake was not different between the snacks (NP: 686 ± 33 kcal vs. HP: 709 ± 34 kcal; p = 0.324). In identifying key factors that predict eating initiation, perceived hunger, fullness, and habitual dinner time accounted for 30% of the variability of time to dinner request (r = 0.55; p < 0.001). Conclusions The additional 9 g of protein contained in the high protein Greek yogurt was insufficient to elicit protein-related improvements in markers of energy intake regulation. PMID:23742659

  8. Granzyme A Cleaves a Mitochondrial Complex I Protein to Initiate Caspase-Independent Cell Death

    Science.gov (United States)

    Martinvalet, Denis; Dykxhoorn, Derek M.; Ferrini, Roger; Lieberman, Judy

    2010-01-01

    SUMMARY The killer lymphocyte protease granzyme A (GzmA) triggers caspase-independent target cell death with morphological features of apoptosis. We previously showed that GzmA acts directly on mitochondria to generate reactive oxygen species (ROS) and disrupt the transmembrane potential (ΔΨm) but does not permeabilize the mitochondrial outer membrane. Mitochondrial damage is critical to GzmA-induced cell death since cells treated with superoxide scavengers are resistant to GzmA. Here we find that GzmA accesses the mitochondrial matrix to cleave the complex I protein NDUFS3, an iron-sulfur subunit of the NADH:ubiquinone oxidoreductase complex I, after Lys56 to interfere with NADH oxidation and generate superoxide anions. Target cells expressing a cleavage site mutant of NDUFS3 are resistant to GzmA-mediated cell death but remain sensitive to GzmB. PMID:18485875

  9. N-MYC down-regulated-like proteins regulate meristem initiation by modulating auxin transport and MAX2 expression.

    Science.gov (United States)

    Mudgil, Yashwanti; Ghawana, Sanjay; Jones, Alan M

    2013-01-01

    N-MYC down-regulated-like (NDL) proteins interact with the Gβ subunit (AGB1) of the heterotrimeric G protein complex and play an important role in AGB1-dependent regulation of lateral root formation by affecting root auxin transport, auxin gradients and the steady-state levels of mRNA encoding the PIN-FORMED 2 and AUXIN 1 auxin transport facilitators. Auxin transport in aerial tissue follows different paths and utilizes different transporters than in roots; therefore, in the present study, we analyzed whether NDL proteins play an important role in AGB1-dependent, auxin-mediated meristem development. Expression levels of NDL gene family members need to be tightly regulated, and altered expression (both over-expression and down-regulation) confers ectopic growth. Over-expression of NDL1 disrupts vegetative and reproductive organ development. Reduced expression of the NDL gene family members results in asymmetric leaf emergence, twinning of rosette leaves, defects in leaf formation, and abnormal silique distribution. Reduced expression of the NDL genes in the agb1-2 (null allele) mutant rescues some of the abnormal phenotypes, such as silique morphology, silique distribution, and peduncle angle, suggesting that proper levels of NDL proteins are maintained by AGB1. We found that all of these abnormal aerial phenotypes due to altered NDL expression were associated with increases in basipetal auxin transport, altered auxin maxima and altered MAX2 expression within the inflorescence stem. NDL proteins, together with AGB1, act as positive regulators of meristem initiation and branching. AGB1 and NDL1 positively regulate basipetal inflorescence auxin transport and modulate MAX2 expression in shoots, which in turn regulates organ and lateral meristem formation by the establishment and maintenance of auxin gradients.

  10. N-MYC down-regulated-like proteins regulate meristem initiation by modulating auxin transport and MAX2 expression.

    Directory of Open Access Journals (Sweden)

    Yashwanti Mudgil

    Full Text Available N-MYC down-regulated-like (NDL proteins interact with the Gβ subunit (AGB1 of the heterotrimeric G protein complex and play an important role in AGB1-dependent regulation of lateral root formation by affecting root auxin transport, auxin gradients and the steady-state levels of mRNA encoding the PIN-FORMED 2 and AUXIN 1 auxin transport facilitators. Auxin transport in aerial tissue follows different paths and utilizes different transporters than in roots; therefore, in the present study, we analyzed whether NDL proteins play an important role in AGB1-dependent, auxin-mediated meristem development.Expression levels of NDL gene family members need to be tightly regulated, and altered expression (both over-expression and down-regulation confers ectopic growth. Over-expression of NDL1 disrupts vegetative and reproductive organ development. Reduced expression of the NDL gene family members results in asymmetric leaf emergence, twinning of rosette leaves, defects in leaf formation, and abnormal silique distribution. Reduced expression of the NDL genes in the agb1-2 (null allele mutant rescues some of the abnormal phenotypes, such as silique morphology, silique distribution, and peduncle angle, suggesting that proper levels of NDL proteins are maintained by AGB1. We found that all of these abnormal aerial phenotypes due to altered NDL expression were associated with increases in basipetal auxin transport, altered auxin maxima and altered MAX2 expression within the inflorescence stem.NDL proteins, together with AGB1, act as positive regulators of meristem initiation and branching. AGB1 and NDL1 positively regulate basipetal inflorescence auxin transport and modulate MAX2 expression in shoots, which in turn regulates organ and lateral meristem formation by the establishment and maintenance of auxin gradients.

  11. Elimination of head and neck cancer initiating cells through targeting glucose regulated protein78 signaling

    Directory of Open Access Journals (Sweden)

    Huang Chih-Yang

    2010-10-01

    Full Text Available Abstract Background Head and neck squamous cell carcinoma (HNSCC is a highly lethal cancer that contains cellular and functional heterogeneity. Previously, we enriched a subpopulation of highly tumorigenic head and neck cancer initiating cells (HN-CICs from HNSCC. However, the molecular mechanisms by which to govern the characteristics of HN-CICs remain unclear. GRP78, a stress-inducible endoplasmic reticulum chaperone, has been reported to play a crucial role in the maintenance of embryonic stem cells, but the role of GRP78 in CICs has not been elucidated. Results Initially, we recognized GRP78 as a putative candidate on mediating the stemness and tumorigenic properties of HN-CICs by differential systemic analyses. Subsequently, cells with GRP78 anchored at the plasma membrane (memGRP78+ exerted cancer stemness properties of self-renewal, differentiation and radioresistance. Of note, xenotransplantation assay indicated merely 100 memGRP78+ HNSCCs resulted in tumor growth. Moreover, knockdown of GRP78 significantly reduced the self-renewal ability, side population cells and expression of stemness genes, but inversely promoted cell differentiation and apoptosis in HN-CICs. Targeting GRP78 also lessened tumorigenicity of HN-CICs both in vitro and in vivo. Clinically, co-expression of GRP78 and Nanog predicted the worse survival prognosis of HNSCC patients by immunohistochemical analyses. Finally, depletion of GRP78 in HN-CICs induced the expression of Bax, Caspase 3, and PTEN. Conclusions In summary, memGRP78 should be a novel surface marker for isolation of HN-CICs, and targeting GRP78 signaling might be a potential therapeutic strategy for HNSCC through eliminating HN-CICs.

  12. Glutamine domain of the chimeric protein, CAD, that initiates pyrimidine biosynthesis in mammalian cells

    International Nuclear Information System (INIS)

    Kelly, R.E.; Kim, H.; Evans, D.R.

    1986-01-01

    Glutamine dependent carbamyl phosphate synthesis, the first step in mammalian de novo pyrimidine biosynthesis, is catalyzed by a 240 kDa chimeric protein, CAD, that also has the aspartate transcarbamylase and dihydroorotase activities. The complex was found to have a separate glutaminase activity of 0.04 μmol/min/mg, that increased five fold in the presence of bicarbonate and ATP. To determine whether the glutaminase activity, which provides ammonia for carbamyl phosphate synthesis, is associated with a separate structural domain (GLN), CAD was subjected to controlled proteolysis with elastase. The glutaminase, glutamine and ammonia dependent carbamyl phosphate synthetase activities, as well as the partial reactions; carbamyl phosphate dependent ATP synthesis and bicarbonate dependent ATPase, were correlated with the concentration of the various proteolytic fragments that accumulated in the digest. While the glutamine dependent carbamyl phosphate synthetase was rapidly inactivated, the glutaminase activity was found to be very resistant to proteolysis. The glutamine binding site of CAD was also specifically modified with 6-diazo-5-oxo-L-norleucine (DON). The modification was accompanied by a loss of both glutaminase and glutamine dependent carbamyl phosphate synthetase activities. Bicarbonate and ATP increased the rate of reaction of CAD with DON, while glutamine protected against inactivation. The stoichiometry of the reaction and the identity of the modified proteolytic fragments was determined using 14 C labelled DON

  13. The NS1 Protein from Influenza Virus Stimulates Translation Initiation by Enhancing Ribosome Recruitment to mRNAs.

    Science.gov (United States)

    Panthu, Baptiste; Terrier, Olivier; Carron, Coralie; Traversier, Aurélien; Corbin, Antoine; Balvay, Laurent; Lina, Bruno; Rosa-Calatrava, Manuel; Ohlmann, Théophile

    2017-10-27

    The non-structural protein NS1 of influenza A viruses exerts pleiotropic functions during infection. Among these functions, NS1 was shown to be involved in the control of both viral and cellular translation; however, the mechanism by which this occurs remains to be determined. Thus, we have revisited the role of NS1 in translation by using a combination of influenza infection, mRNA reporter transfection, and in vitro functional and biochemical assays. Our data show that the NS1 protein is able to enhance the translation of virtually all tested mRNAs with the exception of constructs bearing the Dicistroviruses Internal ribosome entry segment (IRESes) (DCV and CrPV), suggesting a role at the level of translation initiation. The domain of NS1 required for translation stimulation was mapped to the RNA binding amino-terminal motif of the protein with residues R38 and K41 being critical for activity. Although we show that NS1 can bind directly to mRNAs, it does not correlate with its ability to stimulate translation. This activity rather relies on the property of NS1 to associate with ribosomes and to recruit them to target mRNAs. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. An isolated Hda-clamp complex is functional in the regulatory inactivation of DnaA and DNA replication.

    Science.gov (United States)

    Kawakami, Hironori; Su'etsugu, Masayuki; Katayama, Tsutomu

    2006-10-01

    In Escherichia coli, a complex consisting of Hda and the DNA-loaded clamp-subunit of the DNA polymerase III holoenzyme promotes hydrolysis of DnaA-ATP. The resultant ADP-DnaA is inactive for initiation of chromosomal DNA replication, thereby repressing excessive initiations. As the cellular content of the clamp is 10-100 times higher than that of Hda, most Hda molecules might be complexed with the clamp in vivo. Although Hda predominantly forms irregular aggregates when overexpressed, in the present study we found that co-overexpression of the clamp with Hda enhances Hda solubility dramatically and we efficiently isolated the Hda-clamp complex. A single molecule of the complex appears to consist of two Hda molecules and a single clamp. The complex is competent in DnaA-ATP hydrolysis and DNA replication in the presence of DNA and the clamp deficient subassembly of the DNA polymerase III holoenzyme (pol III*). These findings indicate that the clamp contained in the complex is loaded onto DNA through an interaction with the pol III* and that the Hda activity is preserved in these processes. The complex consisting of Hda and the DNA-unloaded clamp may play a specific role in a process proceeding to the DnaA-ATP hydrolysis in vivo.

  15. Effect of a high-protein diet on maintenance of blood pressure levels achieved after initial weight loss : the DiOGenes randomized study

    NARCIS (Netherlands)

    Engberink, M. F.; Geleijnse, J. M.; Bakker, S. J. L.; Larsen, T. M.; Handjieva-Darlesnka, T.; Kafatos, A.; Martinez, J. A.; Pfeiffer, A. F. H.; Kunesova, M.; Jebb, S. A.; Holst, C.; Astrup, A.; Saris, W. H. M.; Brink, E. J.; van Baak, M. A.

    Randomized trials have shown significant blood pressure (BP) reductions after increased protein compared with carbohydrate intake, but the effect on BP maintenance after initial weight loss is unclear. We examined the effect of a high-protein diet on the maintenance of reduced BP after weight loss

  16. Effect of a high-protein diet on maintenance of blood pressure levels achieved after initial weight loss: the DiOGenes randomized study

    NARCIS (Netherlands)

    Engberink, M.F.; Geleijnse, J.M.; Bakker, S.J.L.; Larsen, T.

    2015-01-01

    Randomized trials have shown significant blood pressure (BP) reductions after increased protein compared with carbohydrate intake, but the effect on BP maintenance after initial weight loss is unclear. We examined the effect of a high-protein diet on the maintenance of reduced BP after weight loss

  17. Effect of baculovirus infection on the mRNA and protein levels of the Spodoptera frugiperda eukaryotic initiation factor 4E

    NARCIS (Netherlands)

    Oers, van M.M.; Veken, van der L.T.J.N.; Vlak, J.M.; Thomas, A.A.M.

    2001-01-01

    The cDNA sequence of eukaryotic translation initiation factor eIF4E was derived from a Spodoptera frugiperda cDNA library. Eight tryptophan residues, typical for eIF4E, are strictly conserved in the encoded 210 amino acid protein. A polyclonal antiserum detected a 26 kDa protein in lepidopteran cell

  18. Subcellular Trafficking of the Papillomavirus Genome during Initial Infection: The Remarkable Abilities of Minor Capsid Protein L2

    Directory of Open Access Journals (Sweden)

    Samuel K. Campos

    2017-12-01

    Full Text Available Since 2012, our understanding of human papillomavirus (HPV subcellular trafficking has undergone a drastic paradigm shift. Work from multiple laboratories has revealed that HPV has evolved a unique means to deliver its viral genome (vDNA to the cell nucleus, relying on myriad host cell proteins and processes. The major breakthrough finding from these recent endeavors has been the realization of L2-dependent utilization of cellular sorting factors for the retrograde transport of vDNA away from degradative endo/lysosomal compartments to the Golgi, prior to mitosis-dependent nuclear accumulation of L2/vDNA. An overview of current models of HPV entry, subcellular trafficking, and the role of L2 during initial infection is provided below, highlighting unresolved questions and gaps in knowledge.

  19. A photo-responsive F-box protein FOF2 regulates floral initiation by promoting FLC expression in Arabidopsis.

    Science.gov (United States)

    He, Reqing; Li, Xinmei; Zhong, Ming; Yan, Jindong; Ji, Ronghuan; Li, Xu; Wang, Qin; Wu, Dan; Sun, Mengsi; Tang, Dongying; Lin, Jianzhong; Li, Hongyu; Liu, Bin; Liu, Hongtao; Liu, Xuanming; Zhao, Xiaoying; Lin, Chentao

    2017-09-01

    Floral initiation is regulated by various genetic pathways in response to light, temperature, hormones and developmental status; however, the molecular mechanisms underlying the interactions between different genetic pathways are not fully understood. Here, we show that the photoresponsive gene FOF2 (F-box of flowering 2) negatively regulates flowering. FOF2 encodes a putative F-box protein that interacts specifically with ASK14, and its overexpression results in later flowering under both long-day and short-day photoperiods. Conversely, transgenic plants expressing the F-box domain deletion mutant of FOF2 (FOF2ΔF), or double loss of function mutant of FOF2 and FOL1 (FOF2-LIKE 1) present early flowering phenotypes. The late flowering phenotype of the FOF2 overexpression lines is suppressed by the flc-3 loss-of-function mutation. Furthermore, FOF2 mRNA expression is regulated by autonomous pathway gene FCA, and the repressive effect of FOF2 in flowering can be overcome by vernalization. Interestingly, FOF2 expression is regulated by light. The protein level of FOF2 accumulates in response to light, whereas it is degraded under dark conditions via the 26S proteasome pathway. Our findings suggest a possible mechanistic link between light conditions and the autonomous floral promotion pathway in Arabidopsis. © 2017 The Authors The Plant Journal © 2017 John Wiley & Sons Ltd.

  20. An isoform of eukaryotic initiation factor 4E from Chrysanthemum morifolium interacts with Chrysanthemum virus B coat protein.

    Directory of Open Access Journals (Sweden)

    Aiping Song

    Full Text Available BACKGROUND: Eukaryotic translation initiation factor 4E (eIF4E plays an important role in plant virus infection as well as the regulation of gene translation. METHODOLOGY/PRINCIPAL FINDINGS: Here, we describe the isolation of a cDNA encoding CmeIF(iso4E (GenBank accession no. JQ904592, an isoform of eIF4E from chrysanthemum, using RACE PCR. We used the CmeIF(iso4E cDNA for expression profiling and to analyze the interaction between CmeIF(iso4E and the Chrysanthemum virus B coat protein (CVBCP. Multiple sequence alignment and phylogenetic tree analysis showed that the sequence similarity of CmeIF(iso4E with other reported plant eIF(iso4E sequences varied between 69.12% and 89.18%, indicating that CmeIF(iso4E belongs to the eIF(iso4E subfamily of the eIF4E family. CmeIF(iso4E was present in all chrysanthemum organs, but was particularly abundant in the roots and flowers. Confocal microscopy showed that a transiently transfected CmeIF(iso4E-GFP fusion protein distributed throughout the whole cell in onion epidermis cells. A yeast two hybrid assay showed CVBCP interacted with CmeIF(iso4E but not with CmeIF4E. BiFC assay further demonstrated the interaction between CmeIF(iso4E and CVBCP. Luminescence assay showed that CVBCP increased the RLU of Luc-CVB, suggesting CVBCP might participate in the translation of viral proteins. CONCLUSIONS/SIGNIFICANCE: These results inferred that CmeIF(iso4E as the cap-binding subunit eIF(iso4F may be involved in Chrysanthemum Virus B infection in chrysanthemum through its interaction with CVBCP in spatial.

  1. The Fusion Loops of the Initial Prefusion Conformation of Herpes Simplex Virus 1 Fusion Protein Point Toward the Membrane

    Directory of Open Access Journals (Sweden)

    Juan Fontana

    2017-08-01

    Full Text Available All enveloped viruses, including herpesviruses, must fuse their envelope with the host membrane to deliver their genomes into target cells, making this essential step subject to interference by antibodies and drugs. Viral fusion is mediated by a viral surface protein that transits from an initial prefusion conformation to a final postfusion conformation. Strikingly, the prefusion conformation of the herpesvirus fusion protein, gB, is poorly understood. Herpes simplex virus (HSV, a model system for herpesviruses, causes diseases ranging from mild skin lesions to serious encephalitis and neonatal infections. Using cryo-electron tomography and subtomogram averaging, we have characterized the structure of the prefusion conformation and fusion intermediates of HSV-1 gB. To this end, we have set up a system that generates microvesicles displaying full-length gB on their envelope. We confirmed proper folding of gB by nondenaturing electrophoresis-Western blotting with a panel of monoclonal antibodies (MAbs covering all gB domains. To elucidate the arrangement of gB domains, we labeled them by using (i mutagenesis to insert fluorescent proteins at specific positions, (ii coexpression of gB with Fabs for a neutralizing MAb with known binding sites, and (iii incubation of gB with an antibody directed against the fusion loops. Our results show that gB starts in a compact prefusion conformation with the fusion loops pointing toward the viral membrane and suggest, for the first time, a model for gB’s conformational rearrangements during fusion. These experiments further illustrate how neutralizing antibodies can interfere with the essential gB structural transitions that mediate viral entry and therefore infectivity.

  2. Low-protein diet for chronic kidney disease in the Caserta Local Health Unit: the SaniARP Initiative

    Directory of Open Access Journals (Sweden)

    Chiara Troncone

    2015-06-01

    disease has increased. Currently, these products are not yet contemplated in the essential levels of assistance but the Italian regions deliver, low-protein food discretionally using their own funds. The Campania Region, in 2010, interrupted the distribution of these products. With the exception of Caserta which took up distribution again in 2013. OBJECTIVE: The aim of this paper is to describe an initiative put in place by Caserta which has decided to invest in prevention. MATERIALS AND METHODS: A treatment plan for the distribution designed by a team of in-house nephrologists and data are recorded using Saniarp, a web-based platform. RESULTS: In the observation period patients with a prescription of low-protein food products were 869. The mean age was 61 years. The average cost patient / month for the nutritional treatment was 59 Euro. The average cost patient / month for any type of drug was 632 Euro. In particular, 48 Euro for EPO, 277 Euro for Chelate Agents, 16 Euro for antihypertensive therapy. DISCUSSION AND CONCLUSIONS: The policy put in place by the LHU Caserta improved care of kidney patients. The results available to date are still incomplete and do not enable us to clearly assess the benefits both in clinical and economic terms which can be produced by a low-protein diet in kidney patients. In the perspective of third party payers to budget this expense it appears entirely sustainable especially in view of the fact that this dietary treatment might delay the onset of dialysis therapy and lead to lower comorbidity for the patient.

  3. Viral Genome-Linked Protein (VPg Is Essential for Translation Initiation of Rabbit Hemorrhagic Disease Virus (RHDV.

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    Jie Zhu

    Full Text Available Rabbit hemorrhagic disease virus (RHDV, the causative agent of rabbit hemorrhagic disease, is an important member of the caliciviridae family. Currently, no suitable tissue culture system is available for proliferating RHDV, limiting the study of the pathogenesis of RHDV. In addition, the mechanisms underlying RHDV translation and replication are largely unknown compared with other caliciviridae viruses. The RHDV replicon recently constructed in our laboratory provides an appropriate model to study the pathogenesis of RHDV without in vitro RHDV propagation and culture. Using this RHDV replicon, we demonstrated that the viral genome-linked protein (VPg is essential for RHDV translation in RK-13 cells for the first time. In addition, we showed that VPg interacts with eukaryotic initiation factor 4E (eIF4E in vivo and in vitro and that eIF4E silencing inhibits RHDV translation, suggesting the interaction between VPg and eIF4E is involved in RHDV translation. Our results support the hypothesis that VPg serves as a novel cap substitute during the initiation of RHDV translation.

  4. Paralogous SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) genes differentially regulate leaf initiation and reproductive phase change in petunia.

    Science.gov (United States)

    Preston, Jill C; Jorgensen, Stacy A; Orozco, Rebecca; Hileman, Lena C

    2016-02-01

    Duplicated petunia clade-VI SPL genes differentially promote the timing of inflorescence and flower development, and leaf initiation rate. The timing of plant reproduction relative to favorable environmental conditions is a critical component of plant fitness, and is often associated with variation in plant architecture and habit. Recent studies have shown that overexpression of the microRNA miR156 in distantly related annual species results in plants with perennial characteristics, including late flowering, weak apical dominance, and abundant leaf production. These phenotypes are largely mediated through the negative regulation of a subset of genes belonging to the SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) family of transcription factors. In order to determine how and to what extent paralogous SPL genes have partitioned their roles in plant growth and development, we functionally characterized petunia clade-VI SPL genes under different environmental conditions. Our results demonstrate that PhSBP1and PhSBP2 differentially promote discrete stages of the reproductive transition, and that PhSBP1, and possibly PhCNR, accelerates leaf initiation rate. In contrast to the closest homologs in annual Arabidopsis thaliana and Mimulus guttatus, PhSBP1 and PhSBP2 transcription is not mediated by the gibberellic acid pathway, but is positively correlated with photoperiod and developmental age. The developmental functions of clade-VI SPL genes have, thus, evolved following both gene duplication and speciation within the core eudicots, likely through differential regulation and incomplete sub-functionalization.

  5. Experimental Analysis of Mimivirus Translation Initiation Factor 4a Reveals Its Importance in Viral Protein Translation during Infection of Acanthamoeba polyphaga.

    Science.gov (United States)

    Bekliz, Meriem; Azza, Said; Seligmann, Hervé; Decloquement, Philippe; Raoult, Didier; La Scola, Bernard

    2018-05-15

    The Acanthamoeba polyphaga mimivirus is the first giant virus ever described, with a 1.2-Mb genome which encodes 979 proteins, including central components of the translation apparatus. One of these proteins, R458, was predicted to initiate translation, although its specific role remains unknown. We silenced the R458 gene using small interfering RNA (siRNA) and compared levels of viral fitness and protein expression in silenced versus wild-type mimivirus. Silencing decreased the growth rate, but viral particle production at the end of the viral cycle was unaffected. A comparative proteomic approach using two-dimensional difference-in-gel electrophoresis (2D-DIGE) revealed deregulation of the expression of 32 proteins in silenced mimivirus, which were defined as up- or downregulated. Besides revealing proteins with unknown functions, silencing R458 also revealed deregulation in proteins associated with viral particle structures, transcriptional machinery, oxidative pathways, modification of proteins/lipids, and DNA topology/repair. Most of these proteins belong to genes transcribed at the end of the viral cycle. Overall, our data suggest that the R458 protein regulates the expression of mimivirus proteins and, thus, that mimivirus translational proteins may not be strictly redundant in relation to those from the amoeba host. As is the case for eukaryotic initiation factor 4a (eIF4a), the R458 protein is the prototypical member of the ATP-dependent DEAD box RNA helicase mechanism. We suggest that the R458 protein is required to unwind the secondary structures at the 5' ends of mRNAs and to bind the mRNA to the ribosome, making it possible to scan for the start codon. These data are the first experimental evidence of mimivirus translation-related genes, predicted to initiate protein biosynthesis. IMPORTANCE The presence in the genome of a mimivirus of genes coding for many translational processes, with the exception of ribosome constituents, has been the subject of

  6. Structural Transition and Antibody Binding of EBOV GP and ZIKV E Proteins from Pre-Fusion to Fusion-Initiation State

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    Anna Lappala

    2018-05-01

    Full Text Available Membrane fusion proteins are responsible for viral entry into host cells—a crucial first step in viral infection. These proteins undergo large conformational changes from pre-fusion to fusion-initiation structures, and, despite differences in viral genomes and disease etiology, many fusion proteins are arranged as trimers. Structural information for both pre-fusion and fusion-initiation states is critical for understanding virus neutralization by the host immune system. In the case of Ebola virus glycoprotein (EBOV GP and Zika virus envelope protein (ZIKV E, pre-fusion state structures have been identified experimentally, but only partial structures of fusion-initiation states have been described. While the fusion-initiation structure is in an energetically unfavorable state that is difficult to solve experimentally, the existing structural information combined with computational approaches enabled the modeling of fusion-initiation state structures of both proteins. These structural models provide an improved understanding of four different neutralizing antibodies in the prevention of viral host entry.

  7. Analysis of initial changes in the proteins of soybean root tip under flooding stress using gel-free and gel-based proteomic techniques.

    Science.gov (United States)

    Yin, Xiaojian; Sakata, Katsumi; Nanjo, Yohei; Komatsu, Setsuko

    2014-06-25

    Flooding has a severe negative effect on soybean cultivation in the early stages of growth. To obtain a better understanding of the response mechanisms of soybean to flooding stress, initial changes in root tip proteins under flooding were analyzed using two proteomic techniques. Two-day-old soybeans were treated with flooding for 3, 6, 12, and 24h. The weight of soybeans increased during the first 3h of flooding, but root elongation was not observed. Using gel-based and gel-free proteomic techniques, 115 proteins were identified in root tips, of which 9 proteins were commonly detected by both methods. The 71 proteins identified by the gel-free proteomics were analyzed by a hierarchical clustering method based on induction levels during the flooding, and the proteins were divided into 5 clusters. Additional interaction analysis of the proteins revealed that ten proteins belonging to cluster I formed the center of a protein interaction network. mRNA expression analysis of these ten proteins showed that citrate lyase and heat shock protein 70 were down-regulated, whereas calreticulin was up-regulated in initial phase of flooding. These results suggest that flooding stress to soybean induces calcium-related signal transduction, which might play important roles in the early responses to flooding. Flooding has a severe negative effect on soybean cultivation, particularly in the early stages of growth. To better understand the response mechanisms of soybean to the early stages of flooding stress, two proteomic techniques were used. Two-day-old soybeans were treated without or with flooding for 3, 6, 12, and 24h. The fresh weight of soybeans increased during the first 3h of flooding stress, but the growth then slowed and no root elongation was observed. Using gel-based and gel-free proteomic techniques, 115 proteins were identified in root tips, of which 9 proteins were commonly detected by both methods. The 71 proteins identified by the gel-free proteomics were analyzed

  8. Antiviral RNA silencing initiated in the absence of RDE-4, a double-stranded RNA binding protein, in Caenorhabditis elegans.

    Science.gov (United States)

    Guo, Xunyang; Zhang, Rui; Wang, Jeffrey; Lu, Rui

    2013-10-01

    Small interfering RNAs (siRNAs) processed from double-stranded RNA (dsRNA) of virus origins mediate potent antiviral defense through a process referred to as RNA interference (RNAi) or RNA silencing in diverse organisms. In the simple invertebrate Caenorhabditis elegans, the RNAi process is initiated by a single Dicer, which partners with the dsRNA binding protein RDE-4 to process dsRNA into viral siRNAs (viRNAs). Notably, in C. elegans this RNA-directed viral immunity (RDVI) also requires a number of worm-specific genes for its full antiviral potential. One such gene is rsd-2 (RNAi spreading defective 2), which was implicated in RDVI in our previous studies. In the current study, we first established an antiviral role by showing that rsd-2 null mutants permitted higher levels of viral RNA accumulation, and that this enhanced viral susceptibility was reversed by ectopic expression of RSD-2. We then examined the relationship of rsd-2 with other known components of RNAi pathways and established that rsd-2 functions in a novel pathway that is independent of rde-4 but likely requires the RNA-dependent RNA polymerase RRF-1, suggesting a critical role for RSD-2 in secondary viRNA biogenesis, likely through coordinated action with RRF-1. Together, these results suggest that RDVI in the single-Dicer organism C. elegans depends on the collective actions of both RDE-4-dependent and RDE-4-independent mechanisms to produce RNAi-inducing viRNAs. Our study reveals, for the first time, a novel siRNA-producing mechanism in C. elegans that bypasses the need for a dsRNA-binding protein.

  9. Effect of intravenous administration of D-lysergic acid diethylamide on initiation of protein synthesis in a cell-free system derived from brain.

    Science.gov (United States)

    Cosgrove, J W; Brown, I R

    1984-05-01

    An initiating cell-free protein synthesis system derived from brain was utilized to demonstrate that the intravenous injection of D-lysergic acid diethylamide (LSD) to rabbits resulted in a lesion at the initiation stage of brain protein synthesis. Three inhibitors of initiation, edeine, poly(I), and aurintricarboxylic acid were used to demonstrate a reduction in initiation-dependent amino acid incorporation in the brain cell-free system. One hour after LSD injection, there was also a measurable decrease in the formation of 40S and 80S initiation complexes in vitro, using either [35S]methionine or [35S]Met-tRNAf. Analysis of the methionine pool size after LSD administration indicated there was no change in methionine levels. Analysis of the formation of initiation complexes in the brain cell-free protein synthesis system prepared 6 h after LSD administration indicated that there was a return to control levels at this time. The effects of LSD on steps in the initiation process are thus reversible.

  10. Backbone and sidechain methyl Ile (δ1), Leu and Val chemical shift assignments of RDE-4 (1-243), an RNA interference initiation protein in C. elegans.

    Science.gov (United States)

    Chiliveri, Sai Chaitanya; Kumar, Sonu; Marelli, Udaya Kiran; Deshmukh, Mandar V

    2012-10-01

    The RNAi pathway of several organisms requires presence of double stranded RNA binding proteins for functioning of Dicer in gene regulation. In C. elegans, a double stranded RNA binding protein, RDE-4 (385 aa, 44 kDa) recognizes long exogenous dsRNA and initiates the RNAi pathway. We have achieved complete backbone and stereospecific methyl sidechain Ile (δ1), Leu and Val chemical shifts of first 243 amino acids of RDE-4, namely RDE-4ΔC.

  11. Identification of Plasmodium falciparum translation initiation eIF2β subunit: direct interaction with Protein Phosphatase type 1

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    Géraldine eTellier

    2016-05-01

    Full Text Available Protein phosphatase 1 (PP1c is one of the main phosphatases whose function is shaped by many regulators to confer a specific location and a selective function for this enzyme. Here, we report that eukaryotic initiation factor 2 of P. falciparum (PfeIF2β is an interactor of PfPP1c. Sequence analysis of PfeIF2 revealed a deletion of 111 amino acids when compared to its human counterpart and the presence of two potential binding motifs to PfPP1 (29FGEKKK34, 103KVAW106. As expected, we showed that PfeIF2 binds PfeIF2 and PfeIF5, confirming its canonical interaction with partners of the translation complex. Studies of the PfeIF2-PfPP1 interaction using wild-type, single and double mutated versions of PfeIF2β revealed that both binding motifs are critical. We next showed that PfeIF2 is able to induce Germinal Vesicle BreakDown (GVBD when expressed in Xenopus oocytes, an indicator of its capacity to regulate PP1. Only combined mutations of both binding motifs abolished the interaction with PP1 and the induction of GVBD. In P. falciparum, although the locus is accessible for genetic manipulation, PfeIF2 seems to play an essential role in intraerythrocytic cycle as no viable knockout parasites were detectable. Interestingly, as for PfPP1, the subcellular fractionation of P. falciparum localized PfeIF2β in cytoplasm and nuclear extracts, suggesting a potential effect on PfPP1 in both compartments and raising the question of a non-canonical function of PfeIf2 in the nucleus. Hence, the role played by PfeIF2 in blood stage parasites could occur at multiple levels involving the binding to proteins of the translational complex and to PfPP1.

  12. Identification of Plasmodium falciparum Translation Initiation eIF2β Subunit: Direct Interaction with Protein Phosphatase Type 1

    Czech Academy of Sciences Publication Activity Database

    Tellier, G.; Lenne, A.; Cailliau-Maggio, K.; Cabezas-Cruz, A.; Valdés, James J.; Martoriati, A.; Aliouat, El M.; Gosset, P.; Delaire, B.; Fréville, A.; Pierrot, C.; Khalife, J.

    2016-01-01

    Roč. 7, MAY 26 (2016), č. článku 777. ISSN 1664-302X Institutional support: RVO:60077344 Keywords : Plasmodium falciparum * Protein Phosphatase type1 * eIF2b * protein-protein interaction * translation complex Subject RIV: EE - Microbiology, Virology Impact factor: 4.076, year: 2016

  13. Control of protein synthesis in Escherichia coli: strain differences in control of translational initiation after energy source shift-down.

    Science.gov (United States)

    Jacobson, L A; Jen-Jacobson, L

    1980-06-01

    We have studied the parameters of protein synthesis in a number of Escherichia coli strains after a shift-down from glucose-minimal to succinate-minimal medium. One group of strains, including K-12(lambda) (ATCC 10798) and NF162, showed a postshift translational yield of 50 to 65% and a 2- to 2.5-fold increase in the functional lifetime of general messenger ribonucleic acid. There was no change in the lag time for beta-galactosidase induction in these strains after the shift-down. A second group, including W1 and W2, showed no reduction in translational yield, no change in the functional lifetime of messenger ribonucleic acid, and a 50% increase in the lag time for beta-galactosidase induction. Evidence is presented which indicates that this increased lag time is not the result of a decreased rate of polypeptide chain propagation. A third group of strains, including NF161, CP78, and NF859, showed an intermediate pattern: translational yield was reduced to about 75% of normal, and the messenger ribonucleic acid functional lifetime was increased by about 50%. Calculation of the relative postshift rates of translational initiation gave about 0.2, 1.0, and 0.5, respectively, for the three groups. There was no apparent correlation between the ability to control translation and the genotypes of these strains at the relA, relX, or spoT loci. Measurements of the induction lag for beta-galactosidase during short-term glucose starvation or after a down-shift induced by alpha-methylglucoside indicated that these regimens elicit responses that are physiologically distinct from those elicited by a glucose-to-succinate shift-down.

  14. Initial proteome analysis of caffeine-induced proteins in Aspergillus tamarii using two-dimensional fluorescence difference gel electrophoresis.

    Science.gov (United States)

    Gutiérrez-Sánchez, Gerardo; Atwood, James; Kolli, V S Kumar; Roussos, Sévastianos; Augur, Christopher

    2012-04-01

    Caffeine is toxic to most microorganisms. However, some filamentous fungi, such as Aspergillus tamarii, are able to metabolize this alkaloid when fed caffeine as the sole nitrogen source. The aim of the present work was to identify intracellular A. tamarii proteins, regulated by caffeine, using fluorescence difference two-dimensional gel electrophoresis. Specific proteins from two culture media of A. tamarii grown either on ammonium sulfate or caffeine as the sole nitrogen source were analysed by mass spectrometry. Thirteen out of a total of 85 differentially expressed spots were identified after database search. Identified up-regulated proteins include phosphoglycerate kinase, malate dehydrogenase, dyp-type peroxidase family protein, heat shock protein, Cu, Zn superoxidase dismutase and xanthine dehydrogenase. Some of the proteins identified in this study are involved in the caffeine degradation pathway as well as in stress response, suggesting that stress proteins could be involved in caffeine metabolism in filamentous fungi.

  15. Depletion of elongation initiation factor 4E binding proteins by CRISPR/Cas9 genome editing enhances antiviral response in porcine cells

    Science.gov (United States)

    Type I interferons (IFN) are key mediators of the innate antiviral response in mammalian cells. Elongation initiation factor 4E binding proteins (4E-BPs) are translational controllers of interferon regulatory factor 7 (IRF7), the master regulator of IFN transcription. The role of 4EBPs in the negat...

  16. C-reactive protein implications in new-onset hypertension in a healthy population initially aged 65 years : the Proof study

    NARCIS (Netherlands)

    Dauphinot, Virginie; Roche, Frederic; Kossovsky, Michel P.; Schott, Anne-Marie; Pichot, Vincent; Gaspoz, Jean-Michel; Gosse, Philippe; Barthelemy, Jean-Claude

    Background Because inflammation is known to be related with several cardiovascular diseases, we sought to determine whether C-reactive protein (CRP) might precede the onset of hypertension. Methods The study population was selected from the Proof study cohort including 1011 individuals initially

  17. Crystallization and preliminary crystallographic studies of the W2 domain of Drosophila melanogaster eukaryotic translation initiation factor 5C domain-containing protein

    International Nuclear Information System (INIS)

    Zhao, Hui; Wang, Hong; Liu, Huihui; Teng, Maikun; Li, Xu

    2012-01-01

    The crystallization and preliminary crystallographic studies of the carboxy-terminal domain of D. melanogaster eukaryotic translation initiation factor 5C domain-containing protein are reported. The Drosophila melanogaster eukaryotic translation initiation factor 5C domain-containing protein (ECP) is composed of two independently folded domains which belong to the basic leucine-zipper and W2 domain-containing protein (BZW) family. Based on the sequence similarity between the C-terminal W2 domain of ECP and some eukaryotic translation initiation factors (such as eIF2B∊, eIF4γ, eIF5 etc.), ECP has been speculated to participate in the translation initiation process. Structural information on the C-terminal W2 domain of ECP would be helpful in understanding the specific cellular function of this protein. Here, the W2 domain of ECP was expressed and crystallized. Crystals grown by the hanging-drop vapour-diffusion method diffracted to 2.70 Å resolution and belonged to space group I4, with unit-cell parameters a = b = 81.05, c = 57.44 Å. The Matthews coefficient suggested that there was one molecule per asymmetric unit in the crystal

  18. Modulation of translation-initiation in CHO-K1 cells by rapamycin-induced heterodimerization of engineered eIF4G fusion proteins.

    Science.gov (United States)

    Schlatter, Stefan; Senn, Claudia; Fussenegger, Martin

    2003-07-20

    Translation-initiation is a predominant checkpoint in mammalian cells which controls protein synthesis and fine-tunes the flow of information from gene to protein. In eukaryotes, translation-initiation is typically initiated at a 7-methyl-guanylic acid cap posttranscriptionally linked to the 5' end of mRNAs. Alternative cap-independent translation-initiation involves 5' untranslated regions (UTR) known as internal ribosome entry sites, which adopt a particular secondary structure. Translation-initiating ribosome assembly at cap or IRES elements is mediated by a multiprotein complex of which the initiation factor 4F (eIF4F) consisting of eIF4A (helicase), eIF4E (cap-binding protein), and eIF4G is a major constituent. eIF4G is a key target of picornaviral protease 2A, which cleaves this initiation factor into eIF4G(Delta) and (Delta)eIF4G to redirect the cellular translation machinery exclusively to its own IRES-containing transcripts. We have designed a novel translation control system (TCS) for conditional as well as adjustable translation of cap- and IRES-dependent transgene mRNAs in mammalian cells. eIF4G(Delta) and (Delta)eIF4G were fused C- and N-terminally to the FK506-binding protein (FKBP) and the FKBP-rapamycin-binding domain (FRB) of the human FKBP-rapamycin-associated protein (FRAP), respectively. Rapamycin-induced heterodimerization of eIF4G(Delta)-FKBP and FRB-(Delta)eIF4G fusion proteins reconstituted a functional chimeric elongation factor 4G in a dose-dependent manner. Rigorous quantitative expression analysis of cap- and IRES-dependent SEAP- (human placental secreted alkaline phosphatase) and luc- (Photinus pyralis luciferase) encoding reporter constructs confirmed adjustable translation control and revealed increased production of desired proteins in response to dimerization-induced heterologous eIF4G in Chinese hamster ovary (CHO-K1) cells. Copyright 2003 Wiley Periodicals, Inc. Biotechnol Bioeng 83: 210-225, 2003.

  19. The effects of increased dietary protein yogurt snack in the afternoon on appetite control and eating initiation in healthy women

    OpenAIRE

    Ortinau, Laura C; Culp, Julie M; Hoertel, Heather A; Douglas, Steve M; Leidy, Heather J

    2013-01-01

    Background A large portion of daily intake comes from snacking. One of the increasingly common, healthier snacks includes Greek-style yogurt, which is typically higher in protein than regular yogurt. This study evaluated whether a 160?kcal higher-protein (HP) Greek-style yogurt snack improves appetite control, satiety, and delays subsequent eating compared to an isocaloric normal protein (NP) regular yogurt in healthy women. This study also identified the factors that predict the onset of eat...

  20. Variability of the Cyclin-Dependent Kinase 2 Flexibility Without Significant Change in the Initial Conformation of the Protein or Its Environment; a Computational Study.

    Science.gov (United States)

    Taghizadeh, Mohammad; Goliaei, Bahram; Madadkar-Sobhani, Armin

    2016-06-01

    Protein flexibility, which has been referred as a dynamic behavior has various roles in proteins' functions. Furthermore, for some developed tools in bioinformatics, such as protein-protein docking software, considering the protein flexibility, causes a higher degree of accuracy. Through undertaking the present work, we have accomplished the quantification plus analysis of the variations in the human Cyclin Dependent Kinase 2 (hCDK2) protein flexibility without affecting a significant change in its initial environment or the protein per se. The main goal of the present research was to calculate variations in the flexibility for each residue of the hCDK2, analysis of their flexibility variations through clustering, and to investigate the functional aspects of the residues with high flexibility variations. Using Gromacs package (version 4.5.4), three independent molecular dynamics (MD) simulations of the hCDK2 protein (PDB ID: 1HCL) was accomplished with no significant changes in their initial environments, structures, or conformations, followed by Root Mean Square Fluctuations (RMSF) calculation of these MD trajectories. The amount of variations in these three curves of RMSF was calculated using two formulas. More than 50% of the variation in the flexibility (the distance between the maximum and the minimum amount of the RMSF) was found at the region of Val-154. As well, there are other major flexibility fluctuations in other residues. These residues were mostly positioned in the vicinity of the functional residues. The subsequent works were done, as followed by clustering all hCDK2 residues into four groups considering the amount of their variability with respect to flexibility and their position in the RMSF curves. This work has introduced a new class of flexibility aspect of the proteins' residues. It could also help designing and engineering proteins, with introducing a new dynamic aspect of hCDK2, and accordingly, for the other similar globular proteins. In

  1. SdrF, a Staphylococcus epidermidis surface protein, contributes to the initiation of ventricular assist device driveline-related infections.

    Directory of Open Access Journals (Sweden)

    Carlos Arrecubieta

    2009-05-01

    Full Text Available Staphylococcus epidermidis remains the predominant pathogen in prosthetic-device infections. Ventricular assist devices, a recently developed form of therapy for end-stage congestive heart failure, have had considerable success. However, infections, most often caused by Staphylococcus epidermidis, have limited their long-term use. The transcutaneous driveline entry site acts as a potential portal of entry for bacteria, allowing development of either localized or systemic infections. A novel in vitro binding assay using explanted drivelines obtained from patients undergoing transplantation and a heterologous lactococcal system of surface protein expression were used to identify S. epidermidis surface components involved in the pathogenesis of driveline infections. Of the four components tested, SdrF, SdrG, PIA, and GehD, SdrF was identified as the primary ligand. SdrF adherence was mediated via its B domain attaching to host collagen deposited on the surface of the driveline. Antibodies directed against SdrF reduced adherence of S. epidermidis to the drivelines. SdrF was also found to adhere with high affinity to Dacron, the hydrophobic polymeric outer surface of drivelines. Solid phase binding assays showed that SdrF was also able to adhere to other hydrophobic artificial materials such as polystyrene. A murine model of infection was developed and used to test the role of SdrF during in vivo driveline infection. SdrF alone was able to mediate bacterial adherence to implanted drivelines. Anti-SdrF antibodies reduced S. epidermidis colonization of implanted drivelines. SdrF appears to play a key role in the initiation of ventricular assist device driveline infections caused by S. epidermidis. This pluripotential adherence capacity provides a potential pathway to infection with SdrF-positive commensal staphylococci first adhering to the external Dacron-coated driveline at the transcutaneous entry site, then spreading along the collagen

  2. A Role of Protein Degradation in Memory Consolidation after Initial Learning and Extinction Learning in the Honeybee ("Apis mellifera")

    Science.gov (United States)

    Felsenberg, Johannes; Dombrowski, Vincent; Eisenhardt, Dorothea

    2012-01-01

    Protein degradation is known to affect memory formation after extinction learning. We demonstrate here that an inhibitor of protein degradation, MG132, interferes with memory formation after extinction learning in a classical appetitive conditioning paradigm. In addition, we find an enhancement of memory formation when the same inhibitor is…

  3. Identification, characterization and preliminary X-ray diffraction analysis of the rolling-circle replication initiator protein from plasmid pSTK1

    International Nuclear Information System (INIS)

    Carr, Stephen B.; Mecia, Lauren B.; Phillips, Simon E. V.; Thomas, Christopher D.

    2013-01-01

    A proteolytically stable fragment of a plasmid replication initiation protein from the thermophile G. stearothermophilus has been biochemically characterized, crystallized and diffraction data collected to a resolution of 2.5 Å. Antibiotic resistance in bacterial pathogens poses an ever-increasing risk to human health. In antibiotic-resistant strains of Staphylococcus aureus this resistance often resides in extra-chromosomal plasmids, such as those of the pT181 family, which replicate via a rolling-circle mechanism mediated by a plasmid-encoded replication initiation protein. Currently, there is no structural information available for the pT181-family Rep proteins. Here, the crystallization of a catalytically active fragment of a homologous replication initiation protein from the thermophile Geobacillus stearothermophilus responsible for the replication of plasmid pSTK1 is reported. Crystals of the RepSTK1 fragment diffracted to a resolution of 2.5 Å and belonged to space group P2 1 2 1 2 1

  4. Differential tumor biology effects of double-initiation in a mouse skin chemical carcinogenesis model comparing wild type versus protein kinase Cepsilon overexpression mice.

    Science.gov (United States)

    Li, Yafan; Wheeler, Deric L; Ananthaswamy, Honnavara N; Verma, Ajit K; Oberley, Terry D

    2007-12-01

    Our previous studies showed that protein kinase Cepsilon (PKCepsilon) verexpression in mouse skin resulted in metastatic squamous cell carcinoma (SCC) elicited by single 7,12-dimethylbenz(a)anthracene (DMBA)-initiation and 12-O-tetradecanoylphorbol-13-acetate (TPA)-promotion in the absence of preceding papilloma formation as is typically observed in wild type mice. The present study demonstrates that double-DMBA initiation modulates tumor incidence, multiplicity, and latency period in both wild type and PKCepsilon overexpression transgenic (PKCepsilon-Tg) mice. After 17 weeks (wks) of tumor promotion, a reduction in papilloma multiplicity was observed in double- versus single-DMBA initiated wild type mice. Papilloma multiplicity was inversely correlated with cell death indices of interfollicular keratinocytes, indicating decreased papilloma formation was caused by increased cell death and suggesting the origin of papillomas is in interfollicular epidermis. Double-initiated PKCepsilon-Tg mice had accelerated carcinoma formation and cancer incidence in comparison to single-initiated PKCepsilon-Tg mice. Morphologic analysis of mouse skin following double initiation and tumor promotion showed a similar if not identical series of events to those previously observed following single initiation and tumor promotion: putative preneoplastic cells were observed arising from hyperplastic hair follicles (HFs) with subsequent cancer cell infiltration into the dermis. Single-initiated PKCepsilon-Tg mice exhibited increased mitosis in epidermal cells of HFs during tumor promotion.

  5. A transdisciplinary approach to the initial validation of a single cell protein as an alternative protein source for use in aquafeeds

    Directory of Open Access Journals (Sweden)

    Michael Tlusty

    2017-04-01

    Full Text Available The human population is growing and, globally, we must meet the challenge of increased protein needs required to feed this population. Single cell proteins (SCP, when coupled to aquaculture production, offer a means to ensure future protein needs can be met without direct competition with food for people. To demonstrate a given type of SCP has potential as a protein source for use in aquaculture feed, a number of steps need to be validated including demonstrating that the SCP is accepted by the species in question, leads to equivalent survival and growth, does not result in illness or other maladies, is palatable to the consumer, is cost effective to produce and can easily be incorporated into diets using existing technology. Here we examine white shrimp (Litopenaeus vannamei growth and consumer taste preference, smallmouth grunt (Haemulon chrysargyreum growth, survival, health and gut microbiota, and Atlantic salmon (Salmo salar digestibility when fed diets that substitute the bacterium Methylobacterium extorquens at a level of 30% (grunts, 100% (shrimp, or 55% (salmon of the fishmeal in a compound feed. In each of these tests, animals performed equivalently when fed diets containing M. extorquens as when fed a standard aquaculture diet. This transdisciplinary approach is a first validation of this bacterium as a potential SCP protein substitute in aquafeeds. Given the ease to produce this SCP through an aerobic fermentation process, the broad applicability for use in aquaculture indicates the promise of M. extorquens in leading toward greater food security in the future.

  6. AtLa1 protein initiates IRES-dependent translation of WUSCHEL mRNA and regulates the stem cell homeostasis of Arabidopsis in response to environmental hazards.

    Science.gov (United States)

    Cui, Yuchao; Rao, Shaofei; Chang, Beibei; Wang, Xiaoshuang; Zhang, Kaidian; Hou, Xueliang; Zhu, Xueyi; Wu, Haijun; Tian, Zhaoxia; Zhao, Zhong; Yang, Chengwei; Huang, Tao

    2015-10-01

    Plant stem cells are hypersensitive to environmental hazards throughout their life cycle, but the mechanism by which plants safeguard stem cell homeostasis in response to environmental hazards is largely unknown. The homeodomain transcription factor WUSCHEL (WUS) protein maintains the stem cell pool in the shoot apical meristem of Arabidopsis. Here, we demonstrate that the translation of WUS mRNA is directed by an internal ribosomal entry site (IRES) located in the 5'-untranslated region. The AtLa1 protein, an RNA-binding factor, binds to the 5'-untranslated region and initiates the IRES-dependent translation of WUS mRNA. Knockdown of AtLa1 expression represses the WUS IRES-dependent translation and leads to the arrest of growth and development. The AtLa1 protein is mainly located in the nucleoplasm. However, environmental hazards promote the nuclear-to-cytoplasmic translocation of the AtLa1 protein, which further enhances the IRES-dependent translation of WUS mRNA. Genetic evidence indicates that the WUS protein increases the tolerance of the shoot apical meristem to environmental hazards. Based on these results, we conclude that the stem cell niche in Arabidopsis copes with environmental hazards by enhancing the IRES-dependent translation of WUS mRNA under the control of the AtLa1 protein. © 2015 John Wiley & Sons Ltd.

  7. Generation of a predicted protein database from EST data and application to iTRAQ analyses in grape (Vitis vinifera cv. Cabernet Sauvignon) berries at ripening initiation

    Science.gov (United States)

    Lücker, Joost; Laszczak, Mario; Smith, Derek; Lund, Steven T

    2009-01-01

    Background iTRAQ is a proteomics technique that uses isobaric tags for relative and absolute quantitation of tryptic peptides. In proteomics experiments, the detection and high confidence annotation of proteins and the significance of corresponding expression differences can depend on the quality and the species specificity of the tryptic peptide map database used for analysis of the data. For species for which finished genome sequence data are not available, identification of proteins relies on similarity to proteins from other species using comprehensive peptide map databases such as the MSDB. Results We were interested in characterizing ripening initiation ('veraison') in grape berries at the protein level in order to better define the molecular control of this important process for grape growers and wine makers. We developed a bioinformatic pipeline for processing EST data in order to produce a predicted tryptic peptide database specifically targeted to the wine grape cultivar, Vitis vinifera cv. Cabernet Sauvignon, and lacking truncated N- and C-terminal fragments. By searching iTRAQ MS/MS data generated from berry exocarp and mesocarp samples at ripening initiation, we determined that implementation of the custom database afforded a large improvement in high confidence peptide annotation in comparison to the MSDB. We used iTRAQ MS/MS in conjunction with custom peptide db searches to quantitatively characterize several important pathway components for berry ripening previously described at the transcriptional level and confirmed expression patterns for these at the protein level. Conclusion We determined that a predicted peptide database for MS/MS applications can be derived from EST data using advanced clustering and trimming approaches and successfully implemented for quantitative proteome profiling. Quantitative shotgun proteome profiling holds great promise for characterizing biological processes such as fruit ripening initiation and may be further

  8. Generation of a predicted protein database from EST data and application to iTRAQ analyses in grape (Vitis vinifera cv. Cabernet Sauvignon berries at ripening initiation

    Directory of Open Access Journals (Sweden)

    Smith Derek

    2009-01-01

    Full Text Available Abstract Background iTRAQ is a proteomics technique that uses isobaric tags for relative and absolute quantitation of tryptic peptides. In proteomics experiments, the detection and high confidence annotation of proteins and the significance of corresponding expression differences can depend on the quality and the species specificity of the tryptic peptide map database used for analysis of the data. For species for which finished genome sequence data are not available, identification of proteins relies on similarity to proteins from other species using comprehensive peptide map databases such as the MSDB. Results We were interested in characterizing ripening initiation ('veraison' in grape berries at the protein level in order to better define the molecular control of this important process for grape growers and wine makers. We developed a bioinformatic pipeline for processing EST data in order to produce a predicted tryptic peptide database specifically targeted to the wine grape cultivar, Vitis vinifera cv. Cabernet Sauvignon, and lacking truncated N- and C-terminal fragments. By searching iTRAQ MS/MS data generated from berry exocarp and mesocarp samples at ripening initiation, we determined that implementation of the custom database afforded a large improvement in high confidence peptide annotation in comparison to the MSDB. We used iTRAQ MS/MS in conjunction with custom peptide db searches to quantitatively characterize several important pathway components for berry ripening previously described at the transcriptional level and confirmed expression patterns for these at the protein level. Conclusion We determined that a predicted peptide database for MS/MS applications can be derived from EST data using advanced clustering and trimming approaches and successfully implemented for quantitative proteome profiling. Quantitative shotgun proteome profiling holds great promise for characterizing biological processes such as fruit ripening

  9. Identification and Initial Characterization of the Effectors of an Anther Smut Fungus and Potential Host Target Proteins

    Directory of Open Access Journals (Sweden)

    Venkata S. Kuppireddy

    2017-11-01

    Full Text Available (1 Background: Plant pathogenic fungi often display high levels of host specificity and biotrophic fungi; in particular, they must manipulate their hosts to avoid detection and to complete their obligate pathogenic lifecycles. One important strategy of such fungi is the secretion of small proteins that serve as effectors in this process. Microbotryum violaceum is a species complex whose members infect members of the Caryophyllaceae; M. lychnidis-dioicae, a parasite on Silene latifolia, is one of the best studied interactions. We are interested in identifying and characterizing effectors of the fungus and possible corresponding host targets; (2 Methods: In silico analysis of the M. lychnidis-dioicae genome and transcriptomes allowed us to predict a pool of small secreted proteins (SSPs with the hallmarks of effectors, including a lack of conserved protein family (PFAM domains and also localized regions of disorder. Putative SSPs were tested for secretion using a yeast secretion trap method. We then used yeast two-hybrid analyses for candidate-secreted effectors to probe a cDNA library from a range of growth conditions of the fungus, including infected plants; (3 Results: Roughly 50 SSPs were identified by in silico analysis. Of these, 4 were studied further and shown to be secreted, as well as examined for potential host interactors. One of the putative effectors, MVLG_01732, was found to interact with Arabidopsis thaliana calcium-dependent lipid binding protein (AtCLB and with cellulose synthase interactive protein 1 orthologues; and (4 Conclusions: The identification of a pool of putative effectors provides a resource for functional characterization of fungal proteins that mediate the delicate interaction between pathogen and host. The candidate targets of effectors, e.g., AtCLB, involved in pollen germination suggest tantalizing insights that could drive future studies.

  10. Norovirus translation requires an interaction between the C Terminus of the genome-linked viral protein VPg and eukaryotic translation initiation factor 4G.

    Science.gov (United States)

    Chung, Liliane; Bailey, Dalan; Leen, Eoin N; Emmott, Edward P; Chaudhry, Yasmin; Roberts, Lisa O; Curry, Stephen; Locker, Nicolas; Goodfellow, Ian G

    2014-08-01

    Viruses have evolved a variety of mechanisms to usurp the host cell translation machinery to enable translation of the viral genome in the presence of high levels of cellular mRNAs. Noroviruses, a major cause of gastroenteritis in man, have evolved a mechanism that relies on the interaction of translation initiation factors with the virus-encoded VPg protein covalently linked to the 5' end of the viral RNA. To further characterize this novel mechanism of translation initiation, we have used proteomics to identify the components of the norovirus translation initiation factor complex. This approach revealed that VPg binds directly to the eIF4F complex, with a high affinity interaction occurring between VPg and eIF4G. Mutational analyses indicated that the C-terminal region of VPg is important for the VPg-eIF4G interaction; viruses with mutations that alter or disrupt this interaction are debilitated or non-viable. Our results shed new light on the unusual mechanisms of protein-directed translation initiation. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  11. N-MYC DOWN-REGULATED-LIKE Proteins Regulate Meristem Initiation by Modulating Auxin Transport and MAX2 Expression

    OpenAIRE

    Mudgil, Yashwanti; Ghawana, Sanjay; Jones, Alan M.

    2013-01-01

    Background N-MYC DOWN-REGULATED-LIKE (NDL) proteins interact with the G? subunit (AGB1) of the heterotrimeric G protein complex and play an important role in AGB1-dependent regulation of lateral root formation by affecting root auxin transport, auxin gradients and the steady-state levels of mRNA encoding the PIN-FORMED 2 and AUXIN 1 auxin transport facilitators. Auxin transport in aerial tissue follows different paths and utilizes different transporters than in roots; therefore, in the presen...

  12. Phosphorylation and interactions associated with the control of the Leishmania Poly-A Binding Protein 1 (PABP1) function during translation initiation.

    Science.gov (United States)

    de Melo Neto, Osvaldo P; da Costa Lima, Tamara D C; Merlo, Kleison C; Romão, Tatiany P; Rocha, Pollyanna O; Assis, Ludmila A; Nascimento, Larissa M; Xavier, Camila C; Rezende, Antonio M; Reis, Christian R S; Papadopoulou, Barbara

    2018-03-23

    The Poly-A Binding Protein (PABP) is a conserved eukaryotic polypeptide involved in many aspects of mRNA metabolism. During translation initiation, PABP interacts with the translation initiation complex eIF4F and enhances the translation of polyadenylated mRNAs. Schematically, most PABPs can be divided into an N-terminal RNA-binding region, a non-conserved linker segment and the C-terminal MLLE domain. In pathogenic Leishmania protozoans, three PABP homologues have been identified, with the first one (PABP1) targeted by phosphorylation and shown to co-immunoprecipitate with an eIF4F-like complex (EIF4E4/EIF4G3) implicated in translation initiation. Here, PABP1 phosphorylation was shown to be linked to logarithmic cell growth, reminiscent of EIF4E4 phosphorylation, and coincides with polysomal association. Phosphorylation targets multiple serine-proline (SP) or threonine-proline (TP) residues within the PABP1 linker region. This is an essential protein, but phosphorylation is not needed for its association with polysomes or cell viability. Mutations which do impair PABP1 polysomal association and are required for viability do not prevent phosphorylation, although further mutations lead to a presumed inactive protein largely lacking phosphorylated isoforms. Co-immunoprecipitation experiments were carried out to investigate PABP1 function further, identifying several novel protein partners and the EIF4E4/EIF4G3 complex, but no other eIF4F-like complex or subunit. A novel, direct interaction between PABP1 and EIF4E4 was also investigated and found to be mediated by the PABP1 MLLE binding to PABP Interacting Motifs (PAM2) within the EIF4E4 N-terminus. The results shown here are consistent with phosphorylation of PABP1 being part of a novel pathway controlling its function and possibly translation in Leishmania.

  13. Phosphorylation state of a Tob/BTG protein, FOG-3, regulates initiation and maintenance of the Caenorhabditis elegans sperm fate program.

    Science.gov (United States)

    Lee, Myon-Hee; Kim, Kyung Won; Morgan, Clinton T; Morgan, Dyan E; Kimble, Judith

    2011-05-31

    FOG-3, the single Caenorhabditis elegans Tob/BTG protein, directs germ cells to adopt the sperm fate at the expense of oogenesis. Importantly, FOG-3 activity must be maintained for the continued production of sperm that is typical of the male sex. Vertebrate Tob proteins have antiproliferative activity and ERK phosphorylation of Tob proteins has been proposed to abrogate "antiproliferative" activity. Here we investigate FOG-3 phosphorylation and its effect on sperm fate specification. We found both phosphorylated and unphosphorylated forms of FOG-3 in nematodes. We then interrogated the role of FOG-3 phosphorylation in sperm fate specification. Specifically, we assayed FOG-3 transgenes for rescue of a fog-3 null mutant. Wild-type FOG-3 rescued both initiation and maintenance of sperm fate specification. A FOG-3 mutant with its four consensus ERK phosphorylation sites substituted to alanines, called FOG-3(4A), rescued partially: sperm were made transiently but not continuously in both sexes. A different FOG-3 mutant with its sites substituted to glutamates, called FOG-3(4E), had no rescuing activity on its own, but together with FOG-3(4A) rescue was complete. Thus, when FOG-3(4A) and FOG-3(4E) were both introduced into the same animals, sperm fate specification was not only initiated but also maintained, resulting in continuous spermatogenesis in males. Our findings suggest that unphosphorylated FOG-3 initiates the sperm fate program and that phosphorylated FOG-3 maintains that program for continued sperm production typical of males. We discuss implications of our results for Tob/BTG proteins in vertebrates.

  14. Cooperative Interactions between 480 kDa Ankyrin-G and EB proteins assemble the axon initial segment

    NARCIS (Netherlands)

    Fréal, Amélie|info:eu-repo/dai/nl/412516721; Fassier, Coralie; Le Bras, Barbara; Bullier, Erika; de Gois, Stéphanie; Hazan, Jamilé; Hoogenraad, Casper C.|info:eu-repo/dai/nl/227263502; Couraud, François

    2016-01-01

    The axon initial segment (AIS) is required for generating action potentials and maintaining neuronal polarity. Significant progress has been made in deciphering the basic building blocks composing the AIS, but the underlying mechanisms required for AIS formation remains unclear. The scaffolding

  15. Binding proteins for the regulatory subunit (RII-B) of brain cAMP-dependent protein kinase II: isolation and initial characterization of cDNA clones

    International Nuclear Information System (INIS)

    Bregman, D.B.; Hu, E.; Rubin, C.S.

    1987-01-01

    In mammalian brain several proteins bind RII-B with high affinity. An example is P75, which co-purifies with RII-B and also complexes Ca 2+ -calmodulin. Thus, RII-B binding proteins (RBPs) might play a role in integrating the Ca 2+ and cAMP signalling pathways in the CNS. In order to study the structure and function of these polypeptides they have isolated cloned cDNAs for RBPs by screening brain λgt11 expression libraries using a functional assay: the binding of 32 P-labeled RII to fusion proteins produced by recombinants expressing RII binding domains. Inserts from rat brain recombinant clones λ7B and λ10B both hybridize to a brain mRNA of 7000 nucleotides. Northern gel analyses indicate that the putative RBP mRNA is also expressed in lung, but not in several other tissues. The λ7B insert was subcloned into the expression plasmid pINIA. A 50 kDa high affinity RII-B binding polypeptide accumulated in E. coli transformed with pINIA-7B. Two RBP cDNAs (λ77, λ100A) have been retrieved from a bovine λgt 11 library using a monoclonal antibody directed against P75 and the binding assay respectively. On Southern blots the insert from λ100A hybridizes to the cDNA insert from clones λ77, suggesting that λ 77 cDNA might contain sequences coding for both an RII binding domain and a P75 epitope. The bovine λ100A insert also hybridizes with the rat λ7B clone indicating that an RII binding domain is conserved in the two species

  16. Initiation of lambda DNA replication. The Escherichia coli small heat shock proteins, DnaJ and GrpE, increase DnaK's affinity for the lambda P protein.

    Science.gov (United States)

    Osipiuk, J; Georgopoulos, C; Zylicz, M

    1993-03-05

    It is known that the initiation of bacteriophage lambda replication requires the orderly assembly of the lambda O.lambda P.DnaB helicase protein preprimosomal complex at the ori lambda DNA site. The DnaK, DnaJ, and GrpE heat shock proteins act together to destabilize the lambda P.DnaB complex, thus freeing DnaB and allowing it to unwind lambda DNA near the ori lambda site. The first step of this disassembly reaction is the binding of DnaK to the lambda P protein. In this report, we examined the influence of the DnaJ and GrpE proteins on the stability of the lambda P.DnaK complex. We present evidence for the existence of the following protein-protein complexes: lambda P.DnaK, lambda P.DnaJ, DnaJ.DnaK, DnaK.GrpE, and lambda P.DnaK.GrpE. Our results suggest that the presence of GrpE alone destabilizes the lambda P.DnaK complex, whereas the presence of DnaJ alone stabilizes the lambda P.DnaK complex. Using immunoprecipitation, we show that in the presence of GrpE, DnaK exhibits a higher affinity for the lambda P.DnaJ complex than it does alone. Using cross-linking with glutaraldehyde, we show that oligomeric forms of DnaK exhibit a higher affinity for lambda P than monomeric DnaK. However, in the presence of GrpE, monomeric DnaK can efficiently bind lambda P protein. These findings help explain our previous results, namely that in the GrpE-dependent lambda DNA replication system, the DnaK protein requirement can be reduced up to 10-fold.

  17. Control of protein synthesis in Escherichia coli: strain differences in control of translational initiation after energy source shift-down.

    OpenAIRE

    Jacobson, L A; Jen-Jacobson, L

    1980-01-01

    We have studied the parameters of protein synthesis in a number of Escherichia coli strains after a shift-down from glucose-minimal to succinate-minimal medium. One group of strains, including K-12(lambda) (ATCC 10798) and NF162, showed a postshift translational yield of 50 to 65% and a 2- to 2.5-fold increase in the functional lifetime of general messenger ribonucleic acid. There was no change in the lag time for beta-galactosidase induction in these strains after the shift-down. A second gr...

  18. Signature of protein adaptation to warm deep sea environments: the case of Initiation Factor 6 studied by molecular simulation and neutron scattering

    International Nuclear Information System (INIS)

    Calligari, Paolo

    2008-01-01

    The protein Initiation Factor 6 (IF6) takes part in the protein synthesis regulation of several organisms. It was also found in archeaebacteria such as Methanococcus jannaschii which lives in deep-seas near hydrothermal vents where temperature reaches 80 C and pressure is between 250 bar and 500 bar. The aim of this work was to study for the first time dynamical and structural properties of IF6 produced by M. jannaschii and comparing them with those of the IF6 homologue present in Saccharomyces cerevisiae which lives at 'normal' environmental conditions (27 C and 1 bar). Molecular simulation gave here new insights into the adaptation of these two proteins to their respective physiological conditions and showed that the latter induced similar dynamical and structural properties: in their respective 'natural' conditions, IF6s show very similar structural fluctuations and the characteristic relaxation times which define their dynamical properties shows similar changes when comparing unfavorable conditions to physiological ones. The creation of these corresponding states between the two homologues has been interpreted by the fractional Brownian dynamics model and by a novel method for the characterization of protein secondary structures. The latter is presented here in detail together with some examples of other applications. Experimental data obtained from quasi-elastic neutron scattering seemed to support the results obtained by molecular simulations. (author) [fr

  19. Resolving the paradox for protein aggregation diseases: a common mechanism for aggregated proteins to initially attack membranes without needing aggregates [v1; ref status: indexed, http://f1000r.es/221

    Directory of Open Access Journals (Sweden)

    Haina Qin

    2013-10-01

    Full Text Available Paradoxically, aggregation of specific proteins is characteristic of many human diseases and aging, yet aggregates have been found to be unnecessary for initiating pathogenesis. Here we determined the NMR topology and dynamics of a helical mutant in a membrane environment transformed from the 125-residue cytosolic all-β MSP by the ALS-causing P56S mutation. Unexpectedly, despite its low hydrophobicity, the P56S major sperm protein (MSP domain becomes largely embedded in the membrane environment with high backbone rigidity. Furthermore it is composed of five helices with amphiphilicity comparable to those of the partly-soluble membrane toxin mellitin and α-synuclein causing Parkinson's disease. Consequently, the mechanism underlying this chameleon transformation becomes clear: by disrupting the specific tertiary interaction network stabilizing the native all-β MSP fold to release previously-locked amphiphilic segments, the P56S mutation acts to convert the classic MSP fold into a membrane-active protein that is fundamentally indistinguishable from mellitin and α-synuclein which are disordered in aqueous solution but spontaneously partition into membrane interfaces driven by hydrogen-bond energetics gained from forming α-helix in the membrane environments. As segments with high amphiphilicity exist in all proteins, our study successfully resolves the paradox by deciphering that the proteins with a higher tendency to aggregate have a stronger potential to partition into membranes through the same mechanism as α-synuclein to initially attack membranes to trigger pathogenesis without needing aggregates. This might represent the common first step for various kinds of aggregated proteins to trigger familiar, sporadic and aging diseases. Therefore the homeostasis of aggregated proteins in vivo is the central factor responsible for a variety of human diseases including aging. The number and degree of the membrane attacks by aggregated proteins may

  20. Biological significance of facilitated diffusion in protein-DNA interactions. Applications to T4 endonuclease V-initiated DNA repair

    International Nuclear Information System (INIS)

    Dowd, D.R.; Lloyd, R.S.

    1990-01-01

    Facilitated diffusion along nontarget DNA is employed by numerous DNA-interactive proteins to locate specific targets. Until now, the biological significance of DNA scanning has remained elusive. T4 endonuclease V is a DNA repair enzyme which scans nontarget DNA and processively incises DNA at the site of pyrimidine dimers which are produced by exposure to ultraviolet (UV) light. In this study we tested the hypothesis that there exists a direct correlation between the degree of processivity of wild type and mutant endonuclease V molecules and the degree of enhanced UV resistance which is conferred to repair-deficient Eshcerichia coli. This was accomplished by first creating a series of endonuclease V mutants whose in vitro catalytic activities were shown to be very similar to that of the wild type enzyme. However, when the mechanisms by which these enzymes search nontarget DNA for its substrate were analyzed in vitro and in vivo, the mutants displayed varying degrees of nontarget DNA scanning ranging from being nearly as processive as wild type to randomly incising dimers within the DNA population. The ability of these altered endonuclease V molecules to enhance UV survival in DNA repair-deficient E. coli then was assessed. The degree of enhanced UV survival was directly correlated with the level of facilitated diffusion. This is the first conclusive evidence directly relating a reduction of in vivo facilitated diffusion with a change in an observed phenotype. These results support the assertion that the mechanisms which DNA-interactive proteins employ in locating their target sites are of biological significance

  1. Confined laminar flow on a super-hydrophobic surface drives the initial stages of tau protein aggregation

    KAUST Repository

    Moretti, Manola

    2018-02-01

    Super-hydrophobic micro-patterned surfaces are ideal substrates for the controlled self-assembly and substrate-free characterization of biological molecules. In this device, the tailored surface supports a micro-volume drop containing the molecules of interest. While the quasi-spherical drop is evaporating under controlled conditions, its de-wetting direction is guided by the pillared microstructure on top of the device, leading to the formation of threads between the neighboring pillars. This effect has been exploited here to elucidate the mechanism triggering the formation of amyloid fibers and oligomers in tau related neurodegenerative diseases. By using Raman spectroscopy, we demonstrate that the fiber bridging the pillars contains β-sheets, a characteristic feature of amyloid aggregation. We propose that the combination of laminar flow, shear stress and molecular crowding taking place while the drop is evaporating on the SHMS, induces the reorganization of the tau protein secondary structure and we suggest that this effect could in fact closely mimic the actual mechanism occurring in the human brain environment. Such a straightforward technique opens up new possibilities in the field of self-assembly of biomolecules and their characterization by different methods (SEM, AFM, Raman spectroscopy, TEM), in a single device.

  2. Confined laminar flow on a super-hydrophobic surface drives the initial stages of tau protein aggregation

    KAUST Repository

    Moretti, Manola; Allione, Marco; Marini, Monica; Giugni, Andrea; Torre, Bruno; Das, Gobind; Di Fabrizio, Enzo M.

    2018-01-01

    Super-hydrophobic micro-patterned surfaces are ideal substrates for the controlled self-assembly and substrate-free characterization of biological molecules. In this device, the tailored surface supports a micro-volume drop containing the molecules of interest. While the quasi-spherical drop is evaporating under controlled conditions, its de-wetting direction is guided by the pillared microstructure on top of the device, leading to the formation of threads between the neighboring pillars. This effect has been exploited here to elucidate the mechanism triggering the formation of amyloid fibers and oligomers in tau related neurodegenerative diseases. By using Raman spectroscopy, we demonstrate that the fiber bridging the pillars contains β-sheets, a characteristic feature of amyloid aggregation. We propose that the combination of laminar flow, shear stress and molecular crowding taking place while the drop is evaporating on the SHMS, induces the reorganization of the tau protein secondary structure and we suggest that this effect could in fact closely mimic the actual mechanism occurring in the human brain environment. Such a straightforward technique opens up new possibilities in the field of self-assembly of biomolecules and their characterization by different methods (SEM, AFM, Raman spectroscopy, TEM), in a single device.

  3. The structural proteins of epidemic and historical strains of Zika virus differ in their ability to initiate viral infection in human host cells.

    Science.gov (United States)

    Bos, Sandra; Viranaicken, Wildriss; Turpin, Jonathan; El-Kalamouni, Chaker; Roche, Marjolaine; Krejbich-Trotot, Pascale; Desprès, Philippe; Gadea, Gilles

    2018-03-01

    Mosquito-borne Zika virus (ZIKV) recently emerged in South Pacific islands and Americas where large epidemics were documented. In the present study, we investigated the contribution of the structural proteins C, prM and E in the permissiveness of human host cells to epidemic strains of ZIKV. To this end, we evaluated the capacity of the epidemic strain BeH819015 to infect epithelial A549 and neuronal SH-SY5Y cells in comparison to the African historical MR766 strain. For that purpose, we generated a molecular clone of BeH819015 and a chimeric clone of MR766 which contains the BeH819015 structural protein region. We showed that ZIKV containing BeH819015 structural proteins was much less efficient in cell-attachment leading to a reduced susceptibility of A549 and SH-SY5Y cells to viral infection. Our data illustrate a previously underrated role for C, prM, and E in ZIKV epidemic strain ability to initiate viral infection in human host cells. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

  4. The eclipse period of Escherichia coli

    DEFF Research Database (Denmark)

    von Freiesleben, Ulrik; Krekling, Martin A.; Hansen, Flemming G.

    2000-01-01

    corresponds to the period of origin hemimethylation. The SeqA protein was absolutely required for the eclipse, and DnaA titration studies suggested that the SeqA protein prevented the binding of multiple DnaA molecules on oriC (initial complex formation). No correlation between the amount of SeqA and eclipse...... length was revealed, but increased SeqA levels affected chromosome partitioning and/or cell division. This was corroborated further by an aberrant nucleoid distribution in SeqA-deficient cells. We suggest that the SeqA protein's role in maintaining the eclipse is tied to a function in chromosome...

  5. Roles for Dam methylation in bacterial chromosome replication

    DEFF Research Database (Denmark)

    Charbon, Godefroid; Koch, Birgit; Skovgaard, Ole

    GATC sequences in the DNA of Escherichia coli and related species are methylated at the adenine residue by DNA adenine methyltransferase (DamMT). These methylated residues and/or the level of DamMT influence initiation of chromosome replication from the replication origin, oriC, which contain...... for about one third of the cell cycle. During sequestration at least three mechanisms operate to lower the activity of the initiator protein, DnaA. First, the dnaA promoter, which also contains an excess of GATC sequences, is sequestered for the same period of time as oriC to prevent de novo DnaA synthesis....... Second, new DnaA binding sites outside oriC are generated by replication which serve to titrate free DNA protein. Third, after initiation, DnaA-ATP is converted to inactive DnaA-ADP by a process called RIDA (regulatory inactivation of DnaA), which is dependent on the beta-clamp of DNA polymerase III...

  6. Reduction in C-reactive protein and LDL cholesterol and cardiovascular event rates after initiation of rosuvastatin: a prospective study of the JUPITER trial

    DEFF Research Database (Denmark)

    Ridker, Paul M; Danielson, Eleanor; Fonseca, Francisco Ah

    2009-01-01

    BACKGROUND: Statins lower high-sensitivity C-reactive protein (hsCRP) and cholesterol concentrations, and hypothesis generating analyses suggest that clinical outcomes improve in patients given statins who achieve hsCRP concentrations less than 2 mg/L in addition to LDL cholesterol less than 1.......8 mmol/L (LDL cholesterol and hsCRP after the start of statin therapy is controversial. We prospectively tested this hypothesis. METHODS: In an analysis of 15 548 initially healthy men and women participating in the JUPITER trial (87% of full cohort), we...... to on-treatment concentrations of LDL cholesterol (>/=1.8 mmol/L or /=2 mg/L or

  7. Overexpression of eIF5 or its protein mimic 5MP perturbs eIF2 function and induces ATF4 translation through delayed re-initiation.

    Science.gov (United States)

    Kozel, Caitlin; Thompson, Brytteny; Hustak, Samantha; Moore, Chelsea; Nakashima, Akio; Singh, Chingakham Ranjit; Reid, Megan; Cox, Christian; Papadopoulos, Evangelos; Luna, Rafael E; Anderson, Abbey; Tagami, Hideaki; Hiraishi, Hiroyuki; Slone, Emily Archer; Yoshino, Ken-Ichi; Asano, Masayo; Gillaspie, Sarah; Nietfeld, Jerome; Perchellet, Jean-Pierre; Rothenburg, Stefan; Masai, Hisao; Wagner, Gerhard; Beeser, Alexander; Kikkawa, Ushio; Fleming, Sherry D; Asano, Katsura

    2016-10-14

    ATF4 is a pro-oncogenic transcription factor whose translation is activated by eIF2 phosphorylation through delayed re-initiation involving two uORFs in the mRNA leader. However, in yeast, the effect of eIF2 phosphorylation can be mimicked by eIF5 overexpression, which turns eIF5 into translational inhibitor, thereby promoting translation of GCN4, the yeast ATF4 equivalent. Furthermore, regulatory protein termed eIF5-mimic protein (5MP) can bind eIF2 and inhibit general translation. Here, we show that 5MP1 overexpression in human cells leads to strong formation of 5MP1:eIF2 complex, nearly comparable to that of eIF5:eIF2 complex produced by eIF5 overexpression. Overexpression of eIF5, 5MP1 and 5MP2, the second human paralog, promotes ATF4 expression in certain types of human cells including fibrosarcoma. 5MP overexpression also induces ATF4 expression in Drosophila The knockdown of 5MP1 in fibrosarcoma attenuates ATF4 expression and its tumor formation on nude mice. Since 5MP2 is overproduced in salivary mucoepidermoid carcinoma, we propose that overexpression of eIF5 and 5MP induces translation of ATF4 and potentially other genes with uORFs in their mRNA leaders through delayed re-initiation, thereby enhancing the survival of normal and cancer cells under stress conditions. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  8. C-terminal region of the UV-B photoreceptor UVR8 initiates signaling through interaction with the COP1 protein

    Science.gov (United States)

    Cloix, Catherine; Kaiserli, Eirini; Heilmann, Monika; Baxter, Katherine J.; Brown, Bobby A.; O’Hara, Andrew; Smith, Brian O.; Christie, John M.; Jenkins, Gareth I.

    2012-01-01

    UV-B light initiates photomorphogenic responses in plants. Arabidopsis UV RESISTANCE LOCUS8 (UVR8) specifically mediates these responses by functioning as a UV-B photoreceptor. UV-B exposure converts UVR8 from a dimer to a monomer, stimulates the rapid accumulation of UVR8 in the nucleus, where it binds to chromatin, and induces interaction of UVR8 with CONSTITUTIVELY PHOTOMORPHOGENIC1 (COP1), which functions with UVR8 to control photomorphogenic UV-B responses. Although the crystal structure of UVR8 reveals the basis of photoreception, it does not show how UVR8 initiates signaling through interaction with COP1. Here we report that a region of 27 amino acids from the C terminus of UVR8 (C27) mediates the interaction with COP1. The C27 region is necessary for UVR8 function in the regulation of gene expression and hypocotyl growth suppression in Arabidopsis. However, UVR8 lacking C27 still undergoes UV-B–induced monomerization in both yeast and plant protein extracts, accumulates in the nucleus in response to UV-B, and interacts with chromatin at the UVR8-regulated ELONGATED HYPOCOTYL5 (HY5) gene. The UV-B–dependent interaction of UVR8 and COP1 is reproduced in yeast cells and we show that C27 is both necessary and sufficient for the interaction of UVR8 with the WD40 domain of COP1. Furthermore, we show that C27 interacts in yeast with the REPRESSOR OF UV-B PHOTOMORPHOGENESIS proteins, RUP1 and RUP2, which are negative regulators of UVR8 function. Hence the C27 region has a key role in UVR8 function. PMID:22988111

  9. The eukaryotic translation initiation factor 3 subunit L protein interacts with Flavivirus NS5 and may modulate yellow fever virus replication.

    Science.gov (United States)

    Morais, Ana Ts; Terzian, Ana Cb; Duarte, Danilo Vb; Bronzoni, Roberta Vm; Madrid, Maria Cfs; Gavioli, Arieli F; Gil, Laura Hvg; Oliveira, Amanda G; Zanelli, Cleslei F; Valentini, Sandro R; Rahal, Paula; Nogueira, Mauricio L

    2013-06-22

    Yellow fever virus (YFV) belongs to the Flavivirus genus and causes an important disease. An alarming resurgence of viral circulation and the expansion of YFV-endemic zones have been detected in Africa and South America in recent years. NS5 is a viral protein that contains methyltransferase and RNA-dependent RNA polymerase (RdRp) domains, which are essential for viral replication, and the interactions between NS5 and cellular proteins have been studied to better understand viral replication. The aim of this study was to characterize the interaction of the NS5 protein with eukaryotic translation initiation factor 3 subunit L (eIF3L) and to evaluate the role of eIF3L in yellow fever replication. To identify interactions of YFV NS5 with cellular proteins, we performed a two-hybrid screen using the YFV NS5 RdRp domain as bait with a human cDNA library, and RNApol deletion mutants were generated and analyzed using the two-hybrid system for mapping the interactions. The RNApol region involved was segmented into three fragments and analyzed using an eIF3L-expressing yeast strain. To map the NS5 residues that are critical for the interactions, we performed site-direct mutagenesis in segment 3 of the interaction domain (ID) and confirmed the interaction using in vitro assays and in vivo coimmunoprecipitation. The significance of eIF3L for YFV replication was investigated using eIF3L overexpression and RNA interference. In this work, we describe and characterize the interaction of NS5 with the translation factor eIF3L. The interaction between NS5 and eIF3L was confirmed using in vitro binding and in vivo coimmunoprecipitation assays. This interaction occurs at a region (the interaction domain of the RNApol domain) that is conserved in several flaviviruses and that is, therefore, likely to be relevant to the genus. eIF3L overexpression and plaque reduction assays showed a slight effect on YFV replication, indicating that the interaction of eIF3L with YFV NS5 may play a role

  10. mRNA expression of the DNA replication-initiation proteins in epithelial dysplasia and squamous cell carcinoma of the tongue

    International Nuclear Information System (INIS)

    Li, Jian-na; Feng, Chong-jin; Lu, Yong-jun; Li, Hui-jun; Tu, Zheng; Liao, Gui-qing; Liang, Chun

    2008-01-01

    The tongue squamous cell carcinomas (SCCs) are characterized by high mitotic activity, and early detection is desirable. Overexpression of the DNA replication-initiation proteins has been associated with dysplasia and malignancy. Our aim was to determine whether these proteins are useful biomarkers for assessing the development of tongue SCC. We analyzed the mRNA expression of CDC6, CDT1, MCM2 and CDC45 in formalin-fixed, paraffin-embedded benign and malignant tongue tissues using quantitative real-time PCR followed by statistical analysis. We found that the expression levels are significantly higher in malignant SCC than mild precancerous epithelial dysplasia, and the expression levels in general increase with increasing grade of precancerous lesions from mild, moderate to severe epithelial dysplasia. CDC6 and CDC45 expression is dependent of the dysplasia grade and lymph node status. CDT1 expression is higher in severe dysplasia than in mild and moderate dysplasia. MCM2 expression is dependent of the dysplasia grade, lymph node status and clinical stage. The expression of the four genes is independent of tumor size or histological grade. A simple linear regression analysis revealed a linear increase in the mRNA levels of the four genes from the mild to severe dysplasia and SCC. A strong association was established between CDC6 and CDT1, and between MCM2 and CDC45 expression. The nonparametric receiver operating characteristic analysis suggested that MCM2 and CDC45 had a higher accuracy than CDC6 and CDT1 for distinguishing dysplasia from tongue SCC. These proteins can be used as biomarkers to distinguish precancerous dysplasia from SCC and are useful for early detection and diagnosis of SCC as an adjunct to clinicopathological parameters

  11. Avr4 promotes Cf-4 receptor-like protein association with the BAK1/SERK3 receptor-like kinase to initiate receptor endocytosis and plant immunity.

    Science.gov (United States)

    Postma, Jelle; Liebrand, Thomas W H; Bi, Guozhi; Evrard, Alexandre; Bye, Ruby R; Mbengue, Malick; Kuhn, Hannah; Joosten, Matthieu H A J; Robatzek, Silke

    2016-04-01

    The first layer of plant immunity is activated by cell surface receptor-like kinases (RLKs) and proteins (RLPs) that detect infectious pathogens. Constitutive interaction with the SUPPRESSOR OF BIR1 (SOBIR1) RLK contributes to RLP stability and kinase activity. As RLK activation requires transphosphorylation with a second associated RLK, it remains elusive how RLPs initiate downstream signaling. We employed live-cell imaging, gene silencing and coimmunoprecipitation to investigate the requirement of associated kinases for functioning and ligand-induced subcellular trafficking of Cf RLPs that mediate immunity of tomato against Cladosporium fulvum. Our research shows that after elicitation with matching effector ligands Avr4 and Avr9, BRI1-ASSOCIATED KINASE 1/SOMATIC EMBRYOGENESIS RECEPTOR KINASE 3 (BAK1/SERK3) associates with Cf-4 and Cf-9. BAK1/SERK3 is required for the effector-triggered hypersensitive response and resistance of tomato against C. fulvum. Furthermore, Cf-4 interacts with SOBIR1 at the plasma membrane and is recruited to late endosomes upon Avr4 trigger, also depending on BAK1/SERK3. These observations indicate that RLP-mediated resistance and endocytosis require ligand-induced recruitment of BAK1/SERK3, reminiscent of BAK1/SERK3 interaction and subcellular fate of the FLAGELLIN SENSING 2 (FLS2) RLK. This reveals that diverse classes of cell surface immune receptors share common requirements for initiation of resistance and endocytosis. © 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.

  12. Sequence Classification: 512897 [

    Lifescience Database Archive (English)

    Full Text Available Non-TMB Non-TMH Non-TMB Non-TMB Non-TMB Non-TMB >gi|17548159|ref|NP_521561.1| PROBABLE CHROMOSOMAL REPLICATI...ON INITIATOR PROTEIN DNAA || http://www.ncbi.nlm.nih.gov/protein/17548159 ...

  13. Hda Monomerization by ADP Binding Promotes Replicase Clamp-mediated DnaA-ATP Hydrolysis*S⃞

    OpenAIRE

    Su'etsugu, Masayuki; Nakamura, Kenta; Keyamura, Kenji; Kudo, Yuka; Katayama, Tsutomu

    2008-01-01

    ATP-DnaA is the initiator of chromosomal replication in Escherichia coli, and the activity of DnaA is regulated by the regulatory inactivation of the DnaA (RIDA) system. In this system, the Hda protein promotes DnaA-ATP hydrolysis to produce inactive ADP-DnaA in a mechanism that is mediated by the DNA-loaded form of the replicase sliding clamp. In this study, we first revealed that hda translation uses an unusual initiation codon, CUG, located downstream of the annotat...

  14. Initiation but no execution - modulation of peripheral blood lymphocyte apoptosis in rheumatoid arthritis - a potential role for heat shock protein 70

    Directory of Open Access Journals (Sweden)

    Chuturgoon Anil A

    2011-11-01

    Full Text Available Abstract Background Rheumatoid arthritis (RA is a chronic autoimmune disease, which causes synovial damage. Persistence of lymphocyte infiltrates in the rheumatoid synovium has been attributed to abnormal apoptosis. While not comprehensively investigated, perturbations in peripheral blood lymphocyte (PBL apoptosis may also be involved in perpetuation of autoimmune processes in RA. Methods We investigated total, CD4+ and CD19+ PBL apoptosis in our study cohort by monitoring the translocation of phosphatidylserine using the Annexin-V assay. To examine the role of death receptor mediated apoptosis as well as activation-induced-cell-death (AICD, PBLs were labeled with CD95/Fas and CD69 markers and enumerated by flow cytometry. Proteolytic activity of initiator and executioner caspases was determined by luminometry. DNA fragmentation assays were used to examine whether apoptotic signals were transduced to the nucleus. Quantitative PCR arrays were used to investigate apoptotic pathways associated with RA-PBLs. Since heat-shock-protein-70 (HSP70 is an inducible protein which modulates apoptotic signals, we determined HSP70 levels by intra-cellular flow cytometry and western blots. Results The RA-PBLs showed signs of elevated apoptosis whilst in circulation. These include increases in the loss of plasma membrane asymmetry, indicated by increased externalization of phosphatidylserine (especially in B-lymphocytes. RA-PBLs showed a bias to CD95/Fas mediated apoptotic pathways, but low levels of the CD69 marker suggested that this was not associated with immune activation. Although downstream markers of apoptosis such as caspase-3/7 activity, were increased, no DNA fragmentation was observed in RA-PBLs. Interestingly, elevated levels of apoptosis did not correlate with absolute lymphocyte counts in RA patients. Levels of HSP70 were highly elevated in RA-PBLs compared to controls. Conclusion The results suggest that while apoptosis may be initiated in RA

  15. C-Reactive Protein (CRP), Interferon Gamma-Inducible Protein 10 (IP-10), and Lipopolysaccharide (LPS) Are Associated with Risk of Tuberculosis after Initiation of Antiretroviral Therapy in Resource-Limited Settings

    Science.gov (United States)

    Tenforde, Mark W.; Gupte, Nikhil; Dowdy, David W.; Asmuth, David M.; Balagopal, Ashwin; Pollard, Richard B.; Sugandhavesa, Patcharaphan; Lama, Javier R.; Pillay, Sandy; Cardoso, Sandra W.; Pawar, Jyoti; Santos, Breno; Riviere, Cynthia; Mwelase, Noluthando; Kanyama, Cecilia; Kumwenda, Johnstone; Hakim, James G.; Kumarasamy, Nagalingeswaran; Bollinger, Robert; Semba, Richard D.; Campbell, Thomas B.; Gupta, Amita

    2015-01-01

    Objective The association between pre-antiretroviral (ART) inflammation and immune activation and risk for incident tuberculosis (TB) after ART initiation among adults is uncertain. Design Nested case-control study (n = 332) within ACTG PEARLS trial of three ART regimens among 1571 HIV-infected, treatment-naïve adults in 9 countries. We compared cases (participants with incident TB diagnosed by 96 weeks) to a random sample of controls (participants who did not develop TB, stratified by country and treatment arm). Methods We measured pre-ART C-reactive protein (CRP), EndoCab IgM, ferritin, interferon gamma (IFN-γ), interleukin 6 (IL-6), interferon gamma-inducible protein 10 (IP-10), lipopolysaccharide (LPS), soluble CD14 (sCD14), tumor necrosis factor alpha (TNF-α), and CD4/DR+/38+ and CD8/DR+/38+ T cells. Markers were defined according to established cutoff definitions when available, 75th percentile of measured values when not, and detectable versus undetectable for LPS. Using logistic regression, we measured associations between biomarkers and incident TB, adjusting for age, sex, study site, treatment arm, baseline CD4 and log10 viral load. We assessed the discriminatory value of biomarkers using receiver operating characteristic (ROC) analysis. Results Seventy-seven persons (4.9%) developed incident TB during follow-up. Elevated baseline CRP (aOR 3.25, 95% CI: 1.55–6.81) and IP-10 (aOR 1.89, 95% CI: 1.05–3.39), detectable plasma LPS (aOR 2.39, 95% CI: 1.13–5.06), and the established TB risk factors anemia and hypoalbuminemia were independently associated with incident TB. In ROC analysis, CRP, albumin, and LPS improved discrimination only modestly for TB risk when added to baseline routine patient characteristics including CD4 count, body mass index, and prior TB. Conclusion Incident TB occurs commonly after ART initiation. Although associated with higher post-ART TB risk, baseline CRP, IP-10, and LPS add limited value to routine patient characteristics

  16. C-reactive protein in patients with advanced metastatic renal cell carcinoma: Usefulness in identifying patients most likely to benefit from initial nephrectomy

    International Nuclear Information System (INIS)

    Ito, Hiroki; Kishida, Takeshi; Miura, Takeshi; Kubota, Yoshinobu; Yao, Masahiro; Shioi, Koichi; Murakami, Takayuki; Takizawa, Akitoshi; Sano, Futoshi; Kawahara, Takashi; Mizuno, Nobuhiko; Makiyama, Kazuhide; Nakaigawa, Noboru

    2012-01-01

    C-reactive protein (CRP) is considered a useful serum marker for patients with RCC. However, its clinical utility in advanced metastatic renal cell carcinoma (AM-RCC), particularly in deciding whether to perform nephrectomy at the onset, is not well studied. We retrospectively evaluated 181 patients with AM-RCC, including 18 patients underwent potentially curative surgery, 111 underwent cytoreductive nephrectomy, and 52 received medical treatment only. CRP cutoff points were determined by receiver operating characteristic (ROC) curve analysis. Kaplan-Meier and Cox regression analyses were used for survival tests. ROC analysis suggested that grouping patients according to 3 CRP ranges was a rational model. Patients with highly elevated CRP (≥67.0 mg/L) presented remarkably poor prognosis despite treatment (nephrectomy or medical treatment only). Cox regression models demonstrated that risk factors of overall survival for patients who underwent nephrectomy were the CRP ranges defined in this study (≤18.0 mg/L, >18.0 and <67.0 mg/L, and ≥67.0 mg/L), ECOG PS (0, 1, and ≥2), and number of metastatic organ sites (0–1 and ≥2). The retrospective design is a limitation of this study. Our study demonstrated that the serum CRP level is a statistically significant prognostic parameter for patients with AM-RCC. The data also indicated that pretreatment serum CRP level provides useful prognostic information that helps in deciding whether to perform initial nephrectomy for patients with AM-RCC

  17. DNA double strand break repair in mammalian cells: role of MRE11 and BLM proteins at the initiation of Non Homologous End Joining (NHEJ)

    International Nuclear Information System (INIS)

    Grabarz, Anastazja

    2011-01-01

    DNA double strand breaks (DSBs) are highly cytotoxic lesions, which can lead to genetic rearrangements. Two pathways are responsible for repairing these lesions: homologous recombination (HR) and non homologous end joining (NHEJ). In our laboratory, an intrachromosomal substrate has been established in order to measure the efficiency and the fidelity of NHEJ in living cells (Guirouilh-Barbat 2004). This approach led us to identify a KU-independent alternative pathway, which uses micro homologies in the proximity of the junction to accomplish repair - the alternative NHEJ (Guirouilh-Barbat 2004, Guirouilh-Barbat et Rass 2007). The goal of my thesis consisted in identifying and characterising major actors of this pathway. In the absence of KU, alternative NHEJ would be initiated by ssDNA resection of damaged ends. We showed that the nuclease activity of MRE11 is necessary for this mechanism. MRE11 overexpression leads to a two fold stimulation of NHEJ efficiency, while the extinction of MRE11 by siRNA results in a two fold decrease. Our results demonstrate that the proteins RAD50 and CtIP act in the same pathway as MRE11. Moreover, in cells deficient for XRCC4, MIRIN - an inhibitor of the MRN complex - leads to a decrease in repair efficiency, implicating MRE11 in alternative NHEJ. We also showed that MRE11 can act in an ATM-dependent and independent manner (Rass et Grabarz Nat Struct Mol Biol 2009). The initiation of break resection needs to be pursued by a more extensive degradation of DNA, which is accomplished in yeast by the proteins Exo1 and Sgs1/Dna2. In human cells, in vitro studies have recently proposed a similar model of a two-step break resection. We chose to elucidate the role of one of the human homologs of Sgs1 - the RecQ helicase BLM - in the resection process. Our experiments show, that he absence of BLM decreases the efficiency of end joining by NHEJ, accompanied by an increase in error-prone events, especially long-range deletions (≥200 nt). This

  18. The Jasmonate-ZIM-domain proteins interact with the WD-Repeat/bHLH/MYB complexes to regulate Jasmonate-mediated anthocyanin accumulation and trichome initiation in Arabidopsis thaliana.

    Science.gov (United States)

    Qi, Tiancong; Song, Susheng; Ren, Qingcuo; Wu, Dewei; Huang, Huang; Chen, Yan; Fan, Meng; Peng, Wen; Ren, Chunmei; Xie, Daoxin

    2011-05-01

    Jasmonates (JAs) mediate plant responses to insect attack, wounding, pathogen infection, stress, and UV damage and regulate plant fertility, anthocyanin accumulation, trichome formation, and many other plant developmental processes. Arabidopsis thaliana Jasmonate ZIM-domain (JAZ) proteins, substrates of the CORONATINE INSENSITIVE1 (COI1)-based SCF(COI1) complex, negatively regulate these plant responses. Little is known about the molecular mechanism for JA regulation of anthocyanin accumulation and trichome initiation. In this study, we revealed that JAZ proteins interact with bHLH (Transparent Testa8, Glabra3 [GL3], and Enhancer of Glabra3 [EGL3]) and R2R3 MYB transcription factors (MYB75 and Glabra1), essential components of WD-repeat/bHLH/MYB transcriptional complexes, to repress JA-regulated anthocyanin accumulation and trichome initiation. Genetic and physiological evidence showed that JA regulates WD-repeat/bHLH/MYB complex-mediated anthocyanin accumulation and trichome initiation in a COI1-dependent manner. Overexpression of the MYB transcription factor MYB75 and bHLH factors (GL3 and EGL3) restored anthocyanin accumulation and trichome initiation in the coi1 mutant, respectively. We speculate that the JA-induced degradation of JAZ proteins abolishes the interactions of JAZ proteins with bHLH and MYB factors, allowing the transcriptional function of WD-repeat/bHLH/MYB complexes, which subsequently activate respective downstream signal cascades to modulate anthocyanin accumulation and trichome initiation.

  19. The Jasmonate-ZIM-Domain Proteins Interact with the WD-Repeat/bHLH/MYB Complexes to Regulate Jasmonate-Mediated Anthocyanin Accumulation and Trichome Initiation in Arabidopsis thaliana[C][W

    Science.gov (United States)

    Qi, Tiancong; Song, Susheng; Ren, Qingcuo; Wu, Dewei; Huang, Huang; Chen, Yan; Fan, Meng; Peng, Wen; Ren, Chunmei; Xie, Daoxin

    2011-01-01

    Jasmonates (JAs) mediate plant responses to insect attack, wounding, pathogen infection, stress, and UV damage and regulate plant fertility, anthocyanin accumulation, trichome formation, and many other plant developmental processes. Arabidopsis thaliana Jasmonate ZIM-domain (JAZ) proteins, substrates of the CORONATINE INSENSITIVE1 (COI1)–based SCFCOI1 complex, negatively regulate these plant responses. Little is known about the molecular mechanism for JA regulation of anthocyanin accumulation and trichome initiation. In this study, we revealed that JAZ proteins interact with bHLH (Transparent Testa8, Glabra3 [GL3], and Enhancer of Glabra3 [EGL3]) and R2R3 MYB transcription factors (MYB75 and Glabra1), essential components of WD-repeat/bHLH/MYB transcriptional complexes, to repress JA-regulated anthocyanin accumulation and trichome initiation. Genetic and physiological evidence showed that JA regulates WD-repeat/bHLH/MYB complex-mediated anthocyanin accumulation and trichome initiation in a COI1-dependent manner. Overexpression of the MYB transcription factor MYB75 and bHLH factors (GL3 and EGL3) restored anthocyanin accumulation and trichome initiation in the coi1 mutant, respectively. We speculate that the JA-induced degradation of JAZ proteins abolishes the interactions of JAZ proteins with bHLH and MYB factors, allowing the transcriptional function of WD-repeat/bHLH/MYB complexes, which subsequently activate respective downstream signal cascades to modulate anthocyanin accumulation and trichome initiation. PMID:21551388

  20. Mutations within Four Distinct Gag Proteins Are Required To Restore Replication of Human Immunodeficiency Virus Type 1 after Deletion Mutagenesis within the Dimerization Initiation Site

    Science.gov (United States)

    Liang, Chen; Rong, Liwei; Quan, Yudong; Laughrea, Michael; Kleiman, Lawrence; Wainberg, Mark A.

    1999-01-01

    Human immunodeficiency virus type 1 (HIV-1) genomic RNA segments at nucleotide (nt) positions +240 to +274 are thought to form a stem-loop secondary structure, termed SL1, that serves as a dimerization initiation site for viral genomic RNA. We have generated two distinct deletion mutations within this region, termed BH10-LD3 and BH10-LD4, involving nt positions +238 to +253 and +261 to +274, respectively, and have shown that each of these resulted in significant diminutions in levels of viral infectiousness. However, long-term culture of each of these viruses in MT-2 cells resulted in a restoration of infectiousness, due to a series of compensatory point mutations within four distinct proteins that are normally cleaved from the Gag precursor. In the case of BH10-LD3, these four mutations were MA1, CA1, MP2, and MNC, and they involved changes of amino acid Val-35 to Ile within the matrix protein (MA), Ile-91 to Thr within the capsid (CA), Thr-12 to Ile within p2, and Thr-24 to Ile within the nucleocapsid (NC). The order in which these mutations were acquired by the mutated BH10-LD3 was MNC > CA1 > MP2 > MA1. The results of site-directed mutagenesis studies confirmed that each of these four substitutions contributed to the increased viability of the mutated BH10-LD3 viruses and that the MNC substitution, which was acquired first, played the most important role in this regard. Three point mutations, MP2, MNC, and MA2, were also shown to be sequentially acquired by viruses that had emerged in culture from the BH10-LD4 deletion. The first two of these were identical to those described above, while the last involved a change of Val-35 to Leu. All three of these substitutions were necessary to restore the infectiousness of mutated BH10-LD4 viruses to wild-type levels, although the MP2 mutation alone, but neither of the other two substitutions, was able to confer some viability on BH10-LD4 viruses. Studies of viral RNA packaging showed that the BH10-LD4 deletion only

  1. The RAS Initiative

    Science.gov (United States)

    NCI established the RAS Initiative to explore innovative approaches for attacking the proteins encoded by mutant forms of RAS genes and to ultimately create effective, new therapies for RAS-related cancers.

  2. Dermal matrix proteins initiate re-epithelialization but are not sufficient for coordinated epidermal outgrowth in a new fish skin culture model.

    Science.gov (United States)

    Matsumoto, Reiko; Sugimoto, Masazumi

    2007-02-01

    We have established a new culture system to study re-epithelialization during fish epidermal wound healing. In this culture system, fetal bovine serum (FBS) stimulates the epidermal outgrowth of multi-cellular layers from scale skin mounted on a coverslip, even when cell proliferation is blocked. The rate of outgrowth is about 0.4 mm/h, and at 3 h after incubation, the area occupied by the epidermal sheet is nine times larger than the area of the original scale skin. Cells at the bottom of the outgrowth show a migratory phenotype with lamellipodia, and "purse string"-like actin bundles have been found over the leading-edge cells with polarized lamellipodia. In the superficial cells, re-development of adherens junctions and microridges has been detected, together with the appearance and translocation of phosphorylated p38 MAPK into nuclear areas. Thus, this culture system provides an excellent model to study the mechanisms of epidermal outgrowth accompanied by migration and re-differentiation. We have also examined the role of extracellular matrix proteins in the outgrowth. Type I collagen or fibronectin stimulates moderate outgrowth in the absence of FBS, but development of microridges and the distribution of phosphorylated p38 MAPK are attenuated in the superficial cells. In addition, the leading-edge cells do not have apparent "purse string"-like actin bundles. The outgrowth stimulated by FBS is inhibited by laminin. These results suggest that dermal substrates such as type I collagen and fibronectin are able to initiate epidermal outgrowth but require other factors to enhance such outgrowth, together with coordinated alterations in cellular phenotype.

  3. A Novel Initiation Mechanism of Death in Streptococcus pneumoniae Induced by the Human Milk Protein-Lipid Complex HAMLET and Activated during Physiological Death*

    Science.gov (United States)

    Clementi, Emily A.; Marks, Laura R.; Duffey, Michael E.; Hakansson, Anders P.

    2012-01-01

    To cause colonization or infection, most bacteria grow in biofilms where differentiation and death of subpopulations is critical for optimal survival of the whole population. However, little is known about initiation of bacterial death under physiological conditions. Membrane depolarization has been suggested, but never shown to be involved, due to the difficulty of performing such studies in bacteria and the paucity of information that exists regarding ion transport mechanisms in prokaryotes. In this study, we performed the first extensive investigation of ion transport and membrane depolarization in a bacterial system. We found that HAMLET, a human milk protein-lipid complex, kills Streptococcus pneumoniae (the pneumococcus) in a manner that shares features with activation of physiological death from starvation. Addition of HAMLET to pneumococci dissipated membrane polarity, but depolarization per se was not enough to trigger death. Rather, both HAMLET- and starvation-induced death of pneumococci specifically required a sodium-dependent calcium influx, as shown using calcium and sodium transport inhibitors. This mechanism was verified under low sodium conditions, and in the presence of ionomycin or monensin, which enhanced pneumococcal sensitivity to HAMLET- and starvation-induced death. Pneumococcal death was also inhibited by kinase inhibitors, and indicated the involvement of Ser/Thr kinases in these processes. The importance of this activation mechanism was made evident, as dysregulation and manipulation of physiological death was detrimental to biofilm formation, a hallmark of bacterial colonization. Overall, our findings provide novel information on the role of ion transport during bacterial death, with the potential to uncover future antimicrobial targets. PMID:22700972

  4. A novel initiation mechanism of death in Streptococcus pneumoniae induced by the human milk protein-lipid complex HAMLET and activated during physiological death.

    Science.gov (United States)

    Clementi, Emily A; Marks, Laura R; Duffey, Michael E; Hakansson, Anders P

    2012-08-03

    To cause colonization or infection, most bacteria grow in biofilms where differentiation and death of subpopulations is critical for optimal survival of the whole population. However, little is known about initiation of bacterial death under physiological conditions. Membrane depolarization has been suggested, but never shown to be involved, due to the difficulty of performing such studies in bacteria and the paucity of information that exists regarding ion transport mechanisms in prokaryotes. In this study, we performed the first extensive investigation of ion transport and membrane depolarization in a bacterial system. We found that HAMLET, a human milk protein-lipid complex, kills Streptococcus pneumoniae (the pneumococcus) in a manner that shares features with activation of physiological death from starvation. Addition of HAMLET to pneumococci dissipated membrane polarity, but depolarization per se was not enough to trigger death. Rather, both HAMLET- and starvation-induced death of pneumococci specifically required a sodium-dependent calcium influx, as shown using calcium and sodium transport inhibitors. This mechanism was verified under low sodium conditions, and in the presence of ionomycin or monensin, which enhanced pneumococcal sensitivity to HAMLET- and starvation-induced death. Pneumococcal death was also inhibited by kinase inhibitors, and indicated the involvement of Ser/Thr kinases in these processes. The importance of this activation mechanism was made evident, as dysregulation and manipulation of physiological death was detrimental to biofilm formation, a hallmark of bacterial colonization. Overall, our findings provide novel information on the role of ion transport during bacterial death, with the potential to uncover future antimicrobial targets.

  5. ORF Alignment: NC_003317 [GENIUS II[Archive

    Lifescience Database Archive (English)

    Full Text Available NC_003317 gi|17987645 >1j1vA 4 94 20 109 8e-10 ... gb|AAL52543.1| CHROMOSOMAL REPLICATION... INITIATOR PROTEIN DNAA [Brucella melitensis ... 16M] ref|NP_540279.1| CHROMOSOMAL REPLICATION IN

  6. Fulltext PDF

    Indian Academy of Sciences (India)

    mainly from the studies carried out in the bacterial model organ- ism Escherichia coli. ... locus. DnaA protein is essential for the process of replication initiation at oriC .... using the β-lactam antibiotic mecillinam, cells become spherical, growth ...

  7. Oxidative DNA damage is instrumental in hyperreplication stress-induced inviability of Escherichia coli

    DEFF Research Database (Denmark)

    Charbon, Godefroid; Bjørn, Louise; Mendoza-Chamizo, Belén

    2014-01-01

    In Escherichia coli, an increase in the ATP bound form of the DnaA initiator protein results in hyperinitiation and inviability. Here, we show that such replication stress is tolerated during anaerobic growth. In hyperinitiating cells, a shift from anaerobic to aerobic growth resulted in appearance...

  8. At same leucine intake, a whey/plant protein blend is not as effective as whey to initiate a transient post prandial muscle anabolic response during a catabolic state in mini pigs.

    Directory of Open Access Journals (Sweden)

    Aurélia Revel

    Full Text Available Muscle atrophy has been explained by an anabolic resistance following food intake and an increase of dietary protein intake is recommended. To be optimal, a dietary protein has to be effective not only to initiate but also to prolong a muscle anabolic response in a catabolic state. To our knowledge, whether or not a dairy or a dairy/plant protein blend fulfills these criterions is unknown in a muscle wasting situation.Our aim was, in a control and a catabolic state, to measure continuously muscle anabolism in term of intensity and duration in response to a meal containing casein (CAS, whey (WHEY or a whey/ plant protein blend (BLEND and to evaluate the best protein source to elicit the best post prandial anabolism according to the physio-pathological state.Adult male Yucatan mini pigs were infused with U-13C-Phenylalanine and fed either CAS, WHEY or BLEND. A catabolic state was induced by a glucocorticoid treatment for 8 days (DEX. Muscle protein synthesis, proteolysis and balance were measured with the hind limb arterio-venous differences technique. Repeated time variance analysis were used to assess significant differences.In a catabolic situation, whey proteins were able to initiate muscle anabolism which remained transient in contrast to the stimulated muscle protein accretion with WHEY, CAS or BLEND in healthy conditions. Despite the same leucine intake compared to WHEY, BLEND did not restore a positive protein balance in DEX animals.Even with WHEY, the duration of the anabolic response was not optimal and has to be improved in a catabolic state. The use of BLEND remained of lower efficiency even at same leucine intake than whey.

  9. Initial preclinical safety of non-replicating human endogenous retrovirus envelope protein-coated baculovirus vector-based vaccines against human papillomavirus.

    Science.gov (United States)

    Han, Su-Eun; Kim, Mi-Gyeong; Lee, Soondong; Cho, Hee-Jeong; Byun, Youngro; Kim, Sujeong; Kim, Young Bong; Choi, Yongseok; Oh, Yu-Kyoung

    2013-12-01

    Human endogenous retrovirus (HERV) envelope protein-coated, baculovirus vector-based HPV 16 L1 (AcHERV-HPV16L1) is a non-replicating recombinant baculoviral vaccine. Here, we report an initial evaluation of the preclinical safety of AcHERV-HPV16L1 vaccine. In an acute toxicity study, a single administration of AcHERV-HPV16L1 DNA vaccine given intramuscularly (i.m.) to mice at a dose of 1 × 10(8) plaque-forming units (PFU) did not cause significant changes in body weight compared with vehicle-treated controls. It did cause a brief increase in the weights of some organs on day 15 post-treatment, but by day 30, all organ weights were not significantly different from those in the vehicle-treated control group. No hematological changes were observed on day 30 post-treatment. In a range-finding toxicity study with three doses of 1 × 10(7) , 2 × 10(7) and 5 × 10(7) PFU once daily for 5 days, the group treated with 5 × 10(7) PFU showed a transient decrease in the body weights from day 5 to day 15 post-treatment, but recovery to the levels similar to those in the vehicle-treated control group by post-treatment day 20. Organ weights were slightly higher for lymph nodes, spleen, thymus and liver after repeated dosing with 5 × 10(7) PFU on day 15, but had normalized by day 30. Moreover, repeated administration of AcHERV-HPV16L1 did not induce myosin-specific autoantibody in serum, and did not cause immune complex deposition or tissue damage at injection sites. Taken together, these results provide preliminary evidence of the preclinical safety of AcHERV-based HPV16L1 DNA vaccines in mice. Copyright © 2012 John Wiley & Sons, Ltd.

  10. Acceleration of lipid peroxidation in alpha-tocopherol transfer protein-knockout mice following the consumption of drinking water containing a radical initiator.

    Science.gov (United States)

    Yoshida, Yasukazu; Hayakawa, Mieko; Cynshi, Osamu; Jishage, Kou-ichi; Niki, Etsuo

    2008-01-01

    To assess the antioxidative role of vitamin E (VE) in a mouse model of severe VE deficiency by using biomarkers, alpha-tocopherol transfer protein (alpha-TTP(-/-))-knockout mice were maintained on a VE-deficient diet for 28 weeks [KO group, n = 6]. Wild-type C57BL/6 mice were maintained on a diet containing 0.002% alpha-tocopherol [WT group, n = 6]. The animals were housed individually in a metabolic cage from the age of 9 weeks (Week 0) to 27 weeks. Urine was collected every week, and the levels of total hydroxyoctadecadienoic acid (tHODE), 7-hydroxycholesterol (t7-OHCh), and 8-iso-prostaglandin F(2alpha)(t8-isoPGF(2alpha)), which are biomarkers for lipid peroxidation, were measured by gas chromatography (GC)-mass spectrometry. From the age of 21 weeks (Week 12), three mice in each group were provided drinking water containing the water-soluble radical initiator 2,2'-azobis[2-(2-imidazolin-2-yl)propane] dihydrochloride (AIPH) until the end of the study (Week 19). Blood and tissue samples were collected, and the levels of the abovementioned biomarkers therein were assessed. AIPH consumption clearly elevated the plasma and erythrocyte levels of tHODE and t8-isoPGF(2alpha) in both the WT and KO groups except for the erythrocyte level of tHODE in the WT group. Furthermore, this elevation was more prominent in the KO group than in the WT group. Interestingly, AIPH consumption reduced the stereoisomer ratio of HODE (ZE/EE), which is reflective of the efficacy of a compound as an antioxidant in vivo; this suggests that free radical-mediated oxidation reduces the antioxidant capacity in vivo. The urine levels of tHODE, t7-OHCh, and t8-isoPGF(2alpha) tended to increase with AIPH consumption, but these individual levels fluctuated. It was clearly demonstrated by the proposed biomarkers that maintaining alpha-TTP(-/-) mice on a VE-deficient diet results in a severe VE deficiency and promotes lipid peroxidation.

  11. Sheep scrapie susceptibility-linked polymorphisms do not modulate the initial binding of cellular to disease-associated prion protein prior to conversion

    NARCIS (Netherlands)

    Rigter, A.; Bossers, A.

    2005-01-01

    Conversion of the host-encoded protease-sensitive cellular prion protein (PrPC) into the scrapie-associated protease-resistant isoform (PrPSc) of prion protein (PrP) is the central event in transmissible spongiform encephalopathies or prion diseases. Differences in transmissibility and

  12. Skeletal muscle protein synthesis and the abundance of the mRNA translation initiation repressor PDCD4 are inversely regulated by fasting and refeeding in rats.

    Science.gov (United States)

    Zargar, Sana; Moreira, Tracy S; Samimi-Seisan, Helena; Jeganathan, Senthure; Kakade, Dhanshri; Islam, Nushaba; Campbell, Jonathan; Adegoke, Olasunkanmi A J

    2011-06-01

    Optimal skeletal muscle mass is vital to human health, because defects in muscle protein metabolism underlie or exacerbate human diseases. The mammalian target of rapamycin complex 1 is critical in the regulation of mRNA translation and protein synthesis. These functions are mediated in part by the ribosomal protein S6 kinase 1 (S6K1) through mechanisms that are poorly understood. The tumor suppressor programmed cell death 4 (PDCD4) has been identified as a novel substrate of S6K1. Here, we examined 1) the expression of PDCD4 in skeletal muscle and 2) its regulation by feed deprivation (FD) and refeeding. Male rats (~100 g; n = 6) were subjected to FD for 48 h; some rats were refed for 2 h. FD suppressed muscle fractional rates of protein synthesis and Ser(67) phosphorylation of PDCD4 (-50%) but increased PDCD4 abundance (P muscle fractional rates of protein synthesis and reduced PDCD4 abundance relative to FD. Finally, when myoblasts were grown in amino acid- and serum-free medium, phenylalanine incorporation into proteins in cells depleted of PDCD4 more than doubled the values in cells with a normal level of PDCD4 (P skeletal muscle in parallel with the reduction of the abundance of this mRNA translation inhibitor.

  13. Tomato leaf curl Kerala virus (ToLCKeV AC3 protein forms a higher order oligomer and enhances ATPase activity of replication initiator protein (Rep/AC1

    Directory of Open Access Journals (Sweden)

    Mukherjee Sunil K

    2010-06-01

    Full Text Available Abstract Background Geminiviruses are emerging plant viruses that infect a wide variety of vegetable crops, ornamental plants and cereal crops. They undergo recombination during co-infections by different species of geminiviruses and give rise to more virulent species. Antiviral strategies targeting a broad range of viruses necessitate a detailed understanding of the basic biology of the viruses. ToLCKeV, a virus prevalent in the tomato crop of Kerala state of India and a member of genus Begomovirus has been used as a model system in this study. Results AC3 is a geminiviral protein conserved across all the begomoviral species and is postulated to enhance viral DNA replication. In this work we have successfully expressed and purified the AC3 fusion proteins from E. coli. We demonstrated the higher order oligomerization of AC3 using sucrose gradient ultra-centrifugation and gel-filtration experiments. In addition we also established that ToLCKeV AC3 protein interacted with cognate AC1 protein and enhanced the AC1-mediated ATPase activity in vitro. Conclusions Highly hydrophobic viral protein AC3 can be purified as a fusion protein with either MBP or GST. The purification method of AC3 protein improves scope for the biochemical characterization of the viral protein. The enhancement of AC1-mediated ATPase activity might lead to increased viral DNA replication.

  14. Association between lectin complement pathway initiators, C-reactive protein and left ventricular remodeling in myocardial infarction-a magnetic resonance study

    DEFF Research Database (Denmark)

    Schoos, Mikkel Malby; Munthe-Fog, Lea; Skjoedt, Mikkel-Ole

    2013-01-01

    Lectin complement pathway (LP) activation is an important mechanism in myocardial ischemia reperfusion injury (IRI). LP is activated via the recognition molecules mannose-binding lectin (MBL), ficolins-2 and-3 and is regulated by MBL/Ficolin-associated Protein-1 (MAP-1). Also, C-reactive protein...... (CRP) and ficolin-2 interact in vitro, but the role of the ficolins in IRI is unknown.Methods and results In 55 patients with ST segment elevation myocardial infarction, we investigated the association of LP components and CRP in plasma samples with left ventricular (LV) end systolic and diastolic......-activation in IRI and LV remodeling....

  15. Resonance assignment of disordered protein with repetitive and overlapping sequence using combinatorial approach reveals initial structural propensities and local restrictions in the denatured state

    Energy Technology Data Exchange (ETDEWEB)

    Malik, Nikita; Kumar, Ashutosh, E-mail: askutoshk@iitb.ac.in [Indian Institute of Technology Bombay, Department of Bioscience and Bioengineering (India)

    2016-09-15

    NMR resonance assignment of intrinsically disordered proteins poses a challenge because of the limited dispersion of amide proton chemical shifts. This becomes even more complex with the increase in the size of the system. Residue specific selective labeling/unlabeling experiments have been used to resolve the overlap, but require multiple sample preparations. Here, we demonstrate an assignment strategy requiring only a single sample of uniformly labeled {sup 13}C,{sup 15}N-protein. We have used a combinatorial approach, involving 3D-HNN, CC(CO)NH and 2D-MUSIC, which allowed us to assign a denatured centromeric protein Cse4 of 229 residues. Further, we show that even the less sensitive experiments, when used in an efficient manner can lead to the complete assignment of a complex system without the use of specialized probes in a relatively short time frame. The assignment of the amino acids discloses the presence of local structural propensities even in the denatured state accompanied by restricted motion in certain regions that provides insights into the early folding events of the protein.

  16. Resonance assignment of disordered protein with repetitive and overlapping sequence using combinatorial approach reveals initial structural propensities and local restrictions in the denatured state

    International Nuclear Information System (INIS)

    Malik, Nikita; Kumar, Ashutosh

    2016-01-01

    NMR resonance assignment of intrinsically disordered proteins poses a challenge because of the limited dispersion of amide proton chemical shifts. This becomes even more complex with the increase in the size of the system. Residue specific selective labeling/unlabeling experiments have been used to resolve the overlap, but require multiple sample preparations. Here, we demonstrate an assignment strategy requiring only a single sample of uniformly labeled "1"3C,"1"5N-protein. We have used a combinatorial approach, involving 3D-HNN, CC(CO)NH and 2D-MUSIC, which allowed us to assign a denatured centromeric protein Cse4 of 229 residues. Further, we show that even the less sensitive experiments, when used in an efficient manner can lead to the complete assignment of a complex system without the use of specialized probes in a relatively short time frame. The assignment of the amino acids discloses the presence of local structural propensities even in the denatured state accompanied by restricted motion in certain regions that provides insights into the early folding events of the protein.

  17. Quantitative proteomics identifies Gemin5, a scaffolding protein involved in ribonucleoprotein assembly, as a novel partner for eukaryotic initiation factor 4E

    DEFF Research Database (Denmark)

    Fierro-Monti, Ivo; Mohammed, Shabaz; Matthiesen, Rune

    2006-01-01

    Protein complexes are dynamic entities; identification and quantitation of their components is critical in elucidating functional roles under specific cellular conditions. We report the first quantitative proteomic analysis of the human cap-binding protein complex. Components and proteins......-starved tumorigenic human mesenchymal stromal cells, attested to their activated translational states. The WD-repeat, scaffolding-protein Gemin5 was identified as a novel eIF4E binding partner, which interacted directly with eIF4E through a motif (YXXXXLPhi) present in a number of eIF4E-interacting partners. Elevated...... levels of Gemin5:eIF4E complexes were found in phorbol ester treated HEK293 cells. Gemin5 and eIF4E co-localized to cytoplasmic P-bodies in human osteosarcoma U2OS cells. Interaction between eIF4E and Gemin5 and their co-localization to the P-bodies, may serve to recruit capped mRNAs to these RNP...

  18. Identification of residues within the African swine fever virus DP71L protein required for dephosphorylation of translation initiation factor eIF2α and inhibiting activation of pro-apoptotic CHOP

    Energy Technology Data Exchange (ETDEWEB)

    Barber, Claire; Netherton, Chris; Goatley, Lynnette [The Pirbright Institute, Ash Road, Pirbright, Woking, Surrey GU24 0NF (United Kingdom); Moon, Alice; Goodbourn, Steve [Institute for Infection and Immunity, St. George' s, University of London, London SW17 0RE (United Kingdom); Dixon, Linda, E-mail: linda.dixon@pirbright.ac.uk [The Pirbright Institute, Ash Road, Pirbright, Woking, Surrey GU24 0NF (United Kingdom)

    2017-04-15

    The African swine fever virus DP71L protein recruits protein phosphatase 1 (PP1) to dephosphorylate the translation initiation factor 2α (eIF2α) and avoid shut-off of global protein synthesis and downstream activation of the pro-apoptotic factor CHOP. Residues V16 and F18A were critical for binding of DP71L to PP1. Mutation of this PP1 binding motif or deletion of residues between 52 and 66 reduced the ability of DP71L to cause dephosphorylation of eIF2α and inhibit CHOP induction. The residues LSAVL, between 57 and 61, were also required. PP1 was co-precipitated with wild type DP71L and the mutant lacking residues 52- 66 or the LSAVL motif, but not with the PP1 binding motif mutant. The residues in the LSAVL motif play a critical role in DP71L function but do not interfere with binding to PP1. Instead we propose these residues are important for DP71L binding to eIF2α. - Highlights: •The African swine fever virus DP71L protein recruits protein phosphatase 1 (PP1) to dephosphorylate translation initiation factor eIF2α (eIF2α). •The residues V{sup 16}, F{sup 18} of DP71L are required for binding to the α, β and γ isoforms of PP1 and for DP71L function. •The sequence LSAVL downstream from the PP1 binding site (residues 57–61) are also important for DP71L function. •DP71L mutants of the LSAVL sequence retain ability to co-precipitate with PP1 showing these sequences have a different role to PP1 binding.

  19. Evanescent wave-initiated photopolymerisation as a new way to create monolithic open-tubular capillary columns: use as enzymatic microreactor for on-line protein digestion

    Czech Academy of Sciences Publication Activity Database

    Abele, S.; Smejkal, Petr; Yavorska, O.; Foret, František; Macka, M.

    2010-01-01

    Roč. 135, č. 3 (2010), s. 477-481 ISSN 0003-2654 R&D Projects: GA MŠk LC06023; GA AV ČR KAN400310651 Institutional research plan: CEZ:AV0Z40310501 Keywords : photopolymerisation * light-emitting -diode * on-line protein digestion Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.913, year: 2010

  20. Resistance to Plum pox virus strain C in Arabidopsis thaliana and Chenopodium foetidum involves genome-linked viral protein and other viral determinants and might depend on compatibility with host translation initiation factors.

    Science.gov (United States)

    Calvo, María; Martínez-Turiño, Sandra; García, Juan Antonio

    2014-11-01

    Research performed on model herbaceous hosts has been useful to unravel the molecular mechanisms that control viral infections. The most common Plum pox virus (PPV) strains are able to infect Nicotiana species as well as Chenopodium and Arabidopsis species. However, isolates belonging to strain C (PPV-C) that have been adapted to Nicotiana spp. are not infectious either in Chenopodium foetidum or in Arabidopsis thaliana. In order to determine the mechanism underlying this interesting host-specific behavior, we have constructed chimerical clones derived from Nicotiana-adapted PPV isolates from the D and C strains, which differ in their capacity to infect A. thaliana and C. foetidum. With this approach, we have identified the nuclear inclusion a protein (VPg+Pro) as the major pathogenicity determinant that conditions resistance in the presence of additional secondary determinants, different for each host. Genome-linked viral protein (VPg) mutations similar to those involved in the breakdown of eIF4E-mediated resistance to other potyviruses allow some PPV chimeras to infect A. thaliana. These results point to defective interactions between a translation initiation factor and the viral VPg as the most probable cause of host-specific incompatibility, in which other viral factors also participate, and suggest that complex interactions between multiple viral proteins and translation initiation factors not only define resistance to potyviruses in particular varieties of susceptible hosts but also contribute to establish nonhost resistance.

  1. Manufacturing Initiative

    Data.gov (United States)

    National Aeronautics and Space Administration — The Advanced Manufacturing Technologies (AMT) Project supports multiple activities within the Administration's National Manufacturing Initiative. A key component of...

  2. Heat shock protein 70 promotes coxsackievirus B3 translation initiation and elongation via Akt-mTORC1 pathway depending on activation of p70S6K and Cdc2.

    Science.gov (United States)

    Wang, Fengping; Qiu, Ye; Zhang, Huifang M; Hanson, Paul; Ye, Xin; Zhao, Guangze; Xie, Ronald; Tong, Lei; Yang, Decheng

    2017-07-01

    We previously demonstrated that coxsackievirus B3 (CVB3) infection upregulated heat shock protein 70 (Hsp70) and promoted CVB3 multiplication. Here, we report the underlying mechanism by which Hsp70 enhances viral RNA translation. By using an Hsp70-overexpressing cell line infected with CVB3, we found that Hsp70 enhanced CVB3 VP1 translation at two stages. First, Hsp70 induced upregulation of VP1 translation at the initiation stage via upregulation of internal ribosome entry site trans-acting factor lupus autoantigen protein and activation of eIF4E binding protein 1, a cap-dependent translation suppressor. Second, we found that Hsp70 increased CVB3 VP1 translation by enhancing translation elongation. This was mediated by the Akt-mammalian target of rapamycin complex 1 signal cascade, which led to the activation of eukaryotic elongation factor 2 via p70S6K- and cell division cycle protein 2 homolog (Cdc2)-mediated phosphorylation and inactivation of eukaryotic elongation factor 2 kinase. We also determined the position of Cdc2 in this signal pathway, indicating that Cdc2 is regulated by mammalian target of rapamycin complex 1. This signal transduction pathway was validated using a number of specific pharmacological inhibitors, short interfering RNAs (siRNAs) and a dominant negative Akt plasmid. Because Hsp70 is a central component of the cellular network of molecular chaperones enhancing viral replication, these data may provide new strategies to limit this viral infection. © 2017 John Wiley & Sons Ltd.

  3. Initial infection of roots and leaves reveals different resistance phenotypes associated with coat protein gene-mediated resistance to Potato mop-top virus.

    Science.gov (United States)

    Germundsson, Anna; Sandgren, Maria; Barker, Hugh; Savenkov, Eugene I; Valkonen, Jari P T

    2002-05-01

    Resistance to the pomovirus Potato mop-top virus (PMTV) was studied in potato (Solanum tuberosum cv. Saturna) and Nicotiana benthamiana transformed with the coat protein (CP) gene of PMTV. The incidence of PMTV infections was reduced in tubers of the CP-transgenic potatoes grown in the field in soil infested with the viruliferous vector, Spongospora subterranea. However, in those tubers that were infected, all three virus RNAs were detected and virus titres were high. The CP-transgenic N. benthamiana plants were inoculated with PMTV using two methods. Following mechanical inoculation of leaves, no RNA 3 (the CP-encoding RNA homologous to the transgene) was detected in leaves, but in some plants low amounts of RNA 3 were detected in roots; RNA 2 was readily detected in leaves and roots of several plants. Inoculation of roots using viruliferous S. subterranea resulted in infection of roots in all plants and the three PMTV RNAs were detected. However, no systemic movement of PMTV from roots to the above-ground parts was observed, indicating a novel expression of resistance. These data indicate that the CP gene-mediated resistance to PMTV specifically restricts accumulation of PMTV RNA 3, and is more effective in leaves than roots. Furthermore, expression of resistance is different depending on whether leaves or roots are inoculated. Data do not exclude the possibility that both a protein-mediated and an RNA-mediated resistance mechanism are involved.

  4. The small molecule '1-(4-biphenylylcarbonyl)-4-(5-bromo-2-methoxybenzyl) piperazine oxalate' and its derivatives regulate global protein synthesis by inactivating eukaryotic translation initiation factor 2-alpha.

    Science.gov (United States)

    Hong, Mi-Na; Nam, Ky-Youb; Kim, Kyung Kon; Kim, So-Young; Kim, InKi

    2016-05-01

    By environmental stresses, cells can initiate a signaling pathway in which eukaryotic translation initiation factor 2-alpha (eIF2-α) is involved to regulate the response. Phosphorylation of eIF2-α results in the reduction of overall protein neogenesis, which allows cells to conserve resources and to reprogram energy usage for effective stress control. To investigate the role of eIF2-α in cell stress responses, we conducted a viability-based compound screen under endoplasmic reticulum (ER) stress condition, and identified 1-(4-biphenylylcarbonyl)-4-(5-bromo-2-methoxybenzyl) piperazine oxalate (AMC-01) and its derivatives as eIF2-α-inactivating chemical. Molecular characterization of this signaling pathway revealed that AMC-01 induced inactivation of eIF2-α by phosphorylating serine residue 51 in a dose- and time-dependent manner, while the negative control compounds did not affect eIF2-α phosphorylation. In contrast with ER stress induction by thapsigargin, phosphorylation of eIF2-α persisted for the duration of incubation with AMC-01. By pathway analysis, AMC-01 clearly induced the activation of protein kinase RNA-activated (PKR) kinase and nuclear factor-κB (NF-κB), whereas it did not modulate the activity of PERK or heme-regulated inhibitor (HRI). Finally, we could detect a lower protein translation rate in cells incubated with AMC-01, establishing AMC-01 as a potent chemical probe that can regulate eIF2-α activity. We suggest from these data that AMC-01 and its derivative compounds can be used as chemical probes in future studies of the role of eIF2-α in protein synthesis-related cell physiology.

  5. Unilateral initiatives

    International Nuclear Information System (INIS)

    Anon.

    1988-01-01

    This paper reports on arms control which is generally thought of in terms of formal negotiations with an opponent, with the resulting agreements embodied in a treaty. This is not surprising, since arms control discussions between opponents are both important and politically visible. There are, however, strong reasons for countries to consider and frequently take unilateral initiatives. To do so is entirely consistent with the established major precepts of arms control which state that arms control is designed to reduce the risk of war, the costs of preparing for war, and the death and destruction if war should come. Unilateral initiatives on what weapons are purchased, which ones are eliminated and how forces are deployed can all relate to these objectives. There are two main categories of motives for unilateral initiatives in arms control. In one category, internal national objectives are the dominant, often sole, driving force; the initiative is undertaken for our own good

  6. Ports Initiative

    Science.gov (United States)

    EPA's Ports Initiative works in collaboration with the port industry, communities, and government to improve environmental performance and increase economic prosperity. This effort helps people near ports breath cleaner air and live better lives.

  7. Effect of laser wavelength and protein solder concentration on acute tissue repair using laser welding: initial results in a canine ureter model.

    Science.gov (United States)

    Wright, E J; Poppas, D P

    1997-01-01

    Successful tissue approximation can be performed using low power laser energy combined with human albumin solder. In vitro studies were undertaken to investigate the acute repair strengths achieved using different laser wavelengths. Furthermore, we evaluated the change in repair strength with that resulted from changes in protein solder concentration. Intraluminal bursting pressure following ureterotomy repair was measured for the following laser wavelengths: 532, 808, 1,320, 2,100, and 10,600 nm. The tissue absorption characteristics of the 808-nm diode and the KTP-532-nm lasers required the addition of the exogenous chromophores indocyanine green and fluorescein, respectively. A 40% human albumin solder was incorporated in the repair of a 1.0-cm longitudinal defect in the canine ureter. Following determination of an optimal welding wavelength, human albumin solder of varying concentrations (25%, 38%, 45%, and 50%) were prepared and tested. The 1,320-nm YAG laser achieved the highest acute bursting pressure and was the most effective in this model. Of the concentrations of albumin tested, 50% human albumin yielded the greatest bursting pressures. We conclude that of the laser wavelengths evaluated, the 1,320-nm YAG achieves the strongest tissue weld in the acute ex vivo dog ureter model. In addition, when this laser system is used, the acute strength of a photothermal weld appears to be directly proportional to the concentration of human albumin solder in the range of 25 to 50%.

  8. Immobilization of cross linked Col-I–OPN bone matrix protein on aminolysed PCL surfaces enhances initial biocompatibility of human adipogenic mesenchymal stem cells (hADMSC)

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Young-Hee; Jyoti, Md. Anirban; Song, Ho-Yeon, E-mail: songmic@sch.ac.kr

    2014-06-01

    In bone tissue engineering surface modification is considered as one of the important ways of fabricating successful biocompatible material. Addition of biologically active functionality on the surfaces has been tried for improving the overall biocompatibility of the system. In this study poly-ε-caprolactone film surfaces have been modified through aminolysis and immobilization process. Collagen type I (COL-I) and osteopontin (OPN), which play an important role in osteogenesis, was immobilized onto PCL films followed by aminolysis treatment using 1,6-hexanediamine. Characterization of animolysed and immobilized surfaces were done by a number techniques using scanning electron microscopy (SEM), FT-IR, XPS, ninhydrin staining, SDS-PAGE and confocal microscopy and compared between the modified and un-modified surfaces. Results of the successive experiments showed that aminolysis treatment was homogeneously achieved which helped to entrap or immobilize Col-I–OPN proteins on surfaces of PCL film. In vitro studies with human adipogenic mesenchymal stem cells (hADMSC) also confirmed the attachment and proliferation of cells was better in modified PCL surfaces than the unmodified surfaces. SEM, confocal microscopy and MTT assay showed a significant increase in cell spreading, attachment and proliferations on the biofunctionalized surfaces compared to the unmodified PCL surfaces at all-time points indicating the success of surface biofunctionalization.

  9. Initial Study

    DEFF Research Database (Denmark)

    Torp, Kristian

    2009-01-01

    increased. In the initial study presented here, the time it takes to pass an intersection is studied in details. Two major signal-controlled four-way intersections in the center of the city Aalborg are studied in details to estimate the congestion levels in these intersections, based on the time it takes...

  10. Involvement of three meningococcal surface-exposed proteins, the heparin-binding protein NhbA, the α-peptide of IgA protease and the autotransporter protease NalP, in initiation of biofilm formation

    KAUST Repository

    Arenas, Jesú s; Nijland, Reindert; Rodriguez, Francisco J.; Bosma, Tom N. P.; Tommassen, Jan

    2012-01-01

    Neisseria meningitidis is a common and usually harmless inhabitant of the mucosa of the human nasopharynx, which, in rare cases, can cross the epithelial barrier and cause meningitis and sepsis. Biofilm formation favours the colonization of the host and the subsequent carrier state. Two different strategies of biofilm formation, either dependent or independent on extracellular DNA (eDNA), have been described for meningococcal strains. Here, we demonstrate that the autotransporter protease NalP, the expression of which is phase variable, affects eDNA-dependent biofilm formation in N.meningitidis. The effect of NalP was found in biofilm formation under static and flow conditions and was dependent on its protease activity. Cleavage of the heparin-binding antigen NhbA and the α-peptide of IgA protease, resulting in the release of positively charged polypeptides from the cell surface, was responsible for the reduction in biofilm formation when NalP is expressed. Both NhbA and the α-peptide of IgA protease were shown to bind DNA. We conclude that NhbA and the α-peptide of IgA protease are implicated in biofilm formation by binding eDNA and that NalP is an important regulator of this process through the proteolysis of these surface-exposed proteins. © 2012 Blackwell Publishing Ltd.

  11. Involvement of three meningococcal surface-exposed proteins, the heparin-binding protein NhbA, the α-peptide of IgA protease and the autotransporter protease NalP, in initiation of biofilm formation

    KAUST Repository

    Arenas, Jesús

    2012-12-04

    Neisseria meningitidis is a common and usually harmless inhabitant of the mucosa of the human nasopharynx, which, in rare cases, can cross the epithelial barrier and cause meningitis and sepsis. Biofilm formation favours the colonization of the host and the subsequent carrier state. Two different strategies of biofilm formation, either dependent or independent on extracellular DNA (eDNA), have been described for meningococcal strains. Here, we demonstrate that the autotransporter protease NalP, the expression of which is phase variable, affects eDNA-dependent biofilm formation in N.meningitidis. The effect of NalP was found in biofilm formation under static and flow conditions and was dependent on its protease activity. Cleavage of the heparin-binding antigen NhbA and the α-peptide of IgA protease, resulting in the release of positively charged polypeptides from the cell surface, was responsible for the reduction in biofilm formation when NalP is expressed. Both NhbA and the α-peptide of IgA protease were shown to bind DNA. We conclude that NhbA and the α-peptide of IgA protease are implicated in biofilm formation by binding eDNA and that NalP is an important regulator of this process through the proteolysis of these surface-exposed proteins. © 2012 Blackwell Publishing Ltd.

  12. Initiation of protein synthesis by a labeled derivative of the Lactobacillus casei DN-114 001 strain during transit from the stomach to the cecum in mice harboring human microbiota.

    Science.gov (United States)

    Oozeer, R; Mater, D D G; Goupil-Feuillerat, N; Corthier, G

    2004-12-01

    Although studies on the survival of bacteria in the digestive tract have been reported in the literature, little data are available on the physiological adaptation of probiotics to the digestive environment. In previous work, a transcriptional fusion system (i.e., luciferase genes under the control of a deregulated promoter) was used to demonstrate that a derivative of the Lactobacillus casei DN-114 001 strain, ingested in a fermented milk and thus exhibiting initially a very weak metabolic activity, synthesized proteins de novo after its transit in the digestive tract of mice harboring human microbiota (known as human-microbiota-associated mice). With the same genetic system and animal model, we here investigate for the first time the ability of L. casei to reinitiate synthesis in the different digestive tract compartments. In this study, most ingested L. casei cells transited from the stomach to the duodenum-jejunum within 1 h postingestion. No luciferase activity was observed in these digestive tract compartments after the first hour. At later times, the bulk of bacteria had transited to the ileum and the cecum. Luciferase synthesis was detected between 1.5 and 2.0 h postingestion at the ileal level and from 1.5 h to at least 6.0 h postingestion in the cecum, where the activity remained at a maximum level. These results demonstrate that ingested L. casei (derivative of the DN-114 001 strain) administered via a fermented milk has already reinitiated protein synthesis when it reaches the ileal and cecal compartments.

  13. Mitochondrial Respiration Inhibitors Suppress Protein Translation and Hypoxic Signaling via the Hyperphosphorylation and Inactivation of Translation Initiation Factor eIF2α and Elongation Factor eEF2

    Science.gov (United States)

    Li, Jun; Mahdi, Fakhri; Du, Lin; Datta, Sandipan; Nagle, Dale G.; Zhou, Yu-Dong

    2011-01-01

    Over 20000 lipid extracts of plants and marine organisms were evaluated in a human breast tumor T47D cell-based reporter assay for hypoxia-inducible factor-1 (HIF-1) inhibitory activity. Bioassay-guided isolation and dereplication-based structure elucidation of an active extract from the Bael tree (Aegle marmelos) afforded two protolimonoids, skimmiarepin A (1) and skimmiarepin C (2). In T47D cells, 1 and 2 inhibited hypoxia-induced HIF-1 activation with IC50 values of 0.063 µM and 0.068 µM, respectively. Compounds 1 and 2 also suppressed hypoxic induction of the HIF-1 target genes GLUT-1 and VEGF. Mechanistic studies revealed that 1 and 2 inhibited HIF-1 activation by blocking the hypoxia-induced accumulation of HIF-1α protein. At the range of concentrations that inhibited HIF-1 activation, 1 and 2 suppressed cellular respiration by selectively inhibiting the mitochondrial electron transport chain at complex I (NADH dehydrogenase). Further investigation indicated that mitochondrial respiration inhibitors such as 1 and rotenone induced the rapid hyperphosphorylation and inhibition of translation initiation factor eIF2α and elongation factor eEF2. The inhibition of protein translation may account for the short-term exposure effects exerted by mitochondrial inhibitors on cellular signaling, while the suppression of cellular ATP production may contribute to the inhibitory effects following extended treatment periods. PMID:21875114

  14. Re-wiring of energy metabolism promotes viability during hyperreplication stress in E. coli

    DEFF Research Database (Denmark)

    Charbon, Godefroid; Campion, Christopher; Chan, Siu Hung Joshua

    2017-01-01

    Chromosome replication in Escherichia coli is initiated by DnaA. DnaA binds ATP which is essential for formation of a DnaA-oriC nucleoprotein complex that promotes strand opening, helicase loading and replisome assembly. Following initiation, DnaAATP is converted to DnaAADP primarily by the Regul......Chromosome replication in Escherichia coli is initiated by DnaA. DnaA binds ATP which is essential for formation of a DnaA-oriC nucleoprotein complex that promotes strand opening, helicase loading and replisome assembly. Following initiation, DnaAATP is converted to DnaAADP primarily...

  15. Protein oxidation in aquatic foods

    DEFF Research Database (Denmark)

    Baron, Caroline P.

    2014-01-01

    The chapter discusses general considerations about protein oxidation and reviews the mechanisms involved in protein oxidation and consequences of protein oxidation on fish proteins. It presents two case studies, the first deals with protein and lipid oxidation in frozen rainbow trout......, and the second with oxidation in salted herring. The mechanisms responsible for initiation of protein oxidation are unclear, but it is generally accepted that free radical species initiating lipid oxidation can also initiate protein oxidation. The chapter focuses on interaction between protein and lipid...... oxidation. The protein carbonyl group measurement is the widely used method for estimating protein oxidation in foods and has been used in fish muscle. The chapter also talks about the impact of protein oxidation on protein functionality, fish muscle texture, and food nutritional value. Protein oxidation...

  16. Suppressors of Hyperinitiation in Escherichia coli Couple DNA Replication to Precursor Biosynthesis and Energy Metabolism

    DEFF Research Database (Denmark)

    Bjørn, Louise

    The Hda protein plays an essential role in inactivation of the initiator protein DnaA from its active, ATP bound form to the inactive DnaA-ADP in E. coli. Cells deficient in Hda suffer from overinitiation, asynchronous initiation and cell death as a consequence of an increased DnaAATP/ Dna......A-ADP ratio . E. coli can suppress the growth defects caused by Hda deficiency by several different mechanisms. The focus of this Ph.d. thesis is to understand the mechanisms that underlie suppression of Hda deficiency in E. coli. These approaches are described in two manuscripts and one published paper. Over...... expression of Ribonucleotide reductase encoded by either nrdAB or nrdEF has been shown to suppress Hda deficiency. The nrdAB promoter contains four consensus binding sequences for DnaA and a 45bp inverted repeat important for cell cycle regulation of nrdAB transcription. In manuscript 1 we show...

  17. Openness initiative

    International Nuclear Information System (INIS)

    Duncan, S.S.

    1995-01-01

    Although antinuclear campaigns seem to be effective, public communication and education efforts on low-level radioactive waste have mixed results. Attempts at public information programs on low-level radioactive waste still focus on influencing public opinion. A question then is: open-quotes Is it preferable to have a program focus on public education that will empower individuals to make informed decisions rather than trying to influence them in their decisions?close quotes To address this question, a case study with both quantitative and qualitative data will be used. The Ohio Low-Level Radioactive Waste Education Program has a goal to provide people with information they want/need to make their own decisions. The program initiated its efforts by conducting a statewide survey to determine information needed by people and where they turned for that information. This presentation reports data from the survey and then explores the program development process in which programs were designed and presented using the information. Pre and post data from the programs reveal attitude and knowledge shifts

  18. Openness initiative

    Energy Technology Data Exchange (ETDEWEB)

    Duncan, S.S. [Los Alamos National Lab., NM (United States)

    1995-12-31

    Although antinuclear campaigns seem to be effective, public communication and education efforts on low-level radioactive waste have mixed results. Attempts at public information programs on low-level radioactive waste still focus on influencing public opinion. A question then is: {open_quotes}Is it preferable to have a program focus on public education that will empower individuals to make informed decisions rather than trying to influence them in their decisions?{close_quotes} To address this question, a case study with both quantitative and qualitative data will be used. The Ohio Low-Level Radioactive Waste Education Program has a goal to provide people with information they want/need to make their own decisions. The program initiated its efforts by conducting a statewide survey to determine information needed by people and where they turned for that information. This presentation reports data from the survey and then explores the program development process in which programs were designed and presented using the information. Pre and post data from the programs reveal attitude and knowledge shifts.

  19. Initiative hard coal; Initiative Steinkohle

    Energy Technology Data Exchange (ETDEWEB)

    Leonhardt, J.

    2007-08-02

    In order to decrease the import dependence of hard coal in the European Union, the author has submitted suggestions to the director of conventional sources of energy (directorate general for energy and transport) of the European community, which found a positive resonance. These suggestions are summarized in an elaboration 'Initiative Hard Coal'. After clarifying the starting situation and defining the target the presupposition for a better use of hard coal deposits as raw material in the European Union are pointed out. On that basis concrete suggestions for measures are made. Apart from the conditions of the deposits it concerns thereby also new mining techniques and mining-economical developments, connected with tasks for the mining-machine industry. (orig.)

  20. The time frame of Epstein-Barr virus latent membrane protein-1 gene to disappear in nasopharyngeal swabs after initiation of primary radiotherapy is an independently significant prognostic factor predicting local control for patients with nasopharyngeal carcinoma

    International Nuclear Information System (INIS)

    Lin, S.-Y.; Chang, K.-P.; Hsieh, M.-S.; Ueng, S.-H.; Hao, S.-P.; Tseng, C.-K.; Pai, P.-C.; Chang, F.-T.; Tsai, M.-H.; Tsang, N.-M.

    2005-01-01

    Purpose: The presence of Epstein-Barr virus latent membrane protein-1 (LMP-1) gene in nasopharyngeal swabs indicates the presence of nasopharyngeal carcinoma (NPC) mucosal tumor cells. This study was undertaken to investigate whether the time taken for LMP-1 to disappear after initiation of primary radiotherapy (RT) was inversely associated with NPC local control. Methods and Materials: During July 1999 and October 2002, there were 127 nondisseminated NPC patients receiving serial examinations of nasopharyngeal swabbing with detection of LMP-1 during the RT course. The time for LMP-1 regression was defined as the number of days after initiation of RT for LMP-1 results to turn negative. The primary outcome was local control, which was represented by freedom from local recurrence. Results: The time for LMP-1 regression showed a statistically significant influence on NPC local control both univariately (p < 0.0001) and multivariately (p = 0.004). In multivariate analysis, the administration of chemotherapy conferred a significantly more favorable local control (p = 0.03). Advanced T status (≥ T2b), overall treatment time of external photon radiotherapy longer than 55 days, and older age showed trends toward being poor prognosticators. The time for LMP-1 regression was very heterogeneous. According to the quartiles of the time for LMP-1 regression, we defined the pattern of LMP-1 regression as late regression if it required 40 days or more. Kaplan-Meier plots indicated that the patients with late regression had a significantly worse local control than those with intermediate or early regression (p 0.0129). Conclusion: Among the potential prognostic factors examined in this study, the time for LMP-1 regression was the most independently significant factor that was inversely associated with NPC local control

  1. An initial event in the insect innate immune response: structural and biological studies of interactions between β-1,3-glucan and the N-terminal domain of β-1,3-glucan recognition protein.

    Science.gov (United States)

    Dai, Huaien; Hiromasa, Yasuaki; Takahashi, Daisuke; VanderVelde, David; Fabrick, Jeffrey A; Kanost, Michael R; Krishnamoorthi, Ramaswamy

    2013-01-08

    , and Asp68 between the ligand-bound protein molecules contribute in part to the stability of the N-βGRP-laminarin macro complex and that a decreased stability is accompanied by a reduced level of activation of the proPO pathway. An increased level of β-1,6 branching in laminarin also results in destabilization of the macro complex. These novel findings suggest that ligand-induced self-association of the βGRP-β-1,3-glucan complex may form a platform on a microbial surface for recruitment of downstream proteases, as a means of amplification of the initial signal of pathogen recognition for the activation of the proPO pathway.

  2. Thyroid disorders in patients with newly diagnosed rheumatoid arthritis is associated with poor initial treatment response evaluated by disease activity score in 28 joints-C-reactive protein (DAS28-CRP)

    DEFF Research Database (Denmark)

    Emamifar, Amir; Hangaard, Jørgen; Jensen Hansen, Inger Marie

    2017-01-01

    peptide antibody (anti-CCP), and antinuclear antibody (ANA), and also disease activity score in 28 joints-C-reactive protein at the time of diagnosis and after 4 months (±1–2 months) of treatment initiation were extracted from Danish Danbio Registry. Patients’ electronic hospital records for the past 10...... years were reviewed to reveal if they had been diagnosed with thyroid disorders or they had abnormal thyroid test.In all, 439 patients were included, female 60.1%, mean age 64.6 ± 15.0 years and disease duration 2.6 ± 1.7 years. Prevalence of thyroid disorders was 69/439 (15.7%) and hypothyroidism...... treatment compared with patients with isolated RA after 4 months of treatment (P = .02). There were no associations between thyroid disorders and age, disease duration, and also IgM RF positivity.Presence of thyroid disorders in RA patients is suggestive of a more aggressive disease and poor outcome...

  3. Hda Monomerization by ADP Binding Promotes Replicase Clamp-mediated DnaA-ATP Hydrolysis*S⃞

    Science.gov (United States)

    Su'etsugu, Masayuki; Nakamura, Kenta; Keyamura, Kenji; Kudo, Yuka; Katayama, Tsutomu

    2008-01-01

    ATP-DnaA is the initiator of chromosomal replication in Escherichia coli, and the activity of DnaA is regulated by the regulatory inactivation of the DnaA (RIDA) system. In this system, the Hda protein promotes DnaA-ATP hydrolysis to produce inactive ADP-DnaA in a mechanism that is mediated by the DNA-loaded form of the replicase sliding clamp. In this study, we first revealed that hda translation uses an unusual initiation codon, CUG, located downstream of the annotated initiation codon. The CUG initiation codon could be used for restricting the Hda level, as this initiation codon has a low translation efficiency, and the cellular Hda level is only ∼100 molecules per cell. Hda translated using the correct reading frame was purified and found to have a high RIDA activity in vitro. Moreover, we found that Hda has a high affinity for ADP but not for other nucleotides, including ATP. ADP-Hda was active in the RIDA system in vitro and stable in a monomeric state, whereas apo-Hda formed inactive homomultimers. Both ADP-Hda and apo-Hda could form complexes with the DNA-loaded clamp; however, only ADP-Hda-DNA-clamp complexes were highly functional in the following interaction with DnaA. Formation of ADP-Hda was also observed in vivo, and mutant analysis suggested that ADP binding is crucial for cellular Hda activity. Thus, we propose that ADP is a crucial Hda ligand that promotes the activated conformation of the protein. ADP-dependent monomerization might enable the arginine finger of the Hda AAA+ domain to be accessible to ATP bound to the DnaA AAA+ domain. PMID:18977760

  4. Hda monomerization by ADP binding promotes replicase clamp-mediated DnaA-ATP hydrolysis.

    Science.gov (United States)

    Su'etsugu, Masayuki; Nakamura, Kenta; Keyamura, Kenji; Kudo, Yuka; Katayama, Tsutomu

    2008-12-26

    ATP-DnaA is the initiator of chromosomal replication in Escherichia coli, and the activity of DnaA is regulated by the regulatory inactivation of the DnaA (RIDA) system. In this system, the Hda protein promotes DnaA-ATP hydrolysis to produce inactive ADP-DnaA in a mechanism that is mediated by the DNA-loaded form of the replicase sliding clamp. In this study, we first revealed that hda translation uses an unusual initiation codon, CUG, located downstream of the annotated initiation codon. The CUG initiation codon could be used for restricting the Hda level, as this initiation codon has a low translation efficiency, and the cellular Hda level is only approximately 100 molecules per cell. Hda translated using the correct reading frame was purified and found to have a high RIDA activity in vitro. Moreover, we found that Hda has a high affinity for ADP but not for other nucleotides, including ATP. ADP-Hda was active in the RIDA system in vitro and stable in a monomeric state, whereas apo-Hda formed inactive homomultimers. Both ADP-Hda and apo-Hda could form complexes with the DNA-loaded clamp; however, only ADP-Hda-DNA-clamp complexes were highly functional in the following interaction with DnaA. Formation of ADP-Hda was also observed in vivo, and mutant analysis suggested that ADP binding is crucial for cellular Hda activity. Thus, we propose that ADP is a crucial Hda ligand that promotes the activated conformation of the protein. ADP-dependent monomerization might enable the arginine finger of the Hda AAA+ domain to be accessible to ATP bound to the DnaA AAA+ domain.

  5. Initiation devices, initiation systems including initiation devices and related methods

    Energy Technology Data Exchange (ETDEWEB)

    Daniels, Michael A.; Condit, Reston A.; Rasmussen, Nikki; Wallace, Ronald S.

    2018-04-10

    Initiation devices may include at least one substrate, an initiation element positioned on a first side of the at least one substrate, and a spark gap electrically coupled to the initiation element and positioned on a second side of the at least one substrate. Initiation devices may include a plurality of substrates where at least one substrate of the plurality of substrates is electrically connected to at least one adjacent substrate of the plurality of substrates with at least one via extending through the at least one substrate. Initiation systems may include such initiation devices. Methods of igniting energetic materials include passing a current through a spark gap formed on at least one substrate of the initiation device, passing the current through at least one via formed through the at least one substrate, and passing the current through an explosive bridge wire of the initiation device.

  6. Initial brain aging

    DEFF Research Database (Denmark)

    Thomsen, Kirsten; Yokota, Takashi; Hasan-Olive, Md Mahdi

    2018-01-01

    Brain aging is accompanied by declining mitochondrial respiration. We hypothesized that mitochondrial morphology and dynamics would reflect this decline. Using hippocampus and frontal cortex of a segmental progeroid mouse model lacking Cockayne syndrome protein B (CSB(m/m)) and C57Bl/6 (WT......) controls and comparing young (2-5 months) to middle-aged mice (13-14 months), we found that complex I-linked state 3 respiration (CI) was reduced at middle age in CSB(m/m) hippocampus, but not in CSB(m/m) cortex or WT brain. In hippocampus of both genotypes, mitochondrial size heterogeneity increased....... Mitochondrial DNA content was lower, and hypoxia-induced factor 1α mRNA was greater at both ages in CSB(m/m) compared to WT brain. Our findings show that decreased CI and increased mitochondrial size heterogeneity are highly associated and point to declining mitochondrial quality control as an initial event...

  7. The Enzyme Function Initiative.

    Science.gov (United States)

    Gerlt, John A; Allen, Karen N; Almo, Steven C; Armstrong, Richard N; Babbitt, Patricia C; Cronan, John E; Dunaway-Mariano, Debra; Imker, Heidi J; Jacobson, Matthew P; Minor, Wladek; Poulter, C Dale; Raushel, Frank M; Sali, Andrej; Shoichet, Brian K; Sweedler, Jonathan V

    2011-11-22

    The Enzyme Function Initiative (EFI) was recently established to address the challenge of assigning reliable functions to enzymes discovered in bacterial genome projects; in this Current Topic, we review the structure and operations of the EFI. The EFI includes the Superfamily/Genome, Protein, Structure, Computation, and Data/Dissemination Cores that provide the infrastructure for reliably predicting the in vitro functions of unknown enzymes. The initial targets for functional assignment are selected from five functionally diverse superfamilies (amidohydrolase, enolase, glutathione transferase, haloalkanoic acid dehalogenase, and isoprenoid synthase), with five superfamily specific Bridging Projects experimentally testing the predicted in vitro enzymatic activities. The EFI also includes the Microbiology Core that evaluates the in vivo context of in vitro enzymatic functions and confirms the functional predictions of the EFI. The deliverables of the EFI to the scientific community include (1) development of a large-scale, multidisciplinary sequence/structure-based strategy for functional assignment of unknown enzymes discovered in genome projects (target selection, protein production, structure determination, computation, experimental enzymology, microbiology, and structure-based annotation), (2) dissemination of the strategy to the community via publications, collaborations, workshops, and symposia, (3) computational and bioinformatic tools for using the strategy, (4) provision of experimental protocols and/or reagents for enzyme production and characterization, and (5) dissemination of data via the EFI's Website, http://enzymefunction.org. The realization of multidisciplinary strategies for functional assignment will begin to define the full metabolic diversity that exists in nature and will impact basic biochemical and evolutionary understanding, as well as a wide range of applications of central importance to industrial, medicinal, and pharmaceutical efforts.

  8. Evolving trends in biosciences: Multi-purpose proteins - GFP and GFP-like proteins

    Digital Repository Service at National Institute of Oceanography (India)

    Krishna, K.; Ingole, B.S.

    The sea is considered as holding a clue to many known and unknown biologically active compounds. A family of protein named Green Fluorescent Proteins (GFP)-like proteins, initially isolated from marine organisms, started a trend in biotechnological...

  9. Small Ribosomal Protein RPS0 Stimulates Translation Initiation by Mediating 40S-Binding of eIF3 via Its Direct Contact with the eIF3a/TIF32 Subunit

    Czech Academy of Sciences Publication Activity Database

    Kouba, Tomáš; Dányi, István; Gunišová, Stanislava; Munzarová, Vanda; Vlčková, Vladislava; Cuchalová, Lucie; Neueder, A.; Milkereit, P.; Valášek, Leoš Shivaya

    2012-01-01

    Roč. 7, č. 7 (2012), e40464 E-ISSN 1932-6203 R&D Projects: GA ČR(CZ) GAP305/11/0172 Institutional research plan: CEZ:AV0Z50200510 Keywords : eIF3a/TIF32 * mRNAs * protein Subject RIV: EE - Microbiology, Virology Impact factor: 3.730, year: 2012

  10. Initialized Fractional Calculus

    Science.gov (United States)

    Lorenzo, Carl F.; Hartley, Tom T.

    2000-01-01

    This paper demonstrates the need for a nonconstant initialization for the fractional calculus and establishes a basic definition set for the initialized fractional differintegral. This definition set allows the formalization of an initialized fractional calculus. Two basis calculi are considered; the Riemann-Liouville and the Grunwald fractional calculi. Two forms of initialization, terminal and side are developed.

  11. MONTANA PALLADIUM RESEARCH INITIATIVE

    Energy Technology Data Exchange (ETDEWEB)

    Peters, John; McCloskey, Jay; Douglas, Trevor; Young, Mark; Snyder, Stuart; Gurney, Brian

    2012-05-09

    Project Objective: The overarching objective of the Montana Palladium Research Initiative is to perform scientific research on the properties and uses of palladium in the context of the U.S. Department of Energy's Hydrogen, Fuel Cells and Infrastructure Technologies Program. The purpose of the research will be to explore possible palladium as an alternative to platinum in hydrogen-economy applications. To achieve this objective, the Initiatives activities will focus on several cutting-edge research approaches across a range of disciplines, including metallurgy, biomimetics, instrumentation development, and systems analysis. Background: Platinum-group elements (PGEs) play significant roles in processing hydrogen, an element that shows high potential to address this need in the U.S. and the world for inexpensive, reliable, clean energy. Platinum, however, is a very expensive component of current and planned systems, so less-expensive alternatives that have similar physical properties are being sought. To this end, several tasks have been defined under the rubric of the Montana Palladium Research Iniative. This broad swath of activities will allow progress on several fronts. The membrane-related activities of Task 1 employs state-of-the-art and leading-edge technologies to develop new, ceramic-substrate metallic membranes for the production of high-purity hydrogen, and develop techniques for the production of thin, defect-free platinum group element catalytic membranes for energy production and pollution control. The biomimetic work in Task 2 explores the use of substrate-attached hydrogen-producing enzymes and the encapsulation of palladium in virion-based protein coats to determine their utility for distributed hydrogen production. Task 3 work involves developing laser-induced breakdown spectroscopy (LIBS) as a real-time, in situ diagnostic technique to characterize PGEs nanoparticles for process monitoring and control. The systems engineering work in task 4

  12. Dioxin Exposure Initiative

    Science.gov (United States)

    The Dioxin Exposure Initiative (DEI) is no longer active. This page contains a summary of the dioxin exposure initiative with illustrations, contact and background information.Originally supported by scientist Matthew Lorber, who retired in Mar 2017.

  13. Markers of protein oxidation

    DEFF Research Database (Denmark)

    Headlam, Henrietta A; Davies, Michael Jonathan

    2004-01-01

    Exposure of proteins to radicals in the presence of O2 gives both side-chain oxidation and backbone fragmentation. These processes can be interrelated, with initial side-chain oxidation giving rise to backbone damage via transfer reactions. We have shown previously that alkoxyl radicals formed...... of this process depends on the extent of oxidation at C-3 compared with other sites. HO*, generated by gamma radiolysis, gave the highest total carbonyl yield, with protein-bound carbonyls predominating over released. In contrast, metal ion/H2O2 systems, gave more released than bound carbonyls, with this ratio...... modulated by EDTA. This is ascribed to metal ion-protein interactions affecting the sites of initial oxidation. Hypochlorous acid gave low concentrations of released carbonyls, but high yields of protein-bound material. The peroxyl radical generator 2,2'-azobis(2-amidinopropane) hydrochloride...

  14. Tall Buildings Initiative

    Science.gov (United States)

    Initiative 2017 TBI Guidelines Version 2.03 Now Available Screen Shot 2017-10-10 at 3.05.10 PM PEER has just initiative to develop design criteria that will ensure safe and usable tall buildings following future earthquakes. Download the primary product of this initiative: Guidelines for Performance-Based Seismic Design

  15. Transcription initiation complex structures elucidate DNA opening.

    Science.gov (United States)

    Plaschka, C; Hantsche, M; Dienemann, C; Burzinski, C; Plitzko, J; Cramer, P

    2016-05-19

    Transcription of eukaryotic protein-coding genes begins with assembly of the RNA polymerase (Pol) II initiation complex and promoter DNA opening. Here we report cryo-electron microscopy (cryo-EM) structures of yeast initiation complexes containing closed and open DNA at resolutions of 8.8 Å and 3.6 Å, respectively. DNA is positioned and retained over the Pol II cleft by a network of interactions between the TATA-box-binding protein TBP and transcription factors TFIIA, TFIIB, TFIIE, and TFIIF. DNA opening occurs around the tip of the Pol II clamp and the TFIIE 'extended winged helix' domain, and can occur in the absence of TFIIH. Loading of the DNA template strand into the active centre may be facilitated by movements of obstructing protein elements triggered by allosteric binding of the TFIIE 'E-ribbon' domain. The results suggest a unified model for transcription initiation with a key event, the trapping of open promoter DNA by extended protein-protein and protein-DNA contacts.

  16. Expression of multiple proteins in transgenic plants

    Science.gov (United States)

    Vierstra, Richard D.; Walker, Joseph M.

    2002-01-01

    A method is disclosed for the production of multiple proteins in transgenic plants. A DNA construct for introduction into plants includes a provision to express a fusion protein of two proteins of interest joined by a linking domain including plant ubiquitin. When the fusion protein is produced in the cells of a transgenic plant transformed with the DNA construction, native enzymes present in plant cells cleave the fusion protein to release both proteins of interest into the cells of the transgenic plant. Since the proteins are produced from the same fusion protein, the initial quantities of the proteins in the cells of the plant are approximately equal.

  17. Protein oxidation and peroxidation

    DEFF Research Database (Denmark)

    Davies, Michael Jonathan

    2016-01-01

    Proteins are major targets for radicals and two-electron oxidants in biological systems due to their abundance and high rate constants for reaction. With highly reactive radicals damage occurs at multiple side-chain and backbone sites. Less reactive species show greater selectivity with regard...... to the residues targeted and their spatial location. Modification can result in increased side-chain hydrophilicity, side-chain and backbone fragmentation, aggregation via covalent cross-linking or hydrophobic interactions, protein unfolding and altered conformation, altered interactions with biological partners...... and modified turnover. In the presence of O2, high yields of peroxyl radicals and peroxides (protein peroxidation) are formed; the latter account for up to 70% of the initial oxidant flux. Protein peroxides can oxidize both proteins and other targets. One-electron reduction results in additional radicals...

  18. Physics of protein folding

    Science.gov (United States)

    Finkelstein, A. V.; Galzitskaya, O. V.

    2004-04-01

    Protein physics is grounded on three fundamental experimental facts: protein, this long heteropolymer, has a well defined compact three-dimensional structure; this structure can spontaneously arise from the unfolded protein chain in appropriate environment; and this structure is separated from the unfolded state of the chain by the “all-or-none” phase transition, which ensures robustness of protein structure and therefore of its action. The aim of this review is to consider modern understanding of physical principles of self-organization of protein structures and to overview such important features of this process, as finding out the unique protein structure among zillions alternatives, nucleation of the folding process and metastable folding intermediates. Towards this end we will consider the main experimental facts and simple, mostly phenomenological theoretical models. We will concentrate on relatively small (single-domain) water-soluble globular proteins (whose structure and especially folding are much better studied and understood than those of large or membrane and fibrous proteins) and consider kinetic and structural aspects of transition of initially unfolded protein chains into their final solid (“native”) 3D structures.

  19. Emerging Pathogens Initiative (EPI)

    Data.gov (United States)

    Department of Veterans Affairs — The Emerging Pathogens Initiative (EPI) database contains emerging pathogens information from the local Veterans Affairs Medical Centers (VAMCs). The EPI software...

  20. Eukaryotic translation initiator protein 1A isoform, CCS-3, enhances the transcriptional repression of p21CIP1 by proto-oncogene FBI-1 (Pokemon/ZBTB7A).

    Science.gov (United States)

    Choi, Won-Il; Kim, Youngsoo; Kim, Yuri; Yu, Mi-young; Park, Jungeun; Lee, Choong-Eun; Jeon, Bu-Nam; Koh, Dong-In; Hur, Man-Wook

    2009-01-01

    FBI-1, a member of the POK (POZ and Kruppel) family of transcription factors, plays a role in differentiation, oncogenesis, and adipogenesis. eEF1A is a eukaryotic translation elongation factor involved in several cellular processes including embryogenesis, oncogenic transformation, cell proliferation, and cytoskeletal organization. CCS-3, a potential cervical cancer suppressor, is an isoform of eEF1A. We found that eEF1A forms a complex with FBI-1 by co-immunoprecipitation, SDS-PAGE, and MALDI-TOF Mass analysis of the immunoprecipitate. GST fusion protein pull-downs showed that FBI-1 directly interacts with eEF1A and CCS-3 via the zinc finger and POZ-domain of FBI-1. FBI-1 co-localizes with either eEF1A or CCS-3 at the nuclear periplasm. CCS-3 enhances transcriptional repression of the p21CIP1 gene (hereafter referred to as p21) by FBI-1. The POZ-domain of FBI-1 interacts with the co-repressors, SMRT and BCoR. We found that CCS-3 also interacts with the co-repressors independently. The molecular interaction between the co-repressors and CCS-3 at the POZ-domain of FBI-1 appears to enhance FBI-1 mediated transcriptional repression. Our data suggest that CCS-3 may be important in cell differentiation, tumorigenesis, and oncogenesis by interacting with the proto-oncogene FBI-1 and transcriptional co-repressors. Copyright 2009 S. Karger AG, Basel.

  1. AC Initiation System.

    Science.gov (United States)

    An ac initiation system is described which uses three ac transmission signals interlocked for safety by frequency, phase, and power discrimination...The ac initiation system is pre-armed by the application of two ac signals have the proper phases, and activates a load when an ac power signal of the proper frequency and power level is applied. (Author)

  2. Sustainable Agricultural Marketing Initiatives

    Directory of Open Access Journals (Sweden)

    Hakan Adanacıoğlu

    2015-07-01

    Full Text Available Sustainable marketing is a holistic approach that puts equal emphasis on environmental, social equity, and economic concerns in the development of marketing strategies. The purpose of the study is to examine and discuss the sustainable agricultural marketing initiatives practiced throughout the World and Turkey, and to put forth suggestions to further improve the performance of agricultural marketing initiatives in Turkey. Some of the sustainable agricultural marketing initiatives practiced around the world are carried out through civil organizations. Furthermore; some of these initiatives have also launched by farmers, consumers, food processors and retailers. The long-term strategies to increase these initiatives should be determined due to the fact that examples of successful sustainable agricultural marketing initiatives are inadequate and cannot be spread in Turkey. In this context, first of all, the supports provided by the government to improve agricultural marketing systems, such as EU funds for rural development should be compatible with the goals of sustainable marketing. For this purpose, it should be examined whether all proposed projects related to agricultural marketing meet the social, economic, and environmental principles of sustainable marketing. It is important that supporting organizations, especially civil society organisations, should take an active role for faster dissemination and adoption of sustainable agricultural marketing practices in Turkey. These organizations may provide technical assistance in preparing successful project proposals and training to farm groups. In addition, the other organizations, such as local administrations, producers' associations, cooperatives, can contribute to the success of sustainable agricultural marketing initiatives. The use of direct marketing strategies and vertical integration attempts in sustainable agricultural marketing initiatives that will likely be implemented in Turkey is

  3. Scoring functions for protein-protein interactions.

    Science.gov (United States)

    Moal, Iain H; Moretti, Rocco; Baker, David; Fernández-Recio, Juan

    2013-12-01

    The computational evaluation of protein-protein interactions will play an important role in organising the wealth of data being generated by high-throughput initiatives. Here we discuss future applications, report recent developments and identify areas requiring further investigation. Many functions have been developed to quantify the structural and energetic properties of interacting proteins, finding use in interrelated challenges revolving around the relationship between sequence, structure and binding free energy. These include loop modelling, side-chain refinement, docking, multimer assembly, affinity prediction, affinity change upon mutation, hotspots location and interface design. Information derived from models optimised for one of these challenges can be used to benefit the others, and can be unified within the theoretical frameworks of multi-task learning and Pareto-optimal multi-objective learning. Copyright © 2013 Elsevier Ltd. All rights reserved.

  4. 1996 environmental initiatives report

    International Nuclear Information System (INIS)

    1996-01-01

    Progress by Consumers Gas in addressing environmental challenges were reviewed. Proposed environmental initiatives for the next fiscal year and beyond were introduced. Proposed initiatives were placed into three priority categories, high, medium or low, which together with the environmental management framework form the the utility's overall environmental agenda. High on the list of environmental priorities for the company are atmospheric air emissions, planning and construction practices, energy conservation and efficiency, environmental compliance, and methane emissions. The present state of the initiatives by the various company divisions and regions, compiled from the respective business plans, were reported. 21 figs

  5. P1 plasmid replication: initiator sequestration is inadequate to explain control by initiator-binding sites.

    OpenAIRE

    Pal, S K; Chattoraj, D K

    1988-01-01

    The unit-copy plasmid replicon mini-P1 consists of an origin, a gene for an initiator protein, RepA, and a control locus, incA. Both the origin and the incA locus contain repeat sequences that bind RepA. It has been proposed that the incA repeats control replication by sequestering the rate-limiting RepA initiator protein. Here we show that when the concentration of RepA was increased about fourfold beyond its normal physiological level from an inducible source in trans, the copy number of a ...

  6. Research Programs & Initiatives

    Science.gov (United States)

    CGH develops international initiatives and collaborates with other NCI divisions, NCI-designated Cancer Centers, and other countries to support cancer control planning, encourage capacity building, and support cancer research and research networks.

  7. Nursing Home Quality Initiative

    Data.gov (United States)

    U.S. Department of Health & Human Services — This Nursing Home Quality Initiative (NHQI) website provides consumer and provider information regarding the quality of care in nursing homes. NHQI discusses quality...

  8. Surgical Critical Care Initiative

    Data.gov (United States)

    Federal Laboratory Consortium — The Surgical Critical Care Initiative (SC2i) is a USU research program established in October 2013 to develop, translate, and validate biology-driven critical care....

  9. Global Methane Initiative

    Science.gov (United States)

    The Global Methane Initiative promotes cost-effective, near-term methane recovery through partnerships between developed and developing countries, with participation from the private sector, development banks, and nongovernmental organizations.

  10. Medical Errors Reduction Initiative

    National Research Council Canada - National Science Library

    Mutter, Michael L

    2005-01-01

    The Valley Hospital of Ridgewood, New Jersey, is proposing to extend a limited but highly successful specimen management and medication administration medical errors reduction initiative on a hospital-wide basis...

  11. RAS Initiative - Community Outreach

    Science.gov (United States)

    Through community and technical collaborations, workshops and symposia, and the distribution of reference reagents, the RAS Initiative seeks to increase the sharing of knowledge and resources essential to defeating cancers caused by mutant RAS genes.

  12. RAS Initiative - Events

    Science.gov (United States)

    The NCI RAS Initiative has organized multiple events with outside experts to discuss how the latest scientific and technological breakthroughs can be applied to discover vulnerabilities in RAS-driven cancers.

  13. PESP Landscaping Initiative

    Science.gov (United States)

    Landscaping practices can positively or negatively affect local environments and human health. The Landscaping Initiative seeks to enhance benefits of landscaping while reducing need for pesticides, fertilizers, etc., by working with partners.

  14. About the RAS Initiative

    Science.gov (United States)

    The RAS Initiative, a "hub and spoke" model, connects researchers to better understand and target the more than 30% of cancers driven by mutations in RAS genes. Includes oversight and contact information.

  15. The Yekaterinburg headache initiative

    DEFF Research Database (Denmark)

    Lebedeva, Elena R; Olesen, Jes; Osipova, Vera V

    2013-01-01

    for a demonstrational interventional project in Russia, undertaken within the Global Campaign against Headache. The initiative proposes three actions: 1) raise awareness of need for improvement; 2) design and implement a three-tier model (from primary care to a single highly specialized centre with academic affiliation......) for efficient and equitable delivery of headache-related health care; 3) develop a range of educational initiatives aimed at primary-care physicians, non-specialist neurologists, pharmacists and the general public to support the second action. RESULTS AND CONCLUSION: We set these proposals in a context...... of a health-care needs assessment, and as a model for all Russia. We present and discuss early progress of the initiative, justify the investment of resources required for implementation and call for the political support that full implementation requires. The more that the Yekaterinburg headache initiative...

  16. Piezoelectrically Initiated Pyrotechnic Igniter

    Science.gov (United States)

    Quince, Asia; Dutton, Maureen; Hicks, Robert; Burnham, Karen

    2013-01-01

    This innovation consists of a pyrotechnic initiator and piezoelectric initiation system. The device will be capable of being initiated mechanically; resisting initiation by EMF, RF, and EMI (electromagnetic field, radio frequency, and electromagnetic interference, respectively); and initiating in water environments and space environments. Current devices of this nature are initiated by the mechanical action of a firing pin against a primer. Primers historically are prone to failure. These failures are commonly known as misfires or hang-fires. In many cases, the primer shows the dent where the firing pin struck the primer, but the primer failed to fire. In devices such as "T" handles, which are commonly used to initiate the blowout of canopies, loss of function of the device may result in loss of crew. In devices such as flares or smoke generators, failure can result in failure to spot a downed pilot. The piezoelectrically initiated ignition system consists of a pyrotechnic device that plugs into a mechanical system (activator), which on activation, generates a high-voltage spark. The activator, when released, will strike a stack of electrically linked piezo crystals, generating a high-voltage, low-amperage current that is then conducted to the pyro-initiator. Within the initiator, an electrode releases a spark that passes through a pyrotechnic first-fire mixture, causing it to combust. The combustion of the first-fire initiates a primary pyrotechnic or explosive powder. If used in a "T" handle, the primary would ramp the speed of burn up to the speed of sound, generating a shock wave that would cause a high explosive to go "high order." In a flare or smoke generator, the secondary would produce the heat necessary to ignite the pyrotechnic mixture. The piezo activator subsystem is redundant in that a second stack of crystals would be struck at the same time with the same activation force, doubling the probability of a first strike spark generation. If the first

  17. Supply Chain Initiatives Database

    Energy Technology Data Exchange (ETDEWEB)

    None

    2012-11-01

    The Supply Chain Initiatives Database (SCID) presents innovative approaches to engaging industrial suppliers in efforts to save energy, increase productivity and improve environmental performance. This comprehensive and freely-accessible database was developed by the Institute for Industrial Productivity (IIP). IIP acknowledges Ecofys for their valuable contributions. The database contains case studies searchable according to the types of activities buyers are undertaking to motivate suppliers, target sector, organization leading the initiative, and program or partnership linkages.

  18. Interactive protein manipulation

    Energy Technology Data Exchange (ETDEWEB)

    SNCrivelli@lbl.gov

    2003-07-01

    We describe an interactive visualization and modeling program for the creation of protein structures ''from scratch''. The input to our program is an amino acid sequence -decoded from a gene- and a sequence of predicted secondary structure types for each amino acid-provided by external structure prediction programs. Our program can be used in the set-up phase of a protein structure prediction process; the structures created with it serve as input for a subsequent global internal energy minimization, or another method of protein structure prediction. Our program supports basic visualization methods for protein structures, interactive manipulation based on inverse kinematics, and visualization guides to aid a user in creating ''good'' initial structures.

  19. Interactive protein manipulation

    International Nuclear Information System (INIS)

    2003-01-01

    We describe an interactive visualization and modeling program for the creation of protein structures ''from scratch''. The input to our program is an amino acid sequence -decoded from a gene- and a sequence of predicted secondary structure types for each amino acid-provided by external structure prediction programs. Our program can be used in the set-up phase of a protein structure prediction process; the structures created with it serve as input for a subsequent global internal energy minimization, or another method of protein structure prediction. Our program supports basic visualization methods for protein structures, interactive manipulation based on inverse kinematics, and visualization guides to aid a user in creating ''good'' initial structures

  20. Total protein

    Science.gov (United States)

    ... page: //medlineplus.gov/ency/article/003483.htm Total protein To use the sharing features on this page, please enable JavaScript. The total protein test measures the total amount of two classes ...

  1. Proteins engineering

    International Nuclear Information System (INIS)

    2000-01-01

    At the - Departement d'Ingenierie et d'etudes de proteines (Deip) of the CEA more than seventy researchers are working hard to understand the function of proteins. For that they use the molecular labelling technique (F.M.)

  2. Whey Protein

    Science.gov (United States)

    ... reliable information about the safety of taking whey protein if you are pregnant or breast feeding. Stay on the safe side and avoid use. Milk allergy: If you are allergic to cow's milk, avoid using whey protein.

  3. Suplementação de lisina e metionina em dietas com baixo nível protéico para o crescimento inicial do pacu, Piaractus mesopotamicus (Holmberg Lysine and methionine supplementation in diets with low protein level for the initial growth of pacu Piaractus mesopotamicus (Holmberg

    Directory of Open Access Journals (Sweden)

    Adriana Patrícia Muñoz-Ramírez

    2002-04-01

    Full Text Available O objetivo deste trabalho foi estudar os efeitos da suplementação de metionina ou lisina em dietas com baixo teor protéico para o crescimento do pacu Piaractus mesopotamicus (Holmberg (Characiformes, Characidae. Foram formuladas uma dieta basal com 22% de proteína bruta (PB, 4100kcal de energia bruta (EB/kg, 0,42% de metionina e 1,16% de lisina e outras 6 dietas, com a mesma formulação básica, suplementadas com 0,2, 0,4 ou 0,6% de metionina ou lisina. Uma 8ª dieta (controle continha 26% PB, 4100kcal EB/kg, 0,48% metionina e 1,43% de lisina. As dietas foram administradas à vontade a 144 alevinos com 14,98 ± 1,16g de peso médio inicial. As médias de ganho em peso, eficiência de retenção de energia bruta e dos consumos alimentares da dieta controle mostraram-se maiores (P The objective of this research was to study the effects of methionine or lysine supplementation in diets with low protein level for growth of pacu Piaractus mesopotamicus (Holmberg (Characiformes, Characidae. Diets were formulated as a basal diet presenting 22% crude protein (CP, 4100 kcal gross energy (GE/kg, 0.42% of methionine and 1.16% of lysine and other six diets, with the same basic formulation, supplemented with 0.2%, 0.4% or 0.6% methionine or lysine. An eighth diet (control contained 26% CP, 4100 kcal (GE/kg, 0.48% methionine and 1.43% of lysine. The diets were administered ad libitum to 144 fingerlings with initial medium weight of 14.98 ± 1.16 g. Averages weight gain, gross energy efficiency retention and feed intake for the control treatment were significantly higher (P < 0.05 than those of smaller protein level diets. Averages protein retention efficiency were only higher (P < 0.01 in the diets supplemented with lysine, showing the advantages of lysine supplementation in diets with low crude protein level. A higher growth of pacu was confirmed with diets containing 26% of CP.

  4. Phosphorylation of the Bacillus subtilis Replication Controller YabA Plays a Role in Regulation of Sporulation and Biofilm Formation.

    Science.gov (United States)

    García García, Tránsito; Ventroux, Magali; Derouiche, Abderahmane; Bidnenko, Vladimir; Correia Santos, Sara; Henry, Céline; Mijakovic, Ivan; Noirot-Gros, Marie-Françoise; Poncet, Sandrine

    2018-01-01

    Bacillus subtilis cells can adopt different life-styles in response to various environmental cues, including planktonic cells during vegetative growth, sessile cells during biofilm formation and sporulation. While switching life-styles, bacteria must coordinate the progression of their cell cycle with their physiological status. Our current understanding of the regulatory pathways controlling the decision-making processes and triggering developmental switches highlights a key role of protein phosphorylation. The regulatory mechanisms that integrate the bacterial chromosome replication status with sporulation involve checkpoint proteins that target the replication initiator DnaA or the kinase phosphorelay controlling the master regulator Spo0A. B. subtilis YabA is known to interact with DnaA to prevent over-initiation of replication during vegetative growth. Here, we report that YabA is phosphorylated by YabT, a Ser/Thr kinase expressed during sporulation and biofilm formation. The phosphorylation of YabA has no effect on replication initiation control but hyper-phosphorylation of YabA leads to an increase in sporulation efficiency and a strong inhibition of biofilm formation. We also provide evidence that YabA phosphorylation affects the level of Spo0A-P in cells. These results indicate that YabA is a multifunctional protein with a dual role in regulating replication initiation and life-style switching, thereby providing a potential mechanism for cross-talk and coordination of cellular processes during adaptation to environmental change.

  5. Phosphorylation of the Bacillus subtilis Replication Controller YabA Plays a Role in Regulation of Sporulation and Biofilm Formation

    Directory of Open Access Journals (Sweden)

    Tránsito García García

    2018-03-01

    Full Text Available Bacillus subtilis cells can adopt different life-styles in response to various environmental cues, including planktonic cells during vegetative growth, sessile cells during biofilm formation and sporulation. While switching life-styles, bacteria must coordinate the progression of their cell cycle with their physiological status. Our current understanding of the regulatory pathways controlling the decision-making processes and triggering developmental switches highlights a key role of protein phosphorylation. The regulatory mechanisms that integrate the bacterial chromosome replication status with sporulation involve checkpoint proteins that target the replication initiator DnaA or the kinase phosphorelay controlling the master regulator Spo0A. B. subtilis YabA is known to interact with DnaA to prevent over-initiation of replication during vegetative growth. Here, we report that YabA is phosphorylated by YabT, a Ser/Thr kinase expressed during sporulation and biofilm formation. The phosphorylation of YabA has no effect on replication initiation control but hyper-phosphorylation of YabA leads to an increase in sporulation efficiency and a strong inhibition of biofilm formation. We also provide evidence that YabA phosphorylation affects the level of Spo0A-P in cells. These results indicate that YabA is a multifunctional protein with a dual role in regulating replication initiation and life-style switching, thereby providing a potential mechanism for cross-talk and coordination of cellular processes during adaptation to environmental change.

  6. Rv0004 is a new essential member of the mycobacterial DNA replication machinery.

    Science.gov (United States)

    Mann, Katherine M; Huang, Deborah L; Hooppaw, Anna J; Logsdon, Michelle M; Richardson, Kirill; Lee, Hark Joon; Kimmey, Jacqueline M; Aldridge, Bree B; Stallings, Christina L

    2017-11-01

    DNA replication is fundamental for life, yet a detailed understanding of bacterial DNA replication is limited outside the organisms Escherichia coli and Bacillus subtilis. Many bacteria, including mycobacteria, encode no identified homologs of helicase loaders or regulators of the initiator protein DnaA, despite these factors being essential for DNA replication in E. coli and B. subtilis. In this study we discover that a previously uncharacterized protein, Rv0004, from the human pathogen Mycobacterium tuberculosis is essential for bacterial viability and that depletion of Rv0004 leads to a block in cell cycle progression. Using a combination of genetic and biochemical approaches, we found that Rv0004 has a role in DNA replication, interacts with DNA and the replicative helicase DnaB, and affects DnaB-DnaA complex formation. We also identify a conserved domain in Rv0004 that is predicted to structurally resemble the N-terminal protein-protein interaction domain of DnaA. Mutation of a single conserved tryptophan within Rv0004's DnaA N-terminal-like domain leads to phenotypes similar to those observed upon Rv0004 depletion and can affect the association of Rv0004 with DnaB. In addition, using live cell imaging during depletion of Rv0004, we have uncovered a previously unappreciated role for DNA replication in coordinating mycobacterial cell division and cell size. Together, our data support that Rv0004 encodes a homolog of the recently identified DciA family of proteins found in most bacteria that lack the DnaC-DnaI helicase loaders in E. coli and B. subtilis. Therefore, the mechanisms of Rv0004 elucidated here likely apply to other DciA homologs and reveal insight into the diversity of bacterial strategies in even the most conserved biological processes.

  7. ProteinShop: A tool for interactive protein manipulation and steering

    Energy Technology Data Exchange (ETDEWEB)

    Crivelli, Silvia; Kreylos, Oliver; Max, Nelson; Hamann, Bernd; Bethel, Wes

    2004-05-25

    We describe ProteinShop, a new visualization tool that streamlines and simplifies the process of determining optimal protein folds. ProteinShop may be used at different stages of a protein structure prediction process. First, it can create protein configurations containing secondary structures specified by the user. Second, it can interactively manipulate protein fragments to achieve desired folds by adjusting the dihedral angles of selected coil regions using an Inverse Kinematics method. Last, it serves as a visual framework to monitor and steer a protein structure prediction process that may be running on a remote machine. ProteinShop was used to create initial configurations for a protein structure prediction method developed by a team that competed in CASP5. ProteinShop's use accelerated the process of generating initial configurations, reducing the time required from days to hours. This paper describes the structure of ProteinShop and discusses its main features.

  8. Initiation of Meiotic Recombination in Mammals

    Directory of Open Access Journals (Sweden)

    Rajeev Kumar

    2010-12-01

    Full Text Available Meiotic recombination is initiated by the induction of programmed DNA double strand breaks (DSBs. DSB repair promotes homologous interactions and pairing and leads to the formation of crossovers (COs, which are required for the proper reductional segregation at the first meiotic division. In mammals, several hundred DSBs are generated at the beginning of meiotic prophase by the catalytic activity of SPO11. Currently it is not well understood how the frequency and timing of DSB formation and their localization are regulated. Several approaches in humans and mice have provided an extensive description of the localization of initiation events based on CO mapping, leading to the identification and characterization of preferred sites (hotspots of initiation. This review presents the current knowledge about the proteins known to be involved in this process, the sites where initiation takes place, and the factors that control hotspot localization.

  9. International EUREKA: Initialization Segment

    International Nuclear Information System (INIS)

    1982-02-01

    The Initialization Segment creates the starting description of the uranium market. The starting description includes the international boundaries of trade, the geologic provinces, resources, reserves, production, uranium demand forecasts, and existing market transactions. The Initialization Segment is designed to accept information of various degrees of detail, depending on what is known about each region. It must transform this information into a specific data structure required by the Market Segment of the model, filling in gaps in the information through a predetermined sequence of defaults and built in assumptions. A principal function of the Initialization Segment is to create diagnostic messages indicating any inconsistencies in data and explaining which assumptions were used to organize the data base. This permits the user to manipulate the data base until such time the user is satisfied that all the assumptions used are reasonable and that any inconsistencies are resolved in a satisfactory manner

  10. Initiating events frequency determination

    International Nuclear Information System (INIS)

    Simic, Z.; Mikulicic, V.; Vukovic, I.

    2004-01-01

    The paper describes work performed for the Nuclear Power Station (NPS). Work is related to the periodic initiating events frequency update for the Probabilistic Safety Assessment (PSA). Data for all relevant NPS initiating events (IE) were reviewed. The main focus was on events occurring during most recent operating history (i.e., last four years). The final IE frequencies were estimated by incorporating both NPS experience and nuclear industry experience. Each event was categorized according to NPS individual plant examination (IPE) initiating events grouping approach. For the majority of the IE groups, few, or no events have occurred at the NPS. For those IE groups with few or no NPS events, the final estimate was made by means of a Bayesian update with general nuclear industry values. Exceptions are rare loss-of-coolant-accidents (LOCA) events, where evaluation of engineering aspects is used in order to determine frequency.(author)

  11. Dynamic assembly of Hda and the sliding clamp in the regulation of replication licensing

    OpenAIRE

    Kim, Jin S.; Nanfara, Michael T.; Chodavarapu, Sundari; Jin, Kyeong S.; Babu, Vignesh?M.?P.; Ghazy, Mohamed A.; Chung, Scisung; Kaguni, Jon M.; Sutton, Mark D.; Cho, Yunje

    2017-01-01

    Abstract Regulatory inactivation of DnaA (RIDA) is one of the major regulatory mechanisms of prokaryotic replication licensing. In RIDA, the Hda?sliding clamp complex loaded onto DNA directly interacts with adenosine triphosphate (ATP)-bound DnaA and stimulates the hydrolysis of ATP to inactivate DnaA. A prediction is that the activity of Hda is tightly controlled to ensure that replication initiation occurs only once per cell cycle. Here, we determined the crystal structure of the Hda?? clam...

  12. Suppressors of DnaAATP imposed overinitiation in Escherichia coli

    DEFF Research Database (Denmark)

    Charbon, Godefroid; Riber, Leise; Cohen, Malene

    2011-01-01

    Chromosome replication in Escherichia coli is limited by the supply of DnaA associated with ATP. Cells deficient in RIDA (Regulatory Inactivation of DnaA) due to a deletion of the hda gene accumulate suppressor mutations (hsm) to counteract the overinitiation caused by an elevated DnaAATP level....... Eight spontaneous hda suppressor mutations were identified by whole-genome sequencing, and three of these were analysed further. Two mutations (hsm-2 and hsm-4) mapped in the dnaA gene and led to a reduced ability to initiate replication from oriC. One mutation (hsm-1) mapped to the seqA promoter...

  13. RCRA facility stabilization initiative

    International Nuclear Information System (INIS)

    1995-02-01

    The RCRA Facility Stabilization Initiative was developed as a means of implementing the Corrective Action Program's management goals recommended by the RIS for stabilizing actual or imminent releases from solid waste management units that threaten human health and the environment. The overall goal of stabilization is to, as situations warrant, control or abate threats to human health and/or the environment from releases at RCRA facilities, and/or to prevent or minimize the further spread of contamination while long-term remedies are pursued. The Stabilization initiative is a management philosophy and should not be confused with stabilization technologies

  14. Predicting Translation Initiation Rates for Designing Synthetic Biology

    International Nuclear Information System (INIS)

    Reeve, Benjamin; Hargest, Thomas; Gilbert, Charlie; Ellis, Tom

    2014-01-01

    In synthetic biology, precise control over protein expression is required in order to construct functional biological systems. A core principle of the synthetic biology approach is a model-guided design and based on the biological understanding of the process, models of prokaryotic protein production have been described. Translation initiation rate is a rate-limiting step in protein production from mRNA and is dependent on the sequence of the 5′-untranslated region and the start of the coding sequence. Translation rate calculators are programs that estimate protein translation rates based on the sequence of these regions of an mRNA, and as protein expression is proportional to the rate of translation initiation, such calculators have been shown to give good approximations of protein expression levels. In this review, three currently available translation rate calculators developed for synthetic biology are considered, with limitations and possible future progress discussed.

  15. The SEED Initiative

    Science.gov (United States)

    Teich, Carolyn R.

    2011-01-01

    Committed to fulfilling the promise of the green economy, the American Association of Community Colleges (AACC) launched the Sustainability Education and Economic Development (SEED) initiative (www.theseedcenter.org) in October 2010. The project advances sustainability and clean energy workforce development practices at community colleges by…

  16. Major New Initiatives

    Indian Academy of Sciences (India)

    First page Back Continue Last page Overview Graphics. Major New Initiatives. Multi-party multi-rate video conferencing OOPS. Live Lecture OOPS. Rural ATM Machine Vortex. Finger print detection HP-IITM. Medical Diagnostic kit NeuroSynaptic. LCD projection system TeNeT. Web Terminal MeTeL Midas. Entertainment ...

  17. Mixed-Initiative Clustering

    Science.gov (United States)

    Huang, Yifen

    2010-01-01

    Mixed-initiative clustering is a task where a user and a machine work collaboratively to analyze a large set of documents. We hypothesize that a user and a machine can both learn better clustering models through enriched communication and interactive learning from each other. The first contribution or this thesis is providing a framework of…

  18. Monolithic exploding foil initiator

    Science.gov (United States)

    Welle, Eric J; Vianco, Paul T; Headley, Paul S; Jarrell, Jason A; Garrity, J. Emmett; Shelton, Keegan P; Marley, Stephen K

    2012-10-23

    A monolithic exploding foil initiator (EFI) or slapper detonator and the method for making the monolithic EFI wherein the exploding bridge and the dielectric from which the flyer will be generated are integrated directly onto the header. In some embodiments, the barrel is directly integrated directly onto the header.

  19. Next generation initiation techniques

    Science.gov (United States)

    Warner, Tom; Derber, John; Zupanski, Milija; Cohn, Steve; Verlinde, Hans

    1993-01-01

    Four-dimensional data assimilation strategies can generally be classified as either current or next generation, depending upon whether they are used operationally or not. Current-generation data-assimilation techniques are those that are presently used routinely in operational-forecasting or research applications. They can be classified into the following categories: intermittent assimilation, Newtonian relaxation, and physical initialization. It should be noted that these techniques are the subject of continued research, and their improvement will parallel the development of next generation techniques described by the other speakers. Next generation assimilation techniques are those that are under development but are not yet used operationally. Most of these procedures are derived from control theory or variational methods and primarily represent continuous assimilation approaches, in which the data and model dynamics are 'fitted' to each other in an optimal way. Another 'next generation' category is the initialization of convective-scale models. Intermittent assimilation systems use an objective analysis to combine all observations within a time window that is centered on the analysis time. Continuous first-generation assimilation systems are usually based on the Newtonian-relaxation or 'nudging' techniques. Physical initialization procedures generally involve the use of standard or nonstandard data to force some physical process in the model during an assimilation period. Under the topic of next-generation assimilation techniques, variational approaches are currently being actively developed. Variational approaches seek to minimize a cost or penalty function which measures a model's fit to observations, background fields and other imposed constraints. Alternatively, the Kalman filter technique, which is also under investigation as a data assimilation procedure for numerical weather prediction, can yield acceptable initial conditions for mesoscale models. The

  20. Detection of protein complex from protein-protein interaction network using Markov clustering

    International Nuclear Information System (INIS)

    Ochieng, P J; Kusuma, W A; Haryanto, T

    2017-01-01

    Detection of complexes, or groups of functionally related proteins, is an important challenge while analysing biological networks. However, existing algorithms to identify protein complexes are insufficient when applied to dense networks of experimentally derived interaction data. Therefore, we introduced a graph clustering method based on Markov clustering algorithm to identify protein complex within highly interconnected protein-protein interaction networks. Protein-protein interaction network was first constructed to develop geometrical network, the network was then partitioned using Markov clustering to detect protein complexes. The interest of the proposed method was illustrated by its application to Human Proteins associated to type II diabetes mellitus. Flow simulation of MCL algorithm was initially performed and topological properties of the resultant network were analysed for detection of the protein complex. The results indicated the proposed method successfully detect an overall of 34 complexes with 11 complexes consisting of overlapping modules and 20 non-overlapping modules. The major complex consisted of 102 proteins and 521 interactions with cluster modularity and density of 0.745 and 0.101 respectively. The comparison analysis revealed MCL out perform AP, MCODE and SCPS algorithms with high clustering coefficient (0.751) network density and modularity index (0.630). This demonstrated MCL was the most reliable and efficient graph clustering algorithm for detection of protein complexes from PPI networks. (paper)

  1. Initiative to manufacture and characterize Baculovirus Reference Material

    NARCIS (Netherlands)

    Kamen, A.; Aucoin, M.; Merten, O.W.; Alves, P.C.M.; Hashimoto, Y.; Airenne, K.; Hu, Y.C.; Mezzina, M.; Oers, van M.M.

    2011-01-01

    This letter to the editor brings to the attention of researchers an initiative to develop a baculovirus reference material repository. To be successful this initiative needs the support of a broad panel of researchers working with baculovirus vectors for recombinant protein production and gene

  2. Funding Initiatives | Women in Science | Initiatives | Indian Academy ...

    Indian Academy of Sciences (India)

    Home; Initiatives; Women in Science; Funding Initiatives ... The Fellowship Scheme for Women Scientists for societal programmes is initiative of the ... at a young age of 52, after a valiant battle with cancer, today on 29th March 2016 in Delhi.

  3. Sustaining Participatory Design Initiatives

    DEFF Research Database (Denmark)

    Iversen, Ole Sejer; Dindler, Christian

    2014-01-01

    While many participatory design (PD) projects succeed in establishing new organisational initiatives or creating technology that is attuned to the people affected, the issue of how such results are sustained after the project ends remains an important challenge. We explore the challenge...... these various forms of sustainability may be pursued in PD practice and how they can become a resource in reflecting on PD activities. Finally, we discuss implications for PD practice, suggesting that a nuanced conception of sustainability and how it may relate to PD practice are useful resources for designers...... of sustaining PD initiatives beyond the individual project and discuss implications for PD practice. First, based on current PD literature, we distinguish between four ideal typical forms of sustainability: maintaining, scaling, replicating and evolving. Second, we demonstrate from a case study how...

  4. Initial management of breastfeeding.

    Science.gov (United States)

    Sinusas, K; Gagliardi, A

    2001-09-15

    Breast milk is widely accepted as the ideal source of nutrition for infants. In order to ensure success in breastfeeding, it is important that it be initiated as early as possible during the neonatal period. This is facilitated by skin-to-skin contact between the mother and infant immediately following birth. When possible, the infant should be allowed to root and latch on spontaneously within the first hour of life. Many common nursery routines such as weighing the infant, administration of vitamin K and application of ocular antibiotics can be safely delayed until after the initial breastfeeding. Postpartum care practices that improve breastfeeding rates include rooming-in, anticipatory guidance about breastfeeding problems and the avoidance of formula supplementation and pacifiers.

  5. Self-initiated expatriates

    DEFF Research Database (Denmark)

    Selmer, Jan; Lauring, Jakob

    2014-01-01

    Purpose – As it has been suggested that adult third-culture kids may be more culturally adaptable than others, they have been labelled “the ideal” expatriates. In this article, we explore the adjustment of self-initiated expatriate academics in Hong Kong, comparing adult third-culture kids...... with adult mono-culture kids. Design/methodology/approach – We use survey results from 267 self-initiated expatriate academics in Hong Kong. Findings – Exploratory results show that adult third-culture kids had a higher extent of general adjustment. No significant results were found in relation...... to interaction adjustment and job adjustment. We also found that recent expatriate experiences generally had a positive association with the adjustment of adult mono-culture kids, but this association only existed in terms of general adjustment for adult third-culture kids. Originality/value – Once corroborated...

  6. INITIAL TRAINING OF RESEARCHERS

    Directory of Open Access Journals (Sweden)

    Karina Alejandra Cruz-Pallares

    2015-07-01

    Full Text Available The document presents results of a research that used as strategy a complementary training project with thirty-three students of a Bachelors Degree in Primary School 1997(DPS,1997 of an Education Faculty for the initial training of investigators, applied by four teachers members of the academic research group in Mexico; that develops through process of action research methodology. Highlighted in results is the strengthening of the competition of reading, understanding and writing scientific texts, which is analogous to the first feature of the graduate profile called intellectual skills. Among the conclusions it is emphasized that the initial training of teachers in a task that is quite interesting, challenging and complex, as is the educational complex phenomenon.

  7. Protein politics

    NARCIS (Netherlands)

    Vijver, Marike

    2005-01-01

    This study is part of the program of the interdisciplinary research group Profetas (protein foods, environment, technology and society). Profetas consists of technological, environmental and socio-economic research projects on protein food systems which result in the development of scenarios and

  8. Protein adhesives

    Science.gov (United States)

    Charles R. Frihart; Linda F. Lorenz

    2018-01-01

    Nature uses a wide variety of chemicals for providing adhesion internally (e.g., cell to cell) and externally (e.g., mussels to ships and piers). This adhesive bonding is chemically and mechanically complex, involving a variety of proteins, carbohydrates, and other compounds.Consequently,the effect of protein structures on adhesive properties is only partially...

  9. Tau protein

    DEFF Research Database (Denmark)

    Frederiksen, Jette Lautrup Battistini; Kristensen, Kim; Bahl, Jmc

    2011-01-01

    Background: Tau protein has been proposed as biomarker of axonal damage leading to irreversible neurological impairment in MS. CSF concentrations may be useful when determining risk of progression from ON to MS. Objective: To investigate the association between tau protein concentration and 14......-3-3 protein in the cerebrospinal fluid (CSF) of patients with monosymptomatic optic neuritis (ON) versus patients with monosymptomatic onset who progressed to multiple sclerosis (MS). To evaluate results against data found in a complete literature review. Methods: A total of 66 patients with MS and/or ON from...... the Department of Neurology of Glostrup Hospital, University of Copenhagen, Denmark, were included. CSF samples were analysed for tau protein and 14-3-3 protein, and clinical and paraclinical information was obtained from medical records. Results: The study shows a significantly increased concentration of tau...

  10. Characterization of redox proteins using electrochemical methods

    OpenAIRE

    Verhagen, M.

    1995-01-01

    The use of electrochemical techniques in combination with proteins started approximately a decade ago and has since then developed into a powerfull technique for the study of small redox proteins. In addition to the determination of redox potentials, electrochemistry can be used to obtain information about the kinetics of electron transfer between proteins and about the dynamic behaviour of redox cofactors in proteins. This thesis describes the results of a study, initiated to get a ...

  11. Differential Precipitation and Solubilization of Proteins.

    Science.gov (United States)

    Ryan, Barry J; Kinsella, Gemma K

    2017-01-01

    Differential protein precipitation is a rapid and economical step in protein purification and is based on exploiting the inherent physicochemical properties of the polypeptide. Precipitation of recombinant proteins, lysed from the host cell, is commonly used to concentrate the protein of choice before further polishing steps with more selective purification columns (e.g., His-Tag, Size Exclusion, etc.). Recombinant proteins can also precipitate naturally as inclusion bodies due to various influences during overexpression in the host cell. Although this phenomenon permits easier initial separation from native proteins, these inclusion bodies must carefully be differentially solubilized so as to reform functional, correctly folded proteins. Here, appropriate bioinformatics tools to aid in understanding a protein's propensity to aggregate and solubilize are explored as a backdrop for a typical protein extraction, precipitation, and selective resolubilization procedure, based on a recombinantly expressed protein.

  12. The Biophysics Microgravity Initiative

    Science.gov (United States)

    Gorti, S.

    2016-01-01

    Biophysical microgravity research on the International Space Station using biological materials has been ongoing for several decades. The well-documented substantive effects of long duration microgravity include the facilitation of the assembly of biological macromolecules into large structures, e.g., formation of large protein crystals under micro-gravity. NASA is invested not only in understanding the possible physical mechanisms of crystal growth, but also promoting two flight investigations to determine the influence of µ-gravity on protein crystal quality. In addition to crystal growth, flight investigations to determine the effects of shear on nucleation and subsequent formation of complex structures (e.g., crystals, fibrils, etc.) are also supported. It is now considered that long duration microgravity research aboard the ISS could also make possible the formation of large complex biological and biomimetic materials. Investigations of various materials undergoing complex structure formation in microgravity will not only strengthen NASA science programs, but may also provide invaluable insight towards the construction of large complex tissues, organs, or biomimetic materials on Earth.

  13. Hanford tanks initiative plan

    International Nuclear Information System (INIS)

    McKinney, K.E.

    1997-01-01

    Abstract: The Hanford Tanks Initiative (HTI) is a five-year project resulting from the technical and financial partnership of the U.S. Department of Energy's Office of Waste Management (EM-30) and Office of Science and Technology Development (EM-50). The HTI project accelerates activities to gain key technical, cost performance, and regulatory information on two high-level waste tanks. The HTI will provide a basis for design and regulatory decisions affecting the remainder of the Tank Waste Remediation System's tank waste retrieval Program

  14. UNLV Nuclear Hydrogen Initiative

    Energy Technology Data Exchange (ETDEWEB)

    Hechanova, Anthony E.; Johnson, Allen; O' Toole, Brendan; Trabia, Mohamed; Peterson, Per

    2012-10-25

    Evaluation of the Crack growth rate (CGR) of Alloy 617 and Alloy 276 under constant K at ambient temperature has been completed. Creep deformation of Alloy 230 at different temperature range and load level has been completed and heat to heat variation has been noticed. Creep deformation study of Alloy 276 has been completed under an applied initial stress level of 10% of yield stress at 950ºC. The grain size evaluation of the tested creep specimens of Alloy 276 has been completed.

  15. Feedback stabilization initiative

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1997-06-01

    Much progress has been made in attaining high confinement regimes in magnetic confinement devices. These operating modes tend to be transient, however, due to the onset of MHD instabilities, and their stabilization is critical for improved performance at steady state. This report describes the Feedback Stabilization Initiative (FSI), a broad-based, multi-institutional effort to develop and implement methods for raising the achievable plasma betas through active MHD feedback stabilization. A key element in this proposed effort is the Feedback Stabilization Experiment (FSX), a medium-sized, national facility that would be specifically dedicated to demonstrating beta improvement in reactor relevant plasmas by using a variety of MHD feedback stabilization schemes.

  16. Feedback stabilization initiative

    International Nuclear Information System (INIS)

    1997-06-01

    Much progress has been made in attaining high confinement regimes in magnetic confinement devices. These operating modes tend to be transient, however, due to the onset of MHD instabilities, and their stabilization is critical for improved performance at steady state. This report describes the Feedback Stabilization Initiative (FSI), a broad-based, multi-institutional effort to develop and implement methods for raising the achievable plasma betas through active MHD feedback stabilization. A key element in this proposed effort is the Feedback Stabilization Experiment (FSX), a medium-sized, national facility that would be specifically dedicated to demonstrating beta improvement in reactor relevant plasmas by using a variety of MHD feedback stabilization schemes

  17. Initiation of slug flow

    Energy Technology Data Exchange (ETDEWEB)

    Hanratty, T.J.; Woods, B.D. [Univ. of Illinois, Urbana, IL (United States)

    1995-12-31

    The initiation of slug flow in a horizontal pipe can be predicted either by considering the stability of a slug or by considering the stability of a stratified flow. Measurements of the shedding rate of slugs are used to define necessary conditions for the existence of a slug. Recent results show that slugs develop from an unstable stratified flow through the evolution of small wavelength waves into large wavelength waves that have the possibility of growing to form a slug. The mechanism appears to be quite different for fluids with viscosities close to water than for fluids with large viscosities (20 centipoise).

  18. Stirling to Flight Initiative

    Science.gov (United States)

    Hibbard, Kenneth E.; Mason, Lee S.; Ndu, Obi; Smith, Clayton; Withrow, James P.

    2016-01-01

    Flight (S2F) initiative with the objective of developing a 100-500 We Stirling generator system. Additionally, a different approach is being devised for this initiative to avoid pitfalls of the past, and apply lessons learned from the recent ASRG experience. Two key aspects of this initiative are a Stirling System Technology Maturation Effort, and a Surrogate Mission Team (SMT) intended to provide clear mission pull and requirements context. The S2F project seeks to lead directly into a DOE flight system development of a new SRG. This paper will detail the proposed S2F initiative, and provide specifics on the key efforts designed to pave a forward path for bringing Stirling technology to flight.

  19. Initiating statistical maintenance optimization

    International Nuclear Information System (INIS)

    Doyle, E. Kevin; Tuomi, Vesa; Rowley, Ian

    2007-01-01

    Since the 1980 s maintenance optimization has been centered around various formulations of Reliability Centered Maintenance (RCM). Several such optimization techniques have been implemented at the Bruce Nuclear Station. Further cost refinement of the Station preventive maintenance strategy includes evaluation of statistical optimization techniques. A review of successful pilot efforts in this direction is provided as well as initial work with graphical analysis. The present situation reguarding data sourcing, the principle impediment to use of stochastic methods in previous years, is discussed. The use of Crowe/AMSAA (Army Materials Systems Analysis Activity) plots is demonstrated from the point of view of justifying expenditures in optimization efforts. (author)

  20. The LHCb Starterkit initiative

    CERN Document Server

    Puig Navarro, Albert

    2017-01-01

    The vast majority of high-energy physicists use and produce software every day. Software skills are usually acquired on the go and dedicated training courses are rare. The LHCb Starterkit is a new training format for getting LHCb collaborators started in effectively using software to perform their research. The initiative, combining courses and online tutorials, focuses on teaching basic skills for research computing, as well as LHCb software specifics. Unlike traditional tutorials we focus on starting with basics, performing all the material live, with a high degree of interactivity, giving priority to understanding the tools as opposed to handing out recipes that work “as if by magic”. The LHCb Starterkit was started by young members of the collaboration inspired by the principles of Software Carpentry, and the material is created in a collaborative fashion using the tools we teach. Three successful entry-level workshops, as well as two advanced ones, have taken place since the start of the initiative i...

  1. Combat Wound Initiative program.

    Science.gov (United States)

    Stojadinovic, Alexander; Elster, Eric; Potter, Benjamin K; Davis, Thomas A; Tadaki, Doug K; Brown, Trevor S; Ahlers, Stephen; Attinger, Christopher E; Andersen, Romney C; Burris, David; Centeno, Jose; Champion, Hunter; Crumbley, David R; Denobile, John; Duga, Michael; Dunne, James R; Eberhardt, John; Ennis, William J; Forsberg, Jonathan A; Hawksworth, Jason; Helling, Thomas S; Lazarus, Gerald S; Milner, Stephen M; Mullick, Florabel G; Owner, Christopher R; Pasquina, Paul F; Patel, Chirag R; Peoples, George E; Nissan, Aviram; Ring, Michael; Sandberg, Glenn D; Schaden, Wolfgang; Schultz, Gregory S; Scofield, Tom; Shawen, Scott B; Sheppard, Forest R; Stannard, James P; Weina, Peter J; Zenilman, Jonathan M

    2010-07-01

    The Combat Wound Initiative (CWI) program is a collaborative, multidisciplinary, and interservice public-private partnership that provides personalized, state-of-the-art, and complex wound care via targeted clinical and translational research. The CWI uses a bench-to-bedside approach to translational research, including the rapid development of a human extracorporeal shock wave therapy (ESWT) study in complex wounds after establishing the potential efficacy, biologic mechanisms, and safety of this treatment modality in a murine model. Additional clinical trials include the prospective use of clinical data, serum and wound biomarkers, and wound gene expression profiles to predict wound healing/failure and additional clinical patient outcomes following combat-related trauma. These clinical research data are analyzed using machine-based learning algorithms to develop predictive treatment models to guide clinical decision-making. Future CWI directions include additional clinical trials and study centers and the refinement and deployment of our genetically driven, personalized medicine initiative to provide patient-specific care across multiple medical disciplines, with an emphasis on combat casualty care.

  2. Initiatives for proliferation prevention

    International Nuclear Information System (INIS)

    1997-04-01

    Preventing the proliferation of weapons of mass destruction is a central part of US national security policy. A principal instrument of the Department of Energy's (DOE's) program for securing weapons of mass destruction technology and expertise and removing incentives for scientists, engineers and technicians in the newly independent states (NIS) of the former Soviet Union to go to rogue countries or assist terrorist groups is the Initiatives for Proliferation Prevention (IPP). IPP was initiated pursuant to the 1994 Foreign Operations Appropriations Act. IPP is a nonproliferation program with a commercialization strategy. IPP seeks to enhance US national security and to achieve nonproliferation objectives by engaging scientists, engineers and technicians from former NIS weapons institutes; redirecting their activities in cooperatively-developed, commercially viable non-weapons related projects. These projects lead to commercial and economic benefits for both the NIS and the US IPP projects are funded in Russian, Ukraine, Kazakhstan and Belarus. This booklet offers an overview of the IPP program as well as a sampling of some of the projects which are currently underway

  3. Protein Adsorption in Three Dimensions

    Science.gov (United States)

    Vogler, Erwin A.

    2011-01-01

    initially-adsorbed protein. Interphase protein concentration CI increases as VI decreases, resulting in slow reduction in interfacial energetics. Steady-state is governed by a net partition coefficient P=(/CBCI). In the process of occupying space within the interphase, adsorbing protein molecules must displace an equivalent volume of interphase water. Interphase water is itself associated with surface-bound water through a network of transient hydrogen bonds. Displacement of interphase water thus requires an amount of energy that depends on the adsorbent surface chemistry/energy. This “adsorption-dehydration” step is the significant free-energy cost of adsorption that controls the maximum amount of protein that can be adsorbed at steady state to a unit adsorbent-surface area (the adsorbent capacity). As adsorbent hydrophilicity increases, protein adsorption monotonically decreases because the energetic cost of surface dehydration increases, ultimately leading to no protein adsorption near an adsorbent water wettability (surface energy) characterized by a water contact angle θ → 65°. Consequently, protein does not adsorb (accumulate at interphase concentrations greater than bulk solution) to more hydrophilic adsorbents exhibiting θ protein/surface interactions can be highly favorable, causing protein to adsorb in multilayers in a relatively thick interphase. A straightforward, three-component free energy relationship captures salient features of protein adsorption to all surfaces predicting that the overall free energy of protein adsorption ΔGadso is a relatively small multiple of thermal energy for any surface chemistry (except perhaps for bioengineered surfaces bearing specific ligands for adsorbing protein) because a surface chemistry that interacts chemically with proteins must also interact with water through hydrogen bonding. In this way, water moderates protein adsorption to any surface by competing with adsorbing protein molecules. This Leading Opinion ends

  4. Evaluation Program initiative

    International Nuclear Information System (INIS)

    Rich, B.L.

    1987-01-01

    The purpose of this paper is to provide the Department of Energy's (DOE) safeguards and security community with some insights on an important management initiative by the Office of Security Evaluations (OSE). The paper will present the ''what, where, who, when, and why'' of a new Evaluation Program. The Evaluation Program will be comprised of a continuing series of regular and special evaluations of DOE safeguards and security programs and policies. The evaluations will be integrative and ''crosscutting,'' i.e. will extend across DOE organizational lines. Evaluations will be offered as positive advisories to DOE managers with safeguards and security responsibilities and will not be rated. They will complement the ongoing OSE Inspection Program of inspections conducted by OSE's Inspection Division. The purpose for the evaluations is to establish an accurate and current assessment of the effectiveness and status of safeguards and security programs and policies and to provide DOE managers with required information on program and policy effectiveness

  5. Breckinridge Project, initial effort

    Energy Technology Data Exchange (ETDEWEB)

    None, None

    1982-09-01

    Report III, Volume 1 contains those specifications numbered A through J, as follows: General Specifications (A); Specifications for Pressure Vessels (C); Specifications for Tanks (D); Specifications for Exchangers (E); Specifications for Fired Heaters (F); Specifications for Pumps and Drivers (G); and Specifications for Instrumentation (J). The standard specifications of Bechtel Petroleum Incorporated have been amended as necessary to reflect the specific requirements of the Breckinridge Project, and the more stringent specifications of Ashland Synthetic Fuels, Inc. These standard specifications are available to the Initial Effort (Phase Zero) work performed by all contractors and subcontractors. Report III, Volume 1 also contains the unique specifications prepared for Plants 8, 15, and 27. These specifications will be substantially reviewed during Phase I of the project, and modified as necessary for use during the engineering, procurement, and construction of this project.

  6. The climate technology initiative

    Energy Technology Data Exchange (ETDEWEB)

    Smith, Adam [International Energy Agency, Climate Technology Initiative, Paris (France)

    2000-12-01

    The CTI (Climate Technology Initiative) aims to promote those technologies which cause the minimum of harm to the environment: reducing emissions of greenhouse gases and supporting those countries most vulnerable to climate change are priorities. A strong case for cogeneration is made and it is pointed out that both the European Union and the USA aim to double their cogeneration capacity by 2010. The CTI holds training courses and seminars all over the world where the barriers to the expansion of climate-friendly technology are discussed. The article also mentions the CTI Co-operation Technology Implementation Plan, research and development, its website and search engine, its presence at all UNFCCC events and its awards programme.

  7. The climate technology initiative

    International Nuclear Information System (INIS)

    Smith, Adam

    2000-01-01

    The CTI (Climate Technology Initiative) aims to promote those technologies which cause the minimum of harm to the environment: reducing emissions of greenhouse gases and supporting those countries most vulnerable to climate change are priorities. A strong case for cogeneration is made and it is pointed out that both the European Union and the USA aim to double their cogeneration capacity by 2010. The CTI holds training courses and seminars all over the world where the barriers to the expansion of climate-friendly technology are discussed. The article also mentions the CTI Co-operation Technology Implementation Plan, research and development, its website and search engine, its presence at all UNFCCC events and its awards programme

  8. Materials Genome Initiative

    Science.gov (United States)

    Vickers, John

    2015-01-01

    The Materials Genome Initiative (MGI) project element is a cross-Center effort that is focused on the integration of computational tools to simulate manufacturing processes and materials behavior. These computational simulations will be utilized to gain understanding of processes and materials behavior to accelerate process development and certification to more efficiently integrate new materials in existing NASA projects and to lead to the design of new materials for improved performance. This NASA effort looks to collaborate with efforts at other government agencies and universities working under the national MGI. MGI plans to develop integrated computational/experimental/ processing methodologies for accelerating discovery and insertion of materials to satisfy NASA's unique mission demands. The challenges include validated design tools that incorporate materials properties, processes, and design requirements; and materials process control to rapidly mature emerging manufacturing methods and develop certified manufacturing processes

  9. The Ombudperson Initiative Group

    CERN Multimedia

    Laura Stewart

    Following many discussions that took place at some of the ATLAS Women's Network lunch gatherings, a few ATLAS women joined forces with similarly concerned CERN staff women to form a small group last Fall to discuss the need for a CERN-wide Ombudsperson. This has since evolved into the Ombudsperson Initiative Group (OIG) currently composed of the following members: Barbro Asman, Stockholm University; Pierre Charrue, CERN AB; Anna Cook, CERN IT; Catherine Delamare, CERN and IT Ombudsperson; Paula Eerola, Lund University; Pauline Gagnon, Indiana University; Eugenia Hatziangeli, CERN AB; Doreen Klem, CERN IT; Bertrand Nicquevert, CERN TS and Laura Stewart, CERN AT. On June 12, members of the OIG met with representatives of Human Resources (HR) and the Equal Opportunity Advisory Panel (EOAP) to discuss the proposal drafted by the OIG. The meeting was very positive. Everybody agreed that the current procedures at CERN applicable in the event of conflict required a thorough review, and that a professionnally trai...

  10. Instrumented Pipeline Initiative

    Energy Technology Data Exchange (ETDEWEB)

    Thomas Piro; Michael Ream

    2010-07-31

    This report summarizes technical progress achieved during the cooperative agreement between Concurrent Technologies Corporation (CTC) and U.S. Department of Energy to address the need for a for low-cost monitoring and inspection sensor system as identified in the Department of Energy (DOE) National Gas Infrastructure Research & Development (R&D) Delivery Reliability Program Roadmap.. The Instrumented Pipeline Initiative (IPI) achieved the objective by researching technologies for the monitoring of pipeline delivery integrity, through a ubiquitous network of sensors and controllers to detect and diagnose incipient defects, leaks, and failures. This report is organized by tasks as detailed in the Statement of Project Objectives (SOPO). The sections all state the objective and approach before detailing results of work.

  11. Green Power Initiative

    Energy Technology Data Exchange (ETDEWEB)

    Butler, Patrick Barry [Univ. of Iowa, Iowa City, IA (United States)

    2013-01-28

    National energy policy supports the gathering of more detailed and authoritative data on the introduction of renewable bio-based fuels into new and existing district energy systems via the application of biomass gasification. The University of Iowa developed a biomass-fueled, university-scale steam generation system based on biomass gasification technologies. The system serves as a state-of-the-art research and educational facility in the emerging application of gasification in steam generation. The facility, which includes a smaller down-draft gasifier and a larger multi-stage biomass boiler, was designed to operate primarily on wood-based fuels, but has provisions for testing other biomass fuel sources produced within a 100-mile radius, providing enough flexibility to meet the fluctuating local supply of biomass from industry and Midwest agriculture. The equipment was installed in an existing, staffed facility. The down-draft gasifier unit is operated by College of Engineering staff and students, under the direct technical supervision of qualified Utilities plant staff. The Green Power Initiative also includes a substantial, innovative educational component. In addition to an onsite, graduate-level research program in biomass fuels, the investigators have integrated undergraduate and graduate level teaching – through classroom studies and experiential learning – and applied research into a biomass-based, university-scale, functioning power plant. University of Iowa is unique in that it currently has multiple renewable energy technologies deployed, including significant biomass combustion (oat hulls) at its Main Power Plant and a new reciprocating engine based renewable district energy system. This project complements and supports the national energy policy and State of Iowa initiatives in ethanol and biodiesel. Byproducts of ethanol and biodiesel processes (distiller grains) as well as industry residues (oat hulls, wood chips, construction and demolition

  12. Insulin accelerates global and mitochondrial protein synthesis rates in neonatal muscle during sepsis

    Science.gov (United States)

    In neonatal pigs, sepsis decreases protein synthesis in skeletal muscle by decreasing translation initiation. However, insulin stimulates muscle protein synthesis despite persistent repression of translation initiation signaling. To determine whether the insulin-induced increase in global rates of m...

  13. Chronological protein synthesis in regenerating rat liver.

    Science.gov (United States)

    He, Jinjun; Hao, Shuai; Zhang, Hao; Guo, Fuzheng; Huang, Lingyun; Xiao, Xueyuan; He, Dacheng

    2015-07-01

    Liver regeneration has been studied for decades; however, its regulation remains unclear. In this study, we report a dynamic tracing of protein synthesis in rat regenerating liver with a new proteomic technique, (35) S in vivo labeling analysis for dynamic proteomics (SiLAD). Conventional proteomic techniques typically measure protein alteration in accumulated amounts. The SiLAD technique specifically detects protein synthesis velocity instead of accumulated amounts of protein through (35) S pulse labeling of newly synthesized proteins, providing a direct way for analyzing protein synthesis variations. Consequently, protein synthesis within short as 30 min was visualized and protein regulations in the first 8 h of regenerating liver were dynamically traced. Further, the 3.5-5 h post partial hepatectomy (PHx) was shown to be an important regulatory turning point by acute regulation of many proteins in the initiation of liver regeneration. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Initial Egyptian ECMO experience

    Directory of Open Access Journals (Sweden)

    Akram Abdelbary

    2016-04-01

    Results: A total of twelve patients received ECMO between January 2014 and June 2015. The mean age was 35.9 years. (range 13–65 years, 8 males, with VV ECMO in 10 patients, and VA ECMO in 2 patients. Out of ten patients of VV ECMO, one had H1N1 pneumonia, one had advanced vasculitic lung, four had bacterial pneumonia, two traumatic lung contusions and one with organophosphorus poisoning, and one undiagnosed etiology leading to severe ARDS. Lung injury score range was 3–3.8, PaO2/FiO2 (20–76 mechanical ventilation duration before ECMO 1–14 days, Femoro-jugular cannulation in 7 patients and femoro-femoral in 2 patients and femoro-subclavian in 1 patient; all patients were initially sedated and paralyzed for (2–4 days and ventilated on pressure controlled ventilation with Pmax of 25 cm H2O and PEEP of 10 cm H2O. In VA ECMO patients were cannulated percutaneously using femoro-femoral approach. One patient showed no neurologic recovery and died after 24 h, the other had CABG on ECMO however the heart didn’t recover and died after 9 days. Heparin intravenous infusion was used initially in all patients and changed to Bivalirudin in 2 patients due to possible HIT. Pump flow ranged from 2.6 to 6.5 L/min. Average support time was 12 days (range 2–24 days. Seven patients (63.3% were successfully separated from ECMO and survived to hospital discharge. Hospital length of stay ranged from 3 to 42 days, tracheostomy was done percutaneously in 5 patients and surgically in 3. Gastrointestinal bleeding occurred in 6 patients, VAP in 7 patients, neurologic complications in 1 patient with complete recovery, cardiac arrhythmias in 3 patients, pneumothorax in 9 patients, and deep venous thrombosis in 2 patients.

  15. TFTR initial operations

    International Nuclear Information System (INIS)

    Young, K.M.; Bell, M.; Blanchard, W.R.

    1984-01-01

    TFTR (Tokamak Fusion Test Reactor) has operated since December 1982 with ohmically heated plasmas. Routine operation with feedback control of plasma current, position and density has been obtained for plasmas with Isub(p) approx.= 800 kA, a = 68 cm, R = 250 cm, and Bsub(t) = 27 kG. A maximum plasma current of 1 MA was achieved with q approx.= 2.5. Energy confinement times of approx. 150 msec were measured for hydrogen and deuterium plasmas with n-barsub(e) approx.= 2 x 10 13 cm -3 , Tsub(e)(O) approx.= 1.5 keV, Tsub(i)(O) approx.= 1.5 keV and Zsub(eff) approx.= 3. The preliminary results suggest a size-cubed scaling from PLT, and are consistent with Alcator C scaling where tau approx. nR 2 a. Initial measurements of plasma disruption characteristics indicate current decay rates of approx. 800 kA in 8 ms which is within the TFTR design requirement of 3 MA in 3 ms. (author)

  16. The new childcare initiative

    CERN Multimedia

    Cigdem Issever

    The ATLAS Women's Network recently sent out a general mailing to all ATLAS and CMS members to announce a new initiative aimed at improving childcare facilities for Users coming to CERN. Several people have expressed the need that CERN should provide or facilitate affordable day care for children of temporary visitors at CERN. The ATLAS Women's Network is now forming a child care task force from concerned people and invites all those interested to join this effort. You can do so by either adding your name to the mailing list cern-users-childcare@cern.ch in Simba or by contacting Cigdem.Issever@cern.NOSPAM.ch and Pauline.Gagnon@cern.NOSPAM.ch. More than 50 people have already joined this effort. Those who have joined the mailing list will soon receive all the details about the next conference call meeting which has been scheduled for Thursday October 25th from 16:30 to 18:00 CERN time. The preliminary agenda is the following: Summary of our first contact of ATLAS and CMS (5 min) Discussion about the co-conv...

  17. AECL's new environmental initiatives

    International Nuclear Information System (INIS)

    McDonnell, F.N.

    1993-01-01

    AECL's research and development expenditures in environmental sciences and waste management technology are about $50 M per year. The main focus of these programs is the Nuclear Fuel Waste Management Program. This research is supplemented by activities in support of laboratory, Environmental Authority and internal waste management requirements, as well as provision of non-nuclear services. AECL intends to become more involved in performing environmental research and development with broader application. The goal is to achieve a relationship with Canadian industry that would involve a substantial portion of AECL's environmental research capabilities. The research directions and priorities of the resulting partnership would be set by the private sector in accordance with their needs and requirements. It is expected that the activities associated with this new environmental initiative will start small and grow in response to perceived needs. AECL is now increasing its non-nuclear research efforts by targeting those markets that appear most attractive. The thrust can be divided into three broad categories: environmental research, environmental services, and environmental products. (Author)

  18. TFTR initial operations

    International Nuclear Information System (INIS)

    Young, K.M.; Bell, M.; Blanchard, W.R.

    1983-11-01

    The Tokamak Fusion Test Reactor (TFTR) has operated since December 1982 with ohmically heated plasmas. Routine operation with feedback control of plasma current, position, and density has been obtained for plasmas with I/sub p/ approx. = 800 kA, a = 68 cm, R = 250 cm, and B/sub t/ = 27 kG. A maximum plasma current of 1 MA was achieved with q approx. = 2.5. Energy confinement times of approx. 150 msec were measured for hydrogen and deuterium plasmas with anti n/sub e/ approx. = 2 x 10 13 cm -3 , T/sub e/ (0) approx. = 1.5 keV, T/sub i/ (0) approx. = 1.5 keV, and Z/sub eff/ approx. = 3. The preliminary results suggest a size-cubed scaling from PLT and are consistent with Alcator C scaling where tau approx. nR 2 a. Initial measurements of plasma disruption characteristics indicate current decay rates of approx. 800 kA in 8 ms which is within the TFTR design requirement of 3 MA in 3 ms

  19. Multiple regulatory systems coordinate DNA replication with cell growth in Bacillus subtilis.

    Directory of Open Access Journals (Sweden)

    Heath Murray

    2014-10-01

    Full Text Available In many bacteria the rate of DNA replication is linked with cellular physiology to ensure that genome duplication is coordinated with growth. Nutrient-mediated growth rate control of DNA replication initiation has been appreciated for decades, however the mechanism(s that connects these cell cycle activities has eluded understanding. In order to help address this fundamental question we have investigated regulation of DNA replication in the model organism Bacillus subtilis. Contrary to the prevailing view we find that changes in DnaA protein level are not sufficient to account for nutrient-mediated growth rate control of DNA replication initiation, although this regulation does require both DnaA and the endogenous replication origin. We go on to report connections between DNA replication and several essential cellular activities required for rapid bacterial growth, including respiration, central carbon metabolism, fatty acid synthesis, phospholipid synthesis, and protein synthesis. Unexpectedly, the results indicate that multiple regulatory systems are involved in coordinating DNA replication with cell physiology, with some of the regulatory systems targeting oriC while others act in a oriC-independent manner. We propose that distinct regulatory systems are utilized to control DNA replication in response to diverse physiological and chemical changes.

  20. Multiple regulatory systems coordinate DNA replication with cell growth in Bacillus subtilis.

    Science.gov (United States)

    Murray, Heath; Koh, Alan

    2014-10-01

    In many bacteria the rate of DNA replication is linked with cellular physiology to ensure that genome duplication is coordinated with growth. Nutrient-mediated growth rate control of DNA replication initiation has been appreciated for decades, however the mechanism(s) that connects these cell cycle activities has eluded understanding. In order to help address this fundamental question we have investigated regulation of DNA replication in the model organism Bacillus subtilis. Contrary to the prevailing view we find that changes in DnaA protein level are not sufficient to account for nutrient-mediated growth rate control of DNA replication initiation, although this regulation does require both DnaA and the endogenous replication origin. We go on to report connections between DNA replication and several essential cellular activities required for rapid bacterial growth, including respiration, central carbon metabolism, fatty acid synthesis, phospholipid synthesis, and protein synthesis. Unexpectedly, the results indicate that multiple regulatory systems are involved in coordinating DNA replication with cell physiology, with some of the regulatory systems targeting oriC while others act in a oriC-independent manner. We propose that distinct regulatory systems are utilized to control DNA replication in response to diverse physiological and chemical changes.

  1. Multiple Regulatory Systems Coordinate DNA Replication with Cell Growth in Bacillus subtilis

    Science.gov (United States)

    Murray, Heath; Koh, Alan

    2014-01-01

    In many bacteria the rate of DNA replication is linked with cellular physiology to ensure that genome duplication is coordinated with growth. Nutrient-mediated growth rate control of DNA replication initiation has been appreciated for decades, however the mechanism(s) that connects these cell cycle activities has eluded understanding. In order to help address this fundamental question we have investigated regulation of DNA replication in the model organism Bacillus subtilis. Contrary to the prevailing view we find that changes in DnaA protein level are not sufficient to account for nutrient-mediated growth rate control of DNA replication initiation, although this regulation does require both DnaA and the endogenous replication origin. We go on to report connections between DNA replication and several essential cellular activities required for rapid bacterial growth, including respiration, central carbon metabolism, fatty acid synthesis, phospholipid synthesis, and protein synthesis. Unexpectedly, the results indicate that multiple regulatory systems are involved in coordinating DNA replication with cell physiology, with some of the regulatory systems targeting oriC while others act in a oriC-independent manner. We propose that distinct regulatory systems are utilized to control DNA replication in response to diverse physiological and chemical changes. PMID:25340815

  2. European nuclear education initiatives

    International Nuclear Information System (INIS)

    Glatz, Jean-Paul

    2011-01-01

    Whatever option regarding their future nuclear energy development is chosen by European Union Member States, the availability of a sufficient number of well trained and experienced staff is key for the responsible use of nuclear energy. This is true in all areas including design, construction, operation, decommissioning, fuel cycle and waste management as well as radiation protection. Given the high average age of existing experts leading to a significant retirement induce a real risk of the loss of nuclear competencies in the coming years. Therefore the demand of hiring skilled employees is rising. The challenge of ensuring a sufficient number of qualified staff in the nuclear sector has been acknowledged widely among the different stakeholders, in particular the nuclear industry, national regulatory authorities and Technical Support Organisations (TSOs). Already the EURATOM Treaty refers explicitly to the obligation for the Commission to carry out training actions. Recently initiatives have been launched at EU level to facilitate and strengthen the efforts of national stakeholders. The European Nuclear Education Network (ENEN) Association aims at preservation and further development of expertise in the nuclear field by higher education and training. The goal of the European Nuclear Energy Leadership Academy (ENELA) is to educate future leaders in the nuclear field to ensure the further development of sustainable European nuclear energy solutions The European Nuclear Energy Forum (ENEF) is a platform operated by the European Commission for a broad discussion on the opportunities and risks of nuclear energy. The nuclear programs under investigation in the Joint Research Center (JRC) are increasingly contributing to Education and Training (E and T) initiatives, promoting a better cooperation between key players and universities as well as operators and regulatory bodies in order to mutually optimise their training programmes. Another objective is to increase

  3. The MEGAPIE Initiative

    International Nuclear Information System (INIS)

    Salvatores, M.; Bauer, G.S.; Heusener, G.

    2000-10-01

    MEGAPIE (Megawatt Pilot Experiment) is a joint initiative by Commissariat a l'Energie Atomique (CEA), France, Forschungszentrum Karlsruhe (FZK), Germany, and Paul Scherrer Institut (PSI), Switzerland, to design, build, operate and explore a liquid lead-bismuth spallation target for 1MW of beam power, taking advantage of the existing spallation neutron facility SINQ at PSI. Such a target based on an eutectic mixture with a melting point as low as 125 o C and a boiling point as high as 1670 o C is the preferred concept in several studies aiming at utilising accelerators to drive subcritical assemblies in order to transmute long lived nuclear waste into shorter lived isotopes in an effort to ease problems of long term storage and final disposal. MEGAPIE will be an essential step towards demonstrating the feasibility of the coupling of a high power accelerator, a spallation target and a subcritical assembly. It will specifically address one of the most critical issues, namely the behaviour of a liquid metal target under realistic operating conditions. As an intensely instrumented pilot experiment it will provide valuable data for benchmarking of frequently used computer codes and will allow to gain important experience in the safe handling of components that have been irradiated with PbBi. It will be installed at the ring cyclotron at PSI with 590 MeV proton energy and a continuous current of 1.8 mA. The basic concept of the MEGAPIE target as well as the definition of the project phases and of the supporting research and development activities at the participating laboratories are described in the present report

  4. Initial Cladding Condition

    International Nuclear Information System (INIS)

    Siegmann, E.

    2000-01-01

    The purpose of this analysis is to describe the condition of commercial Zircaloy clad fuel as it is received at the Yucca Mountain Project (YMP) site. Most commercial nuclear fuel is encased in Zircaloy cladding. This analysis is developed to describe cladding degradation from the expected failure modes. This includes reactor operation impacts including incipient failures, potential degradation after reactor operation during spent fuel storage in pool and dry storage and impacts due to transportation. Degradation modes include cladding creep, and delayed hydride cracking during dry storage and transportation. Mechanical stresses from fuel handling and transportation vibrations are also included. This Analysis and Model Report (AMR) does not address any potential damage to assemblies that might occur at the YMP surface facilities. Ranges and uncertainties have been defined. This analysis will be the initial boundary condition for the analysis of cladding degradation inside the repository. In accordance with AP-2.13Q, ''Technical Product Development Planning'', a work plan (CRWMS M andO 2000c) was developed, issued, and utilized in the preparation of this document. There are constraints, caveats and limitations to this analysis. This cladding degradation analysis is based on commercial Pressurized Water Reactor (PWR) fuel with Zircaloy cladding but is applicable to Boiling Water Reactor (BWR) fuel. Reactor operating experience for both PWRs and BWRs is used to establish fuel reliability from reactor operation. It is limited to fuel exposed to normal operation and anticipated operational occurrences (i.e. events which are anticipated to occur within a reactor lifetime), and not to fuel that has been exposed to severe accidents. Fuel burnup projections have been limited to the current commercial reactor licensing environment with restrictions on fuel enrichment, oxide coating thickness and rod plenum pressures. The information provided in this analysis will be used in

  5. Hazardous material reduction initiative

    International Nuclear Information System (INIS)

    Nichols, D.H.

    1995-02-01

    The Hazardous Material Reduction Initiative (HMRI) explores using the review of purchase requisitions to reduce both the use of hazardous materials and the generation of regulated and nonregulated wastes. Based on an 11-month program implemented at the Hanford Site, hazardous material use and waste generation was effectively reduced by using a centralized procurement control program known as HMRI. As expected, several changes to the original proposal were needed during the development/testing phase of the program to accommodate changing and actual conditions found at the Hanford Site. The current method requires a central receiving point within the Procurement Organization to review all purchase requisitions for potentially Occupational Safety and Health Administration (OSHA) hazardous products. Those requisitions (approximately 4% to 6% of the total) are then forwarded to Pollution Prevention personnel for evaluation under HMRI. The first step is to determine if the requested item can be filled by existing or surplus material. The requisitions that cannot filled by existing or surplus material are then sorted into two groups based on applicability to the HMRI project. For example, laboratory requests for analytical reagents or standards are excluded and the purchase requisitions are returned to Procurement for normal processing because, although regulated, there is little opportunity for source reduction due to the strict protocols followed. Each item is then checked to determine if it is regulated or not. Regulated items are prioritized based on hazardous contents, quantity requested, and end use. Copies of these requisitions are made and the originals are returned to Procurement within 1-hr. Since changes to the requisition can be made at later stages during procurement, the HMRI fulfills one of its original premises in that it does not slow the procurement process

  6. Nursing Facility Initiative Annual Report

    Data.gov (United States)

    U.S. Department of Health & Human Services — This annual report summarizes impacts from the Initiative to Reduce Avoidable Hospitalizations among Nursing Facility Residents in 2014. This initiative is designed...

  7. National Take-Back Initiative

    Science.gov (United States)

    ... Disposal Information Drug and Chemical Information E-commerce Initiatives Federal Agencies & Related Links Federal Register Notices National ... Disposal Information Drug and Chemical Information E-commerce Initiatives Federal Agencies & Related Links Federal Register Notices National ...

  8. The national geomagnetic initiative

    Science.gov (United States)

    1993-01-01

    The Earth's magnetic field, through its variability over a spectrum of spatial and temporal scales, contains fundamental information on the solid Earth and geospace environment (the latter comprising the atmosphere, ionosphere, and magnetosphere). Integrated studies of the geomagnetic field have the potential to address a wide range of important processes in the deep mantle and core, asthenosphere, lithosphere, oceans, and the solar-terrestrial environment. These studies have direct applications to important societal problems, including resource assessment and exploration, natural hazard mitigation, safe navigation, and the maintenance and survivability of communications and power systems on the ground and in space. Studies of the Earth's magnetic field are supported by a variety of federal and state agencies as well as by private industry. Both basic and applied research is presently supported by several federal agencies, including the National Science Foundation (NSF), U.S. Geological Survey (USGS), U.S. Department of Energy (DOE), National Oceanic and Atmospheric Administration (NOAA), National Aeronautics and Space Administration (NASA), and U.S. Department of Defense (DOD) (through the Navy, Air Force, and Defense Mapping Agency). Although each agency has a unique, well-defined mission in geomagnetic studies, many areas of interest overlap. For example, NASA, the Navy, and USGS collaborate closely in the development of main field reference models. NASA, NSF, and the Air Force collaborate in space physics. These interagency linkages need to be strengthened. Over the past decade, new opportunities for fundamental advances in geomagnetic research have emerged as a result of three factors: well-posed, first-order scientific questions; increased interrelation of research activities dealing with geomagnetic phenomena; and recent developments in technology. These new opportunities can be exploited through a national geomagnetic initiative to define objectives and

  9. Initial Radionuclide Inventories

    Energy Technology Data Exchange (ETDEWEB)

    H. Miller

    2004-09-19

    The purpose of this analysis is to provide an initial radionuclide inventory (in grams per waste package) and associated uncertainty distributions for use in the Total System Performance Assessment for the License Application (TSPA-LA) in support of the license application for the repository at Yucca Mountain, Nevada. This document is intended for use in postclosure analysis only. Bounding waste stream information and data were collected that capture probable limits. For commercially generated waste, this analysis considers alternative waste stream projections to bound the characteristics of wastes likely to be encountered using arrival scenarios that potentially impact the commercial spent nuclear fuel (CSNF) waste stream. For TSPA-LA, this radionuclide inventory analysis considers U.S. Department of Energy (DOE) high-level radioactive waste (DHLW) glass and two types of spent nuclear fuel (SNF): CSNF and DOE-owned (DSNF). These wastes are placed in two groups of waste packages: the CSNF waste package and the codisposal waste package (CDSP), which are designated to contain DHLW glass and DSNF, or DHLW glass only. The radionuclide inventory for naval SNF is provided separately in the classified ''Naval Nuclear Propulsion Program Technical Support Document'' for the License Application. As noted previously, the radionuclide inventory data presented here is intended only for TSPA-LA postclosure calculations. It is not applicable to preclosure safety calculations. Safe storage, transportation, and ultimate disposal of these wastes require safety analyses to support the design and licensing of repository equipment and facilities. These analyses will require radionuclide inventories to represent the radioactive source term that must be accommodated during handling, storage and disposition of these wastes. This analysis uses the best available information to identify the radionuclide inventory that is expected at the last year of last emplacement

  10. Citizen participation and citizen initiatives

    International Nuclear Information System (INIS)

    Matthoefer, H.

    1977-01-01

    Contents: Social conditions for citizen initiatives - technical change and employment - crisis behaviour - socio-psychological analysis of political planning; legitimation - presentation and criticism - conditions for citizen initiatives coming into being within the field of tension citizen : administration - legal problems of citizen initiatives - environmental protection in the energy discussion; participation; models. (HP) [de

  11. NLM Emergency Access Initiative: FAQs

    Science.gov (United States)

    Facebook Visit us on Twitter Visit us on Youtube Emergency Access Initiative Home | Journals | Books | Online Databases | FAQs Take Short Survey FAQ What is the Emergency Access Initiative? The Emergency Access Initiative (EAI) is a collaborative partnership between NLM and participating publishers to

  12. Protein nanoparticles for therapeutic protein delivery.

    Science.gov (United States)

    Herrera Estrada, L P; Champion, J A

    2015-06-01

    Therapeutic proteins can face substantial challenges to their activity, requiring protein modification or use of a delivery vehicle. Nanoparticles can significantly enhance delivery of encapsulated cargo, but traditional small molecule carriers have some limitations in their use for protein delivery. Nanoparticles made from protein have been proposed as alternative carriers and have benefits specific to therapeutic protein delivery. This review describes protein nanoparticles made by self-assembly, including protein cages, protein polymers, and charged or amphipathic peptides, and by desolvation. It presents particle fabrication and delivery characterization for a variety of therapeutic and model proteins, as well as comparison of the features of different protein nanoparticles.

  13. Protein supplementation with sports protein bars in renal patients.

    Science.gov (United States)

    Meade, Anthony

    2007-05-01

    Malnutrition prevalence in patients on dialysis is well established. The protein requirements for both hemodialysis and peritoneal dialysis have been documented elsewhere, including the Kidney Disease Outcomes Quality Initiative Clinical Practice Guidelines for Nutrition in Chronic Renal Failure. The clinical challenge is to assist patients in meeting these targets, especially in those with anorexia. Traditional supplements have included fluid, which is an issue for patients who are fluid restricted. The study objectives were to (1) investigate the range of sports protein supplements that may be suitable for patients on hemodialysis to use and (2) trial nonfluid protein supplements in patients on hemodialysis. Known manufacturers of sports protein bars and other sports supplements available in Australia were contacted for the nutrient breakdown of high-protein products, specifically potassium, protein, and phosphorus contents. As a result, selected high-protein sports bars (Protein FX, Aussie Bodies, Port Melbourne, Victoria, Australia) were used as an alternative to the more commonly used renal-specific fluid supplements (Nepro, Abbott Laboratories, Abbott Park, IL; Novasource Renal, Novartis Nutrition Corporation, Fremont, MI; and Renilon, Nutricia, Wiltshire, UK) in patients with poor nutritional status requiring supplementation. Patient satisfaction and clinical nutrition markers were investigated. The study took place at inpatient, in-center, and satellite hemodialysis settings in Adelaide, South Australia. A total of 32 patients (16 females and 16 males) with an average age of 62.9 years (range 32-86 years) undergoing hemodialysis (acute and maintenance) were included. Subjects were selected by the author as part of routine clinical nutrition care. Patients trialed sports protein bars as a protein supplement alone or in conjunction with other supplementary products. All patients were in favor of the trial, with 22 of 32 patients continuing with the protein

  14. Community Rates of Breastfeeding Initiation.

    Science.gov (United States)

    Grubesic, Tony H; Durbin, Kelly M

    2016-11-01

    Breastfeeding initiation rates vary considerably across racial and ethnic groups, maternal age, and education level, yet there are limited data concerning the influence of geography on community rates of breastfeeding initiation. This study aimed to describe how community rates of breastfeeding initiation vary in geographic space, highlighting "hot spots" and "cool spots" of initiation and exploring the potential connections between race, socioeconomic status, and urbanization levels on these patterns. Birth certificate data from the Kentucky Department of Health for 2004-2010 were combined with county-level geographic base files, Census 2010 demographic and socioeconomic data, and Rural-Urban Continuum Codes to conduct a spatial statistical analysis of community rates of breastfeeding initiation. Between 2004 and 2010, the average rate of breastfeeding initiation for Kentucky increased from 43.84% to 49.22%. Simultaneously, the number of counties identified as breastfeeding initiation hot spots also increased, displaying a systematic geographic pattern in doing so. Cool spots of breastfeeding initiation persisted in rural, Appalachian Kentucky. Spatial regression results suggested that unemployment, income, race, education, location, and the availability of International Board Certified Lactation Consultants are connected to breastfeeding initiation. Not only do spatial analytics facilitate the identification of breastfeeding initiation hot spots and cool spots, but they can be used to better understand the landscape of breastfeeding initiation and help target breastfeeding education and/or support efforts.

  15. Protein-Protein Interaction Databases

    DEFF Research Database (Denmark)

    Szklarczyk, Damian; Jensen, Lars Juhl

    2015-01-01

    Years of meticulous curation of scientific literature and increasingly reliable computational predictions have resulted in creation of vast databases of protein interaction data. Over the years, these repositories have become a basic framework in which experiments are analyzed and new directions...

  16. Florida Hydrogen Initiative

    Energy Technology Data Exchange (ETDEWEB)

    Block, David L

    2013-06-30

    The Florida Hydrogen Initiative (FHI) was a research, development and demonstration hydrogen and fuel cell program. The FHI program objectives were to develop Florida?s hydrogen and fuel cell infrastructure and to assist DOE in its hydrogen and fuel cell activities The FHI program funded 12 RD&D projects as follows: Hydrogen Refueling Infrastructure and Rental Car Strategies -- L. Lines, Rollins College This project analyzes strategies for Florida's early stage adaptation of hydrogen-powered public transportation. In particular, the report investigates urban and statewide network of refueling stations and the feasibility of establishing a hydrogen rental-car fleet based in Orlando. Methanol Fuel Cell Vehicle Charging Station at Florida Atlantic University ? M. Fuchs, EnerFuel, Inc. The project objectives were to design, and demonstrate a 10 kWnet proton exchange membrane fuel cell stationary power plant operating on methanol, to achieve an electrical energy efficiency of 32% and to demonstrate transient response time of less than 3 milliseconds. Assessment of Public Understanding of the Hydrogen Economy Through Science Center Exhibits, J. Newman, Orlando Science Center The project objective was to design and build an interactive Science Center exhibit called: ?H2Now: the Great Hydrogen Xchange?. On-site Reformation of Diesel Fuel for Hydrogen Fueling Station Applications ? A. Raissi, Florida Solar Energy Center This project developed an on-demand forecourt hydrogen production technology by catalytically converting high-sulfur hydrocarbon fuels to an essentially sulfur-free gas. The removal of sulfur from reformate is critical since most catalysts used for the steam reformation have limited sulfur tolerance. Chemochromic Hydrogen Leak Detectors for Safety Monitoring ? N. Mohajeri and N. Muradov, Florida Solar Energy Center This project developed and demonstrated a cost-effective and highly selective chemochromic (visual) hydrogen leak detector for safety

  17. Initiation of DNA replication: functional and evolutionary aspects

    Science.gov (United States)

    Bryant, John A.; Aves, Stephen J.

    2011-01-01

    Background The initiation of DNA replication is a very important and highly regulated step in the cell division cycle. It is of interest to compare different groups of eukaryotic organisms (a) to identify the essential molecular events that occur in all eukaryotes, (b) to start to identify higher-level regulatory mechanisms that are specific to particular groups and (c) to gain insights into the evolution of initiation mechanisms. Scope This review features a wide-ranging literature survey covering replication origins, origin recognition and usage, modification of origin usage (especially in response to plant hormones), assembly of the pre-replication complex, loading of the replisome, genomics, and the likely origin of these mechanisms and proteins in Archaea. Conclusions In all eukaryotes, chromatin is organized for DNA replication as multiple replicons. In each replicon, replication is initiated at an origin. With the exception of those in budding yeast, replication origins, including the only one to be isolated so far from a plant, do not appear to embody a specific sequence; rather, they are AT-rich, with short tracts of locally bent DNA. The proteins involved in initiation are remarkably similar across the range of eukaryotes. Nevertheless, their activity may be modified by plant-specific mechanisms, including regulation by plant hormones. The molecular features of initiation are seen in a much simpler form in the Archaea. In particular, where eukaryotes possess a number of closely related proteins that form ‘hetero-complexes’ (such as the origin recognition complex and the MCM complex), archaeans typically possess one type of protein (e.g. one MCM) that forms a homo-complex. This suggests that several eukaryotic initiation proteins have evolved from archaeal ancestors by gene duplication and divergence. PMID:21508040

  18. Dynamic assembly of Hda and the sliding clamp in the regulation of replication licensing.

    Science.gov (United States)

    Kim, Jin S; Nanfara, Michael T; Chodavarapu, Sundari; Jin, Kyeong S; Babu, Vignesh M P; Ghazy, Mohamed A; Chung, Scisung; Kaguni, Jon M; Sutton, Mark D; Cho, Yunje

    2017-04-20

    Regulatory inactivation of DnaA (RIDA) is one of the major regulatory mechanisms of prokaryotic replication licensing. In RIDA, the Hda-sliding clamp complex loaded onto DNA directly interacts with adenosine triphosphate (ATP)-bound DnaA and stimulates the hydrolysis of ATP to inactivate DnaA. A prediction is that the activity of Hda is tightly controlled to ensure that replication initiation occurs only once per cell cycle. Here, we determined the crystal structure of the Hda-β clamp complex. This complex contains two pairs of Hda dimers sandwiched between two β clamp rings to form an octamer that is stabilized by three discrete interfaces. Two separate surfaces of Hda make contact with the β clamp, which is essential for Hda function in RIDA. The third interface between Hda monomers occludes the active site arginine finger, blocking its access to DnaA. Taken together, our structural and mutational analyses of the Hda-β clamp complex indicate that the interaction of the β clamp with Hda controls the ability of Hda to interact with DnaA. In the octameric Hda-β clamp complex, the inability of Hda to interact with DnaA is a novel mechanism that may regulate Hda function. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  19. Inflation with generalized initial conditions

    International Nuclear Information System (INIS)

    Albrecht, A.; Brandenberger, R.; Matzner, R.

    1987-01-01

    In many current models of the early Universe a scalar field phi which is only very weakly coupled to other quantum fields is used to generate inflation. In such models there are no forces which could thermalize the scalar field, and previous assumptions about its preinflation ''initial'' conditions must be abandoned. In this paper the onset of inflation is studied classically for more general initial conditions of the scalar field configuration. In particular, initial conditions with a nonvanishing spatial average of phi, with phi chosen at random in each initial horizon volume, and with random initial momenta are considered. We identify and discuss several mechanisms that can drive these more general initial conditions toward an inflationary state. The analysis is done in one spatial dimension

  20. EXPERIENCES WITH IDEA PROMOTING INITIATIVES

    DEFF Research Database (Denmark)

    Gish, Liv

    2011-01-01

    In new product development a central activity is to provide new ideas. Over the last decades experiences with stimulating employee creativity and establishing idea promoting initiatives have been made in industrial practice. Such initiatives are often labeled Idea Management – a research field...... with a growing interest. In this paper I examine three different idea promoting initiatives carried out in Grundfos, a leading pump manufacturer. In the analysis I address what understandings of idea work are inscribed in the initiatives and what role these initiatives play in the organization with respect...... understandings of idea work are inscribed in the idea promoting initiatives as they to some degree have to fit with the understandings embedded in practice in order to work....

  1. DNA replication initiator Cdc6 also regulates ribosomal DNA transcription initiation.

    Science.gov (United States)

    Huang, Shijiao; Xu, Xiaowei; Wang, Guopeng; Lu, Guoliang; Xie, Wenbing; Tao, Wei; Zhang, Hongyin; Jiang, Qing; Zhang, Chuanmao

    2016-04-01

    RNA-polymerase-I-dependent ribosomal DNA (rDNA) transcription is fundamental to rRNA processing, ribosome assembly and protein synthesis. However, how this process is initiated during the cell cycle is not fully understood. By performing a proteomic analysis of transcription factors that bind RNA polymerase I during rDNA transcription initiation, we identified that the DNA replication initiator Cdc6 interacts with RNA polymerase I and its co-factors, and promotes rDNA transcription in G1 phase in an ATPase-activity-dependent manner. We further showed that Cdc6 is targeted to the nucleolus during late mitosis and G1 phase in a manner that is dependent on B23 (also known as nucleophosmin, NPM1), and preferentially binds to the rDNA promoter through its ATP-binding domain. Overexpression of Cdc6 increases rDNA transcription, whereas knockdown of Cdc6 results in a decreased association of both RNA polymerase I and the RNA polymerase I transcription factor RRN3 with rDNA, and a reduction of rDNA transcription. Furthermore, depletion of Cdc6 impairs the interaction between RRN3 and RNA polymerase I. Taken together, our data demonstrate that Cdc6 also serves as a regulator of rDNA transcription initiation, and indicate a mechanism by which initiation of rDNA transcription and DNA replication can be coordinated in cells. © 2016. Published by The Company of Biologists Ltd.

  2. DnaC inactivation in Escherichia coli K-12 induces the SOS response and expression of nucleotide biosynthesis genes

    DEFF Research Database (Denmark)

    Løbner-Olesen, Anders; Slominska-Wojewodzka, Monika; Hansen, Flemming G.

    2008-01-01

    Background: Initiation of chromosome replication in E. coli requires the DnaA and DnaC proteins and conditionally-lethal dnaA and dnaC mutants are often used to synchronize cell populations. Methodology/Principal Findings: DNA microarrays were used to measure mRNA steady-state levels in initiatio......C genes was increased at the non-permissive temperature in the respective mutant strains indicating auto-regulation of both genes. Induction of the SOS regulon was observed in dnaC2 cells at 38 degrees C and 42 degrees C. Flow cytometric analysis revealed that dnaC2 mutant cells at non......-permissive temperature had completed the early stages of chromosome replication initiation. Conclusion/Significance: We suggest that in dnaC2 cells the SOS response is triggered by persistent open-complex formation at oriC and/or by arrested forks that require DnaC for replication restart....

  3. Initiation into Adolescent Marijuana Use.

    Science.gov (United States)

    Brook, Judith S.; And Others

    1980-01-01

    This longitudinal study examined the relationship of three domains (personality/attitudinal orientations, peer relationships, and family socialization factors) with initiation into adolescent marihuana use. (Author/DB)

  4. Aquaporin Protein-Protein Interactions

    Directory of Open Access Journals (Sweden)

    Jennifer Virginia Roche

    2017-10-01

    Full Text Available Aquaporins are tetrameric membrane-bound channels that facilitate transport of water and other small solutes across cell membranes. In eukaryotes, they are frequently regulated by gating or trafficking, allowing for the cell to control membrane permeability in a specific manner. Protein–protein interactions play crucial roles in both regulatory processes and also mediate alternative functions such as cell adhesion. In this review, we summarize recent knowledge about aquaporin protein–protein interactions; dividing the interactions into three types: (1 interactions between aquaporin tetramers; (2 interactions between aquaporin monomers within a tetramer (hetero-tetramerization; and (3 transient interactions with regulatory proteins. We particularly focus on the structural aspects of the interactions, discussing the small differences within a conserved overall fold that allow for aquaporins to be differentially regulated in an organism-, tissue- and trigger-specific manner. A deep knowledge about these differences is needed to fully understand aquaporin function and regulation in many physiological processes, and may enable design of compounds targeting specific aquaporins for treatment of human disease.

  5. Origins of Protein Functions in Cells

    Science.gov (United States)

    Seelig, Burchard; Pohorille, Andrzej

    2011-01-01

    In modern organisms proteins perform a majority of cellular functions, such as chemical catalysis, energy transduction and transport of material across cell walls. Although great strides have been made towards understanding protein evolution, a meaningful extrapolation from contemporary proteins to their earliest ancestors is virtually impossible. In an alternative approach, the origin of water-soluble proteins was probed through the synthesis and in vitro evolution of very large libraries of random amino acid sequences. In combination with computer modeling and simulations, these experiments allow us to address a number of fundamental questions about the origins of proteins. Can functionality emerge from random sequences of proteins? How did the initial repertoire of functional proteins diversify to facilitate new functions? Did this diversification proceed primarily through drawing novel functionalities from random sequences or through evolution of already existing proto-enzymes? Did protein evolution start from a pool of proteins defined by a frozen accident and other collections of proteins could start a different evolutionary pathway? Although we do not have definitive answers to these questions yet, important clues have been uncovered. In one example (Keefe and Szostak, 2001), novel ATP binding proteins were identified that appear to be unrelated in both sequence and structure to any known ATP binding proteins. One of these proteins was subsequently redesigned computationally to bind GTP through introducing several mutations that introduce targeted structural changes to the protein, improve its binding to guanine and prevent water from accessing the active center. This study facilitates further investigations of individual evolutionary steps that lead to a change of function in primordial proteins. In a second study (Seelig and Szostak, 2007), novel enzymes were generated that can join two pieces of RNA in a reaction for which no natural enzymes are known

  6. Protein immobilization strategies for protein biochips

    NARCIS (Netherlands)

    Rusmini, F.; Rusmini, Federica; Zhong, Zhiyuan; Feijen, Jan

    2007-01-01

    In the past few years, protein biochips have emerged as promising proteomic and diagnostic tools for obtaining information about protein functions and interactions. Important technological innovations have been made. However, considerable development is still required, especially regarding protein

  7. Effects of Whey, Caseinate, or Milk Protein Ingestion on Muscle Protein Synthesis after Exercise.

    Science.gov (United States)

    Kanda, Atsushi; Nakayama, Kyosuke; Sanbongi, Chiaki; Nagata, Masashi; Ikegami, Shuji; Itoh, Hiroyuki

    2016-06-03

    Whey protein (WP) is characterized as a "fast" protein and caseinate (CA) as a "slow" protein according to their digestion and absorption rates. We hypothesized that co-ingestion of milk proteins (WP and CA) may be effective for prolonging the muscle protein synthesis response compared to either protein alone. We therefore compared the effect of ingesting milk protein (MP) to either WP or CA alone on muscle protein synthesis after exercise in rats. We also compared the effects of these milk-derived proteins to a control, soy protein (SP). Male Sprague-Dawley rats swam for two hours. Immediately after exercise, one of the following four solutions was administered: WP, CA, MP, or SP. Individual rats were euthanized at designated postprandial time points and triceps muscle samples collected for measurement of the protein fractional synthesis rate (FSR). FSR tended to increase in all groups post-ingestion, although the initial peaks of FSR occurred at different times (WP, peak time = 60 min, FSR = 7.76%/day; MP, peak time = 90 min, FSR = 8.34%/day; CA, peak time = 120 min, FSR = 7.85%/day). Milk-derived proteins caused significantly greater increases (p protein synthesis to occur at different times (WP, fast; MP, intermediate; CA, slow) and the dairy proteins have a superior effect on muscle protein synthesis after exercise compared with SP.

  8. The clinical usefulness of initial serum procalcitonin as an ...

    African Journals Online (AJOL)

    The serum levels of WBC counts and C‑reactive protein in the aggravation group were elevated. However, the median value (interquartile range) of procalcitonin was relatively increased at 2.28 (0.41–7.84 ng/ml), demonstrating a significant difference. Conclusions: In conclusion, initial serum levels of procalcitonin might be ...

  9. Student initiative: A conceptual analysis

    Directory of Open Access Journals (Sweden)

    Polovina Nada

    2014-01-01

    Full Text Available In the description and scientific consideration of the attitude of children and youth towards their education and development, the concept of student initiative has been gaining ground lately, and it is hence the subject of analysis in this paper. The analysis is important because of the discrepancy between the increased efforts of the key educational policy holders to promote the idea about the importance of the development of student initiative and rare acceptance of this idea among theoreticians, researchers and practitioners dealing with the education and development of children and youth. By concretising the features of initiative student behaviour, our aim was, on the one hand, to observe the structural determinants and scientific status of the very concept of an initiative student, and, on the other, to contribute to the understanding of the initiative behaviour in practice. In the first part of the paper we deal with different notions and concretisations of the features of initiative behaviour of children and youth, which includes the consideration of: basic student initiative, academic student initiative, individual student initiative, the capacity for initiative and personal development initiative. In the second part of the paper, we discuss the relations of the concept of student initiative with the similar general concepts (activity/passivity, proactivity, agency and the concepts immediately related to school environment (student involvement, student participation. The results of our analysis indicate that the concept of student initiative has: particular features that differentiate it from similar concepts; the potential to reach the status of a scientific concept, bearing in mind the initial empirical specifications and general empirical verifiability of the yet unverified determinants of the concept. In the concluding part of the paper, we discuss the implications of the conceptual analysis for further research, as well as for

  10. Proliferative signaling initiated in ACTH receptors

    Directory of Open Access Journals (Sweden)

    C.F.P. Lotfi

    2000-10-01

    Full Text Available This article reviews recent results of studies aiming to elucidate modes of integrating signals initiated in ACTH receptors and FGF2 receptors, within the network system of signal transduction found in Y1 adrenocortical cells. These modes of signal integration should be central to the mechanisms underlying the regulation of the G0->G1->S transition in the adrenal cell cycle. FGF2 elicits a strong mitogenic response in G0/G1-arrested Y1 adrenocortical cells, that includes a rapid and transient activation of extracellular signal-regulated kinases-mitogen-activated protein kinases (ERK-MAPK (2 to 10 min, b transcription activation of c-fos, c-jun and c-myc genes (10 to 30 min, c induction of c-Fos and c-Myc proteins by 1 h and cyclin D1 protein by 5 h, and d onset of DNA synthesis stimulation within 8 h. ACTH, itself a weak mitogen, interacts with FGF2 in a complex manner, blocking the FGF2 mitogenic response during the early and middle G1 phase, keeping ERK-MAPK activation and c-Fos and cyclin D1 induction at maximal levels, but post-transcriptionally inhibiting c-Myc expression. c-Fos and c-Jun proteins are mediators in both the strong and the weak mitogenic responses respectively triggered by FGF2 and ACTH. Induction of c-Fos and stimulation of DNA synthesis by ACTH are independent of PKA and are inhibited by the PKC inhibitor GF109203X. In addition, ACTH is a poor activator of ERK-MAPK, but c-Fos induction and DNA synthesis stimulation by ACTH are strongly inhibited by the inhibitor of MEK1 PD98059.

  11. Career Technical Education Pathways Initiative

    Science.gov (United States)

    California Community Colleges, Chancellor's Office, 2013

    2013-01-01

    California's education system--the largest in the United States--is an essential resource for ensuring strong economic growth in the state. The Career Technical Education Pathways Initiative (referred to as the Initiative in this report), which became law in 2005, brings together community colleges, K-12 school districts, employers, organized…

  12. The metabolomics standards initiative (MSI)

    NARCIS (Netherlands)

    Fiehn, O.; Robertson, D.; Griffin, J.; Werf, M. van der; Nikolau, B.; Morrison, N.; Sumner, L.W.; Goodacre, R.; Hardy, N.W.; Taylor, C.; Fostel, J.; Kristal, B.; Kaddurah-Daouk, R.; Mendes, P.; Ommen, B. van; Lindon, J.C.; Sansone, S.-A.

    2007-01-01

    In 2005, the Metabolomics Standards Initiative has been formed. An outline and general introduction is provided to inform about the history, structure, working plan and intentions of this initiative. Comments on any of the suggested minimal reporting standards are welcome to be sent to the open

  13. Behavioral Initiatives in Broad Perspective.

    Science.gov (United States)

    California Univ., Los Angeles. Center for Mental Health in Schools.

    This booklet is a technical assistance sampler addressing the issues of student misbehavior, discipline problems, and behavioral initiatives. The term behavioral initiative is defined, disciplining children with disabilities is discussed, and a cautionary note concerning ignoring students' reasons for misbehavior is presented. A brief entitled…

  14. Local initiative extrapolated to nation

    DEFF Research Database (Denmark)

    Wittchen, Kim Bjarne; Kragh, Jesper; Brøgger, Morten

    In the municipality of Sønderborg, in the southern part of Jutland, there is a shining example initiated in 2007, ProjectZero, of a local initiative that have resulted in extensive energy savings in residential buildings and at the same time created local workplaces. The intension with the pilot...

  15. Northern Eurasia Future Initiative (NEFI)

    DEFF Research Database (Denmark)

    Groisman, Pavel; Shugart, Herman; Kicklighter, David

    2017-01-01

    . The Northern Eurasia Future Initiative (NEFI) has been designed as an essential continuation of the Northern Eurasia Earth Science Partnership Initiative (NEESPI), which was launched in 2004. NEESPI sought to elucidate all aspects of ongoing environmental change, to inform societies and, thus, to better...

  16. Comprehensive School Safety Initiative Report

    Science.gov (United States)

    National Institute of Justice, 2014

    2014-01-01

    The National Institute of Justice (NIJ) developed the Comprehensive School Safety Initiative in consultation with federal partners and Congress. It is a research-focused initiative designed to increase the safety of schools nationwide through the development of knowledge regarding the most effective and sustainable school safety interventions and…

  17. Initial conditions for chaotic inflation

    International Nuclear Information System (INIS)

    Brandenberger, R.; Kung, J.; Feldman, H.

    1991-01-01

    In contrast to many other inflationary Universe models, chaotic inflation does not depend on fine tuning initial conditions. Within the context of linear perturbation theory, it is shown that chaotic inflation is stable towards both metric and matter perturbations. Neglecting gravitational perturbations, it is shown that chaotic inflation is an attractor in initial condition space. (orig.)

  18. Self-initiated expatriate academics

    DEFF Research Database (Denmark)

    Selmer, Jan; Lauring, Jakob

    2013-01-01

    In this chapter we examine self-initiated expatriate academics. Universities are to an increasing extent looking for talent beyond national boundaries. Accordingly, self-initiated expatriate academics represent a fast growing group of highly educated professionals who gain employment abroad...

  19. Synaptogenic proteins and synaptic organizers: "many hands make light work".

    Science.gov (United States)

    Brose, Nils

    2009-03-12

    Synaptogenesis is thought to be mediated by cell adhesion proteins, which induce the initial contact between an axon and its target cell and subsequently recruit and organize the presynaptic and postsynaptic protein machinery required for synaptic transmission. A new study by Linhoff and colleagues in this issue of Neuron identifies adhesion proteins of the LRRTM family as novel synaptic organizers.

  20. Defense Acquisition Initiatives Review: An Assessment of Extant Initiatives

    National Research Council Canada - National Science Library

    Porter, Gene; Berteau, David; Christle, Gary; Mandelbaum, Jay; Diehl, Richard

    2005-01-01

    ...) to identify and analyze a subset of initiatives that the team finds to have potential for near term management emphasis that could provide visible improvements to the much criticized Defense acquisition system...

  1. A method for investigating protein-protein interactions related to Salmonella typhimurium pathogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Chowdhury, Saiful M. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Shi, Liang [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Yoon, Hyunjin [Dartmouth College, Hanover, NH (United States); Ansong, Charles [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Rommereim, Leah M. [Dartmouth College, Hanover, NH (United States); Norbeck, Angela D. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Auberry, Kenneth J. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Moore, R. J. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Adkins, Joshua N. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Heffron, Fred [Oregon Health and Science Univ., Portland, OR (United States); Smith, Richard D. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States)

    2009-02-10

    We successfully modified an existing method to investigate protein-protein interactions in the pathogenic bacterium Salmonella typhimurium (STM). This method includes i) addition of a histidine-biotin-histidine tag to the bait proteins via recombinant DNA techniques; ii) in vivo cross-linking with formaldehyde; iii) tandem affinity purification of bait proteins under fully denaturing conditions; and iv) identification of the proteins cross-linked to the bait proteins by liquid-chromatography in conjunction with tandem mass-spectrometry. In vivo cross-linking stabilized protein interactions permitted the subsequent two-step purification step conducted under denaturing conditions. The two-step purification greatly reduced nonspecific binding of non-cross-linked proteins to bait proteins. Two different negative controls were employed to reduce false-positive identification. In an initial demonstration of this approach, we tagged three selected STM proteins- HimD, PduB and PhoP- with known binding partners that ranged from stable (e.g., HimD) to transient (i.e., PhoP). Distinct sets of interacting proteins were identified with each bait protein, including the known binding partners such as HimA for HimD, as well as anticipated and unexpected binding partners. Our results suggest that novel protein-protein interactions may be critical to pathogenesis by Salmonella typhimurium. .

  2. Initial Assessment of Whiplash Patients

    Directory of Open Access Journals (Sweden)

    R Gunzburg

    2003-01-01

    Full Text Available The article looks at how for severe trauma, the outcome of treatment depends on the initial medical care. This has now also been accepted for whiplash associated disorders, underlining the importance of a proper initial assessment. Once major injury has been excluded and the diagnosis of whiplash associated disorder has been established, the initial treatment of whiplash in the emergency room can be started. The four key points to remember are described, including reassuring the patient about evolution, no soft collar, nonsteroidal anti-inflammatory drugs and early mobilisation.

  3. Canada's family violence initiative: partnerships

    Directory of Open Access Journals (Sweden)

    Elaine Scott

    1994-01-01

    Full Text Available Under Canada's four-year, $136 million Family Violence Initiative, the federal government is calling upon all Canadians to work in partnerships towards the elimination of family violence - child abuse, violence against women, and elder (senior abuse. Family violence is a complex problem and requires the efforts of all Canadians to resolve it. One of the key themes of the Initiative - a multidisciplinary approach to the problem of family violence - is reflected in the selection and development of projects. Activities funded by the seven federal departments and agencies involved in the Initiative emphasize partnerships with the professional, voluntary, corporate, non-government and government sectors.

  4. Learning from the Ilulissat Initiative

    DEFF Research Database (Denmark)

    Rahbek-Clemmensen, Jon; Thomasen, Gry

    In May 2018, ten years will have passed since the representatives from the five Arctic coastal states (Canada, Denmark, Norway, Russia) and the Home Rule government of Greenland met in Ilulissat. Learning from the Ilulissat Initiative examines how the initiative came about, and how it has affected...... strong economic interests in maintaining peaceful Arctic relations. Northern forums therefore give policymakers a rare opportunity to meet, communicate, and influence a key region. The Ilulissat meeting was the result of a joint Danish‒Greenlandic initiative and is often hailed as one of the most...

  5. New protection initiatives announced for coral reefs

    Science.gov (United States)

    Showstack, Randy

    Off the coasts of some of the South Pacific's most idyllic-sounding atolls, Austin Bowden-Kerby has seen first-hand the heavy damage to coral reefs from dynamite and cyanide fishing. For instance, while snorkeling near Chuuk, an island in Micronesia, he has observed craters and rubble beds of coral, which locals have told him date to World War II ordnance.A marine biologist and project scientist for the Coral Gardens Initiative of the Foundation for the Peoples of the South Pacific, Bowden-Kerby has also identified what he says are some public health effects related to destroyed coral reefs and their dying fisheries. These problems include protein and vitamin A deficiency and blindness, all of which may—in some instances—be linked to poor nutrition resulting from lower reef fish consumption by islanders, according to Bowden-Kerby.

  6. The Luxembourg Space Resources Initiative

    Science.gov (United States)

    Link, M.

    2017-09-01

    This keynote talk by M. Link from the Directorate of ICT and Space Affairs, Ministry of the Economy, The Government of the Grand Duchy of Luxembourg, will provide an overview of Luxembourg's ins-space resource utilization initiative.

  7. Integrated landscape initiatives in Europe

    DEFF Research Database (Denmark)

    García-Martín, María; Bieling, Claudia; Hart, Abigail

    2016-01-01

    Landscapes are linked to human well-being in a multitude of ways, some of which are challenged by global market forces and traditional management approaches. In response to this situation there has been a rise in local initiatives to sustain the values of landscape. The aim of this paper is to pr......Landscapes are linked to human well-being in a multitude of ways, some of which are challenged by global market forces and traditional management approaches. In response to this situation there has been a rise in local initiatives to sustain the values of landscape. The aim of this paper...... searches and canvassing of European umbrella organisations; followed by an online survey of representatives from the identified initiatives (n??=??71). Our results show that the most relevant characteristics of integrated landscape initiatives in Europe are: a holistic approach to landscape management...

  8. FY 10 Multifamily Initial Endorsements

    Data.gov (United States)

    Department of Housing and Urban Development — In FY 2010, HUD's Multifamily's 18 Hubs initially endorsed 1011 loans totaling $11.3 billion and providing 170,672 units/ beds. FY 10's $11.3 billion is the highest...

  9. Smart roadside initiative : user manual.

    Science.gov (United States)

    2015-09-01

    This document provides the user instructions for the Smart Roadside Initiative (SRI) applications including mobile and web-based SRI applications. These applications include smartphone-enabled information exchange and notification, and software compo...

  10. Smart roadside initiative : final report.

    Science.gov (United States)

    2015-09-01

    This is the Final Report for the Smart Roadside Initiative (SRI) prototype system deployment project. The SRI prototype was implemented at weigh stations in Grass Lake, Michigan and West Friendship, Maryland. The prototype was developed to integrate ...

  11. Global Peace Operations Initiative (GPOI)

    National Research Council Canada - National Science Library

    Kim-Mitchell, Elena

    2005-01-01

    .... In terms of manpower, the initiative aims at deploying 75,000 peace support operations (PSO) troops worldwide over the next 5 years, primarily to Africa, but also to Latin America, Europe, and Asia...

  12. Canada's family violence initiative: partnerships

    OpenAIRE

    Scott,Elaine

    1994-01-01

    Under Canada's four-year, $136 million Family Violence Initiative, the federal government is calling upon all Canadians to work in partnerships towards the elimination of family violence - child abuse, violence against women, and elder (senior) abuse. Family violence is a complex problem and requires the efforts of all Canadians to resolve it. One of the key themes of the Initiative - a multidisciplinary approach to the problem of family violence - is reflected in the selection and developmen...

  13. Student initiative: A conceptual analysis

    OpenAIRE

    Polovina Nada

    2014-01-01

    In the description and scientific consideration of the attitude of children and youth towards their education and development, the concept of student initiative has been gaining ground lately, and it is hence the subject of analysis in this paper. The analysis is important because of the discrepancy between the increased efforts of the key educational policy holders to promote the idea about the importance of the development of student initiative and rare a...

  14. Teachers’ perceptions of their own initiative: Collective initiative vs. personal initiative

    Directory of Open Access Journals (Sweden)

    Džinović Vladimir

    2013-01-01

    Full Text Available Current trends in education demand from teachers to exhibit proactive behaviour and assume responsibility for the implementation of changes in school practice. In that sense, it is important to study how teachers perceive their own initiative and to gain insight into the activities where such initiative is demonstrated. This study has been conceived as a mixed-methods research. The qualitative study implied forming four focus groups with subject teachers and class teachers (N=38, while the quantitative study entailed surveying 1441 teachers in forty primary schools in Serbia using the questionnaire constructed based on qualitative data. Data from focus groups were processed by qualitative thematic analysis, while the questionnaire data were processed by principal component analysis and univariate analysis of variance. The findings of the study have shown that teachers mostly demonstrate initiative through co­operative activities that include planning of joint teaching as well as conducting joint projects within school and with the local community actors. Teachers are least ready to demonstrate personal initiative and the initiative aimed at accomplishing considerable changes in school work. The concluding part includes the recommendations for encouraging teachers’ personal initiative and building organizational culture that would support such initiative. [Projekat Ministarstva nauke Republike Srbije, br. br. 47008: Unapređivanje kvaliteta i dostupnosti obrazovanja u procesima modernizacije Srbije i br. 179034: Od podsticanja inicijative, saradnje i stvaralaštva u obrazovanju do novih uloga i identiteta u društvu

  15. Effects of ionizing radiations on proteins

    International Nuclear Information System (INIS)

    Maire, M. le; Foresta, B. de; Viel, A.; Thauvette, L.; Beauregard, G.; Potier, M.

    1990-01-01

    We have reinvestigated the use of ionizing radiations to measure the molecular mass of water-soluble or membrane proteins. Exposure of purified standard proteins to increasing doses of ionizing radiation causes progressive fragmentation of the native protein into defined peptide patterns. The coloured band corresponding to the intact protein was measured on the SDS gel as a function of dose to determine the dose (D 37.t ) corresponding to 37% of the initial amount of unfragmented protein deposited on the gel. This led to a calibration curve and the known molecular mass of the standard proteins. However, we have to conclude that this method is useless to determine the state of aggregation of a protein, since, for all the oligomers tested, the best fit was obtained by using the protomeric molecular mass, suggesting that there is no energy transfer between protomers. Furthermore, SDS greatly increases the fragmentation rate of proteins, which suggests additional calibration problems for membrane proteins in detergent or in the lipid bilayer. The main drawback of the technique is that some proteins behaved anomalously, leading to very large errors in the apparent target size as compared with true molecular mass. It is thus unreliable to apply the radiation method for absolute molecular-mass determination. We then focused on the novel finding that discrete fragmentation of proteins occurs at preferential sites, and this was studied with aspartate transcarbamylase. (author)

  16. The continuing conundrum of the LEA proteins.

    Science.gov (United States)

    Tunnacliffe, Alan; Wise, Michael J

    2007-10-01

    Research into late embryogenesis abundant (LEA) proteins has been ongoing for more than 20 years but, although there is a strong association of LEA proteins with abiotic stress tolerance particularly dehydration and cold stress, for most of that time, their function has been entirely obscure. After their initial discovery in plant seeds, three major groups (numbered 1, 2 and 3) of LEA proteins have been described in a range of different plants and plant tissues. Homologues of groups 1 and 3 proteins have also been found in bacteria and in certain invertebrates. In this review, we present some new data, survey the biochemistry, biophysics and bioinformatics of the LEA proteins and highlight several possible functions. These include roles as antioxidants and as membrane and protein stabilisers during water stress, either by direct interaction or by acting as molecular shields. Along with other hydrophilic proteins and compatible solutes, LEA proteins might also serve as "space fillers" to prevent cellular collapse at low water activities. This multifunctional capacity of the LEA proteins is probably attributable in part to their structural plasticity, as they are largely lacking in secondary structure in the fully hydrated state, but can become more folded during water stress and/or through association with membrane surfaces. The challenge now facing researchers investigating these enigmatic proteins is to make sense of the various in vitro defined functions in the living cell: Are the LEA proteins truly multi-talented, or are they still just misunderstood?

  17. PROTEIN ANTIMIKROB DARI TANAMAN TRICHOSANTHES

    Directory of Open Access Journals (Sweden)

    Sukma D

    2008-08-01

    Full Text Available The research was aimed to study morphology, growth, development, pest and disease of 3 Trichosanthes species, initiate shoots, callus and hairy root culture in vitro, analyze chitinase and peroxidase activities and the effect of salicylic acid (SA and etefon (ETF on the chitinase and peroxidase activities of crude protein extract from Trichosanthes, and evaluate in vitro antifungal activity of crude protein extract of Trichosanthes. The results of the research showed the differences of morphological characters, growth habit of T. cucumerina var. anguina, T.tricuspidata and the differences of pest and diseases problem of T. quinquangulata. T. cucumerina var. anguina and T. quinquangulata. T. tricuspidata had the highest chitinase activity in crude protein extract of in vitro shoots, calli and plant roots and peroxidase activity in plant roots grown in field. T. cucumerina var. anguina showed the highest chitinase and peroxidase activities in crude protein extract of plant roots grown in field and calli. Chitinase and peroxidase activities of calli crude protein extract of T. tricuspidata could be increased by SA and ETF. Adversely, ETF decreased the peroxidase activity of calli crude protein exract ofT. tricuspidata. In T. cucumerina var. anguina, SA could not increase the chitinase activity but increase the peroxidase activity. The crude protein from in vitro shoots of T. tricuspidata could inhibited the spore germination of Fusarium sp. from T. cucumerina var. anguina, Fusarium oxysporum from shallot, Puccinia arachidis from peanut and Pseudoperonospora cubensis from cucumber. The protein could not inhibit spore germination of Curvularia eragrostidis from Dendrobium orchids

  18. My 65 years in protein chemistry.

    Science.gov (United States)

    Scheraga, Harold A

    2015-05-01

    This is a tour of a physical chemist through 65 years of protein chemistry from the time when emphasis was placed on the determination of the size and shape of the protein molecule as a colloidal particle, with an early breakthrough by James Sumner, followed by Linus Pauling and Fred Sanger, that a protein was a real molecule, albeit a macromolecule. It deals with the recognition of the nature and importance of hydrogen bonds and hydrophobic interactions in determining the structure, properties, and biological function of proteins until the present acquisition of an understanding of the structure, thermodynamics, and folding pathways from a linear array of amino acids to a biological entity. Along the way, with a combination of experiment and theoretical interpretation, a mechanism was elucidated for the thrombin-induced conversion of fibrinogen to a fibrin blood clot and for the oxidative-folding pathways of ribonuclease A. Before the atomic structure of a protein molecule was determined by x-ray diffraction or nuclear magnetic resonance spectroscopy, experimental studies of the fundamental interactions underlying protein structure led to several distance constraints which motivated the theoretical approach to determine protein structure, and culminated in the Empirical Conformational Energy Program for Peptides (ECEPP), an all-atom force field, with which the structures of fibrous collagen-like proteins and the 46-residue globular staphylococcal protein A were determined. To undertake the study of larger globular proteins, a physics-based coarse-grained UNited-RESidue (UNRES) force field was developed, and applied to the protein-folding problem in terms of structure, thermodynamics, dynamics, and folding pathways. Initially, single-chain and, ultimately, multiple-chain proteins were examined, and the methodology was extended to protein-protein interactions and to nucleic acids and to protein-nucleic acid interactions. The ultimate results led to an understanding

  19. Effects of atmospheric pressure plasmas on isolated and cellular DNA-a review.

    Science.gov (United States)

    Arjunan, Krishna Priya; Sharma, Virender K; Ptasinska, Sylwia

    2015-01-29

    Atmospheric Pressure Plasma (APP) is being used widely in a variety of biomedical applications. Extensive research in the field of plasma medicine has shown the induction of DNA damage by APP in a dose-dependent manner in both prokaryotic and eukaryotic systems. Recent evidence suggests that APP-induced DNA damage shows potential benefits in many applications, such as sterilization and cancer therapy. However, in several other applications, such as wound healing and dentistry, DNA damage can be detrimental. This review reports on the extensive investigations devoted to APP interactions with DNA, with an emphasis on the critical role of reactive species in plasma-induced damage to DNA. The review consists of three main sections dedicated to fundamental knowledge of the interactions of reactive oxygen species (ROS)/reactive nitrogen species (RNS) with DNA and its components, as well as the effects of APP on isolated and cellular DNA in prokaryotes and eukaryotes.

  20. IGF binding proteins.

    Science.gov (United States)

    Bach, Leon A

    2017-12-18

    Insulin-like growth factor binding proteins (IGFBPs) 1-6 bind IGFs but not insulin with high affinity. They were initially identified as serum carriers and passive inhibitors of IGF actions. However, subsequent studies showed that, although IGFBPs inhibit IGF actions in many circumstances, they may also potentiate these actions. IGFBPs are widely expressed in most tissues, and they are flexible endocrine and autocrine/paracrine regulators of IGF activity, which is essential for this important physiological system. More recently, individual IGFBPs have been shown to have IGF-independent actions. Mechanisms underlying these actions include (i) interaction with non-IGF proteins in compartments including the extracellular space and matrix, the cell surface and intracellularly; (ii) interaction with and modulation of other growth factor pathways including EGF, TGF- and VEGF; and (iii) direct or indirect transcriptional effects following nuclear entry of IGFBPs. Through these IGF-dependent and IGF-independent actions, IGFBPs modulate essential cellular processes including proliferation, survival, migration, senescence, autophagy and angiogenesis. They have been implicated in a range of disorders including malignant, metabolic, neurological and immune diseases. A more complete understanding of their cellular roles may lead to the development of novel IGFBP-based therapeutic opportunities.

  1. The E5 Proteins

    OpenAIRE

    DiMaio, Daniel; Petti, Lisa

    2013-01-01

    The E5 proteins are short transmembrane proteins encoded by many animal and human papillomaviruses. These proteins display transforming activity in cultured cells and animals, and they presumably also play a role in the productive virus life cycle. The E5 proteins are thought to act by modulating the activity of cellular proteins. Here, we describe the biological activities of the best-studied E5 proteins and discuss the evidence implicating specific protein targets and pathways in mediating ...

  2. Spider Silk Glue Proteins BAA 8.1

    Science.gov (United States)

    2017-09-14

    model for each. These genes were all sequenced at the ends to confirm the sequences were correct prior to using them for...protein expression. 3. The genes were then used in flask fermentation expression studies to insure that protein of the correct size was being...one of the lengths (3X, roughly a 140kD protein) for initial studies. We were able to detect protein production in these fermentations but

  3. Cellular Handling of Protein Aggregates by Disaggregation Machines.

    Science.gov (United States)

    Mogk, Axel; Bukau, Bernd; Kampinga, Harm H

    2018-01-18

    Both acute proteotoxic stresses that unfold proteins and expression of disease-causing mutant proteins that expose aggregation-prone regions can promote protein aggregation. Protein aggregates can interfere with cellular processes and deplete factors crucial for protein homeostasis. To cope with these challenges, cells are equipped with diverse folding and degradation activities to rescue or eliminate aggregated proteins. Here, we review the different chaperone disaggregation machines and their mechanisms of action. In all these machines, the coating of protein aggregates by Hsp70 chaperones represents the conserved, initializing step. In bacteria, fungi, and plants, Hsp70 recruits and activates Hsp100 disaggregases to extract aggregated proteins. In the cytosol of metazoa, Hsp70 is empowered by a specific cast of J-protein and Hsp110 co-chaperones allowing for standalone disaggregation activity. Both types of disaggregation machines are supported by small Hsps that sequester misfolded proteins. Copyright © 2018 Elsevier Inc. All rights reserved.

  4. EDITORIAL: Precision proteins Precision proteins

    Science.gov (United States)

    Demming, Anna

    2010-06-01

    Since the birth of modern day medicine, during the times of Hippocrates in ancient Greece, the profession has developed from the rudimentary classification of disease into a rigorous science with an inspiring capability to treat and cure. Scientific methodology has distilled clinical diagnostic tools from the early arts of prognosis, which used to rely as much on revelation and prophecy, as intuition and judgement [1]. Over the past decade, research into the interactions between proteins and nanosystems has provided some ingenious and apt techniques for delving into the intricacies of anatomical systems. In vivo biosensing has emerged as a vibrant field of research, as much of medical diagnosis relies on the detection of substances or an imbalance in the chemicals in the body. The inherent properties of nanoscale structures, such as cantilevers, make them well suited to biosensing applications that demand the detection of molecules at very low concentrations. Measurable deflections in cantilevers functionalised with antibodies provide quantitative indicators of the presence of specific antigens when the two react. Such developments have roused mounting interest in the interactions of proteins with nanostructures, such as carbon nanotubes [3], which have demonstrated great potential as generic biomarkers. Plasmonic properties are also being exploited in sensing applications, such as the molecular sentinel recently devised by researchers in the US. The device uses the plasmonic properties of a silver nanoparticle linked to a Raman labelled hairpin DNA probe to signal changes in the probe geometry resulting from interactions with substances in the environment. Success stories so far include the detection of two specific genes associated with breast cancer [4]. A greater understanding of how RNA interference regulates gene expression has highlighted the potential of using this natural process as another agent for combating disease in personalized medicine. However, the

  5. Integrated Initiating Event Performance Indicators

    International Nuclear Information System (INIS)

    S. A. Eide; Dale M. Rasmuson; Corwin L. Atwood

    2005-01-01

    The U.S. Nuclear Regulatory Commission Industry Trends Program (ITP) collects and analyses industry-wide data, assesses the safety significance of results, and communicates results to Congress and other stakeholders. This paper outlines potential enhancements in the ITP to comprehensively cover the Initiating Events Cornerstone of Safety. Future work will address other cornerstones of safety. The proposed Tier 1 activity involves collecting data on ten categories of risk-significant initiating events, trending the results, and comparing early performance with prediction limits (allowable numbers of events, above which NRC action may occur). Tier 1 results would be used to monitor industry performance at the level of individual categories of initiating events. The proposed Tier 2 activity involves integrating the information for individual categories of initiating events into a single risk-based indicator, termed the Baseline Risk Index for Initiating Events or BRIIE. The BRIIE would be evaluated yearly and compared against a threshold. BRIIE results would be reported to Congress on a yearly basis

  6. Two Chimeric Regulators of G-protein Signaling (RGS) Proteins Differentially Modulate Soybean Heterotrimeric G-protein Cycle*

    Science.gov (United States)

    Roy Choudhury, Swarup; Westfall, Corey S.; Laborde, John P.; Bisht, Naveen C.; Jez, Joseph M.; Pandey, Sona

    2012-01-01

    Heterotrimeric G-proteins and the regulator of G-protein signaling (RGS) proteins, which accelerate the inherent GTPase activity of Gα proteins, are common in animals and encoded by large gene families; however, in plants G-protein signaling is thought to be more limited in scope. For example, Arabidopsis thaliana contains one Gα, one Gβ, three Gγ, and one RGS protein. Recent examination of the Glycine max (soybean) genome reveals a larger set of G-protein-related genes and raises the possibility of more intricate G-protein networks than previously observed in plants. Stopped-flow analysis of GTP-binding and GDP/GTP exchange for the four soybean Gα proteins (GmGα1–4) reveals differences in their kinetic properties. The soybean genome encodes two chimeric RGS proteins with an N-terminal seven transmembrane domain and a C-terminal RGS box. Both GmRGS interact with each of the four GmGα and regulate their GTPase activity. The GTPase-accelerating activities of GmRGS1 and -2 differ for each GmGα, suggesting more than one possible rate of the G-protein cycle initiated by each of the Gα proteins. The differential effects of GmRGS1 and GmRGS2 on GmGα1–4 result from a single valine versus alanine difference. The emerging picture suggests complex regulation of the G-protein cycle in soybean and in other plants with expanded G-protein networks. PMID:22474294

  7. Characteristics of patients initiating raloxifene compared to those initiating bisphosphonates

    Directory of Open Access Journals (Sweden)

    Wang Sara

    2008-12-01

    Full Text Available Abstract Background Both raloxifene and bisphosphonates are indicated for the prevention and treatment of postmenopausal osteoporosis, however these medications have different efficacy and safety profiles. It is plausible that physicians would prescribe these agents to optimize the benefit/risk profile for individual patients. The objective of this study was to compare demographic and clinical characteristics of patients initiating raloxifene with those of patients initiating bisphosphonates for the prevention and treatment of osteoporosis. Methods This study was conducted using a retrospective cohort design. Female beneficiaries (45 years and older with at least one claim for raloxifene or a bisphosphonate in 2003 through 2005 and continuous enrollment in the previous 12 months and subsequent 6 months were identified using a collection of large national commercial, Medicare supplemental, and Medicaid administrative claims databases (MarketScan®. Patients were divided into two cohorts, a combined commercial/Medicare cohort and a Medicaid cohort. Within each cohort, characteristics (demographic, clinical, and resource utilization of patients initiating raloxifene were compared to those of patients initiating bisphosphonate therapy. Group comparisons were made using chi-square tests for proportions of categorical measures and Wilcoxon rank-sum tests for continuous variables. Logistic regression was used to simultaneously examine factors independently associated with initiation of raloxifene versus a bisphosphonate. Results Within both the commercial/Medicare and Medicaid cohorts, raloxifene patients were younger, had fewer comorbid conditions, and fewer pre-existing fractures than bisphosphonate patients. Raloxifene patients in both cohorts were less likely to have had a bone mineral density (BMD screening in the previous year than were bisphosphonate patients, and were also more likely to have used estrogen or estrogen/progestin therapy in the

  8. Protein docking prediction using predicted protein-protein interface

    Directory of Open Access Journals (Sweden)

    Li Bin

    2012-01-01

    Full Text Available Abstract Background Many important cellular processes are carried out by protein complexes. To provide physical pictures of interacting proteins, many computational protein-protein prediction methods have been developed in the past. However, it is still difficult to identify the correct docking complex structure within top ranks among alternative conformations. Results We present a novel protein docking algorithm that utilizes imperfect protein-protein binding interface prediction for guiding protein docking. Since the accuracy of protein binding site prediction varies depending on cases, the challenge is to develop a method which does not deteriorate but improves docking results by using a binding site prediction which may not be 100% accurate. The algorithm, named PI-LZerD (using Predicted Interface with Local 3D Zernike descriptor-based Docking algorithm, is based on a pair wise protein docking prediction algorithm, LZerD, which we have developed earlier. PI-LZerD starts from performing docking prediction using the provided protein-protein binding interface prediction as constraints, which is followed by the second round of docking with updated docking interface information to further improve docking conformation. Benchmark results on bound and unbound cases show that PI-LZerD consistently improves the docking prediction accuracy as compared with docking without using binding site prediction or using the binding site prediction as post-filtering. Conclusion We have developed PI-LZerD, a pairwise docking algorithm, which uses imperfect protein-protein binding interface prediction to improve docking accuracy. PI-LZerD consistently showed better prediction accuracy over alternative methods in the series of benchmark experiments including docking using actual docking interface site predictions as well as unbound docking cases.

  9. Protein docking prediction using predicted protein-protein interface.

    Science.gov (United States)

    Li, Bin; Kihara, Daisuke

    2012-01-10

    Many important cellular processes are carried out by protein complexes. To provide physical pictures of interacting proteins, many computational protein-protein prediction methods have been developed in the past. However, it is still difficult to identify the correct docking complex structure within top ranks among alternative conformations. We present a novel protein docking algorithm that utilizes imperfect protein-protein binding interface prediction for guiding protein docking. Since the accuracy of protein binding site prediction varies depending on cases, the challenge is to develop a method which does not deteriorate but improves docking results by using a binding site prediction which may not be 100% accurate. The algorithm, named PI-LZerD (using Predicted Interface with Local 3D Zernike descriptor-based Docking algorithm), is based on a pair wise protein docking prediction algorithm, LZerD, which we have developed earlier. PI-LZerD starts from performing docking prediction using the provided protein-protein binding interface prediction as constraints, which is followed by the second round of docking with updated docking interface information to further improve docking conformation. Benchmark results on bound and unbound cases show that PI-LZerD consistently improves the docking prediction accuracy as compared with docking without using binding site prediction or using the binding site prediction as post-filtering. We have developed PI-LZerD, a pairwise docking algorithm, which uses imperfect protein-protein binding interface prediction to improve docking accuracy. PI-LZerD consistently showed better prediction accuracy over alternative methods in the series of benchmark experiments including docking using actual docking interface site predictions as well as unbound docking cases.

  10. Shotgun protein sequencing.

    Energy Technology Data Exchange (ETDEWEB)

    Faulon, Jean-Loup Michel; Heffelfinger, Grant S.

    2009-06-01

    A novel experimental and computational technique based on multiple enzymatic digestion of a protein or protein mixture that reconstructs protein sequences from sequences of overlapping peptides is described in this SAND report. This approach, analogous to shotgun sequencing of DNA, is to be used to sequence alternative spliced proteins, to identify post-translational modifications, and to sequence genetically engineered proteins.

  11. Introduction to protein blotting.

    Science.gov (United States)

    Kurien, Biji T; Scofield, R Hal

    2009-01-01

    Protein blotting is a powerful and important procedure for the immunodetection of proteins following electrophoresis, particularly proteins that are of low abundance. Since the inception of the protocol for protein transfer from an electrophoresed gel to a membrane in 1979, protein blotting has evolved greatly. The scientific community is now confronted with a variety of ways and means to carry out this transfer.

  12. Our interests in protein-protein interactions

    Indian Academy of Sciences (India)

    protein interactions. Evolution of P-P partnerships. Evolution of P-P structures. Evolutionary dynamics of P-P interactions. Dynamics of P-P interaction network. Host-pathogen interactions. CryoEM mapping of gigantic protein assemblies.

  13. Metabolic behavior of cell surface biotinylated proteins

    International Nuclear Information System (INIS)

    Hare, J.F.; Lee, E.

    1989-01-01

    The turnover of proteins on the surface of cultured mammalian cells was measured by a new approach. Reactive free amino or sulfhydryl groups on surface-accessible proteins were derivatized with biotinyl reagents and the proteins solubilized from culture dishes with detergent. Solubilized, biotinylated proteins were then adsorbed onto streptavidin-agarose, released with sodium dodecyl sulfate and mercaptoethanol, and separated on polyacrylamide gels. Biotin-epsilon-aminocaproic acid N-hydroxysuccinimide ester (BNHS) or N-biotinoyl-N'-(maleimidohexanoyl)hydrazine (BM) were the derivatizing agents. Only 10-12 bands were adsorbed onto streptavidin-agarose from undervatized cells or from derivatized cells treated with free avidin at 4 degrees C. Two-dimensional isoelectric focusing-sodium dodecyl sulfate gel electrophoresis resolved greater than 100 BNHS-derivatized proteins and greater than 40 BM-derivatized proteins. There appeared to be little overlap between the two groups of derivatized proteins. Short-term pulse-chase studies showed an accumulation of label into both groups of biotinylated proteins up until 1-2 h of chase and a rapid decrease over the next 1-5 h. Delayed appearance of labeled protein at the cell surface was attributed to transit time from site of synthesis. The unexpected and unexplained rapid disappearance of pulse-labeled proteins from the cell surface was invariant for all two-dimensionally resolved proteins and was sensitive to temperature reduction to 18 degrees C. Long-term pulse-chase experiments beginning 4-8 h after the initiation of chase showed the disappearance of derivatized proteins to be a simple first-order process having a half-life of 115 h in the case of BNHS-derivatized proteins and 30 h in the case of BM-derivatized proteins

  14. Evolution of protein-protein interactions

    Indian Academy of Sciences (India)

    Evolution of protein-protein interactions · Our interests in protein-protein interactions · Slide 3 · Slide 4 · Slide 5 · Slide 6 · Slide 7 · Slide 8 · Slide 9 · Slide 10 · Slide 11 · Slide 12 · Slide 13 · Slide 14 · Slide 15 · Slide 16 · Slide 17 · Slide 18 · Slide 19 · Slide 20.

  15. The sustainable wood production initiative.

    Science.gov (United States)

    Robert. Deal

    2004-01-01

    To address concerns about sustainable forestry in the region, the Focused Science Delivery Program is sponsoring a three year Sustainable Wood Production Initiative. The Pacific Northwest is one of the world's major timber producing regions, and the ability of this region to produce wood on a sustained yield basis is widely recognized. Concerns relating to the...

  16. Intelligent vehicle initiative : business plan

    Science.gov (United States)

    1997-11-01

    The U.S. Department of Transportation (USDOT) is embarking on a new program called the Intelligent Vehicle Initiative (IVI). The USDOT intends to jointly define the program plan and conduct the IVI in cooperation with the motor vehicle, trucking, and...

  17. Nuclear materials facility safety initiative

    International Nuclear Information System (INIS)

    Peddicord, K.L.; Nelson, P.; Roundhill, M.; Jardine, L.J.; Lazarev, L.; Moshkov, M.; Khromov, V.V.; Kruchkov, E.; Bolyatko, V.; Kazanskij, Yu.; Vorobeva, I.; Lash, T.R.; Newton, D.; Harris, B.

    2000-01-01

    Safety in any facility in the nuclear fuel cycle is a fundamental goal. However, it is recognized that, for example, should an accident occur in either the U.S. or Russia, the results could seriously delay joint activities to store and disposition weapons fissile materials in both countries. To address this, plans are underway jointly to develop a nuclear materials facility safety initiative. The focus of the initiative would be to share expertise which would lead in improvements in safety and safe practices in the nuclear fuel cycle.The program has two components. The first is a lab-to-lab initiative. The second involves university-to-university collaboration.The lab-to-lab and university-to-university programs will contribute to increased safety in facilities dealing with nuclear materials and related processes. These programs will support important bilateral initiatives, develop the next generation of scientists and engineers which will deal with these challenges, and foster the development of a safety culture

  18. Initial heating in cold cars

    NARCIS (Netherlands)

    Daanen, H.A.M.; Teunissen, L.P.J.; Hoogh, I.M. de

    2012-01-01

    During the initial minutes after entering a cold car, people feel uncomfortably cold. Six different warming systems were investigated in a small car in order to find out how to improve the feeling of comfort using 16 volunteers. The methods were: no additional warming next to a standard heating

  19. Does the cerebellum initiate movement?

    Science.gov (United States)

    Thach, W T

    2014-02-01

    Opinion is divided on what the exact function of the cerebellum is. Experiments are summarized that support the following views: (1) the cerebellum is a combiner of multiple movement factors; (2) it contains anatomically fixed permanent focal representation of individual body parts (muscles and segments) and movement modes (e.g., vestibular driven vs. cognitive driven); (3) it contains flexible changing representations/memory of physical properties of the body parts including muscle strength, segment inertia, joint viscosity, and segmental interaction torques (dynamics); (4) it contains mechanisms for learning and storage of the properties in item no. 3 through trial-and-error practice; (5) it provides for linkage of body parts, motor modes, and motordynamics via the parallel fiber system; (6) it combines and integrates the many factors so as to initiate coordinated movements of the many body parts; (7) it is thus enabled to play the unique role of initiating coordinated movements; and (8) this unique causative role is evidenced by the fact that: (a) electrical stimulation of the cerebellum can initiate compound coordinated movements; (b) in naturally initiated compound movements, cerebellar discharge precedes that in downstream target structures such as motor cerebral cortex; and (c) cerebellar ablation abolishes the natural production of compound movements in the awake alert individuals.

  20. Light-initiated detonation systems

    Science.gov (United States)

    Cooper, Stafford S.; Malone, Philip G.; Bartholomew, Stephen W.; Necker, William J.

    1986-09-01

    Numerous light sources could be employed in detonation systems, but lasers have the most efficient coupling to optical fibers and can generate energetic light pulses required for detonation. Flash lamp-pumped, solid state lasers are presently the most useful light source for explosive initiation. Laser diodes in current production cannot generate enough energy for practical applications. The most useful optical fiber for blast line application is a step index fiber with a large core-to-cladding ratio. The large core minimizes energy losses due to misalignment core of fibers in connectors. Couplers that involve mechanically crimped connectors and cleaved fibers, rather than the epoxy-cemented connectors with polished fibers, provide superior energy transmission due to the reduced carbonization at the fiber end. Detonators for optical initiation systems are similar in basic construction to those employed in electrical initiation systems. Explosive and pyrotechnic charges can also be similar. Either primary or secondary explosives can be initiated in present laser-based systems. Two laser detonation systems are presently accessible; a multiple-shot laser with a single-shot, single fiber system designed for use with detonators containing primary explosives. Additional research related to development of low-energy, photoreactive detonators, continuity checking techniques and improved connectors and fibers can produce significant improvements in presently fielded systems.

  1. Big Data Initiatives for Agroecosystems

    Science.gov (United States)

    NAL has developed a workspace for research groups associated with the i5k initiative, which aims to sequence the genomes of all insesct species known to be important to worldwide agriculture, food safety, medicine, and energy production; all those used as models in biology; the most abundant in worl...

  2. Smoking Initiation: Peers and Personality

    NARCIS (Netherlands)

    C-S. Hsieh (Chih-Sheng); J.L.W. van Kippersluis (Hans)

    2016-01-01

    textabstractSocial interactions are widely recognized to play an important role in smoking initiation among adolescents. In this paper we hypothesize that individual with `stronger' personalities (i.e. emotionally stable, conscientious individuals) are better able to resist peer pressure in the

  3. Vacuum type D initial data

    Science.gov (United States)

    García-Parrado Gómez-Lobo, Alfonso

    2016-09-01

    A vacuum type D initial data set is a vacuum initial data set of the Einstein field equations whose data development contains a region where the space–time is of Petrov type D. In this paper we give a systematic characterisation of a vacuum type D initial data set. By systematic we mean that the only quantities involved are those appearing in the vacuum constraints, namely the first fundamental form (Riemannian metric) and the second fundamental form. Our characterisation is a set of conditions consisting of the vacuum constraints and some additional differential equations for the first and second fundamental forms These conditions can be regarded as a system of partial differential equations on a Riemannian manifold and the solutions of the system contain all possible regular vacuum type D initial data sets. As an application we particularise our conditions for the case of vacuum data whose data development is a subset of the Kerr solution. This has applications in the formulation of the nonlinear stability problem of the Kerr black hole.

  4. Initial drug resistance in India

    Indian Academy of Sciences (India)

    First page Back Continue Last page Overview Graphics. Initial drug resistance in India. There is gradual increase in primary MDR all over India : Pondi= Pondicherry 1985; Bangalore =1986; Jaipur = 1991; Jaipur =2000. Overall the MDR is less than 3% (TRC studies).

  5. Parenting and Adolescents' Sexual Initiation

    Science.gov (United States)

    Longmore, Monica A.; Eng, Abbey L.; Giordano, Peggy C.; Manning, Wendy D.

    2009-01-01

    This study draws on social control and social learning theories to examine the role of dating-specific attitudes and practices as predictors of adolescents' sexual initiation. We include attention to the adolescent's reaction to control attempts as a further means of assessing family dynamics (i.e., frequency of dating disagreements). The study…

  6. Russian Initiatives in Internet Governance

    Directory of Open Access Journals (Sweden)

    Sergey Sergeevich Shirin

    2014-01-01

    Full Text Available The article is devoted to the research of Russian initiatives concerning its participation in international system of Internet governance. In the research, conceptual and theoretic basis for participation of state governments in Internet governance is given, academic discourse of this issue is assessed, the review of main Russian initiatives made from 2005 (since the second stage of the World Summit on Information Society till the present day is carried out. The conclusion is made that the most successful Internet governance initiative of Russia is the implementation of cyrillic top level domains. Much less successful are Russian attempts to reconstruct existing organizational structure of Internet governance. Russian policy is assessed as a policy of preferring loud statements and PR rather than real influence on development of Internet technologies. Russia consideres Internet as political resource and tries to increase its influence on decision making process on a political, not operational level of Internet governance. The author states that federal government of the United States of America has fully lost the control on the domain name systems. These functions are going to be given to the private sector, while ICANN, which was established according to the Memorandum of Understanding/Joint Project Agreement with U.S. Department of Commerce, is becoming a truly independent international organization. Considering this, the author comes to the conclusion that political transformations for the sake of Russia are already made. He also forecast that next initiatives of Russia will be directed to the segmentation of the Internet.

  7. Fayette County Better Buildings Initiative

    Energy Technology Data Exchange (ETDEWEB)

    Capella, Arthur [County of Fayette, Uniontown, PA (United States)

    2015-03-04

    The Fayette County Better Buildings Initiative represented a comprehensive and collaborative approach to promoting and implementing energy efficiency improvements. The initiative was designed to focus on implementing energy efficiency improvements in residential units, while simultaneously supporting general marketing of the benefits of implementing energy efficiency measures. The ultimate goal of Fayette County’s Better Buildings Initiative was to implement a total of 1,067 residential energy efficiency retrofits with a minimum 15% estimated energy efficiency savings per unit. Program partners included: United States Department of Energy, Allegheny Power, and Private Industry Council of Westmoreland-Fayette, Fayette County Redevelopment Authority, and various local partners. The program was open to any Fayette County residents who own their home and meet the prequalifying conditions. The level of assistance offered depended upon household income and commitment to undergo a BPI – Certified Audit and implement energy efficiency measures, which aimed to result in at least a 15% reduction in energy usage. The initiative was designed to focus on implementing energy efficiency improvements in residential units, while simultaneously supporting general marketing of the benefits of implementing energy efficiency measures. Additionally, the program had components that involved recruitment and training for employment of persons in the energy sector (green jobs), as well as marketing and implementation of a commercial or community facilities component. The residential component of Fayette County’s Better Buildings Initiative involved a comprehensive approach, providing assistance to low- moderate- and market-rate homeowners. The initiative will also coordinate activities with local utility providers to further incentivize energy efficiency improvements among qualifying homeowners. The commercial component of Fayette County’s Better Building Initiative involved grants

  8. Initiation disruptor systems and methods of initiation disruption

    Science.gov (United States)

    Baum, Dennis W

    2014-09-23

    A system that may be used as an initiation disruption system (IDS) according to one embodiment includes an explosive charge; a plurality of particles in a layer at least partially surrounding the explosive charge; and a fire suppressant adjacent the plurality of particles. A method for disabling an object according to one embodiment includes placing the system as recited above near an object; and causing the explosive charge to initiate, thereby applying mechanical loading to the object such that the object becomes disabled. Additional systems and methods are also presented. A device according to another embodiment includes a plurality of particles bound by a binder thereby defining a sidewall having an interior for receiving an explosive; and a fire suppressant adjacent the plurality of particles and binder. Additional systems and methods are also presented.

  9. Protein in diet

    Science.gov (United States)

    Diet - protein ... Protein foods are broken down into parts called amino acids during digestion. The human body needs a ... to eat animal products to get all the protein you need in your diet. Amino acids are ...

  10. Protein-losing enteropathy

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/007338.htm Protein-losing enteropathy To use the sharing features on this page, please enable JavaScript. Protein-losing enteropathy is an abnormal loss of protein ...

  11. Energy efficiency initiatives: Indian experience

    Energy Technology Data Exchange (ETDEWEB)

    Dey, Dipankar [ICFAI Business School, Kolkata, (IBS-K) (India)

    2007-07-01

    India, with a population of over 1.10 billion is one of the fastest growing economies of the world. As domestic sources of different conventional commercial energy are drying up, dependence on foreign energy sources is increasing. There exists a huge potential for saving energy in India. After the first 'oil shock' (1973), the government of India realized the need for conservation of energy and a 'Petroleum Conservation Action Group' was formed in 1976. Since then many initiatives aiming at energy conservation and improving energy efficiency, have been undertaken (the establishment of Petroleum Conservation Research Association in 1978; the notification of Eco labelling scheme in 1991; the formation of Bureau of Energy Efficiency in 2002). But no such initiative was successful. In this paper an attempt has been made to analyze the changing importance of energy conservation/efficiency measures which have been initiated in India between 1970 and 2005.The present study tries to analyze the limitations and the reasons of failure of those initiatives. The probable reasons are: fuel pricing mechanism (including subsidies), political factors, corruption and unethical practices, influence of oil and related industry lobbies - both internal and external, the economic situation and the prolonged protection of domestic industries. Further, as India is opening its economy, the study explores the opportunities that the globally competitive market would offer to improve the overall energy efficiency of the economy. The study suggests that the Bureau of Energy Efficiency (BEE) - the newly formed nodal agency for improving energy efficiency of the economy may be made an autonomous institution where intervention from the politicians would be very low. For proper implementation of different initiatives to improve energy efficiency, BEE should involve more the civil societies (NGO) from the inception to the implementation stage of the programs. The paper also

  12. Oligomeric protein structure networks: insights into protein-protein interactions

    Directory of Open Access Journals (Sweden)

    Brinda KV

    2005-12-01

    Full Text Available Abstract Background Protein-protein association is essential for a variety of cellular processes and hence a large number of investigations are being carried out to understand the principles of protein-protein interactions. In this study, oligomeric protein structures are viewed from a network perspective to obtain new insights into protein association. Structure graphs of proteins have been constructed from a non-redundant set of protein oligomer crystal structures by considering amino acid residues as nodes and the edges are based on the strength of the non-covalent interactions between the residues. The analysis of such networks has been carried out in terms of amino acid clusters and hubs (highly connected residues with special emphasis to protein interfaces. Results A variety of interactions such as hydrogen bond, salt bridges, aromatic and hydrophobic interactions, which occur at the interfaces are identified in a consolidated manner as amino acid clusters at the interface, from this study. Moreover, the characterization of the highly connected hub-forming residues at the interfaces and their comparison with the hubs from the non-interface regions and the non-hubs in the interface regions show that there is a predominance of charged interactions at the interfaces. Further, strong and weak interfaces are identified on the basis of the interaction strength between amino acid residues and the sizes of the interface clusters, which also show that many protein interfaces are stronger than their monomeric protein cores. The interface strengths evaluated based on the interface clusters and hubs also correlate well with experimentally determined dissociation constants for known complexes. Finally, the interface hubs identified using the present method correlate very well with experimentally determined hotspots in the interfaces of protein complexes obtained from the Alanine Scanning Energetics database (ASEdb. A few predictions of interface hot

  13. Alteration of the ATG start codon of the A* protein of bacteriophage [phi]X174 into an ATT codon yields a viable phage indicating that A* protein is not essential for [phi]X174 reproduction

    NARCIS (Netherlands)

    Baas, P.D.; Liewerink, H.; Teeffelen, H.A.A.M. van; Mansfeld, A.D.M. van; Boom, J.M. van; Jansz, H.S.

    1987-01-01

    Bacteriophage X174 gene A encodes two proteins: the gene A protein and the smaller A* protein, which is synthesized from a translational start signal within the A gene in the same reading frame as the gene A protein. The gene A protein is involved in initiation, elongation and termination of rolling

  14. Sox2 is translationally activated by eukaryotic initiation factor 4E in human glioma-initiating cells

    Energy Technology Data Exchange (ETDEWEB)

    Ge, Yuqing; Zhou, Fengbiao; Chen, Hong; Cui, Chunhong; Liu, Dan [Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Shanghai Medical College of Fudan University, Shanghai 200032 (China); Li, Qiuping [Zhongshan Hospital of Fudan University, Shanghai 200032 (China); Yang, Zhiyuan; Wu, Guoqiang [Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Shanghai Medical College of Fudan University, Shanghai 200032 (China); Sun, Shuhui [Key Laboratory of Medical Molecular Virology, Ministry of Education and Health, Shanghai Medical College of Fudan University, Shanghai 200032 (China); Gu, Jianxin [Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Shanghai Medical College of Fudan University, Shanghai 200032 (China); Institutes of Biomedical Sciences of Fudan University, Shanghai 200032 (China); Wei, Yuanyan, E-mail: yywei@fudan.edu.cn [Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Shanghai Medical College of Fudan University, Shanghai 200032 (China); Jiang, Jianhai, E-mail: jianhaijiang@fudan.edu.cn [Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Shanghai Medical College of Fudan University, Shanghai 200032 (China)

    2010-07-09

    Sox2, a master transcription factor, contributes to the generation of induced pluripotent stem cells and plays significant roles in sustaining the self-renewal of neural stem cells and glioma-initiating cells. Understanding the functional differences of Sox2 between glioma-initiating cells and normal neural stem cells would contribute to therapeutic approach for treatment of brain tumors. Here, we first demonstrated that Sox2 could contribute to the self-renewal and proliferation of glioma-initiating cells. The following experiments showed that Sox2 was activated at translational level in a subset of human glioma-initiating cells compared with the normal neural stem cells. Further investigation revealed there was a positive correlation between Sox2 and eukaryotic initiation factor 4E (eIF4E) in glioma tissues. Down-regulation of eIF4E decreased Sox2 protein level without altering its mRNA level in glioma-initiating cells, indicating that Sox2 was activated by eIF4E at translational level. Furthermore, eIF4E was presumed to regulate the expression of Sox2 by its 5' untranslated region (5' UTR) sequence. Our results suggest that the eIF4E-Sox2 axis is a novel mechanism of unregulated self-renewal of glioma-initiating cells, providing a potential therapeutic target for glioma.

  15. Sox2 is translationally activated by eukaryotic initiation factor 4E in human glioma-initiating cells

    International Nuclear Information System (INIS)

    Ge, Yuqing; Zhou, Fengbiao; Chen, Hong; Cui, Chunhong; Liu, Dan; Li, Qiuping; Yang, Zhiyuan; Wu, Guoqiang; Sun, Shuhui; Gu, Jianxin; Wei, Yuanyan; Jiang, Jianhai

    2010-01-01

    Sox2, a master transcription factor, contributes to the generation of induced pluripotent stem cells and plays significant roles in sustaining the self-renewal of neural stem cells and glioma-initiating cells. Understanding the functional differences of Sox2 between glioma-initiating cells and normal neural stem cells would contribute to therapeutic approach for treatment of brain tumors. Here, we first demonstrated that Sox2 could contribute to the self-renewal and proliferation of glioma-initiating cells. The following experiments showed that Sox2 was activated at translational level in a subset of human glioma-initiating cells compared with the normal neural stem cells. Further investigation revealed there was a positive correlation between Sox2 and eukaryotic initiation factor 4E (eIF4E) in glioma tissues. Down-regulation of eIF4E decreased Sox2 protein level without altering its mRNA level in glioma-initiating cells, indicating that Sox2 was activated by eIF4E at translational level. Furthermore, eIF4E was presumed to regulate the expression of Sox2 by its 5' untranslated region (5' UTR) sequence. Our results suggest that the eIF4E-Sox2 axis is a novel mechanism of unregulated self-renewal of glioma-initiating cells, providing a potential therapeutic target for glioma.

  16. Host Proteins Determine MRSA Biofilm Structure and Integrity

    DEFF Research Database (Denmark)

    Dreier, Cindy; Nielsen, Astrid; Jørgensen, Nis Pedersen

    Human extracellular matrix (hECM) proteins aids the initial attachment and initiation of an infection, by specific binding to bacterial cell surface proteins. However, the importance of hECM proteins in structure, integrity and antibiotic resilience of a biofilm is unknown. This study aims...... to determine how specific hECM proteins affect S. aureus USA300 JE2 biofilms. Biofilms were grown in the presence of synovial fluid from rheumatoid arteritis patients to mimic in vivo conditions, where bacteria incorporate hECM proteins into the biofilm matrix. Difference in biofilm structure, with and without...... addition of hECM to growth media, was visualized by confocal laser scanning microscopy. Two enzymatic degradation experiments were used to study biofilm matrix composition and importance of hECM proteins: enzymatic removal of specific hECM proteins from growth media, before biofilm formation, and enzymatic...

  17. Stress proteins, autoimmunity, and autoimmune disease.

    Science.gov (United States)

    Winfield, J B; Jarjour, W N

    1991-01-01

    At birth, the immune system is biased toward recognition of microbial antigens in order to protect the host from infection. Recent data suggest that an important initial line of defense in this regard involves autologous stress proteins, especially conserved peptides of hsp60, which are presented to T cells bearing gamma delta receptors by relatively nonpolymorphic class lb molecules. Natural antibodies may represent a parallel B cell mechanism. Through an evolving process of "physiological" autoreactivity and selection by immunodominant stress proteins common to all prokaryotes, B and T cell repertoires expand during life to meet the continuing challenge of infection. Because stress proteins of bacteria are homologous with stress proteins of the host, there exists in genetically susceptible individuals a constant risk of autoimmune disease due to failure of mechanisms for self-nonself discrimination. That stress proteins actually play a role in autoimmune processes is supported by a growing body of evidence which, collectively, suggests that autoreactivity in chronic inflammatory arthritis involves, at least initially, gamma delta cells which recognize epitopes of the stress protein hsp60. Alternate mechanisms for T cell stimulation by stress proteins undoubtedly also exist, e.g., molecular mimicry of the DR beta third hypervariable region susceptibility locus for rheumatoid arthritis by a DnaJ stress protein epitope in gram-negative bacteria. While there still is confusion with respect to the most relevant stress protein epitopes, a central role for stress proteins in the etiology of arthritis appears likely. Furthermore, insight derived from the work thus far in adjuvant-induced arthritis already is stimulating analyses of related phenomena in autoimmune diseases other than those involving joints. Only limited data are available in the area of humoral autoimmunity to stress proteins. Autoantibodies to a number of stress proteins have been identified in SLE and

  18. Protein surface shielding agents in protein crystallization

    International Nuclear Information System (INIS)

    Hašek, J.

    2011-01-01

    The crystallization process can be controlled by protein surface shielding agents blocking undesirable competitive adhesion modes during non-equilibrium processes of deposition of protein molecules on the surface of growing crystalline blocks. The hypothesis is based on a number of experimental proofs from diffraction experiments and also retrieved from the Protein Data Bank. The molecules adhering temporarily on the surface of protein molecules change the propensity of protein molecules to deposit on the crystal surface in a definite position and orientation. The concepts of competitive adhesion modes and protein surface shielding agents acting on the surface of molecules in a non-equilibrium process of protein crystallization provide a useful platform for the control of crystallization. The desirable goal, i.e. a transient preference of a single dominating adhesion mode between protein molecules during crystallization, leads to uniform deposition of proteins in a crystal. This condition is the most important factor for diffraction quality and thus also for the accuracy of protein structure determination. The presented hypothesis is a generalization of the experimentally well proven behaviour of hydrophilic polymers on the surface of protein molecules of other compounds

  19. Biophysical EPR Studies Applied to Membrane Proteins

    Science.gov (United States)

    Sahu, Indra D; Lorigan, Gary A

    2015-01-01

    Membrane proteins are very important in controlling bioenergetics, functional activity, and initializing signal pathways in a wide variety of complicated biological systems. They also represent approximately 50% of the potential drug targets. EPR spectroscopy is a very popular and powerful biophysical tool that is used to study the structural and dynamic properties of membrane proteins. In this article, a basic overview of the most commonly used EPR techniques and examples of recent applications to answer pertinent structural and dynamic related questions on membrane protein systems will be presented. PMID:26855825

  20. Diversity and evolution of ABC proteins in basidiomycetes.

    Science.gov (United States)

    Kovalchuk, Andriy; Lee, Yong-Hwan; Asiegbu, Fred O

    2013-01-01

    ABC proteins constitute one of the largest families of proteins. They are implicated in wide variety of cellular processes ranging from ribosome biogenesis to multidrug resistance. With the advance of fungal genomics, the number of known fungal ABC proteins increases rapidly but the information on their biological functions remains scarce. In this work we extended the previous analysis of fungal ABC proteins to include recently sequenced species of basidiomycetes. We performed an identification and initial cataloging of ABC proteins from 23 fungal species representing 10 orders within class Agaricomycotina. We identified more than 1000 genes coding for ABC proteins. Comparison of sets of ABC proteins present in basidiomycetes and ascomycetes revealed the existence of two groups of ABC proteins specific for basidiomycetes. Results of survey should contribute to the better understanding of evolution of ABC proteins in fungi and support further experimental work on their characterization.

  1. Initial Semantics for Strengthened Signatures

    Directory of Open Access Journals (Sweden)

    André Hirschowitz

    2012-02-01

    Full Text Available We give a new general definition of arity, yielding the companion notions of signature and associated syntax. This setting is modular in the sense requested by Ghani and Uustalu: merging two extensions of syntax corresponds to building an amalgamated sum. These signatures are too general in the sense that we are not able to prove the existence of an associated syntax in this general context. So we have to select arities and signatures for which there exists the desired initial monad. For this, we follow a track opened by Matthes and Uustalu: we introduce a notion of strengthened arity and prove that the corresponding signatures have initial semantics (i.e. associated syntax. Our strengthened arities admit colimits, which allows the treatment of the λ-calculus with explicit substitution.

  2. Blasting agents and initiation systems

    Energy Technology Data Exchange (ETDEWEB)

    Fiscor, S.

    2000-01-01

    Although blasting differs between and within each industry, as a whole, the mines and quarries are making a shift from a purely ammonium nitrate/fuel oil (ANFO) mixture to a blend of emulsion and ANFO on a straight emulsion. Non-electric (shock tube) initiation systems have provided a viable alternative to the electric detonator (blasting cap). Explosives manufacturers are seeing their roles changes to being blasting contractors or consultants rather than just suppliers. The article discusses these trends and gives examples of typical blasting techniques and amounts of blasting agent used at large USA surface coal mines. Electric caps are still used in blasting underground coal. The Ensign Bickford Co. (EBCo) is developing electronic detonators and has been field testing an electronic initiator, the DIGIDET detonator, for the last four years. When commercially available, electronic detonators will be accurate but will come with a hefty price tag. 2 photos.

  3. Global nuclear cleanout initiative 2004

    International Nuclear Information System (INIS)

    Edlow, J.; Gruber, G.

    2004-01-01

    Full text: During more than 50 years of Atoms for Peace programmes nuclear materials were spread out worldwide. Stranded nuclear materials from nuclear research are left over without any safe back-end solution. 'Dirty Bombs' or so-called 'Radioactive Dispersal Devices (RDD)' are no longer science fiction since the world experienced the 9/11 attack. Governmental, NGO's and private industry organizations having discussed Global Nuclear Cleanout since then and start to take actions. The US Department of Energy (DOE) has announced to establish a dedicated organization in cooperation with IAEA and start the 'Global Threat Reduction Initiative (GTRI)'. The US government will allocate to that program USD 450 M over the next 10 years. Besides the historical development the paper will focus on the progress of the different initiatives and perspectives to threat reduction. (author)

  4. Initial conditions for cosmological perturbations

    Science.gov (United States)

    Ashtekar, Abhay; Gupt, Brajesh

    2017-02-01

    Penrose proposed that the big bang singularity should be constrained by requiring that the Weyl curvature vanishes there. The idea behind this past hypothesis is attractive because it constrains the initial conditions for the universe in geometric terms and is not confined to a specific early universe paradigm. However, the precise statement of Penrose’s hypothesis is tied to classical space-times and furthermore restricts only the gravitational degrees of freedom. These are encapsulated only in the tensor modes of the commonly used cosmological perturbation theory. Drawing inspiration from the underlying idea, we propose a quantum generalization of Penrose’s hypothesis using the Planck regime in place of the big bang, and simultaneously incorporating tensor as well as scalar modes. Initial conditions selected by this generalization constrain the universe to be as homogeneous and isotropic in the Planck regime as permitted by the Heisenberg uncertainty relations.

  5. Initial conditions for cosmological perturbations

    International Nuclear Information System (INIS)

    Ashtekar, Abhay; Gupt, Brajesh

    2017-01-01

    Penrose proposed that the big bang singularity should be constrained by requiring that the Weyl curvature vanishes there. The idea behind this past hypothesis is attractive because it constrains the initial conditions for the universe in geometric terms and is not confined to a specific early universe paradigm. However, the precise statement of Penrose’s hypothesis is tied to classical space-times and furthermore restricts only the gravitational degrees of freedom. These are encapsulated only in the tensor modes of the commonly used cosmological perturbation theory. Drawing inspiration from the underlying idea, we propose a quantum generalization of Penrose’s hypothesis using the Planck regime in place of the big bang, and simultaneously incorporating tensor as well as scalar modes. Initial conditions selected by this generalization constrain the universe to be as homogeneous and isotropic in the Planck regime as permitted by the Heisenberg uncertainty relations . (paper)

  6. Initial occupational exposure to lead

    Energy Technology Data Exchange (ETDEWEB)

    Forni, A.; Cambiaghi, G.; Secchi, G.C.

    1976-01-01

    Serial chromosome and biochemical studies were carried out in 11 subjects before and during initial occupational exposure to moderate quantities of lead fumes in a storage battery plant. The rate of abnormal metaphases, mostly with chromatid and one-break chromosome aberrations, was approximately doubled after one month of work; it further increased after two months of work; remained in this range up to seven months of exposure; and then tended to decrease somewhat. Blood lead levels increased progressively in the first few months, then reached a steady state. Urinary lead and coproporphyrin levels increased sharply after one month of work, while urinary delta-aminolevulinic acid (ALA) levels increased moderately. The ALA dehydratase (ALAD) activity of red blood cells (RBCs) was reduced to almost 50 percent of the initial values after one month, decreased further in subsequent months, and remained decreased through the remainder of the study.

  7. Unraveling the proteomic profile of mice testis during the initiation of meiosis.

    Science.gov (United States)

    Shao, Binbin; Guo, Yueshuai; Wang, Lei; Zhou, Quan; Gao, Tingting; Zheng, Bo; Zheng, Haoyu; Zhou, Tao; Zhou, Zuomin; Guo, Xuejiang; Huang, Xiaoyan; Sha, Jiahao

    2015-04-29

    In mice, once primordial germ cells (PGCs) are generated, they continue to proliferate and migrate to eventually reach the future gonads. They initiate sexual differentiation after their colonization of the gonads. During this process, retinoic acid (RA) induces meiosis in the female germ cells, which proceeds to the diplotene stage of meiotic prophase I, whereas the male germ cells initiate growth arrest. After birth, meiosis is initiated in mice spermatogonia by their conversion to preleptotene spermatocytes. There are evidences showing the roles of RA in the regulation of spermatogonial differentiation and meiosis initiation. However, it is still not well known on what responds to RA and how RA signaling engages meiosis. Thus, we constructed a proteomic profile of proteins associated with meiosis onset during testis development in mouse and identified 104 differentially expressed proteins (≥1.5 folds). Bioinformatic analysis showed proteins functioning in specific cell processes. The expression patterns of five selected proteins were verified via Western blot, of which we found that Tfrc gene was RA responsive, with a RA responsive element, and could be up regulated by RA in spermatogonial stem cell (SSC) line. Taken together, the results provide an important reference profile for further functional study of meiosis initiation. Spermatogenesis involves mitosis of spermatogonia, meiosis of spermatocytes and spermiogenesis, in which meiosis is a unique event to germ cells, and not in the somatic cells. Till now, the detailed molecular mechanisms of the transition from mitosis to meiosis are still not elucidated. With high-throughput proteomic technology, it is now possible to systemically identify proteins possibly involved. With TMT-6plex based quantification, we identified 104 proteins differentially between testes without meiosis (day 8.5) and those that were meiosis initiated (day 10.5). And a well-known protein essential for meiosis initiation, stra8, was

  8. Guam Initial Technical Assessment Report

    Energy Technology Data Exchange (ETDEWEB)

    Baring-Gould, I.; Conrad, M.; Haase, S.; Hotchkiss, E.; McNutt, P.

    2011-04-01

    Under an interagency agreement, funded by the Department of Interior's (DOI) Office of Insular Affairs (OIA), the National Renewable Energy Laboratory (NREL) was tasked to deliver technical assistance to the island of Guam by conducting an island initial technical assessment that would lay out energy consumption and production data and establish a baseline. This assessment will be used to conduct future analysis and studies by NREL that will estimate energy efficiency and renewable energy potential for the island of Guam.

  9. CubeSat Launch Initiative

    Science.gov (United States)

    Higginbotham, Scott

    2016-01-01

    The National Aeronautics and Space Administration (NASA) recognizes the tremendous potential that CubeSats (very small satellites) have to inexpensively demonstrate advanced technologies, collect scientific data, and enhance student engagement in Science, Technology, Engineering, and Mathematics (STEM). The CubeSat Launch Initiative (CSLI) was created to provide launch opportunities for CubeSats developed by academic institutions, non-profit entities, and NASA centers. This presentation will provide an overview of the CSLI, its benefits, and its results.

  10. Air sea ratio reduction initiative

    Energy Technology Data Exchange (ETDEWEB)

    Oberle, Jean

    2010-09-15

    Airfreight is the most expensive mode of transportation as well as the most impacting in terms of CO{sup 2} emissions. It is 7 times more expensive on average to ship by air than shipping by sea 1. Airfreight transportation mode emits 30 times more CO{sup 2} than sea freight mode 2. These elements provided a compelling platform to design a global logistics program to initiate a modal shift from air to sea freight without compromising service to customers.

  11. A Library of Plasmodium vivax Recombinant Merozoite Proteins Reveals New Vaccine Candidates and Protein-Protein Interactions

    Science.gov (United States)

    Hostetler, Jessica B.; Sharma, Sumana; Bartholdson, S. Josefin; Wright, Gavin J.; Fairhurst, Rick M.; Rayner, Julian C.

    2015-01-01

    Background A vaccine targeting Plasmodium vivax will be an essential component of any comprehensive malaria elimination program, but major gaps in our understanding of P. vivax biology, including the protein-protein interactions that mediate merozoite invasion of reticulocytes, hinder the search for candidate antigens. Only one ligand-receptor interaction has been identified, that between P. vivax Duffy Binding Protein (PvDBP) and the erythrocyte Duffy Antigen Receptor for Chemokines (DARC), and strain-specific immune responses to PvDBP make it a complex vaccine target. To broaden the repertoire of potential P. vivax merozoite-stage vaccine targets, we exploited a recent breakthrough in expressing full-length ectodomains of Plasmodium proteins in a functionally-active form in mammalian cells and initiated a large-scale study of P. vivax merozoite proteins that are potentially involved in reticulocyte binding and invasion. Methodology/Principal Findings We selected 39 P. vivax proteins that are predicted to localize to the merozoite surface or invasive secretory organelles, some of which show homology to P. falciparum vaccine candidates. Of these, we were able to express 37 full-length protein ectodomains in a mammalian expression system, which has been previously used to express P. falciparum invasion ligands such as PfRH5. To establish whether the expressed proteins were correctly folded, we assessed whether they were recognized by antibodies from Cambodian patients with acute vivax malaria. IgG from these samples showed at least a two-fold change in reactivity over naïve controls in 27 of 34 antigens tested, and the majority showed heat-labile IgG immunoreactivity, suggesting the presence of conformation-sensitive epitopes and native tertiary protein structures. Using a method specifically designed to detect low-affinity, extracellular protein-protein interactions, we confirmed a predicted interaction between P. vivax 6-cysteine proteins P12 and P41, further

  12. Industry's Commercial Initiatives on ISS

    Science.gov (United States)

    Shields, C. E.; Kessler, C.; Lavitola, M. S.

    2002-01-01

    For more than ten years, private industry has worked to develop a commercial human space market and to create a sustainable ISS commercial utilization customer base. Before ISS assembly was underway - and long before NASA and the international space agencies began to craft ISS commercial business terms and conditions - industry planted and nurtured the seeds of interest in exploiting human space utilization for commerce. These early initiatives have yielded the impetus and framework for industry approaches to ISS commercial utilization today and for NASA's and the International Partners' planned accommodation of private sector interests and desires on the ISS. This paper chronicles major industry initiatives for commercial ISS utilization, emphasizing successful marketing and business approaches and why these approaches have a higher likelihood of success than others. It provides an overview of individual companies' initiatives, as well as collaborative efforts that cross company lines and country borders; and it assesses the relative success of each. Rather than emphasize negative issues and barriers, this paper characterizes and prioritizes actionable success factors for industry and government to make ISS commercial utilization a sustainable reality.

  13. Multiple graph regularized protein domain ranking

    KAUST Repository

    Wang, Jim Jing-Yan

    2012-11-19

    Background: Protein domain ranking is a fundamental task in structural biology. Most protein domain ranking methods rely on the pairwise comparison of protein domains while neglecting the global manifold structure of the protein domain database. Recently, graph regularized ranking that exploits the global structure of the graph defined by the pairwise similarities has been proposed. However, the existing graph regularized ranking methods are very sensitive to the choice of the graph model and parameters, and this remains a difficult problem for most of the protein domain ranking methods.Results: To tackle this problem, we have developed the Multiple Graph regularized Ranking algorithm, MultiG-Rank. Instead of using a single graph to regularize the ranking scores, MultiG-Rank approximates the intrinsic manifold of protein domain distribution by combining multiple initial graphs for the regularization. Graph weights are learned with ranking scores jointly and automatically, by alternately minimizing an objective function in an iterative algorithm. Experimental results on a subset of the ASTRAL SCOP protein domain database demonstrate that MultiG-Rank achieves a better ranking performance than single graph regularized ranking methods and pairwise similarity based ranking methods.Conclusion: The problem of graph model and parameter selection in graph regularized protein domain ranking can be solved effectively by combining multiple graphs. This aspect of generalization introduces a new frontier in applying multiple graphs to solving protein domain ranking applications. 2012 Wang et al; licensee BioMed Central Ltd.

  14. Building blocks for protein interaction devices

    Science.gov (United States)

    Grünberg, Raik; Ferrar, Tony S.; van der Sloot, Almer M.; Constante, Marco; Serrano, Luis

    2010-01-01

    Here, we propose a framework for the design of synthetic protein networks from modular protein–protein or protein–peptide interactions and provide a starter toolkit of protein building blocks. Our proof of concept experiments outline a general work flow for part–based protein systems engineering. We streamlined the iterative BioBrick cloning protocol and assembled 25 synthetic multidomain proteins each from seven standardized DNA fragments. A systematic screen revealed two main factors controlling protein expression in Escherichia coli: obstruction of translation initiation by mRNA secondary structure or toxicity of individual domains. Eventually, 13 proteins were purified for further characterization. Starting from well-established biotechnological tools, two general–purpose interaction input and two readout devices were built and characterized in vitro. Constitutive interaction input was achieved with a pair of synthetic leucine zippers. The second interaction was drug-controlled utilizing the rapamycin-induced binding of FRB(T2098L) to FKBP12. The interaction kinetics of both devices were analyzed by surface plasmon resonance. Readout was based on Förster resonance energy transfer between fluorescent proteins and was quantified for various combinations of input and output devices. Our results demonstrate the feasibility of parts-based protein synthetic biology. Additionally, we identify future challenges and limitations of modular design along with approaches to address them. PMID:20215443

  15. Multiple graph regularized protein domain ranking

    KAUST Repository

    Wang, Jim Jing-Yan; Bensmail, Halima; Gao, Xin

    2012-01-01

    Background: Protein domain ranking is a fundamental task in structural biology. Most protein domain ranking methods rely on the pairwise comparison of protein domains while neglecting the global manifold structure of the protein domain database. Recently, graph regularized ranking that exploits the global structure of the graph defined by the pairwise similarities has been proposed. However, the existing graph regularized ranking methods are very sensitive to the choice of the graph model and parameters, and this remains a difficult problem for most of the protein domain ranking methods.Results: To tackle this problem, we have developed the Multiple Graph regularized Ranking algorithm, MultiG-Rank. Instead of using a single graph to regularize the ranking scores, MultiG-Rank approximates the intrinsic manifold of protein domain distribution by combining multiple initial graphs for the regularization. Graph weights are learned with ranking scores jointly and automatically, by alternately minimizing an objective function in an iterative algorithm. Experimental results on a subset of the ASTRAL SCOP protein domain database demonstrate that MultiG-Rank achieves a better ranking performance than single graph regularized ranking methods and pairwise similarity based ranking methods.Conclusion: The problem of graph model and parameter selection in graph regularized protein domain ranking can be solved effectively by combining multiple graphs. This aspect of generalization introduces a new frontier in applying multiple graphs to solving protein domain ranking applications. 2012 Wang et al; licensee BioMed Central Ltd.

  16. Multiple graph regularized protein domain ranking.

    Science.gov (United States)

    Wang, Jim Jing-Yan; Bensmail, Halima; Gao, Xin

    2012-11-19

    Protein domain ranking is a fundamental task in structural biology. Most protein domain ranking methods rely on the pairwise comparison of protein domains while neglecting the global manifold structure of the protein domain database. Recently, graph regularized ranking that exploits the global structure of the graph defined by the pairwise similarities has been proposed. However, the existing graph regularized ranking methods are very sensitive to the choice of the graph model and parameters, and this remains a difficult problem for most of the protein domain ranking methods. To tackle this problem, we have developed the Multiple Graph regularized Ranking algorithm, MultiG-Rank. Instead of using a single graph to regularize the ranking scores, MultiG-Rank approximates the intrinsic manifold of protein domain distribution by combining multiple initial graphs for the regularization. Graph weights are learned with ranking scores jointly and automatically, by alternately minimizing an objective function in an iterative algorithm. Experimental results on a subset of the ASTRAL SCOP protein domain database demonstrate that MultiG-Rank achieves a better ranking performance than single graph regularized ranking methods and pairwise similarity based ranking methods. The problem of graph model and parameter selection in graph regularized protein domain ranking can be solved effectively by combining multiple graphs. This aspect of generalization introduces a new frontier in applying multiple graphs to solving protein domain ranking applications.

  17. Dietary protein intake and chronic kidney disease.

    Science.gov (United States)

    Ko, Gang Jee; Obi, Yoshitsugu; Tortorici, Amanda R; Kalantar-Zadeh, Kamyar

    2017-01-01

    High-protein intake may lead to increased intraglomerular pressure and glomerular hyperfiltration. This can cause damage to glomerular structure leading to or aggravating chronic kidney disease (CKD). Hence, a low-protein diet (LPD) of 0.6-0.8 g/kg/day is often recommended for the management of CKD. We reviewed the effect of protein intake on incidence and progression of CKD and the role of LPD in the CKD management. Actual dietary protein consumption in CKD patients remains substantially higher than the recommendations for LPD. Notwithstanding the inconclusive results of the 'Modification of Diet in Renal Disease' (MDRD) study, the largest randomized controlled trial to examine protein restriction in CKD, several prior and subsequent studies and meta-analyses appear to support the role of LPD on retarding progression of CKD and delaying initiation of maintenance dialysis therapy. LPD can also be used to control metabolic derangements in CKD. Supplemented LPD with essential amino acids or their ketoanalogs may be used for incremental transition to dialysis especially on nondialysis days. The LPD management in lieu of dialysis therapy can reduce costs, enhance psychological adaptation, and preserve residual renal function upon transition to dialysis. Adherence and adequate protein and energy intake should be ensured to avoid protein-energy wasting. A balanced and individualized dietary approach based on LPD should be elaborated with periodic dietitian counseling and surveillance to optimize management of CKD, to assure adequate protein and energy intake, and to avoid or correct protein-energy wasting.

  18. Structural analysis of recombinant human protein QM

    International Nuclear Information System (INIS)

    Gualberto, D.C.H.; Fernandes, J.L.; Silva, F.S.; Saraiva, K.W.; Affonso, R.; Pereira, L.M.; Silva, I.D.C.G.

    2012-01-01

    Full text: The ribosomal protein QM belongs to a family of ribosomal proteins, which is highly conserved from yeast to humans. The presence of the QM protein is necessary for joining the 60S and 40S subunits in a late step of the initiation of mRNA translation. Although the exact extra-ribosomal functions of QM are not yet fully understood, it has been identified as a putative tumor suppressor. This protein was reported to interact with the transcription factor c-Jun and thereby prevent c-Jun actives genes of the cellular growth. In this study, the human QM protein was expressed in bacterial system, in the soluble form and this structure was analyzed by Circular Dichroism and Fluorescence. The results of Circular Dichroism showed that this protein has less alpha helix than beta sheet, as described in the literature. QM protein does not contain a leucine zipper region; however the ion zinc is necessary for binding of QM to c-Jun. Then we analyzed the relationship between the removal of zinc ions and folding of protein. Preliminary results obtained by the technique Fluorescence showed a gradual increase in fluorescence with the addition of increasing concentration of EDTA. This suggests that the zinc is important in the tertiary structure of the protein. More studies are being made for better understand these results. (author)

  19. Multiple graph regularized protein domain ranking

    Directory of Open Access Journals (Sweden)

    Wang Jim

    2012-11-01

    Full Text Available Abstract Background Protein domain ranking is a fundamental task in structural biology. Most protein domain ranking methods rely on the pairwise comparison of protein domains while neglecting the global manifold structure of the protein domain database. Recently, graph regularized ranking that exploits the global structure of the graph defined by the pairwise similarities has been proposed. However, the existing graph regularized ranking methods are very sensitive to the choice of the graph model and parameters, and this remains a difficult problem for most of the protein domain ranking methods. Results To tackle this problem, we have developed the Multiple Graph regularized Ranking algorithm, MultiG-Rank. Instead of using a single graph to regularize the ranking scores, MultiG-Rank approximates the intrinsic manifold of protein domain distribution by combining multiple initial graphs for the regularization. Graph weights are learned with ranking scores jointly and automatically, by alternately minimizing an objective function in an iterative algorithm. Experimental results on a subset of the ASTRAL SCOP protein domain database demonstrate that MultiG-Rank achieves a better ranking performance than single graph regularized ranking methods and pairwise similarity based ranking methods. Conclusion The problem of graph model and parameter selection in graph regularized protein domain ranking can be solved effectively by combining multiple graphs. This aspect of generalization introduces a new frontier in applying multiple graphs to solving protein domain ranking applications.

  20. The next generation safeguards initiative

    International Nuclear Information System (INIS)

    Tobey, William

    2008-01-01

    NGSI or the Next Generation Safeguards Initiative is designed to revitalize the U.S. safeguards technical base, as well as invest in human resources, and to mobilize our primary asset - the U.S. National Laboratories - as well as industry and academia to restore capabilities. While NGSI is a U.S. effort it is intended to serve as a catalyst for a much broader commitment to international safeguards in partnership with the IAEA and other countries. Initiatives over the last years include such as the Proliferation Security Initiative, UN Security Council Resolution 1540, the Global Initiative to Combat Nuclear Terrorism, and initiatives of the G-8 and NSG to discourage the spread of enrichment and reprocessing. NGSI augments this agenda by providing a means to strengthen the technical and political underpinnings of IAEA safeguards. Priorities and envisioned activities under NGSI are the following. (1) Cooperation with IAEA and others to promote universal adoption of safeguards agreements and the Additional Protocol including greater information sharing between member states and the IAEA, investigation of weaponization and procurement activities, and options to strengthen the state-level approach to safeguards. (2) NGSI anticipates the deployment of new types of reactors and fuel cycle facilities, as well as the need to use limited safeguards resources effectively and efficiently, especially in plants that pose the largest burden specifically complex, bulk-handling facilities. (3) NGSI will encourage a generational improvement in current safeguards technologies including improvement of precision and speed of nuclear measurements, performance of real-time process monitoring and surveillance in unattended mode, enabling in-field, pre-screening and analysis of nuclear and environmental samples, and collection, integration, analysis and archiving safeguards-relevant information from all available sources.(4) NGSI will address human capital management. Training and

  1. Protein sequence comparison and protein evolution

    Energy Technology Data Exchange (ETDEWEB)

    Pearson, W.R. [Univ. of Virginia, Charlottesville, VA (United States). Dept. of Biochemistry

    1995-12-31

    This tutorial was one of eight tutorials selected to be presented at the Third International Conference on Intelligent Systems for Molecular Biology which was held in the United Kingdom from July 16 to 19, 1995. This tutorial examines how the information conserved during the evolution of a protein molecule can be used to infer reliably homology, and thus a shared proteinfold and possibly a shared active site or function. The authors start by reviewing a geological/evolutionary time scale. Next they look at the evolution of several protein families. During the tutorial, these families will be used to demonstrate that homologous protein ancestry can be inferred with confidence. They also examine different modes of protein evolution and consider some hypotheses that have been presented to explain the very earliest events in protein evolution. The next part of the tutorial will examine the technical aspects of protein sequence comparison. Both optimal and heuristic algorithms and their associated parameters that are used to characterize protein sequence similarities are discussed. Perhaps more importantly, they survey the statistics of local similarity scores, and how these statistics can both be used to improve the selectivity of a search and to evaluate the significance of a match. They them examine distantly related members of three protein families, the serine proteases, the glutathione transferases, and the G-protein-coupled receptors (GCRs). Finally, the discuss how sequence similarity can be used to examine internal repeated or mosaic structures in proteins.

  2. Protein Structure Prediction by Protein Threading

    Science.gov (United States)

    Xu, Ying; Liu, Zhijie; Cai, Liming; Xu, Dong

    The seminal work of Bowie, Lüthy, and Eisenberg (Bowie et al., 1991) on "the inverse protein folding problem" laid the foundation of protein structure prediction by protein threading. By using simple measures for fitness of different amino acid types to local structural environments defined in terms of solvent accessibility and protein secondary structure, the authors derived a simple and yet profoundly novel approach to assessing if a protein sequence fits well with a given protein structural fold. Their follow-up work (Elofsson et al., 1996; Fischer and Eisenberg, 1996; Fischer et al., 1996a,b) and the work by Jones, Taylor, and Thornton (Jones et al., 1992) on protein fold recognition led to the development of a new brand of powerful tools for protein structure prediction, which we now term "protein threading." These computational tools have played a key role in extending the utility of all the experimentally solved structures by X-ray crystallography and nuclear magnetic resonance (NMR), providing structural models and functional predictions for many of the proteins encoded in the hundreds of genomes that have been sequenced up to now.

  3. Eukaryotic Initiation Factor 4H Is under Transcriptional Control of p65/NF-κB

    Science.gov (United States)

    Fiume, Giuseppe; Rossi, Annalisa; de Laurentiis, Annamaria; Falcone, Cristina; Pisano, Antonio; Vecchio, Eleonora; Pontoriero, Marilena; Scala, Iris; Scialdone, Annarita; Masci, Francesca Fasanella; Mimmi, Selena; Palmieri, Camillo; Scala, Giuseppe; Quinto, Ileana

    2013-01-01

    Protein synthesis is mainly regulated at the initiation step, allowing the fast, reversible and spatial control of gene expression. Initiation of protein synthesis requires at least 13 translation initiation factors to assemble the 80S ribosomal initiation complex. Loss of translation control may result in cell malignant transformation. Here, we asked whether translational initiation factors could be regulated by NF-κB transcription factor, a major regulator of genes involved in cell proliferation, survival, and inflammatory response. We show that the p65 subunit of NF-κB activates the transcription of eIF4H gene, which is the regulatory subunit of eIF4A, the most relevant RNA helicase in translation initiation. The p65-dependent transcriptional activation of eIF4H increased the eIF4H protein content augmenting the rate of global protein synthesis. In this context, our results provide novel insights into protein synthesis regulation in response to NF-κB activation signalling, suggesting a transcription-translation coupled mechanism of control. PMID:23776612

  4. Silibinin inhibits translation initiation: implications for anticancer therapy.

    Science.gov (United States)

    Lin, Chen-Ju; Sukarieh, Rami; Pelletier, Jerry

    2009-06-01

    Silibinin is a nontoxic flavonoid reported to have anticancer properties. In this study, we show that silibinin exhibits antiproliferative activity on MCF-7 breast cancer cells. Exposure to silibinin leads to a concentration-dependent decrease in global protein synthesis associated with reduced levels of eukaryotic initiation factor 4F complex. Moreover, polysome profile analysis of silibinin-treated cells shows a decrease in polysome content and translation of cyclin D1 mRNA. Silibinin exerts its effects on translation initiation by inhibiting the mammalian target of rapamycin signaling pathway by acting upstream of TSC2. Our results show that silibinin blocks mammalian target of rapamycin signaling with a concomitant reduction in translation initiation, thus providing a possible molecular mechanism of how silibinin can inhibit growth of transformed cells.

  5. Microsoft Biology Initiative: .NET Bioinformatics Platform and Tools

    Science.gov (United States)

    Diaz Acosta, B.

    2011-01-01

    The Microsoft Biology Initiative (MBI) is an effort in Microsoft Research to bring new technology and tools to the area of bioinformatics and biology. This initiative is comprised of two primary components, the Microsoft Biology Foundation (MBF) and the Microsoft Biology Tools (MBT). MBF is a language-neutral bioinformatics toolkit built as an extension to the Microsoft .NET Framework—initially aimed at the area of Genomics research. Currently, it implements a range of parsers for common bioinformatics file formats; a range of algorithms for manipulating DNA, RNA, and protein sequences; and a set of connectors to biological web services such as NCBI BLAST. MBF is available under an open source license, and executables, source code, demo applications, documentation and training materials are freely downloadable from http://research.microsoft.com/bio. MBT is a collection of tools that enable biology and bioinformatics researchers to be more productive in making scientific discoveries.

  6. Women's Health Initiative (WHI) Background and Overview

    Science.gov (United States)

    ... Back To The Science / Women’s Health Initiative (WHI) Women’s Health Initiative (WHI) Project began 1991 Point of ... What is the goal of the WHI? The Women's Health Initiative (WHI), sponsored by the National Heart, ...

  7. 75 FR 11837 - Chesapeake Bay Watershed Initiative

    Science.gov (United States)

    2010-03-12

    ... DEPARTMENT OF AGRICULTURE Commodity Credit Corporation Chesapeake Bay Watershed Initiative AGENCY...: Notice of availability of program funds for the Chesapeake Bay Watershed Initiative. SUMMARY: The... through the Chesapeake Bay Watershed Initiative for agricultural producers in the Chesapeake Bay watershed...

  8. Polymer Directed Protein Assemblies

    NARCIS (Netherlands)

    van Rijn, Patrick

    2013-01-01

    Protein aggregation and protein self-assembly is an important occurrence in natural systems, and is in some form or other dictated by biopolymers. Very obvious influences of biopolymers on protein assemblies are, e. g., virus particles. Viruses are a multi-protein assembly of which the morphology is

  9. Amino acids and proteins

    Science.gov (United States)

    A balanced, safe diet with proteins is important to meet nutritional requirements. Proteins occur in animal as well as vegetable products in important quantities. In some countries, many people obtain much of their protein from animal products. In other regions, the major portion of dietary protein ...

  10. Characterization of DNA-binding proteins from pea mitochondria

    DEFF Research Database (Denmark)

    Hatzack, F.A.; Dombrowski, S.; Brennicke, A.

    1998-01-01

    We studied transcription initiation in the mitochondria of higher plants, with particular respect to promoter structures. Conserved elements of these promoters have been successfully identified by in vitro transcription systems in different species, whereas the involved protein components are still...

  11. Hepatitis C virus internal ribosome entry site initiates protein synthesis at the authentic initiation codon in yeast

    Czech Academy of Sciences Publication Activity Database

    Mašek, T.; Vopálenský, V.; Horváth, Ondřej; Vortelová, L.; Feketová, Z.; Pospíšek, M.

    2007-01-01

    Roč. 88, č. 7 (2007), s. 1992-2002 ISSN 0022-1317 Grant - others:Grantová agentura UK(CZ) 251/2004/B-BIO/PrF; GA ČR(CZ) GA204/03/1487; GA ČR(CZ) GA301/07/0607 Institutional research plan: CEZ:AV0Z50520514 Keywords : IRES * hepatitida C * kvasinky Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.120, year: 2007

  12. Complement Activation by Ceramide Transporter Proteins

    NARCIS (Netherlands)

    Bode, G.H.; Losen, M.; Buurman, W.A.; Veerhuis, R.; Molenaar, P.C.; Steinbusch, H.W.M.; De Baets, M.H.; Daha, MR; Martinez-Martinez, P.

    2014-01-01

    C1q is the initiator of the classical complement pathway and, as such, is essential for efficient opsonization and clearance of pathogens, altered self-structures, and apoptotic cells. The ceramide transporter protein (CERT) and its longer splicing isoform CERTL are known to interact with

  13. The Protein Model Portal

    OpenAIRE

    Arnold, Konstantin; Kiefer, Florian; Kopp, J?rgen; Battey, James N. D.; Podvinec, Michael; Westbrook, John D.; Berman, Helen M.; Bordoli, Lorenza; Schwede, Torsten

    2008-01-01

    Structural Genomics has been successful in determining the structures of many unique proteins in a high throughput manner. Still, the number of known protein sequences is much larger than the number of experimentally solved protein structures. Homology (or comparative) modeling methods make use of experimental protein structures to build models for evolutionary related proteins. Thereby, experimental structure determination efforts and homology modeling complement each other in the exploratio...

  14. HST Replacement Battery Initial Performance

    Science.gov (United States)

    Krol, Stan; Waldo, Greg; Hollandsworth, Roger

    2009-01-01

    The Hubble Space Telescope (HST) original Nickel-Hydrogen (NiH2) batteries were replaced during the Servicing Mission 4 (SM4) after 19 years and one month on orbit.The purpose of this presentation is to highlight the findings from the assessment of the initial sm4 replacement battery performance. The batteries are described, the 0 C capacity is reviewed, descriptions, charts and tables reviewing the State Of Charge (SOC) Performance, the Battery Voltage Performance, the battery impedance, the minimum voltage performance, the thermal performance, the battery current, and the battery system recharge ratio,

  15. Initial CAD investigations for NET

    International Nuclear Information System (INIS)

    Katz, F.; Leinemann, K.; Ludwig, A.; Marek, U.; Olbrich, W.; Schlechtendahl, E.G.

    1985-11-01

    This report summarizes the work done under contract no. 164/84-7/FU-D-/NET between the Commission of the European Communities and KfK during the period from June 1, 1984, through May 31, 1985. The following topics are covered in this report: Initial modelling of NET version NET2A, CAD system extension for remote handling studies, analysis of the CAD information structure, work related to the transfer of CAD information between KfK and the NET team. (orig.) [de

  16. Powering the Space Exploration Initiative

    International Nuclear Information System (INIS)

    Bennett, G.L.

    1991-01-01

    The Space Exploration Initiative (SEI) establishes the long-term goal of returning to the Moon and then exploring Mars. One of the prerequisites of SEI is the Exploration Technology Program which includes program elements on space nuclear power and surface solar power. These program elements in turn build upon the ongoing NASA research and technology base program in space energy conversion. There is a wide range of missions in NASA's strategic planning and most would benefit from power sources with improved efficiency, lighter weight and reduced cost

  17. Initial treatment of Parkinson's disease.

    Science.gov (United States)

    Tarsy, Daniel

    2006-05-01

    Initial treatment of early idiopathic Parkinson's disease (PD) begins with diagnosis based on clinical evaluation supplemented by laboratory studies and brain imaging to exclude causes of secondary parkinsonism. In most cases, testing is normal and the diagnosis of PD rests on clinical criteria. In patients with mild symptoms and signs, the diagnosis of PD may not initially be apparent, and follow-up evaluation is needed to arrive at a diagnosis. Once the diagnosis is made, pharmacologic treatment may not be the first step. First, patient education is essential, especially because PD is a high-profile disease for which information and misinformation are readily available to patients and families. Counseling concerning prognosis, future symptoms, future disability, and treatment must be provided. Questions from patients concerning diet, lifestyle, and exercise are especially common at this point. The decision of when to initiate treatment is the next major consideration. Much controversy but relatively little light has been brought to bear on this issue. L-dopa was the first major antiparkinson medication to be introduced and remains the "gold standard" of treatment. Next in efficacy are the dopamine agonists (DAs). A debate has raged concerning whether initial dopaminergic treatment should be with L-dopa or DAs. Physicians have been concerned about forestalling the appearance of dyskinesias and motor fluctuations, whereas patients have incorrectly understood that L-dopa and possibly other antiparkinson drugs have a finite duration of usefulness, making it important to defer treatment for as long as possible. This has created "L-dopa phobia," which may stand in the way of useful treatment. In spite of this controversy, there is uniform agreement that the appropriate time to treat is when the patient is beginning to be disabled. This varies from patient to patient and depends on age, employment status, nature of job, level of physical activity, concern about

  18. Interstellar Initiative Web Page Design

    Science.gov (United States)

    Mehta, Alkesh

    1999-01-01

    This summer at NASA/MSFC, I have contributed to two projects: Interstellar Initiative Web Page Design and Lenz's Law Relative Motion Demonstration. In the Web Design Project, I worked on an Outline. The Web Design Outline was developed to provide a foundation for a Hierarchy Tree Structure. The Outline would help design a Website information base for future and near-term missions. The Website would give in-depth information on Propulsion Systems and Interstellar Travel. The Lenz's Law Relative Motion Demonstrator is discussed in this volume by Russell Lee.

  19. [Initial stages of steel biocorrosion].

    Science.gov (United States)

    Zhigletsova, S K; Rodin, V B; Kobelev, V S; Aleksandrova, N V; Rasulova, G E; Kholodenko, V P

    2000-01-01

    Initial stages of corrosion of mild steel induced by Klebsiela rhinoscleromatis BO2 were studied in various media. The effect of the microorganism was detected 8-10 h after inoculation. The number of viable cells were virtually unchanged within one month in all media, but the corrosive activity of the strain decreased. The corrosive activity of microorganisms can be determined by spectrophotometry even only after incubation for 24 h. At a low level of organic substrate, even strong colonization with microorganisms does not inevitably result in a significant damage to metals.

  20. Radiation safety without borders initiative

    International Nuclear Information System (INIS)

    Dibblee, Martha; Dickson, Howard; Krieger, Ken; Lopez, Jose; Waite, David; Weaver, Ken

    2008-01-01

    The Radiation Safety Without Borders (RSWB) initiative provides peer support to radiation safety professionals in developing countries, which bolsters the country's infrastructure and may lead the way for IRPA Associate membership. The Health Physics Society (HPS) recognizes that many nations do not possess the infrastructure to adequately control and beneficially use ionizing radiation. In a substantial number of countries, organized radiation protection programs are minimal. The RSWB initiative relies on HPS volunteers to assist their counterparts in developing countries with emerging health physics and radiation safety programs, but whose resources are limited, to provide tools that promote and support infrastructure and help these professionals help themselves. RSWB experience to date has shown that by providing refurbished instruments, promoting visits to a HPS venue, or visiting a country just to look provide valuable technical and social infrastructure experiences often missing in the developing nation's cadre of radiation safety professionals. HPS/RSWB with the assistance of the International Atomic Energy Agency (IAEA) pairs chapters with a country, with the expectation that the country's professional radiation safety personnel will form a foreign HPS chapter, and the country eventually will become an IRPA Associate. Although still in its formative stage, RSWB nonetheless has gotten valuable information in spite of the small number of missions. The RSWB initiative continues to have significant beneficial impacts, including: Improving the radiation safety infrastructure of the countries that participate; Assisting those countries without professional radiation safety societies to form one; Strengthening the humanitarian efforts of the United States; Enhancing Homeland Security efforts through improved control of radioactive material internationally. Developing countries, including those in Latin America, underwritten by IAEA, may take advantage of resources

  1. Protein- protein interaction detection system using fluorescent protein microdomains

    Science.gov (United States)

    Waldo, Geoffrey S.; Cabantous, Stephanie

    2010-02-23

    The invention provides a protein labeling and interaction detection system based on engineered fragments of fluorescent and chromophoric proteins that require fused interacting polypeptides to drive the association of the fragments, and further are soluble and stable, and do not change the solubility of polypeptides to which they are fused. In one embodiment, a test protein X is fused to a sixteen amino acid fragment of GFP (.beta.-strand 10, amino acids 198-214), engineered to not perturb fusion protein solubility. A second test protein Y is fused to a sixteen amino acid fragment of GFP (.beta.-strand 11, amino acids 215-230), engineered to not perturb fusion protein solubility. When X and Y interact, they bring the GFP strands into proximity, and are detected by complementation with a third GFP fragment consisting of GFP amino acids 1-198 (strands 1-9). When GFP strands 10 and 11 are held together by interaction of protein X and Y, they spontaneous association with GFP strands 1-9, resulting in structural complementation, folding, and concomitant GFP fluorescence.

  2. University involvement in sustainability initiatives

    DEFF Research Database (Denmark)

    Lehmann, Martin; Christensen, Per; Thrane, Mikkel

    2007-01-01

    With an outset in the case of Aalborg University, the linkages (actual, potential and missing) between academic work in research & education and sustainability initiatives are explored. The focus is both on the university's core activities as a provider of research and education and on the univer......With an outset in the case of Aalborg University, the linkages (actual, potential and missing) between academic work in research & education and sustainability initiatives are explored. The focus is both on the university's core activities as a provider of research and education...... and on the university's activities as an organisation, i.e. its own operation and its accounting for this. Sustainability is defined as a continuous process requiring balance between (the emergence of) problems and our ability and capability to solve them. Some core questions that this paper seeks to answer are: "How...... is Sustainable Development understood at executive level at the university? How (if at all) is sustainable development integrated in the core activities of the university? How is the university attached to ‘real life outside the ivory tower', e.g. through the establishing of Public-Private-Academic Partnerships...

  3. Phillips Laboratory small satellite initiatives

    Science.gov (United States)

    Lutey, Mark K.; Imler, Thomas A.; Davis, Robert J.

    1993-09-01

    The Phillips Laboratory Space Experiments Directorate in conjunction with the Air Force Space Test Program (AF STP), Defense Advanced Research and Projects Agency (DARPA) and Strategic Defense Initiative Organization (SDIO), are managing five small satellite program initiatives: Lightweight Exo-Atmospheric Projectile (LEAP) sponsored by SDIO, Miniature Sensor Technology Integration (MSTI) sponsored by SDIO, Technology for Autonomous Operational Survivability (TAOS) sponsored by Phillips Laboratory, TechSat sponsored by SDIO, and the Advanced Technology Standard Satellite Bus (ATSSB) sponsored by DARPA. Each of these spacecraft fulfills a unique set of program requirements. These program requirements range from a short-lived `one-of-a-kind' mission to the robust multi- mission role. Because of these diverging requirements, each program is driven to use a different design philosophy. But regardless of their design, there is the underlying fact that small satellites do not always equate to small missions. These spacecraft with their use of or ability to insert new technologies provide more capabilities and services for their respective payloads which allows the expansion of their mission role. These varying program efforts culminate in an ATSSB spacecraft bus approach that will support moderate size payloads, up to 500 pounds, in a large set of orbits while satisfying the `cheaper, faster, better' method of doing business. This technical paper provides an overview of each of the five spacecraft, focusing on the objectives, payoffs, technologies demonstrated, and program status.

  4. The Saudi Initiative for asthma

    Directory of Open Access Journals (Sweden)

    Al-Moamary Mohamed

    2009-01-01

    Full Text Available The Saudi Initiative for Asthma (SINA provides up-to-date guidelines for healthcare workers managing patients with asthma. SINA was developed by a panel of Saudi experts with respectable academic backgrounds and long-standing experience in the field. SINA is founded on the latest available evidence, local literature, and knowledge of the current setting in Saudi Arabia. Emphasis is placed on understanding the epidemiology, pathophysiology, medications, and clinical presentation. SINA elaborates on the development of patient-doctor partnership, self-management, and control of precipitating factors. Approaches to asthma treatment in SINA are based on disease control by the utilization of Asthma Control Test for the initiation and adjustment of asthma treatment. This guideline is established for the treatment of asthma in both children and adults, with special attention to children 5 years and younger. It is expected that the implementation of these guidelines for treating asthma will lead to better asthma control and decrease patient utilization of the health care system.

  5. Comparing side chain packing in soluble proteins, protein-protein interfaces, and transmembrane proteins.

    Science.gov (United States)

    Gaines, J C; Acebes, S; Virrueta, A; Butler, M; Regan, L; O'Hern, C S

    2018-05-01

    We compare side chain prediction and packing of core and non-core regions of soluble proteins, protein-protein interfaces, and transmembrane proteins. We first identified or created comparable databases of high-resolution crystal structures of these 3 protein classes. We show that the solvent-inaccessible cores of the 3 classes of proteins are equally densely packed. As a result, the side chains of core residues at protein-protein interfaces and in the membrane-exposed regions of transmembrane proteins can be predicted by the hard-sphere plus stereochemical constraint model with the same high prediction accuracies (>90%) as core residues in soluble proteins. We also find that for all 3 classes of proteins, as one moves away from the solvent-inaccessible core, the packing fraction decreases as the solvent accessibility increases. However, the side chain predictability remains high (80% within 30°) up to a relative solvent accessibility, rSASA≲0.3, for all 3 protein classes. Our results show that ≈40% of the interface regions in protein complexes are "core", that is, densely packed with side chain conformations that can be accurately predicted using the hard-sphere model. We propose packing fraction as a metric that can be used to distinguish real protein-protein interactions from designed, non-binding, decoys. Our results also show that cores of membrane proteins are the same as cores of soluble proteins. Thus, the computational methods we are developing for the analysis of the effect of hydrophobic core mutations in soluble proteins will be equally applicable to analyses of mutations in membrane proteins. © 2018 Wiley Periodicals, Inc.

  6. Stress and Protein Turnover in Lemna minor1

    Science.gov (United States)

    Cooke, Robert J.; Oliver, Jane; Davies, David D.

    1979-01-01

    Transfer of fronds of Lemna minor L. to adverse growth conditions or stress situations causes a lowering of the growth rate and a loss of soluble protein per frond, the extent of the loss being dependent on the nature of the stress. The loss or protein is due to two factors: (a) a decrease in the rate constant of protein synthesis (ks); (b) an increase in the rate constant of protein degradation (kd). In plants adapted to the stresses, protein synthesis increases and the initially rapid rate of proteolysis is reduced. Addition of abscisic acid both lowers ks and increases kd, whereas benzyladenine seems to alleviate the effects of stress on protein content by decreasing kd rather than by altering ks. Based on the measurement of enzyme activities, stress-induced protein degradation appears to be a general phenomenon, affecting many soluble proteins. The adaptive significance of stress-induced proteolysis is discussed. PMID:16661102

  7. On the analysis of protein-protein interactions via knowledge-based potentials for the prediction of protein-protein docking

    DEFF Research Database (Denmark)

    Feliu, Elisenda; Aloy, Patrick; Oliva, Baldo

    2011-01-01

    Development of effective methods to screen binary interactions obtained by rigid-body protein-protein docking is key for structure prediction of complexes and for elucidating physicochemical principles of protein-protein binding. We have derived empirical knowledge-based potential functions for s...... and with independence of the partner. This information is encoded at the residue level and could be easily incorporated in the initial grid scoring for Fast Fourier Transform rigid-body docking methods.......Development of effective methods to screen binary interactions obtained by rigid-body protein-protein docking is key for structure prediction of complexes and for elucidating physicochemical principles of protein-protein binding. We have derived empirical knowledge-based potential functions...... for selecting rigid-body docking poses. These potentials include the energetic component that provides the residues with a particular secondary structure and surface accessibility. These scoring functions have been tested on a state-of-art benchmark dataset and on a decoy dataset of permanent interactions. Our...

  8. IGSF9 Family Proteins

    DEFF Research Database (Denmark)

    Hansen, Maria; Walmod, Peter Schledermann

    2013-01-01

    The Drosophila protein Turtle and the vertebrate proteins immunoglobulin superfamily (IgSF), member 9 (IGSF9/Dasm1) and IGSF9B are members of an evolutionarily ancient protein family. A bioinformatics analysis of the protein family revealed that invertebrates contain only a single IGSF9 family gene......, the longest isoforms of the proteins have the same general organization as the neural cell adhesion molecule family of cell adhesion molecule proteins, and like this family of proteins, IGSF9 family members are expressed in the nervous system. A review of the literature revealed that Drosophila Turtle...... facilitates homophilic cell adhesion. Moreover, IGSF9 family proteins have been implicated in the outgrowth and branching of neurites, axon guidance, synapse maturation, self-avoidance, and tiling. However, despite the few published studies on IGSF9 family proteins, reports on the functions of both Turtle...

  9. Personalizing Protein Nourishment

    Science.gov (United States)

    DALLAS, DAVID C.; SANCTUARY, MEGAN R.; QU, YUNYAO; KHAJAVI, SHABNAM HAGHIGHAT; VAN ZANDT, ALEXANDRIA E.; DYANDRA, MELISSA; FRESE, STEVEN A.; BARILE, DANIELA; GERMAN, J. BRUCE

    2016-01-01

    Proteins are not equally digestible—their proteolytic susceptibility varies by their source and processing method. Incomplete digestion increases colonic microbial protein fermentation (putrefaction), which produces toxic metabolites that can induce inflammation in vitro and have been associated with inflammation in vivo. Individual humans differ in protein digestive capacity based on phenotypes, particularly disease states. To avoid putrefaction-induced intestinal inflammation, protein sources and processing methods must be tailored to the consumer’s digestive capacity. This review explores how food processing techniques alter protein digestibility and examines how physiological conditions alter digestive capacity. Possible solutions to improving digestive function or matching low digestive capacity with more digestible protein sources are explored. Beyond the ileal digestibility measurements of protein digestibility, less invasive, quicker and cheaper techniques for monitoring the extent of protein digestion and fermentation are needed to personalize protein nourishment. Biomarkers of protein digestive capacity and efficiency can be identified with the toolsets of peptidomics, metabolomics, microbial sequencing and multiplexed protein analysis of fecal and urine samples. By monitoring individual protein digestive function, the protein component of diets can be tailored via protein source and processing selection to match individual needs to minimize colonic putrefaction and, thus, optimize gut health. PMID:26713355

  10. Prediction of Protein-Protein Interactions Related to Protein Complexes Based on Protein Interaction Networks

    Directory of Open Access Journals (Sweden)

    Peng Liu

    2015-01-01

    Full Text Available A method for predicting protein-protein interactions based on detected protein complexes is proposed to repair deficient interactions derived from high-throughput biological experiments. Protein complexes are pruned and decomposed into small parts based on the adaptive k-cores method to predict protein-protein interactions associated with the complexes. The proposed method is adaptive to protein complexes with different structure, number, and size of nodes in a protein-protein interaction network. Based on different complex sets detected by various algorithms, we can obtain different prediction sets of protein-protein interactions. The reliability of the predicted interaction sets is proved by using estimations with statistical tests and direct confirmation of the biological data. In comparison with the approaches which predict the interactions based on the cliques, the overlap of the predictions is small. Similarly, the overlaps among the predicted sets of interactions derived from various complex sets are also small. Thus, every predicted set of interactions may complement and improve the quality of the original network data. Meanwhile, the predictions from the proposed method replenish protein-protein interactions associated with protein complexes using only the network topology.

  11. The international surface temperature initiative

    Science.gov (United States)

    Thorne, P. W.; Lawrimore, J. H.; Willett, K. M.; Allan, R.; Chandler, R. E.; Mhanda, A.; de Podesta, M.; Possolo, A.; Revadekar, J.; Rusticucci, M.; Stott, P. A.; Strouse, G. F.; Trewin, B.; Wang, X. L.; Yatagai, A.; Merchant, C.; Merlone, A.; Peterson, T. C.; Scott, E. M.

    2013-09-01

    The aim of International Surface Temperature Initiative is to create an end-to-end process for analysis of air temperature data taken over the land surface of the Earth. The foundation of any analysis is the source data. Land surface air temperature records have traditionally been stored in local, organizational, national and international holdings, some of which have been available digitally but many of which are available solely on paper or as imaged files. Further, economic and geopolitical realities have often precluded open sharing of these data. The necessary first step therefore is to collate readily available holdings and augment these over time either through gaining access to previously unavailable digital data or through data rescue and digitization activities. Next, it must be recognized that these historical measurements were made primarily in support of real-time weather applications where timeliness and coverage are key. At almost every long-term station it is virtually certain that changes in instrumentation, siting or observing practices have occurred. Because none of the historical measures were made in a metrologically traceable manner there is no unambiguous way to retrieve the true climate evolution from the heterogeneous raw data holdings. Therefore it is desirable for multiple independent groups to produce adjusted data sets (so-called homogenized data) to adequately understand the data characteristics and estimate uncertainties. Then it is necessary to benchmark the performance of the contributed algorithms (equivalent to metrological software validation) through development of realistic benchmark datasets. In support of this, a series of successive benchmarking and assessment cycles are envisaged, allowing continual improvement while avoiding over-tuning of algorithms. Finally, a portal is proposed giving access to related data-products, utilizing the assessment results to provide guidance to end-users on which product is the most suited to

  12. Biochemistry and pathology of radical-mediated protein oxidation

    DEFF Research Database (Denmark)

    Dean, R T; Fu, S; Stocker, R

    1997-01-01

    Radical-mediated damage to proteins may be initiated by electron leakage, metal-ion-dependent reactions and autoxidation of lipids and sugars. The consequent protein oxidation is O2-dependent, and involves several propagating radicals, notably alkoxyl radicals. Its products include several catego...

  13. Understanding of Protein Synthesis in a Living Cell

    Science.gov (United States)

    Mustapha, Y.; Muhammad, S.

    2006-01-01

    The assembly of proteins takes place in the cytoplasm of a cell. There are three main steps. In initiation, far left, all the necessary parts of the process are brought together by a small molecule called a ribosome. During elongation, amino acids, the building blocks of proteins, are joined to one another in a long chain. The sequence in which…

  14. Mass spectrometry allows direct identification of proteins in large genomes

    DEFF Research Database (Denmark)

    Küster, B; Mortensen, Peter V.; Andersen, Jens S.

    2001-01-01

    Proteome projects seek to provide systematic functional analysis of the genes uncovered by genome sequencing initiatives. Mass spectrometric protein identification is a key requirement in these studies but to date, database searching tools rely on the availability of protein sequences derived fro...

  15. Athoropometric measurements and plasma proteins in protein ...

    African Journals Online (AJOL)

    Athoropometric measurements and plasma proteins in protein energy malnutrition. MH Etukudo, EO Agbedana, OO Akinyinka, BOA Osifo. Abstract. No Abstract. Global Journal of Medical Sciences Vol. 5(1) 2006: 7-11. Full Text: EMAIL FREE FULL TEXT EMAIL FREE FULL TEXT · DOWNLOAD FULL TEXT DOWNLOAD ...

  16. 36 CFR 902.60 - Initial determination.

    Science.gov (United States)

    2010-07-01

    ... 36 Parks, Forests, and Public Property 3 2010-07-01 2010-07-01 false Initial determination. 902.60... INFORMATION ACT Time Limitations § 902.60 Initial determination. (a) An initial determination whether or not... workdays in accordance with § 902.62. (b) Upon making initial determination, the Administrative Officer...

  17. 42 CFR 405.803 - Initial determination.

    Science.gov (United States)

    2010-10-01

    ... 42 Public Health 2 2010-10-01 2010-10-01 false Initial determination. 405.803 Section 405.803....803 Initial determination. (a) Carriers make initial determinations regarding claims for benefits under Medicare Part B. (b) An initial determination for purposes of this subpart includes determinations...

  18. ACHP | Summary of the Preserve America Initiative

    Science.gov (United States)

    Initiative Preserve America: Explore and Enjoy Our Heritage (logo) Summary of the Preserve America Initiative Preserve America is a White House initiative that encourages and supports community efforts to preserve and enjoy our priceless cultural and natural heritage. The goals of the initiative include a greater shared

  19. Global threat reduction initiative (GTRI)

    International Nuclear Information System (INIS)

    Chamberlain, Travis

    2009-01-01

    The Global Threat Reduction Initiative (GTRI) is a vital part of the global efforts to combat nuclear terrorism. GTRI's unique mission to reduce and protect vulnerable nuclear and radiological material located at civilian sites both in the United States and abroad directly addresses recommendations of the bipartisan 9/11 Commission. GTRI efforts are focused on the first line of defense, namely securing or removing vulnerable nuclear and radiological material at the source. The international community has promulgated guidance on the best practice on the technical and administrative aspects of radiological source security, and the GTRI seeks to provide technical assistance to national bodies and individual facilities to adopt this best practice. This presentation will discuss security concepts that are implemented by the GTRI in cooperation with the Australian Nuclear Science and Technology Organization's Regional Security of Radioactive Sources Project. (author)

  20. Teaching Evaluation: Waiting for Initiatives

    Directory of Open Access Journals (Sweden)

    Alejandro Canales Sánchez

    2008-08-01

    Full Text Available In this text, it is sustained that, despite the fact that the teaching activity is one of the main functions of higher education institutions or even the only one in most of them, it hasn’t been reflected in the leading initiatives that have been set in motion in this area for the last two decades. In particular, it points out that the wide evaluation politics established in the education system during the late eighties, didn’t consider the teaching activity as a concern issue for the mechanisms or rewards in the evaluation system. Even though the implementation of new actions tried to repair the situation, mainly by improving the quality of working time and the qualifications of the personnel performing these activities; teaching, in strict sense, and the design or application of a new evaluation scheme to strengthen it, didn’t get better.