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Sample records for assigning large proteins

  1. A Semiautomated Assignment Protocol for Methyl Group Side Chains in Large Proteins.

    Science.gov (United States)

    Kim, Jonggul; Wang, Yingjie; Li, Geoffrey; Veglia, Gianluigi

    2016-01-01

    The developments of biosynthetic specific labeling strategies for side-chain methyl groups have allowed structural and dynamic characterization of very large proteins and protein complexes. However, the assignment of the methyl-group resonances remains an Achilles' heel for NMR, as the experiments designed to correlate side chains to the protein backbone become rather insensitive with the increase of the transverse relaxation rates. In this chapter, we outline a semiempirical approach to assign the resonances of methyl-group side chains in large proteins. This method requires a crystal structure or an NMR ensemble of conformers as an input, together with NMR data sets such as nuclear Overhauser effects (NOEs) and paramagnetic relaxation enhancements (PREs), to be implemented in a computational protocol that provides a probabilistic assignment of methyl-group resonances. As an example, we report the protocol used in our laboratory to assign the side chains of the 42-kDa catalytic subunit of the cAMP-dependent protein kinase A. Although we emphasize the labeling of isoleucine, leucine, and valine residues, this method is applicable to other methyl group side chains such as those of alanine, methionine, and threonine, as well as reductively methylated cysteine side chains.

  2. Assigning large proteins in the solid state: a MAS NMR resonance assignment strategy using selectively and extensively {sup 13}C-labelled proteins

    Energy Technology Data Exchange (ETDEWEB)

    Higman, Victoria A. [Leibniz-Institut fuer Molekulare Pharmakologie (Germany); Flinders, Jeremy [Genentech, Inc., Structural Biology Department (United States); Hiller, Matthias; Jehle, Stefan; Markovic, Stefan; Fiedler, Sebastian; Rossum, Barth-Jan van; Oschkinat, Hartmut [Leibniz-Institut fuer Molekulare Pharmakologie (Germany)], E-mail: oschkinat@fmp-berlin.de

    2009-08-15

    In recent years, solid-state magic-angle spinning nuclear magnetic resonance spectroscopy (MAS NMR) has been growing into an important technique to study the structure of membrane proteins, amyloid fibrils and other protein preparations which do not form crystals or are insoluble. Currently, a key bottleneck is the assignment process due to the absence of the resolving power of proton chemical shifts. Particularly for large proteins (approximately >150 residues) it is difficult to obtain a full set of resonance assignments. In order to address this problem, we present an assignment method based upon samples prepared using [1,3-{sup 13}C]- and [2-{sup 13}C]-glycerol as the sole carbon source in the bacterial growth medium (so-called selectively and extensively labelled protein). Such samples give rise to higher quality spectra than uniformly [{sup 13}C]-labelled protein samples, and have previously been used to obtain long-range restraints for use in structure calculations. Our method exploits the characteristic cross-peak patterns observed for the different amino acid types in {sup 13}C-{sup 13}C correlation and 3D NCACX and NCOCX spectra. An in-depth analysis of the patterns and how they can be used to aid assignment is presented, using spectra of the chicken {alpha}-spectrin SH3 domain (62 residues), {alpha}B-crystallin (175 residues) and outer membrane protein G (OmpG, 281 residues) as examples. Using this procedure, over 90% of the C{alpha}, C{beta}, C' and N resonances in the core domain of {alpha}B-crystallin and around 73% in the flanking domains could be assigned (excluding 24 residues at the extreme termini of the protein)

  3. BEST-TROSY experiments for time-efficient sequential resonance assignment of large disordered proteins

    Energy Technology Data Exchange (ETDEWEB)

    Solyom, Zsofia; Schwarten, Melanie [Universite Grenoble 1, Institut de Biologie Structurale (France); Geist, Leonhard; Konrat, Robert [Max F. Perutz Laboratories, Department of Computational and Structural Biology (Austria); Willbold, Dieter [Commissariat a l' Energie Atomique et aux Energies Alternatives (CEA) (France); Brutscher, Bernhard, E-mail: bernhard.brutscher@ibs.fr [Universite Grenoble 1, Institut de Biologie Structurale (France)

    2013-04-15

    The characterization of the conformational properties of intrinsically disordered proteins (IDPs), and their interaction modes with physiological partners has recently become a major research topic for understanding biological function on the molecular level. Although multidimensional NMR spectroscopy is the technique of choice for the study of IDPs at atomic resolution, the intrinsically low resolution, and the large peak intensity variations often observed in NMR spectra of IDPs call for resolution- and sensitivity-optimized pulse schemes. We present here a set of amide proton-detected 3D BEST-TROSY correlation experiments that yield the required sensitivity and spectral resolution for time-efficient sequential resonance assignment of large IDPs. In addition, we introduce two proline-edited 2D experiments that allow unambiguous identification of residues adjacent to proline that is one of the most abundant amino acids in IDPs. The performance of these experiments, and the advantages of BEST-TROSY pulse schemes are discussed and illustrated for two IDPs of similar length ({approx}270 residues) but with different conformational sampling properties.

  4. Protein secondary structure: category assignment and predictability

    DEFF Research Database (Denmark)

    Andersen, Claus A.; Bohr, Henrik; Brunak, Søren

    2001-01-01

    In the last decade, the prediction of protein secondary structure has been optimized using essentially one and the same assignment scheme known as DSSP. We present here a different scheme, which is more predictable. This scheme predicts directly the hydrogen bonds, which stabilize the secondary...... structures. Single sequence prediction of the new three category assignment gives an overall prediction improvement of 3.1% and 5.1%, compared to the DSSP assignment and schemes where the helix category consists of a-helix and 3(10)-helix, respectively. These results were achieved using a standard feed-forward...

  5. Protein side-chain resonance assignment and NOE assignment using RDC-defined backbones without TOCSY data

    Energy Technology Data Exchange (ETDEWEB)

    Zeng Jianyang [Duke University, Department of Computer Science (United States); Zhou Pei [Duke University Medical Center, Department of Biochemistry (United States); Donald, Bruce Randall [Duke University, Department of Computer Science (United States)

    2011-08-15

    One bottleneck in NMR structure determination lies in the laborious and time-consuming process of side-chain resonance and NOE assignments. Compared to the well-studied backbone resonance assignment problem, automated side-chain resonance and NOE assignments are relatively less explored. Most NOE assignment algorithms require nearly complete side-chain resonance assignments from a series of through-bond experiments such as HCCH-TOCSY or HCCCONH. Unfortunately, these TOCSY experiments perform poorly on large proteins. To overcome this deficiency, we present a novel algorithm, called Nasca (NOE Assignment and Side-Chain Assignment), to automate both side-chain resonance and NOE assignments and to perform high-resolution protein structure determination in the absence of any explicit through-bond experiment to facilitate side-chain resonance assignment, such as HCCH-TOCSY. After casting the assignment problem into a Markov Random Field (MRF), Nasca extends and applies combinatorial protein design algorithms to compute optimal assignments that best interpret the NMR data. The MRF captures the contact map information of the protein derived from NOESY spectra, exploits the backbone structural information determined by RDCs, and considers all possible side-chain rotamers. The complexity of the combinatorial search is reduced by using a dead-end elimination (DEE) algorithm, which prunes side-chain resonance assignments that are provably not part of the optimal solution. Then an A* search algorithm is employed to find a set of optimal side-chain resonance assignments that best fit the NMR data. These side-chain resonance assignments are then used to resolve the NOE assignment ambiguity and compute high-resolution protein structures. Tests on five proteins show that Nasca assigns resonances for more than 90% of side-chain protons, and achieves about 80% correct assignments. The final structures computed using the NOE distance restraints assigned by Nasca have backbone RMSD 0

  6. Protein secondary structure assignment revisited: a detailed analysis of different assignment methods

    Directory of Open Access Journals (Sweden)

    de Brevern Alexandre G

    2005-09-01

    Full Text Available Abstract Background A number of methods are now available to perform automatic assignment of periodic secondary structures from atomic coordinates, based on different characteristics of the secondary structures. In general these methods exhibit a broad consensus as to the location of most helix and strand core segments in protein structures. However the termini of the segments are often ill-defined and it is difficult to decide unambiguously which residues at the edge of the segments have to be included. In addition, there is a "twilight zone" where secondary structure segments depart significantly from the idealized models of Pauling and Corey. For these segments, one has to decide whether the observed structural variations are merely distorsions or whether they constitute a break in the secondary structure. Methods To address these problems, we have developed a method for secondary structure assignment, called KAKSI. Assignments made by KAKSI are compared with assignments given by DSSP, STRIDE, XTLSSTR, PSEA and SECSTR, as well as secondary structures found in PDB files, on 4 datasets (X-ray structures with different resolution range, NMR structures. Results A detailed comparison of KAKSI assignments with those of STRIDE and PSEA reveals that KAKSI assigns slightly longer helices and strands than STRIDE in case of one-to-one correspondence between the segments. However, KAKSI tends also to favor the assignment of several short helices when STRIDE and PSEA assign longer, kinked, helices. Helices assigned by KAKSI have geometrical characteristics close to those described in the PDB. They are more linear than helices assigned by other methods. The same tendency to split long segments is observed for strands, although less systematically. We present a number of cases of secondary structure assignments that illustrate this behavior. Conclusion Our method provides valuable assignments which favor the regularity of secondary structure segments.

  7. Graphical interpretation of Boolean operators for protein NMR assignments.

    Science.gov (United States)

    Verdegem, Dries; Dijkstra, Klaas; Hanoulle, Xavier; Lippens, Guy

    2008-09-01

    We have developed a graphics based algorithm for semi-automated protein NMR assignments. Using the basic sequential triple resonance assignment strategy, the method is inspired by the Boolean operators as it applies "AND"-, "OR"- and "NOT"-like operations on planes pulled out of the classical three-dimensional spectra to obtain its functionality. The method's strength lies in the continuous graphical presentation of the spectra, allowing both a semi-automatic peaklist construction and sequential assignment. We demonstrate here its general use for the case of a folded protein with a well-dispersed spectrum, but equally for a natively unfolded protein where spectral resolution is minimal. PMID:18762868

  8. Graphical interpretation of Boolean operators for protein NMR assignments

    NARCIS (Netherlands)

    Verdegem, Dries; Dijkstra, Klaas; Hanoulle, Xavier; Lippens, Guy

    2008-01-01

    We have developed a graphics based algorithm for semi-automated protein NMR assignments. Using the basic sequential triple resonance assignment strategy, the method is inspired by the Boolean operators as it applies "AND"-, "OR"- and "NOT"-like operations on planes pulled out of the classical three-

  9. Traffic assignment models in large-scale applications

    DEFF Research Database (Denmark)

    Rasmussen, Thomas Kjær

    Transport models are becoming more and more disaggregate to facilitate a realistic representation of individuals and their travel patterns. In line with this development, the PhD study focuses on facilitating the deployment of traffic assignment models in fully disaggregate activity-based model...... focuses on large-scale applications and contributes with methods to actualise the true potential of disaggregate models. To achieve this target, contributions are given to several components of traffic assignment modelling, by (i) enabling the utilisation of the increasingly available data sources......-perceptions in the choice set generation for complex multi-modal networks, and (iv) addressing the difficulty of choice set generation by making available a theoretical framework, and corresponding operational solution methods, which consistently distinguishes between used and unused paths. The availability of data...

  10. Computational Assignment of Chemical Shifts for Protein Residues

    CERN Document Server

    Bratholm, Lars A

    2013-01-01

    Fast and accurate protein structure prediction is one of the major challenges in structural biology, biotechnology and molecular biomedicine. These fields require 3D protein structures for rational design of proteins with improved or novel properties. X-ray crystallography is the most common approach even with its low success rate, but lately NMR based approaches have gained popularity. The general approach involves a set of distance restraints used to guide a structure prediction, but simple NMR triple-resonance experiments often provide enough structural information to predict the structure of small proteins. Previous protein folding simulations that have utilised experimental data have weighted the experimental data and physical force field terms more or less arbitrarily, and the method is thus not generally applicable to new proteins. Furthermore a complete and near error-free assignment of chemical shifts obtained by the NMR experiments is needed, due to the static, or deterministic, assignment. In this ...

  11. A tracked approach for automated NMR assignments in proteins (TATAPRO)

    Energy Technology Data Exchange (ETDEWEB)

    Atreya, H.S.; Sahu, S.C.; Chary, K.V.R.; Govil, Girjesh [Tata Institute of Fundamental Research, Department of Chemical Sciences (India)

    2000-06-15

    A novel automated approach for the sequence specific NMR assignments of {sup 1}H{sup N}, {sup 13}C{sup {alpha}}, {sup 13}C{sup {beta}}, {sup 13}C'/{sup 1}H{sup {alpha}} and {sup 15}N spins in proteins, using triple resonance experimental data, is presented. The algorithm, TATAPRO (Tracked AuTomated Assignments in Proteins) utilizes the protein primary sequence and peak lists from a set of triple resonance spectra which correlate {sup 1}H{sup N} and {sup 15}N chemical shifts with those of {sup 13}C{sup {alpha}}, {sup 13}C{sup {beta}} and {sup 13}C'/{sup 1}H{sup {alpha}}. The information derived from such correlations is used to create a 'master{sub l}ist' consisting of all possible sets of {sup 1}H{sup N}{sub i}, {sup 15}N{sub i}, {sup 13}C{sup {alpha}}{sub i}, {sup 13}C{sup {beta}}{sub i}, {sup 13}C'{sub i}/{sup 1}H{sup {alpha}}{sub i}, {sup 13}C{sup {alpha}}{sub i-1}, {sup 13}C{sup {beta}}{sub i-1} and {sup 13}C'{sub i-1}/ {sup 1}H{sup {alpha}}{sub i-1} chemical shifts. On the basis of an extensive statistical analysis of {sup 13}C{sup {alpha}} and {sup 13}C{sup {beta}} chemical shift data of proteins derived from the BioMagResBank (BMRB), it is shown that the 20 amino acid residues can be grouped into eight distinct categories, each of which is assigned a unique two-digit code. Such a code is used to tag individual sets of chemical shifts in the master{sub l}ist and also to translate the protein primary sequence into an array called pps{sub a}rray. The program then uses the master{sub l}ist to search for neighbouring partners of a given amino acid residue along the polypeptide chain and sequentially assigns a maximum possible stretch of residues on either side. While doing so, each assigned residue is tracked in an array called assig{sub a}rray, with the two-digit code assigned earlier. The assig{sub a}rray is then mapped onto the pps{sub a}rray for sequence specific resonance assignment. The program has been tested using

  12. Assigning protein functions by comparative genome analysis protein phylogenetic profiles

    Science.gov (United States)

    Pellegrini, Matteo; Marcotte, Edward M.; Thompson, Michael J.; Eisenberg, David; Grothe, Robert; Yeates, Todd O.

    2003-05-13

    A computational method system, and computer program are provided for inferring functional links from genome sequences. One method is based on the observation that some pairs of proteins A' and B' have homologs in another organism fused into a single protein chain AB. A trans-genome comparison of sequences can reveal these AB sequences, which are Rosetta Stone sequences because they decipher an interaction between A' and B. Another method compares the genomic sequence of two or more organisms to create a phylogenetic profile for each protein indicating its presence or absence across all the genomes. The profile provides information regarding functional links between different families of proteins. In yet another method a combination of the above two methods is used to predict functional links.

  13. Root-mean-square-deviation-based rapid backbone resonance assignments in proteins.

    Science.gov (United States)

    Rout, Ashok K; Barnwal, Ravi P; Agarwal, Geetika; Chary, Kandala V R

    2010-10-01

    We have shown that the methodology based on the estimation of root-mean-square deviation (RMSD) between two sets of chemical shifts is very useful to rapidly assign the spectral signatures of (1)H(N), (13)C(α), (13)C(β), (13)C', (1)H(α) and (15)N spins of a given protein in one state from the knowledge of its resonance assignments in a different state, without resorting to routine established procedures (manual and automated). We demonstrate the utility of this methodology to rapidly assign the 3D spectra of a metal-binding protein in its holo-state from the knowledge of its assignments in apo-state, the spectra of a protein in its paramagnetic state from the knowledge of its assignments in diamagnetic state and, finally, the spectra of a mutant protein from the knowledge of the chemical shifts of the corresponding wild-type protein. The underlying assumption of this methodology is that, it is impossible for any two amino acid residues in a given protein to have all the six chemical shifts degenerate and that the protein under consideration does not undergo large conformational changes in going from one conformational state to another. The methodology has been tested using experimental data on three proteins, M-crystallin (8.5 kDa, predominantly β-sheet, for apo- to holo-state), Calbindin (7.5 kDa, predominantly α-helical, for diamagnetic to paramagnetic state and apo to holo) and EhCaBP1 (14.3 kDa, α-helical, the wild-type protein with one of its mutant). In all the cases, the extent of assignment is found to be greater than 85%.

  14. MONTE: An automated Monte Carlo based approach to nuclear magnetic resonance assignment of proteins

    Energy Technology Data Exchange (ETDEWEB)

    Hitchens, T. Kevin; Lukin, Jonathan A.; Zhan Yiping; McCallum, Scott A.; Rule, Gordon S. [Carnegie Mellon University, Department of Biological Sciences (United States)], E-mail: rule@andrew.cmu.edu

    2003-01-15

    A general-purpose Monte Carlo assignment program has been developed to aid in the assignment of NMR resonances from proteins. By virtue of its flexible data requirements the program is capable of obtaining assignments of both heavily deuterated and fully protonated proteins. A wide variety of source data, such as inter-residue scalar connectivity, inter-residue dipolar (NOE) connectivity, and residue specific information, can be utilized in the assignment process. The program can also use known assignments from one form of a protein to facilitate the assignment of another form of the protein. This attribute is useful for assigning protein-ligand complexes when the assignments of the unliganded protein are known. The program can be also be used as an interactive research tool to assist in the choice of additional experimental data to facilitate completion of assignments. The assignment of a deuterated 45 kDa homodimeric Glutathione-S-transferase illustrates the principal features of the program.

  15. Automating unambiguous NOE data usage in NVR for NMR protein structure-based assignments.

    Science.gov (United States)

    Akhmedov, Murodzhon; Çatay, Bülent; Apaydın, Mehmet Serkan

    2015-12-01

    Nuclear Magnetic Resonance (NMR) Spectroscopy is an important technique that allows determining protein structure in solution. An important problem in protein structure determination using NMR spectroscopy is the mapping of peaks to corresponding amino acids, also known as the assignment problem. Structure-Based Assignment (SBA) is an approach to solve this problem using a template structure that is homologous to the target. Our previously developed approach Nuclear Vector Replacement-Binary Integer Programming (NVR-BIP) computed the optimal solution for small proteins, but was unable to solve the assignments of large proteins. NVR-Ant Colony Optimization (ACO) extended the applicability of the NVR approach for such proteins. One of the input data utilized in these approaches is the Nuclear Overhauser Effect (NOE) data. NOE is an interaction observed between two protons if the protons are located close in space. These protons could be amide protons, protons attached to the alpha-carbon atom in the backbone of the protein, or side chain protons. NVR only uses backbone protons. In this paper, we reformulate the NVR-BIP model to distinguish the type of proton in NOE data and use the corresponding proton coordinates in the extended formulation. In addition, the threshold value over interproton distances is set in a standard manner for all proteins by extracting the NOE upper bound distance information from the data. We also convert NOE intensities into distance thresholds. Our new approach thus handles the NOE data correctly and without manually determined parameters. We accordingly adapt NVR-ACO solution methodology to these changes. Computational results show that our approaches obtain optimal solutions for small proteins. For the large proteins our ant colony optimization-based approach obtains promising results. PMID:26260854

  16. Solving Large Quadratic|Assignment Problems in Parallel

    DEFF Research Database (Denmark)

    Clausen, Jens; Perregaard, Michael

    1997-01-01

    and recalculation of bounds between branchings when used in a parallel Branch-and-Bound algorithm. The algorithm has been implemented on a 16-processor MEIKO Computing Surface with Intel i860 processors. Computational results from the solution of a number of large QAPs, including the classical Nugent 20...... searched. Much work has been done to identify such functions for the QAP, but with limited success.Parallel processing has also been used in order to increase the size of problems solvable to optimality. The systems used have, however, often been systems with relatively few, but very powerful vector...... processors, and have hence not been ideally suited for computations essentially involving non-vectorizable computations on integers.In this paper we investigate the combination of one of the best bound functions for a Branch-and-Bound algorithm (the Gilmore-Lawler bound) and various testing, variable binding...

  17. Organic Chemistry YouTube Writing Assignment for Large Lecture Classes

    Science.gov (United States)

    Franz, Annaliese K.

    2012-01-01

    This work describes efforts to incorporate and evaluate the use of a YouTube writing assignment in large lecture classes to personalize learning and improve conceptual understanding of chemistry through peer- and self-explanation strategies. Although writing assignments can be a method to incorporate peer- and self-explanation strategies, this…

  18. Towards an intelligent system for the automatic assignment of domains in globular proteins.

    Science.gov (United States)

    Sternberg, M J; Hegyi, H; Islam, S A; Luo, J; Russell, R B

    1995-01-01

    The automatic identification of protein domains from coordinates is the first step in the classification of protein folds and hence is required for databases to guide structure prediction. Most algorithms encode a single concept based and sometimes do not yield assignments that are consistent with the generally accepted perception. Our development of an automatic approach to identify reliably domains from protein coordinates is described. The algorithm is benchmarked against a manual identification of the domains in 284 representative protein chains. The first step is the domain assignment by distance (DAD) algorithm that considers the density of inter-residue contacts represented in a contact matrix. The algorithm yields 85% agreement with the manual assignment. The paper then considers how the reliability of these assignments could be evaluated. Finally the use of structural comparisons using the STAMP algorithm to validate domain assignment is reported on a test case. PMID:7584461

  19. CASA: An Efficient Automated Assignment of Protein Mainchain NMR Data Using an Ordered Tree Search Algorithm

    International Nuclear Information System (INIS)

    Rapid analysis of protein structure, interaction, and dynamics requires fast and automated assignments of 3D protein backbone triple-resonance NMR spectra. We introduce a new depth-first ordered tree search method of automated assignment, CASA, which uses hand-edited peak-pick lists of a flexible number of triple resonance experiments. The computer program was tested on 13 artificially simulated peak lists for proteins up to 723 residues, as well as on the experimental data for four proteins. Under reasonable tolerances, it generated assignments that correspond to the ones reported in the literature within a few minutes of CPU time. The program was also tested on the proteins analyzed by other methods, with both simulated and experimental peaklists, and it could generate good assignments in all relevant cases. The robustness was further tested under various situations

  20. CONSERVATION. Genetic assignment of large seizures of elephant ivory reveals Africa's major poaching hotspots.

    Science.gov (United States)

    Wasser, S K; Brown, L; Mailand, C; Mondol, S; Clark, W; Laurie, C; Weir, B S

    2015-07-01

    Poaching of elephants is now occurring at rates that threaten African populations with extinction. Identifying the number and location of Africa's major poaching hotspots may assist efforts to end poaching and facilitate recovery of elephant populations. We genetically assign origin to 28 large ivory seizures (≥0.5 metric tons) made between 1996 and 2014, also testing assignment accuracy. Results suggest that the major poaching hotspots in Africa may be currently concentrated in as few as two areas. Increasing law enforcement in these two hotspots could help curtail future elephant losses across Africa and disrupt this organized transnational crime.

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

    Energy Technology Data Exchange (ETDEWEB)

    Hansen, A.P.; Curley, R.W. Jr.; Panigot, M.J.; Fesik, S.W. [Ohio State Univ., Columbus, OH (United States)

    1994-12-01

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

  2. Toward Efficient Task Assignment and Motion Planning for Large Scale Underwater Mission

    OpenAIRE

    Zadeh, Somaiyeh Mahmoud.; Powers, David MW; Sammut, Karl; Yazdani, Amirmehdi

    2016-01-01

    An Autonomous Underwater Vehicle (AUV) needs to acquire a certain degree of autonomy for any particular underwater mission to fulfill the mission objectives successfully and ensure its safety in all stages of the mission in a large scale operating filed. In this paper, a novel combinatorial conflict-free-task assignment strategy consisting an interactive engagement of a local path planner and an adaptive global route planner, is introduced. The method is established upon the heuristic search ...

  3. NMR assignment of the arenaviral protein Z from Lassa fever virus.

    Science.gov (United States)

    Volpon, Laurent; Osborne, Michael J; Borden, Katherine L B

    2008-06-01

    The arenavirus protein Z from Lassa fever virus was recently found to inhibit mRNA translation through direct interaction with eIF4E. Here, we report the NMR assignment of this RING-containing protein that was determined by triple resonance NMR techniques. PMID:18958179

  4. NMR assignment of the arenaviral protein Z from Lassa fever virus

    OpenAIRE

    Volpon, Laurent; Osborne, Michael J.; Borden, Katherine L. B.

    2008-01-01

    The arenavirus protein Z from Lassa fever virus was recently found to inhibit mRNA translation through direct interaction with eIF4E. Here, we report the NMR assignment of this RING-containing protein that was determined by triple resonance NMR techniques.

  5. NMR assignment of the arenaviral protein Z from Lassa fever virus

    Science.gov (United States)

    Osborne, Michael J.; Borden, Katherine L.B.

    2008-01-01

    The arenavirus protein Z from Lassa fever virus was recently found to inhibit mRNA translation through direct interaction with eIF4E. Here, we report the NMR assignment of this RING-containing protein that was determined by triple resonance NMR techniques. PMID:18958179

  6. Automated solid-state NMR resonance assignment of protein microcrystals and amyloids

    International Nuclear Information System (INIS)

    Solid-state NMR is an emerging structure determination technique for crystalline and non-crystalline protein assemblies, e.g., amyloids. Resonance assignment constitutes the first and often very time-consuming step to a structure. We present ssFLYA, a generally applicable algorithm for automatic assignment of protein solid-state NMR spectra. Application to microcrystals of ubiquitin and the Ure2 prion C-terminal domain, as well as amyloids of HET-s(218–289) and α-synuclein yielded 88–97 % correctness for the backbone and side-chain assignments that are classified as self-consistent by the algorithm, and 77–90 % correctness if also assignments classified as tentative by the algorithm are included

  7. Automated solid-state NMR resonance assignment of protein microcrystals and amyloids

    Energy Technology Data Exchange (ETDEWEB)

    Schmidt, Elena [Goethe University Frankfurt am Main, Center for Biomolecular Magnetic Resonance, Institute of Biophysical Chemistry (Germany); Gath, Julia [ETH Zurich, Physical Chemistry (Switzerland); Habenstein, Birgit [UMR 5086 CNRS/Universite de Lyon 1, Institut de Biologie et Chimie des Proteines (France); Ravotti, Francesco; Szekely, Kathrin; Huber, Matthias [ETH Zurich, Physical Chemistry (Switzerland); Buchner, Lena [Goethe University Frankfurt am Main, Center for Biomolecular Magnetic Resonance, Institute of Biophysical Chemistry (Germany); Boeckmann, Anja, E-mail: a.bockmann@ibcp.fr [UMR 5086 CNRS/Universite de Lyon 1, Institut de Biologie et Chimie des Proteines (France); Meier, Beat H., E-mail: beme@ethz.ch [ETH Zurich, Physical Chemistry (Switzerland); Guentert, Peter, E-mail: guentert@em.uni-frankfurt.de [Goethe University Frankfurt am Main, Center for Biomolecular Magnetic Resonance, Institute of Biophysical Chemistry (Germany)

    2013-07-15

    Solid-state NMR is an emerging structure determination technique for crystalline and non-crystalline protein assemblies, e.g., amyloids. Resonance assignment constitutes the first and often very time-consuming step to a structure. We present ssFLYA, a generally applicable algorithm for automatic assignment of protein solid-state NMR spectra. Application to microcrystals of ubiquitin and the Ure2 prion C-terminal domain, as well as amyloids of HET-s(218-289) and {alpha}-synuclein yielded 88-97 % correctness for the backbone and side-chain assignments that are classified as self-consistent by the algorithm, and 77-90 % correctness if also assignments classified as tentative by the algorithm are included.

  8. Amino acid selective unlabeling for sequence specific resonance assignments in proteins

    Energy Technology Data Exchange (ETDEWEB)

    Krishnarjuna, B.; Jaipuria, Garima; Thakur, Anushikha [Indian Institute of Science, NMR Research Centre (India); D' Silva, Patrick, E-mail: patrick@biochem.iisc.ernet.in [Indian Institute of Science, Department of Biochemistry (India); Atreya, Hanudatta S., E-mail: hsatreya@sif.iisc.ernet.in [Indian Institute of Science, NMR Research Centre (India)

    2011-01-15

    Sequence specific resonance assignment constitutes an important step towards high-resolution structure determination of proteins by NMR and is aided by selective identification and assignment of amino acid types. The traditional approach to selective labeling yields only the chemical shifts of the particular amino acid being selected and does not help in establishing a link between adjacent residues along the polypeptide chain, which is important for sequential assignments. An alternative approach is the method of amino acid selective 'unlabeling' or reverse labeling, which involves selective unlabeling of specific amino acid types against a uniformly {sup 13}C/{sup 15}N labeled background. Based on this method, we present a novel approach for sequential assignments in proteins. The method involves a new NMR experiment named, {l_brace}{sup 12}CO{sub i}-{sup 15}N{sub i+1}{r_brace}-filtered HSQC, which aids in linking the {sup 1}H{sup N}/{sup 15}N resonances of the selectively unlabeled residue, i, and its C-terminal neighbor, i + 1, in HN-detected double and triple resonance spectra. This leads to the assignment of a tri-peptide segment from the knowledge of the amino acid types of residues: i - 1, i and i + 1, thereby speeding up the sequential assignment process. The method has the advantage of being relatively inexpensive, applicable to {sup 2}H labeled protein and can be coupled with cell-free synthesis and/or automated assignment approaches. A detailed survey involving unlabeling of different amino acid types individually or in pairs reveals that the proposed approach is also robust to misincorporation of {sup 14}N at undesired sites. Taken together, this study represents the first application of selective unlabeling for sequence specific resonance assignments and opens up new avenues to using this methodology in protein structural studies.

  9. Dominant-Negative Proteins in Herpesviruses – From Assigning Gene Function to Intracellular Immunization

    Directory of Open Access Journals (Sweden)

    Zsolt Ruzsics

    2009-10-01

    Full Text Available Investigating and assigning gene functions of herpesviruses is a process, which profits from consistent technical innovation. Cloning of bacterial artificial chromosomes encoding herpesvirus genomes permits nearly unlimited possibilities in the construction of genetically modified viruses. Targeted or randomized screening approaches allow rapid identification of essential viral proteins. Nevertheless, mapping of essential genes reveals only limited insight into function. The usage of dominant-negative (DN proteins has been the tool of choice to dissect functions of proteins during the viral life cycle. DN proteins also facilitate the analysis of host-virus interactions. Finally, DNs serve as starting-point for design of new antiviral strategies.

  10. NMR assignment of the amylase-binding protein A from Streptococcus parasanguinis.

    Science.gov (United States)

    Liu, Bing; Zhu, Fan; Wu, Hui; Matthews, Stephen

    2015-04-01

    Streptococcus parasanguinis is a primary colonizer of tooth surfaces in the oral cavity. Amylase-binding protein A (AbpA) from S. parasanguinis is responsible for the recruitment of salivary amylase to bacterial surface, which plays an important role in the development of oral biofilms. Here, we describe the essentially complete NMR assignments for AbpA. PMID:25016927

  11. Benders' Decomposition Based Heuristics for Large-Scale Dynamic Quadratic Assignment Problems

    Directory of Open Access Journals (Sweden)

    Sirirat Muenvanichakul

    2009-01-01

    Full Text Available Problem statement: Dynamic Quadratic Assignment Problem (DQAP is NP hard problem. Benders decomposition based heuristics method is applied to the equivalent mixed-integer linear programming problem of the original DQAP. Approach: Approximate Benders Decomposition (ABD generates the ensemble of a subset of feasible layout for Approximate Dynamic Programming (ADP to determine the sub-optimal optimal solution. A Trust-Region Constraint (TRC for the master problem in ABD and a Successive Adaptation Procedure (SAP were implemented to accelerate the convergence rate of the method. Results: The sub-optimal solutions of large-scales DQAPs from the method and its variants were compared well with other metaheuristic methods. Conclusion: Overall performance of the method is comparable to other metaheuristic methods for large-scale DQAPs.

  12. Automated analysis of protein NMR assignments using methods from artificial intelligence.

    Science.gov (United States)

    Zimmerman, D E; Kulikowski, C A; Huang, Y; Feng, W; Tashiro, M; Shimotakahara, S; Chien, C; Powers, R; Montelione, G T

    1997-06-20

    An expert system for determining resonance assignments from NMR spectra of proteins is described. Given the amino acid sequence, a two-dimensional 15N-1H heteronuclear correlation spectrum and seven to eight three-dimensional triple-resonance NMR spectra for seven proteins, AUTOASSIGN obtained an average of 98% of sequence-specific spin-system assignments with an error rate of less than 0.5%. Execution times on a Sparc 10 workstation varied from 16 seconds for smaller proteins with simple spectra to one to nine minutes for medium size proteins exhibiting numerous extra spin systems attributed to conformational isomerization. AUTOASSIGN combines symbolic constraint satisfaction methods with a domain-specific knowledge base to exploit the logical structure of the sequential assignment problem, the specific features of the various NMR experiments, and the expected chemical shift frequencies of different amino acids. The current implementation specializes in the analysis of data derived from the most sensitive of the currently available triple-resonance experiments. Potential extensions of the system for analysis of additional types of protein NMR data are also discussed.

  13. APSY-NMR for protein backbone assignment in high-throughput structural biology

    Energy Technology Data Exchange (ETDEWEB)

    Dutta, Samit Kumar; Serrano, Pedro; Proudfoot, Andrew; Geralt, Michael [The Scripps Research Institute, Department of Integrative Structural and Computational Biology (United States); Pedrini, Bill [Paul Scherrer Institute (PSI), SwissFEL Project (Switzerland); Herrmann, Torsten [Université de Lyon, Institut des Sciences Analytiques, Centre de RMN à Très Hauts Champs, UMR 5280 CNRS, ENS Lyon, UCB Lyon 1 (France); Wüthrich, Kurt, E-mail: wuthrich@scripps.edu [The Scripps Research Institute, Department of Integrative Structural and Computational Biology (United States)

    2015-01-15

    A standard set of three APSY-NMR experiments has been used in daily practice to obtain polypeptide backbone NMR assignments in globular proteins with sizes up to about 150 residues, which had been identified as targets for structure determination by the Joint Center for Structural Genomics (JCSG) under the auspices of the Protein Structure Initiative (PSI). In a representative sample of 30 proteins, initial fully automated data analysis with the software UNIO-MATCH-2014 yielded complete or partial assignments for over 90 % of the residues. For most proteins the APSY data acquisition was completed in less than 30 h. The results of the automated procedure provided a basis for efficient interactive validation and extension to near-completion of the assignments by reference to the same 3D heteronuclear-resolved [{sup 1}H,{sup 1}H]-NOESY spectra that were subsequently used for the collection of conformational constraints. High-quality structures were obtained for all 30 proteins, using the J-UNIO protocol, which includes extensive automation of NMR structure determination.

  14. Protein residue linking in a single spectrum for magic-angle spinning NMR assignment

    International Nuclear Information System (INIS)

    Here we introduce a new pulse sequence for resonance assignment that halves the number of data sets required for sequential linking by directly correlating sequential amide resonances in a single diagonal-free spectrum. The method is demonstrated with both microcrystalline and sedimented deuterated proteins spinning at 60 and 111 kHz, and a fully protonated microcrystalline protein spinning at 111 kHz, with as little as 0.5 mg protein sample. We find that amide signals have a low chance of ambiguous linkage, which is further improved by linking in both forward and backward directions. The spectra obtained are amenable to automated resonance assignment using general-purpose software such as UNIO-MATCH

  15. Protein residue linking in a single spectrum for magic-angle spinning NMR assignment

    Energy Technology Data Exchange (ETDEWEB)

    Andreas, Loren B.; Stanek, Jan; Marchand, Tanguy Le; Bertarello, Andrea; Paepe, Diane Cala-De; Lalli, Daniela; Krejčíková, Magdaléna; Doyen, Camille; Öster, Carl [Université de Lyon, Centre de RMN à Très Hauts Champs, Institut des Sciences Analytiques (CNRS, ENS Lyon, UCB Lyon 1) (France); Knott, Benno; Wegner, Sebastian; Engelke, Frank [Bruker Biospin (Germany); Felli, Isabella C.; Pierattelli, Roberta [University of Florence, Department of Chemistry “Ugo Schiff“and Magnetic Resonance Center (CERM) (Italy); Dixon, Nicholas E. [University of Wollongong, School of Chemistry (Australia); Emsley, Lyndon; Herrmann, Torsten; Pintacuda, Guido, E-mail: guido.pintacuda@ens-lyon.fr [Université de Lyon, Centre de RMN à Très Hauts Champs, Institut des Sciences Analytiques (CNRS, ENS Lyon, UCB Lyon 1) (France)

    2015-07-15

    Here we introduce a new pulse sequence for resonance assignment that halves the number of data sets required for sequential linking by directly correlating sequential amide resonances in a single diagonal-free spectrum. The method is demonstrated with both microcrystalline and sedimented deuterated proteins spinning at 60 and 111 kHz, and a fully protonated microcrystalline protein spinning at 111 kHz, with as little as 0.5 mg protein sample. We find that amide signals have a low chance of ambiguous linkage, which is further improved by linking in both forward and backward directions. The spectra obtained are amenable to automated resonance assignment using general-purpose software such as UNIO-MATCH.

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

    Energy Technology Data Exchange (ETDEWEB)

    Strzelecka, T.; Gronenborn, A.M.; Clore, G.M. [National Institutes of Health, Bethesda, MD (United States)

    1994-12-01

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

  17. A spectroscopic assignment technique for membrane proteins reconstituted in magnetically aligned bicelles

    International Nuclear Information System (INIS)

    Oriented-sample NMR (OS-NMR) has emerged as a powerful tool for the structure determination of membrane proteins in their physiological environments. However, the traditional spectroscopic assignment method in OS NMR that uses the “shotgun” approach, though effective, is quite labor- and time-consuming as it is based on the preparation of multiple selectively labeled samples. Here we demonstrate that, by using a combination of the spin exchange under mismatched Hartmann-Hahn conditions and a recent sensitivity-enhancement REP-CP sequence, spectroscopic assignment of solid-state NMR spectra of Pf1 coat protein reconstituted in magnetically aligned bicelles can be significantly improved. This method yields a two-dimensional spin-exchanged version of the SAMPI4 spectrum correlating the 15N chemical shift and 15N–1H dipolar couplings, as well as spin-correlations between the (i, i ± 1) amide sites. Combining the spin-exchanged SAMPI4 spectrum with the original SAMPI4 experiment makes it possible to establish sequential assignments, and this technique is generally applicable to other uniaxially aligned membrane proteins. Inclusion of an 15N–15N correlation spectrum into the assignment process helps establish correlations between the peaks in crowded or ambiguous spectral regions of the spin-exchanged SAMPI4 experiment. Notably, unlike the traditional method, only a uniformly labeled protein sample is required for spectroscopic assignment with perhaps only a few selectively labeled “seed” spectra. Simulations for the magnetization transfer between the dilute spins under mismatched Hartmann Hahn conditions for various B1 fields have also been performed. The results adequately describe the optimal conditions for establishing the cross peaks, thus eliminating the need for lengthy experimental optimizations.

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

    Science.gov (United States)

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

    2013-11-01

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

  19. Using Computer-Based Writing Software to Facilitate Writing Assignments in Large Political Science Classes

    Science.gov (United States)

    Ishiyama, John; Watson, Wendy L.

    2014-01-01

    It is generally accepted in the literature that writing assignments, even short ones, increase both student writing ability and comprehension of the material covered in the assignments. As class enrollments increase, particularly at the introductory level, however, instructors often sacrifice writing assignments because of the difficulty in…

  20. A novel strategy for NMR resonance assignment and protein structure determination

    Energy Technology Data Exchange (ETDEWEB)

    Lemak, Alexander [University of Toronto, Ontario Cancer Institute and Campbell Family Cancer Research Institute, Department of Medical Biophysics (Canada); Gutmanas, Aleksandras [European Bioinformatics Institute, Protein Data Bank Europe (United Kingdom); Chitayat, Seth; Karra, Murthy [University of Toronto, Ontario Cancer Institute and Campbell Family Cancer Research Institute, Department of Medical Biophysics (Canada); Fares, Christophe [Max-Planck-Institut f. Kohlenforschung (Germany); Sunnerhagen, Maria [Linkoeping University, Division of Molecular Biotechnology, Department of Physics, Chemistry and Biology (Sweden); Arrowsmith, Cheryl H., E-mail: carrow@uhnres.utoronto.ca [University of Toronto, Ontario Cancer Institute and Campbell Family Cancer Research Institute, Department of Medical Biophysics (Canada)

    2011-01-15

    The quality of protein structures determined by nuclear magnetic resonance (NMR) spectroscopy is contingent on the number and quality of experimentally-derived resonance assignments, distance and angular restraints. Two key features of protein NMR data have posed challenges for the routine and automated structure determination of small to medium sized proteins; (1) spectral resolution - especially of crowded nuclear Overhauser effect spectroscopy (NOESY) spectra, and (2) the reliance on a continuous network of weak scalar couplings as part of most common assignment protocols. In order to facilitate NMR structure determination, we developed a semi-automated strategy that utilizes non-uniform sampling (NUS) and multidimensional decomposition (MDD) for optimal data collection and processing of selected, high resolution multidimensional NMR experiments, combined it with an ABACUS protocol for sequential and side chain resonance assignments, and streamlined this procedure to execute structure and refinement calculations in CYANA and CNS, respectively. Two graphical user interfaces (GUIs) were developed to facilitate efficient analysis and compilation of the data and to guide automated structure determination. This integrated method was implemented and refined on over 30 high quality structures of proteins ranging from 5.5 to 16.5 kDa in size.

  1. A novel strategy for NMR resonance assignment and protein structure determination

    International Nuclear Information System (INIS)

    The quality of protein structures determined by nuclear magnetic resonance (NMR) spectroscopy is contingent on the number and quality of experimentally-derived resonance assignments, distance and angular restraints. Two key features of protein NMR data have posed challenges for the routine and automated structure determination of small to medium sized proteins; (1) spectral resolution – especially of crowded nuclear Overhauser effect spectroscopy (NOESY) spectra, and (2) the reliance on a continuous network of weak scalar couplings as part of most common assignment protocols. In order to facilitate NMR structure determination, we developed a semi-automated strategy that utilizes non-uniform sampling (NUS) and multidimensional decomposition (MDD) for optimal data collection and processing of selected, high resolution multidimensional NMR experiments, combined it with an ABACUS protocol for sequential and side chain resonance assignments, and streamlined this procedure to execute structure and refinement calculations in CYANA and CNS, respectively. Two graphical user interfaces (GUIs) were developed to facilitate efficient analysis and compilation of the data and to guide automated structure determination. This integrated method was implemented and refined on over 30 high quality structures of proteins ranging from 5.5 to 16.5 kDa in size.

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

    Science.gov (United States)

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

    2002-01-01

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

  3. "CON-CON" assignment strategy for highly flexible intrinsically disordered proteins.

    Science.gov (United States)

    Piai, Alessandro; Hošek, Tomáš; Gonnelli, Leonardo; Zawadzka-Kazimierczuk, Anna; Koźmiński, Wiktor; Brutscher, Bernhard; Bermel, Wolfgang; Pierattelli, Roberta; Felli, Isabella C

    2014-12-01

    Intrinsically disordered proteins (IDPs) are a class of highly flexible proteins whose characterization by NMR spectroscopy is complicated by severe spectral overlaps. The development of experiments designed to facilitate the sequence-specific assignment procedure is thus very important to improve the tools for the characterization of IDPs and thus to be able to focus on IDPs of increasing size and complexity. Here, we present and describe the implementation of a set of novel ¹H-detected 5D experiments, (HACA)CON(CACO)NCO(CA)HA, BT-(H)NCO(CAN)CONNH and BT-HN(COCAN)CONNH, optimized for the study of highly flexible IDPs that exploit the best resolved correlations, those involving the carbonyl and nitrogen nuclei of neighboring amino acids, to achieve sequence-specific resonance assignment. Together with the analogous recently proposed pulse schemes based on ¹³C detection, they form a complete set of experiments for sequence-specific assignment of highly flexible IDPs. Depending on the particular sample conditions (concentration, lifetime, pH, temperature, etc.), these experiments present certain advantages and disadvantages that will be discussed. Needless to say, that the availability of a variety of complementary experiments will be important for accurate determination of resonance frequencies in complex IDPs.

  4. Chemical shift assignment of the alternative scaffold protein IscA.

    Science.gov (United States)

    Popovic, Matija; Pastore, Annalisa

    2016-04-01

    The IscA protein (11.5 kDa) is an essential component of the iron sulphur cluster biogenesis machine. In bacteria, the machine components are clustered in operons, amongst which the most important is the isc operon. Bacterial IscA has direct homologues also in eukaryotes. Like the protein IscU, IscA is thought to assist cluster formation as an alternative scaffold protein which receives the cluster before transferring it further to the final acceptors. Several crystal structures have been published. They all report an IscA dimeric form, although the packing of the protomers in the dimers differs amongst structures. No solution studies have currently been reported. Here we report the (1)H, (13)C and (15)N backbone and side-chain chemical shift assignments of the cluster-free E. coli IscA as a starting point for further studies of the structure and functions of this still poorly characterized protein. We show that IscA exists in solution as an equilibrium between different species. Spectrum assignment was thus challenging given the heterogeneous nature of the sample but doable through judicious choice of selective labelling and concentration dependent studies.

  5. Writing Assignments with a Metacognitive Component Enhance Learning in a Large Introductory Biology Course

    OpenAIRE

    Mynlieff, Michelle; Manogaran, Anita L.; St. Maurice, Martin; Eddinger, Thomas J.

    2014-01-01

    Writing assignments, including note taking and written recall, should enhance retention of knowledge, whereas analytical writing tasks with metacognitive aspects should enhance higher-order thinking. In this study, we assessed how certain writing-intensive “interventions,” such as written exam corrections and peer-reviewed writing assignments using Calibrated Peer Review and including a metacognitive component, improve student learning. We designed and tested the possible benefits of these ap...

  6. Specific labeling and assignment strategies of valine methyl groups for NMR studies of high molecular weight proteins

    Energy Technology Data Exchange (ETDEWEB)

    Mas, Guillaume; Crublet, Elodie [Univ. Grenoble Alpes, Institut de Biologie Structurale (IBS) (France); Hamelin, Olivier [CNRS (France); Gans, Pierre; Boisbouvier, Jérôme, E-mail: jerome.boisbouvier@ibs.fr [Univ. Grenoble Alpes, Institut de Biologie Structurale (IBS) (France)

    2013-09-28

    The specific protonation of valine and leucine methyl groups in proteins is typically achieved by overexpressing proteins in M9/D{sub 2}O medium supplemented with either labeled α-ketoisovalerate for the labeling of the four prochiral methyl groups or with 2-acetolactate for the stereospecific labeling of the valine and leucine side chains. However, when these labeling schemes are applied to large protein assemblies, significant overlap between the correlations of the valine and leucine methyl groups occurs, hampering the analysis of 2D methyl-TROSY spectra. Analysis of the leucine and valine biosynthesis pathways revealed that the incorporation of labeled precursors in the leucine pathway can be inhibited by the addition of exogenous l-leucine-d{sub 10}. We exploited this property to label stereospecifically the pro-R and pro-S methyl groups of valine with minimal scrambling to the leucine residues. This new labeling protocol was applied to the 468 kDa homododecameric peptidase TET2 to decrease the complexity of its NMR spectra. All of the pro-S valine methyl resonances of TET2 were assigned by combining mutagenesis with this innovative labeling approach. The assignments were transferred to the pro-R groups using an optimally labeled sample and a set of triple resonance experiments. This improved labeling scheme enables us to overcome the main limitation of overcrowding in the NMR spectra of prochiral methyl groups, which is a prerequisite for the site-specific measurement of the structural and dynamic parameters or for the study of interactions in very large protein assemblies.

  7. Specific labeling and assignment strategies of valine methyl groups for NMR studies of high molecular weight proteins.

    Science.gov (United States)

    Mas, Guillaume; Crublet, Elodie; Hamelin, Olivier; Gans, Pierre; Boisbouvier, Jérôme

    2013-11-01

    The specific protonation of valine and leucine methyl groups in proteins is typically achieved by overexpressing proteins in M9/D2O medium supplemented with either labeled α-ketoisovalerate for the labeling of the four prochiral methyl groups or with 2-acetolactate for the stereospecific labeling of the valine and leucine side chains. However, when these labeling schemes are applied to large protein assemblies, significant overlap between the correlations of the valine and leucine methyl groups occurs, hampering the analysis of 2D methyl-TROSY spectra. Analysis of the leucine and valine biosynthesis pathways revealed that the incorporation of labeled precursors in the leucine pathway can be inhibited by the addition of exogenous l-leucine-d10. We exploited this property to label stereospecifically the pro-R and pro-S methyl groups of valine with minimal scrambling to the leucine residues. This new labeling protocol was applied to the 468 kDa homododecameric peptidase TET2 to decrease the complexity of its NMR spectra. All of the pro-S valine methyl resonances of TET2 were assigned by combining mutagenesis with this innovative labeling approach. The assignments were transferred to the pro-R groups using an optimally labeled sample and a set of triple resonance experiments. This improved labeling scheme enables us to overcome the main limitation of overcrowding in the NMR spectra of prochiral methyl groups, which is a prerequisite for the site-specific measurement of the structural and dynamic parameters or for the study of interactions in very large protein assemblies.

  8. Toward a better understanding of structural divergences in proteins using different secondary structure assignment methods

    Science.gov (United States)

    Rocha, L. F. O.

    2014-04-01

    Structural disagreements on the location and quantity of secondary structure segments comprise a current challenging problem leading to several limitations for theoretical and applied research. This paper presents 116 structural evaluations by steric and hydrophobic interactions in secondary structures within a specific template group; determines simple prediction rules that calculate 88 occurrence frequencies of large and hydrophobic residues into target intra- and inter-subgroups with structure disagreements; and utilizes 42 comparisons between the methods PROMOTIF, DSSP and STRIDE. In the stereochemical predictions inside the subgroups there are predominantly excellent and/or good success amounts with their expected values, and the disclosure of a triple molecular mechanism by residue volumetric and hydrophobic ingredients. The method comparisons show high compatibility scores between them, therefore validating their seemingly incompatible assignments. Thus, the nonconsensual ascriptions are better understood and appreciated. Furthermore, such results suggest a broad utility of our assignment method for other benchmark datasets and known methods.

  9. Using MUSIC and CC(CO)NH for backbone assignment of two medium-sized proteins not fully accessible to standard 3D NMR.

    Science.gov (United States)

    Brenner, Annette K; Frøystein, Nils Åge

    2014-01-01

    The backbone assignment of medium-sized proteins is rarely as straightforward as that of small proteins, and thus often requires creative solutions. Here, we describe the application of a combination of standard 3D heteronuclear methods with CC(CO)NH and a variety of MUltiplicity Selective In-phase Coherence transfer (MUSIC) experiments. Both CC(CO)NH and MUSIC are, in theory, very powerful methods for the backbone assignment of proteins. Due to low sensitivity, their use has usually been linked to small proteins only. However, we found that combining CC(CO)NH and MUSIC experiments simplified the assignment of two challenging medium-sized proteins of 13 and 19.5 kDa, respectively. These methods are to some extent complementary to each other: CC(CO)NH acquired with a long isotropic mixing time can identify amino acids with large aliphatic side chains. Whereas the most sensitive MUSIC experiments identify amino acid types that cannot be detected by CC(CO)NH, comprising the residues with acid and amide groups, and aromatic rings in their side chains. Together these methods provide a means of identifying the majority of peaks in the 2D 15N HSQC spectrum which simplifies the backbone assignment work even for proteins, e.g., small kinases, whose standard spectra resulted in little spectral resolution and low signal intensities.

  10. CONNJUR R: an annotation strategy for fostering reproducibility in bio-NMR—protein spectral assignment

    Energy Technology Data Exchange (ETDEWEB)

    Fenwick, Matthew; Hoch, Jeffrey C. [UConn Health, Department of Molecular Biology and Biophysics (United States); Ulrich, Eldon [University of Wisconsin-Madison, Department of Biochemistry (United States); Gryk, Michael R., E-mail: gryk@uchc.edu [UConn Health, Department of Molecular Biology and Biophysics (United States)

    2015-10-15

    Reproducibility is a cornerstone of the scientific method, essential for validation of results by independent laboratories and the sine qua non of scientific progress. A key step toward reproducibility of biomolecular NMR studies was the establishment of public data repositories (PDB and BMRB). Nevertheless, bio-NMR studies routinely fall short of the requirement for reproducibility that all the data needed to reproduce the results are published. A key limitation is that considerable metadata goes unpublished, notably manual interventions that are typically applied during the assignment of multidimensional NMR spectra. A general solution to this problem has been elusive, in part because of the wide range of approaches and software packages employed in the analysis of protein NMR spectra. Here we describe an approach for capturing missing metadata during the assignment of protein NMR spectra that can be generalized to arbitrary workflows, different software packages, other biomolecules, or other stages of data analysis in bio-NMR. We also present extensions to the NMR-STAR data dictionary that enable machine archival and retrieval of the “missing” metadata.

  11. Enhanced biosynthetically directed fractional carbon-13 enrichment of proteins for backbone NMR assignments.

    Science.gov (United States)

    Wenrich, Broc R; Sonstrom, Reilly E; Gupta, Riju A; Rovnyak, David

    2015-11-01

    Routes to carbon-13 enrichment of bacterially expressed proteins include achieving uniform or positionally selective (e.g. ILV-Me, or (13)C', etc.) enrichment. We consider the potential for biosynthetically directed fractional enrichment (e.g. carbon-13 incorporation in the protein less than 100%) for performing routine n-(D)dimensional NMR spectroscopy of proteins. First, we demonstrate an approach to fractional isotope addition where the initial growth media containing natural abundance glucose is replenished at induction with a small amount (e.g. 10%(w/w)u-(13)C-glucose) of enriched nutrient. The approach considered here is to add 10% (e.g. 200mg for a 2g/L culture) u-(13)C-glucose at the induction time (OD600=0.8), resulting in a protein with enhanced (13)C incorporation that gives almost the same NMR signal levels as an exact 20% (13)C sample. Second, whereas fractional enrichment is used for obtaining stereospecific methyl assignments, we find that (13)C incorporation levels no greater than 20%(w/w) yield (13)C and (13)C-(13)C spin pair incorporation sufficient to conduct typical 3D-bioNMR backbone experiments on moderate instrumentation (600 MHz, RT probe). Typical 3D-bioNMR experiments of a fractionally enriched protein yield expected backbone connectivities, and did not show amino acid biases in this work, with one exception. When adding 10% u-(13)C glucose to expression media at induction, there is poor preservation of (13)Cα-(13)Cβ spin pairs in the amino acids ILV, leading to the absence of Cβ signals in HNCACB spectra for ILV, a potentially useful editing effect. Enhanced fractional carbon-13 enrichment provides lower-cost routes to high throughput protein NMR studies, and makes modern protein NMR more cost-accessible.

  12. Automated sequence- and stereo-specific assignment of methyl-labeled proteins by paramagnetic relaxation and methyl-methyl nuclear overhauser enhancement spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Venditti, Vincenzo; Fawzi, Nicolas L.; Clore, G. Marius, E-mail: mariusc@mail.nih.gov [National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Laboratory of Chemical Physics (United States)

    2011-11-15

    Methyl-transverse relaxation optimized spectroscopy is rapidly becoming the preferred NMR technique for probing structure and dynamics of very large proteins up to {approx}1 MDa in molecular size. Data interpretation, however, necessitates assignment of methyl groups which still presents a very challenging and time-consuming process. Here we demonstrate that, in combination with a known 3D structure, paramagnetic relaxation enhancement (PRE), induced by nitroxide spin-labels incorporated at only a few surface-exposed engineered cysteines, provides fast, straightforward and robust access to methyl group resonance assignments, including stereoassignments for the methyl groups of leucine and valine. Neither prior assignments, including backbone assignments, for the protein, nor experiments that transfer magnetization between methyl groups and the protein backbone, are required. PRE-derived assignments are refined by 4D methyl-methyl nuclear Overhauser enhancement data, eliminating ambiguities and errors that may arise due to the high sensitivity of PREs to the potential presence of sparsely-populated transient states.

  13. Reliable resonance assignments of selected residues of proteins with known structure based on empirical NMR chemical shift prediction

    Science.gov (United States)

    Li, Da-Wei; Meng, Dan; Brüschweiler, Rafael

    2015-05-01

    A robust NMR resonance assignment method is introduced for proteins whose 3D structure has previously been determined by X-ray crystallography. The goal of the method is to obtain a subset of correct assignments from a parsimonious set of 3D NMR experiments of 15N, 13C labeled proteins. Chemical shifts of sequential residue pairs are predicted from static protein structures using PPM_One, which are then compared with the corresponding experimental shifts. Globally optimized weighted matching identifies the assignments that are robust with respect to small changes in NMR cross-peak positions. The method, termed PASSPORT, is demonstrated for 4 proteins with 100-250 amino acids using 3D NHCA and a 3D CBCA(CO)NH experiments as input producing correct assignments with high reliability for 22% of the residues. The method, which works best for Gly, Ala, Ser, and Thr residues, provides assignments that serve as anchor points for additional assignments by both manual and semi-automated methods or they can be directly used for further studies, e.g. on ligand binding, protein dynamics, or post-translational modification, such as phosphorylation.

  14. Writing Assignments with a Metacognitive Component Enhance Learning in a Large Introductory Biology Course.

    Science.gov (United States)

    Mynlieff, Michelle; Manogaran, Anita L; St Maurice, Martin; Eddinger, Thomas J

    2014-01-01

    Writing assignments, including note taking and written recall, should enhance retention of knowledge, whereas analytical writing tasks with metacognitive aspects should enhance higher-order thinking. In this study, we assessed how certain writing-intensive "interventions," such as written exam corrections and peer-reviewed writing assignments using Calibrated Peer Review and including a metacognitive component, improve student learning. We designed and tested the possible benefits of these approaches using control and experimental variables across and between our three-section introductory biology course. Based on assessment, students who corrected exam questions showed significant improvement on postexam assessment compared with their nonparticipating peers. Differences were also observed between students participating in written and discussion-based exercises. Students with low ACT scores benefited equally from written and discussion-based exam corrections, whereas students with midrange to high ACT scores benefited more from written than discussion-based exam corrections. Students scored higher on topics learned via peer-reviewed writing assignments relative to learning in an active classroom discussion or traditional lecture. However, students with low ACT scores (17-23) did not show the same benefit from peer-reviewed written essays as the other students. These changes offer significant student learning benefits with minimal additional effort by the instructors.

  15. Towards fully automated structure-based NMR resonance assignment of 15N-labeled proteins from automatically picked peaks

    KAUST Repository

    Jang, Richard

    2011-03-01

    In NMR resonance assignment, an indispensable step in NMR protein studies, manually processed peaks from both N-labeled and C-labeled spectra are typically used as inputs. However, the use of homologous structures can allow one to use only N-labeled NMR data and avoid the added expense of using C-labeled data. We propose a novel integer programming framework for structure-based backbone resonance assignment using N-labeled data. The core consists of a pair of integer programming models: one for spin system forming and amino acid typing, and the other for backbone resonance assignment. The goal is to perform the assignment directly from spectra without any manual intervention via automatically picked peaks, which are much noisier than manually picked peaks, so methods must be error-tolerant. In the case of semi-automated/manually processed peak data, we compare our system with the Xiong-Pandurangan-Bailey- Kellogg\\'s contact replacement (CR) method, which is the most error-tolerant method for structure-based resonance assignment. Our system, on average, reduces the error rate of the CR method by five folds on their data set. In addition, by using an iterative algorithm, our system has the added capability of using the NOESY data to correct assignment errors due to errors in predicting the amino acid and secondary structure type of each spin system. On a publicly available data set for human ubiquitin, where the typing accuracy is 83%, we achieve 91% accuracy, compared to the 59% accuracy obtained without correcting for such errors. In the case of automatically picked peaks, using assignment information from yeast ubiquitin, we achieve a fully automatic assignment with 97% accuracy. To our knowledge, this is the first system that can achieve fully automatic structure-based assignment directly from spectra. This has implications in NMR protein mutant studies, where the assignment step is repeated for each mutant. © Copyright 2011, Mary Ann Liebert, Inc.

  16. Quadratic partial eigenvalue assignment in large-scale stochastic dynamic systems for resilient and economic design

    Science.gov (United States)

    Das, S.; Goswami, K.; Datta, B. N.

    2016-05-01

    Failure of structural systems under dynamic loading can be prevented via active vibration control which shifts the damped natural frequencies of the systems away from the dominant range of a loading spectrum. The damped natural frequencies and the dynamic load typically show significant variations in practice. A computationally efficient methodology based on quadratic partial eigenvalue assignment technique and optimization under uncertainty has been formulated in the present work that will rigorously account for these variations and result in economic and resilient design of structures. A novel scheme based on hierarchical clustering and importance sampling is also developed in this work for accurate and efficient estimation of probability of failure to guarantee the desired resilience level of the designed system. Finally the most robust set of feedback matrices is selected from the set of probabilistically characterized optimal closed-loop system to implement the new methodology for design of active controlled structures. Numerical examples are presented to illustrate the proposed methodology.

  17. A new strategy for sequential assignment of intrinsically unstructured proteins based on 15N single isotope labelling

    Science.gov (United States)

    Lopez, Juan; Ahuja, Puneet; Gerard, Melanie; Wieruszeski, Jean-Michel; Lippens, Guy

    2013-11-01

    We describe a new efficient strategy for the sequential assignment of amide resonances of a conventional 15N-1H HSQC spectrum of intrinsically unfolded proteins, based on composite NOESY-TOCSY and TOCSY-NOESY mixing times. These composite mixing times lead to a Hα-proton mediated unidirectional transfer of amide to amide proton. We have implemented the composite mixing times in an HSQC-NOESY-HSQC manner to obtain directional connectivity between amides of neighbouring residues. We experimentally determine the optimal mixing times for both transfer schemes, and demonstrate its use in the assignment for both a fragment of the neuronal tau protein and for α-synuclein.

  18. CH3-specific NMR assignment of alanine, isoleucine, leucine and valine methyl groups in high molecular weight proteins using a single sample.

    Science.gov (United States)

    Kerfah, Rime; Hamelin, Olivier; Boisbouvier, Jérôme; Marion, Dominique

    2015-12-01

    A new strategy for the NMR assignment of aliphatic side-chains in large perdeuterated proteins is proposed. It involves an alternative isotopic labeling protocol, the use of an out-and-back (13)C-(13)C TOCSY experiment ((H)C-TOCSY-C-TOCSY-(C)H) and an optimized non-uniform sampling protocol. It has long been known that the non-linearity of an aliphatic spin-system (for example Ile, Val, or Leu) substantially compromises the efficiency of the TOCSY transfers. To permit the use of this efficient pulse scheme, a series of optimized precursors were designed to yield linear (13)C perdeuterated side-chains with a single protonated CH3 group in these three residues. These precursors were added to the culture medium for incorporation into expressed proteins. For Val and Leu residues, the topologically different spin-systems introduced for the pro-R and pro-S methyl groups enable stereospecific assignment. All CH3 can be simultaneously assigned on a single sample using a TOCSY experiment. It only requires the tuning of a mixing delay and is thus more versatile than the relayed COSY experiment. Enhanced resolution and sensi-tivity can be achieved by non-uniform sampling combined with the removal of the large JCC coupling by deconvolution prior to the processing by iterative soft thresholding. This strategy has been used on malate synthase G where a large percentage of the CH3 groups could be correlated directly up to the backbone Ca. It is anticipated that this robust combined strategy can be routinely applied to large proteins.

  19. CH{sub 3}-specific NMR assignment of alanine, isoleucine, leucine and valine methyl groups in high molecular weight proteins using a single sample

    Energy Technology Data Exchange (ETDEWEB)

    Kerfah, Rime [Université Grenoble Alpes, IBS (France); Hamelin, Olivier [University Grenoble Alpes, Chemistry and Biology of Metals Laboratory (France); Boisbouvier, Jérôme; Marion, Dominique, E-mail: Dominique.Marion@ibs.fr [Université Grenoble Alpes, IBS (France)

    2015-12-15

    A new strategy for the NMR assignment of aliphatic side-chains in large perdeuterated proteins is proposed. It involves an alternative isotopic labeling protocol, the use of an out-and-back {sup 13}C–{sup 13}C TOCSY experiment ((H)C-TOCSY-C-TOCSY-(C)H) and an optimized non-uniform sampling protocol. It has long been known that the non-linearity of an aliphatic spin-system (for example Ile, Val, or Leu) substantially compromises the efficiency of the TOCSY transfers. To permit the use of this efficient pulse scheme, a series of optimized precursors were designed to yield linear {sup 13}C perdeuterated side-chains with a single protonated CH{sub 3} group in these three residues. These precursors were added to the culture medium for incorporation into expressed proteins. For Val and Leu residues, the topologically different spin-systems introduced for the pro-R and pro-S methyl groups enable stereospecific assignment. All CH{sub 3} can be simultaneously assigned on a single sample using a TOCSY experiment. It only requires the tuning of a mixing delay and is thus more versatile than the relayed COSY experiment. Enhanced resolution and sensi-tivity can be achieved by non-uniform sampling combined with the removal of the large J{sub CC} coupling by deconvolution prior to the processing by iterative soft thresholding. This strategy has been used on malate synthase G where a large percentage of the CH{sub 3} groups could be correlated directly up to the backbone Ca. It is anticipated that this robust combined strategy can be routinely applied to large proteins.

  20. METHODOLOGY AND CALCULATIONS FOR THE ASSIGNMENT OF WASTE GROUPS FOR THE LARGE UNDERGROUND WASTE STORAGE TANKS AT THE HANFORD SITE

    International Nuclear Information System (INIS)

    This document categorizes each of the large waste storage tanks into one of several categories based on each tank's waste characteristics. These waste group assignments reflect a tank's propensity to retain a significant volume of flammable gases and the potential of the waste to release retained gas by a buoyant displacement event. Revision 7 is the annual update of the calculations of the flammable gas Waste Groups for DSTs and SSTs. The Hanford Site contains 177 large underground radioactive waste storage tanks (28 double-shell tanks and 149 single-shell tanks). These tanks are categorized into one of three waste groups (A, B, and C) based on their waste and tank characteristics. These waste group assignments reflect a tank's propensity to retain a significant volume of flammable gases and the potential of the waste to release retained gas by a buoyant displacement gas release event. Assignments of waste groups to the 177 double-shell tanks and single-shell tanks, as reported in this document, are based on a Monte Carlo analysis of three criteria. The first criterion is the headspace flammable gas concentration following release of retained gas. This criterion determines whether the tank contains sufficient retained gas such that the well-mixed headspace flammable gas concentration would reach 100% of the lower flammability limit if the entire tank's retained gas were released. If the volume of retained gas is not sufficient to reach 100% of the lower flammability limit, then flammable conditions cannot be reached and the tank is classified as a waste group C tank independent of the method the gas is released. The second criterion is the energy ratio and considers whether there is sufficient supernatant on top of the saturated solids such that gas-bearing solids have the potential energy required to break up the material and release gas. Tanks that are not waste group C tanks and that have an energy ratio < 3.0 do not have sufficient potential energy to break up

  1. METHODOLOGY AND CALCULATIONS FOR THE ASSIGNMENT OF WASTE GROUPS FOR THE LARGE UNDERGROUND WASTE STORAGE TANKS AT THE HANFORD SITE

    Energy Technology Data Exchange (ETDEWEB)

    FOWLER KD

    2007-12-27

    This document categorizes each of the large waste storage tanks into one of several categories based on each tank's waste characteristics. These waste group assignments reflect a tank's propensity to retain a significant volume of flammable gases and the potential of the waste to release retained gas by a buoyant displacement event. Revision 7 is the annual update of the calculations of the flammable gas Waste Groups for DSTs and SSTs. The Hanford Site contains 177 large underground radioactive waste storage tanks (28 double-shell tanks and 149 single-shell tanks). These tanks are categorized into one of three waste groups (A, B, and C) based on their waste and tank characteristics. These waste group assignments reflect a tank's propensity to retain a significant volume of flammable gases and the potential of the waste to release retained gas by a buoyant displacement gas release event. Assignments of waste groups to the 177 double-shell tanks and single-shell tanks, as reported in this document, are based on a Monte Carlo analysis of three criteria. The first criterion is the headspace flammable gas concentration following release of retained gas. This criterion determines whether the tank contains sufficient retained gas such that the well-mixed headspace flammable gas concentration would reach 100% of the lower flammability limit if the entire tank's retained gas were released. If the volume of retained gas is not sufficient to reach 100% of the lower flammability limit, then flammable conditions cannot be reached and the tank is classified as a waste group C tank independent of the method the gas is released. The second criterion is the energy ratio and considers whether there is sufficient supernatant on top of the saturated solids such that gas-bearing solids have the potential energy required to break up the material and release gas. Tanks that are not waste group C tanks and that have an energy ratio < 3.0 do not have sufficient

  2. METHODOLOGY AND CALCULATIONS FOR THE ASSIGNMENT OF WASTE GROUPS FOR THE LARGE UNDERGROUND WASTE STORAGE TANKS AT THE HANFORD SITE

    Energy Technology Data Exchange (ETDEWEB)

    WEBER RA

    2009-01-16

    The Hanford Site contains 177 large underground radioactive waste storage tanks (28 double-shell tanks and 149 single-shell tanks). These tanks are categorized into one of three waste groups (A, B, and C) based on their waste and tank characteristics. These waste group assignments reflect a tank's propensity to retain a significant volume of flammable gases and the potential of the waste to release retained gas by a buoyant displacement gas release event. Assignments of waste groups to the 177 double-shell tanks and single-shell tanks, as reported in this document, are based on a Monte Carlo analysis of three criteria. The first criterion is the headspace flammable gas concentration following release of retained gas. This criterion determines whether the tank contains sufficient retained gas such that the well-mixed headspace flammable gas concentration would reach 100% of the lower flammability limit if the entire tank's retained gas were released. If the volume of retained gas is not sufficient to reach 100% of the lower flammability limit, then flammable conditions cannot be reached and the tank is classified as a waste group C tank independent of the method the gas is released. The second criterion is the energy ratio and considers whether there is sufficient supernatant on top of the saturated solids such that gas-bearing solids have the potential energy required to break up the material and release gas. Tanks that are not waste group C tanks and that have an energy ratio < 3.0 do not have sufficient potential energy to break up material and release gas and are assigned to waste group B. These tanks are considered to represent a potential induced flammable gas release hazard, but no spontaneous buoyant displacement flammable gas release hazard. Tanks that are not waste group C tanks and have an energy ratio {ge} 3.0, but that pass the third criterion (buoyancy ratio < 1.0, see below) are also assigned to waste group B. Even though the designation as

  3. 13C, 15N Resonance Assignment of Parts of the HET-s Prion Protein in its Amyloid Form

    International Nuclear Information System (INIS)

    The partial 15N and 13C solid-state NMR resonance assignment of the HET-s prion protein fragment 218-289 in its amyloid form is presented. It is based on experiments measured at MAS frequencies in the range of 20-40 kHz using exclusively adiabatic polarization-transfer schemes. The resonance assignment within each residue is based on two-dimensional 13C--13C correlation spectra utilizing the DREAM mixing scheme. The sequential linking of the assigned residues used a set of two- and three-dimensional 15N--13C correlation experiments. Almost all cross peaks visible in the spectra are assigned, but only resonances from 43 of the 78 amino-acid residues could be detected. The missing residues are thought to be highly disordered and/or highly dynamic giving rise to broad resonance lines that escaped detection in the experiments applied. The line widths of the observed resonances are narrow and comparable to line widths observed in micro-crystalline samples. The 43 assigned residues are located in two fragments of about 20 residues

  4. Assignment strategies for aliphatic protons in the solid-state in randomly protonated proteins

    Energy Technology Data Exchange (ETDEWEB)

    Asami, Sam; Reif, Bernd, E-mail: reif@tum.de [Leibniz-Institut fuer Molekulare Pharmakologie (FMP) (Germany)

    2012-01-15

    Biological solid-state nuclear magnetic resonance spectroscopy developed rapidly in the past two decades and emerged as an important tool for structural biology. Resonance assignment is an essential prerequisite for structure determination and the characterization of motional properties of a molecule. Experiments, which rely on carbon or nitrogen detection, suffer, however, from low sensitivity. Recently, we introduced the RAP (Reduced Adjoining Protonation) labeling scheme, which allows to detect backbone and sidechain protons with high sensitivity and resolution. We present here a {sup 1}H-detected 3D (H)CCH experiment for assignment of backbone and sidechain proton resonances. Resolution is significantly improved by employing simultaneous {sup 13}CO and {sup 13}C{beta} J-decoupling during evolution of the {sup 13}C{alpha} chemical shift. In total, {approx}90% of the {sup 1}H{alpha}-{sup 13}C{alpha} backbone resonances of chicken {alpha}-spectrin SH3 could be assigned.

  5. Reduced Dimensionality (4,3)D-hnCOCANH Experiment: An Efficient Backbone Assignment tool for NMR studies of Proteins

    CERN Document Server

    Kumar, Dinesh

    2013-01-01

    Sequence specific resonance assignment and secondary structure determination of proteins form the basis for variety of structural and functional proteomics studies by NMR. In this context, an efficient standalone method for rapid assignment of backbone (1H, 15N, 13Ca and 13C') resonances and secondary structure determination of proteins has been presented here. Compared to currently available strategies used for the purpose, the method employs only a single reduced dimensionality (RD) experiment -(4,3)D-hnCOCANH and exploits the linear combinations of backbone (13Ca and 13C') chemical shifts to achieve a dispersion relatively better compared to those of individual chemical shifts (see the text) for efficient and rapid data analysis. Further, the experiment leads to the spectrum with direct distinction of self (intra-residue) and sequential (inter-residue) carbon correlation peaks; these appear opposite in signs and therefore can easily be discriminated without using an additional complementary experiment. On ...

  6. Luminescent conjugated oligothiophenes for sensitive fluorescent assignment of protein inclusion bodies.

    Science.gov (United States)

    Klingstedt, Therése; Blechschmidt, Cristiane; Nogalska, Anna; Prokop, Stefan; Häggqvist, Bo; Danielsson, Olof; Engel, W King; Askanas, Valerie; Heppner, Frank L; Nilsson, K Peter R

    2013-03-18

    Small hydrophobic ligands identifying intracellular protein deposits are of great interest, as protein inclusion bodies are the pathological hallmark of several degenerative diseases. Here we report that fluorescent amyloid ligands, termed luminescent conjugated oligothiophenes (LCOs), rapidly and with high sensitivity detect protein inclusion bodies in skeletal muscle tissue from patients with sporadic inclusion body myositis (s-IBM). LCOs having a conjugated backbone of at least five thiophene units emitted strong fluorescence upon binding, and showed co-localization with proteins reported to accumulate in s-IBM protein inclusion bodies. Compared with conventional amyloid ligands, LCOs identified a larger fraction of immunopositive inclusion bodies. When the conjugated thiophene backbone was extended with terminal carboxyl groups, the LCO revealed striking spectral differences between distinct protein inclusion bodies. We conclude that 1) LCOs are sensitive, rapid and powerful tools for identifying protein inclusion bodies and 2) LCOs identify a wider range of protein inclusion bodies than conventional amyloid ligands.

  7. Identification and RNA segment assignment of six structural proteins of Scylla serrata reovirus.

    Science.gov (United States)

    Yuan, Yangyang; Fan, Dongyang; Zhang, Zhao; Yang, Jifang; Liu, Jingwen; Chen, Jigang

    2016-08-01

    Scylla serrata reovirus (SsRV) is one of the most prevalent viral pathogens of the mud crab (S. serrata). The virus represents an unassigned novel genus in the Reoviridae family, and contains 12 double-stranded RNA genomic segments. Previous analysis of virion proteins concluded that SsRV contains at least eight structural proteins, ranging from 25 to 160 kDa. Here, tandem time-of-flight mass spectrometry and Western blotting were used to re-identify the structural proteins. The results indicate that proteins encoded by SsRV segments S1, S3, S6, S9, S11, and S12 are structural proteins.

  8. Backbone resonance assignments for G protein α(i3) subunit in the GDP-bound state.

    Science.gov (United States)

    Mase, Yoko; Yokogawa, Mariko; Osawa, Masanori; Shimada, Ichio

    2014-10-01

    Guanine-nucleotide binding proteins (G proteins) serve as molecular switches in signaling pathways, by coupling the activation of G protein-coupled receptors (GPCRs) at the cell surface to intracellular responses. In the resting state, G protein forms a heterotrimer, consisting of the G protein α subunit with GDP (Gα·GDP) and the G protein βγ subunit (Gβγ). Ligand binding to GPCRs promotes the GDP-GTP exchange on Gα, leading to the dissociation of the GTP-bound form of Gα (Gα·GTP) and Gβγ. Then, Gα·GTP and Gβγ bind to their downstream effector enzymes or ion channels and regulate their activities, leading to a variety of cellular responses. Finally, Gα hydrolyzes the bound GTP to GDP and returns to the resting state by re-associating with Gβγ. The G proteins are classified with four major families based on the amino acid sequences of Gα: i/o, s, q/11, and 12/13. Here, we established the backbone resonance assignments of human Gαi3, a member of the i/o family with a molecular weight of 41 K, in complex with GDP. The chemical shifts were compared with those of Gα(i3) in complex with a GTP-analogue, GTPγS, which we recently reported, indicating that the residues with significant chemical shift differences are mostly consistent with the regions with the structural differences between the GDP- and GTPγS-bound states, as indicated in the crystal structures. The assignments of Gα(i3)·GDP would be useful for the analyses of the dynamics of Gα(i3) and its interactions with various target molecules.

  9. The Personal Response: A Novel Writing Assignment to Engage First Year Students in Large Human Biology Classes

    Science.gov (United States)

    Moni, Roger W.; Moni, Karen B.; Poronnik, Philip

    2007-01-01

    The teaching of highly valued scientific writing skills in the first year of university is challenging. This report describes the design, implementation, and evaluation of a novel written assignment, "The Personal Response" and accompanying Peer Review, in the course, Human Biology (BIOL1015) at The University of Queensland. These assignments were…

  10. CATH FunFHMMer web server: protein functional annotations using functional family assignments.

    Science.gov (United States)

    Das, Sayoni; Sillitoe, Ian; Lee, David; Lees, Jonathan G; Dawson, Natalie L; Ward, John; Orengo, Christine A

    2015-07-01

    The widening function annotation gap in protein databases and the increasing number and diversity of the proteins being sequenced presents new challenges to protein function prediction methods. Multidomain proteins complicate the protein sequence-structure-function relationship further as new combinations of domains can expand the functional repertoire, creating new proteins and functions. Here, we present the FunFHMMer web server, which provides Gene Ontology (GO) annotations for query protein sequences based on the functional classification of the domain-based CATH-Gene3D resource. Our server also provides valuable information for the prediction of functional sites. The predictive power of FunFHMMer has been validated on a set of 95 proteins where FunFHMMer performs better than BLAST, Pfam and CDD. Recent validation by an independent international competition ranks FunFHMMer as one of the top function prediction methods in predicting GO annotations for both the Biological Process and Molecular Function Ontology. The FunFHMMer web server is available at http://www.cathdb.info/search/by_funfhmmer.

  11. METHODOLOGY & CALCULATIONS FOR THE ASSIGNMENT OF WASTE GROUPS FOR THE LARGE UNDERGROUND WASTE STORAGE TANKS AT THE HANFORD SITE

    Energy Technology Data Exchange (ETDEWEB)

    BARKER, S.A.

    2006-07-27

    Waste stored within tank farm double-shell tanks (DST) and single-shell tanks (SST) generates flammable gas (principally hydrogen) to varying degrees depending on the type, amount, geometry, and condition of the waste. The waste generates hydrogen through the radiolysis of water and organic compounds, thermolytic decomposition of organic compounds, and corrosion of a tank's carbon steel walls. Radiolysis and thermolytic decomposition also generates ammonia. Nonflammable gases, which act as dilutents (such as nitrous oxide), are also produced. Additional flammable gases (e.g., methane) are generated by chemical reactions between various degradation products of organic chemicals present in the tanks. Volatile and semi-volatile organic chemicals in tanks also produce organic vapors. The generated gases in tank waste are either released continuously to the tank headspace or are retained in the waste matrix. Retained gas may be released in a spontaneous or induced gas release event (GRE) that can significantly increase the flammable gas concentration in the tank headspace as described in RPP-7771. The document categorizes each of the large waste storage tanks into one of several categories based on each tank's waste characteristics. These waste group assignments reflect a tank's propensity to retain a significant volume of flammable gases and the potential of the waste to release retained gas by a buoyant displacement event. Revision 5 is the annual update of the methodology and calculations of the flammable gas Waste Groups for DSTs and SSTs.

  12. Revisiting date and party hubs: novel approaches to role assignment in protein interaction networks.

    Directory of Open Access Journals (Sweden)

    Sumeet Agarwal

    2010-06-01

    Full Text Available The idea of "date" and "party" hubs has been influential in the study of protein-protein interaction networks. Date hubs display low co-expression with their partners, whilst party hubs have high co-expression. It was proposed that party hubs are local coordinators whereas date hubs are global connectors. Here, we show that the reported importance of date hubs to network connectivity can in fact be attributed to a tiny subset of them. Crucially, these few, extremely central, hubs do not display particularly low expression correlation, undermining the idea of a link between this quantity and hub function. The date/party distinction was originally motivated by an approximately bimodal distribution of hub co-expression; we show that this feature is not always robust to methodological changes. Additionally, topological properties of hubs do not in general correlate with co-expression. However, we find significant correlations between interaction centrality and the functional similarity of the interacting proteins. We suggest that thinking in terms of a date/party dichotomy for hubs in protein interaction networks is not meaningful, and it might be more useful to conceive of roles for protein-protein interactions rather than for individual proteins.

  13. Nano-mole scale sequential signal assignment by 1 H-detected protein solid-state NMR

    KAUST Repository

    Wang, Songlin

    2015-01-01

    We present a 3D 1H-detected solid-state NMR (SSNMR) approach for main-chain signal assignments of 10-100 nmol of fully protonated proteins using ultra-fast magic-angle spinning (MAS) at ∼80 kHz by a novel spectral-editing method, which permits drastic spectral simplification. The approach offers ∼110 fold time saving over a traditional 3D 13C-detected SSNMR approach. This journal is © The Royal Society of Chemistry 2015.

  14. ProDomAs, protein domain assignment algorithm using center-based clustering and independent dominating set.

    Science.gov (United States)

    Ansari, Elnaz Saberi; Eslahchi, Changiz; Pezeshk, Hamid; Sadeghi, Mehdi

    2014-09-01

    Decomposition of structural domains is an essential task in classifying protein structures, predicting protein function, and many other proteomics problems. As the number of known protein structures in PDB grows exponentially, the need for accurate automatic domain decomposition methods becomes more essential. In this article, we introduce a bottom-up algorithm for assigning protein domains using a graph theoretical approach. This algorithm is based on a center-based clustering approach. For constructing initial clusters, members of an independent dominating set for the graph representation of a protein are considered as the centers. A distance matrix is then defined for these clusters. To obtain final domains, these clusters are merged using the compactness principle of domains and a method similar to the neighbor-joining algorithm considering some thresholds. The thresholds are computed using a training set consisting of 50 protein chains. The algorithm is implemented using C++ language and is named ProDomAs. To assess the performance of ProDomAs, its results are compared with seven automatic methods, against five publicly available benchmarks. The results show that ProDomAs outperforms other methods applied on the mentioned benchmarks. The performance of ProDomAs is also evaluated against 6342 chains obtained from ASTRAL SCOP 1.71. ProDomAs is freely available at http://www.bioinf.cs.ipm.ir/software/prodomas. PMID:24596179

  15. Chemical shifts assignments of the archaeal MC1 protein and a strongly bent 15 base pairs DNA duplex in complex.

    Science.gov (United States)

    Loth, Karine; Landon, Céline; Paquet, Françoise

    2015-04-01

    MC1 is the most abundant architectural protein present in Methanosarcina thermophila CHTI55 in laboratory growth conditions and is structurally unrelated to other DNA-binding proteins. MC1 functions are to shape and to protect DNA against thermal denaturation by binding to it. Therefore, MC1 has a strong affinity for any double-stranded DNA. However, it recognizes and preferentially binds to bent DNA, such as four-way junctions and negatively supercoiled DNA minicircles. Combining NMR data, electron microscopy data, biochemistry, molecular modelisation and docking approaches, we proposed recently a new type of DNA/protein complex, in which the monomeric protein MC1 binds on the concave side of a strongly bent 15 base pairs DNA. We present here the NMR chemical shifts assignments of each partner in the complex, (1)H (15)N MC1 protein and (1)H (13)C (15)N bent duplex DNA, as first step towards the first experimental 3D structure of this new type of DNA/protein complex.

  16. Charge site assignment in native proteins by ultraviolet photodissociation (UVPD) mass spectrometry.

    Science.gov (United States)

    Morrison, Lindsay J; Brodbelt, Jennifer S

    2016-01-01

    Characterization of all gas-phase charge sites of natively sprayed proteins and peptides is demonstrated using 193 nm UVPD. The high sequence coverage offered by UVPD is exploited for the accurate determination of charge sites in protein systems up to 18 kDa, allowing charge site to be studied as a function of protein conformation and the presence of disulfide bonds. Charging protons are found on both basic sidechains and on the amide backbone of less basic amino acids such as serine, glutamine, and proline. UVPD analysis was performed on the 3+ charge state of melittin, the 5+ to 8+ charge states of ubiquitin, and the 8+ charge state of reduced and oxidized β-lactoglobulin. The location of charges in gas-phase proteins is known to impact structure; molecular modeling of different charge site motifs of 3+ melittin demonstrates how placement of protons in simulations can dramatically impact the predicted structure of the molecule. The location of positive charge sites in ubiquitin and β-lactoglobulin are additionally found to depend on the presence or absence of salt-bridges, columbic repulsion across the length of the peptide, and protein conformation. Charge site isomers are demonstrated for ubiquitin and β-lactoglobulin but found to be much less numerous than previously predicted. PMID:26596460

  17. METHODOLOGY & CALCULATIONS FOR THE ASSIGNMENT OF WASTE FOR THE LARGE UNDERGROUND WASTE STORAGE TANKS AT THE HANFORD SITE

    Energy Technology Data Exchange (ETDEWEB)

    TU, T.A.

    2007-01-04

    Waste stored within tank farm double-shell tanks (DST) and single-shell tanks (SST) generates flammable gas (principally hydrogen) to varying degrees depending on the type, amount, geometry, and condition of the waste. The waste generates hydrogen through the radiolysis of water and organic compounds, thermolytic decomposition of organic compounds, and corrosion of a tank's carbon steel walls. Radiolysis and thermolytic decomposition also generates ammonia. Nonflammable gases, which act as dilutents (such as nitrous oxide), are also produced. Additional flammable gases (e.g., methane) are generated by chemical reactions between various degradation products of organic chemicals present in the tanks. Volatile and semi-volatile organic chemicals in tanks also produce organic vapors. The generated gases in tank waste are either released continuously to the tank headspace or are retained in the waste matrix. Retained gas may be released in a spontaneous or induced gas release event (GRE) that can significantly increase the flammable gas concentration in the tank headspace as described in RPP-7771, Flammable Gas Safety Isme Resolution. Appendices A through I provide supporting information. The document categorizes each of the large waste storage tanks into one of several categories based on each tank's waste and characteristics. These waste group assignments reflect a tank's propensity to retain a significant volume of flammable gases and the potential of the waste to release retained gas by a buoyant displacement event. Revision 6 is the annual update of the flammable gas Waste Groups for DSTs and SSTs.

  18. Assignment of the gene for human sphingolipid activator protein-2 (SAP-2) to chromosome 10.

    OpenAIRE

    Fujibayashi, S; Kao, F T; Jones, C.; Morse, H; Law, M; Wenger, D A

    1985-01-01

    Sphingolipid activator protein-2 (SAP-2) has been found to stimulate the enzymatic hydrolysis of glucosylceramide, galactosylceramide, and sphingomyelin. When human skin fibroblast extracts were subjected to sodium dodecylsulfate-polyacrylamide gel electrophoresis followed by electroblotting and immunochemical staining using monospecific antibodies against SAP-2, two or three major bands with estimated mol. wts. of 9,000-10,000 were found. These antibodies did not crossreact with purified SAP...

  19. Selective 'unlabeling' of amino acids in fractionally 13C labeled proteins: An approach for stereospecific NMR assignments of CH3 groups in Val and Leu residues

    Energy Technology Data Exchange (ETDEWEB)

    Atreya, H.S.; Chary, K.V.R. [Tata Institute of Fundamental Research, Department of Chemical Sciences (India)

    2001-03-15

    A novel methodology for stereospecific NMR assignments of methyl (CH{sub 3}) groups of Val and Leu residues in fractionally {sup 13}C-labeled proteins is presented. The approach is based on selective 'unlabeling' of specific amino acids in proteins while fractionally {sup 13}C-labeling the rest. A 2D [{sup 13}C-{sup 1}H] HSQC spectrum recorded on such a sample is devoid of peaks belonging to the 'unlabeled' amino acid residues. Such spectral simplification aids in unambiguous stereospecific assignment of diastereotopic CH{sub 3} groups in Val and Leu residues in large proteins. This methodology has been demonstrated on a 15 kDa calcium binding protein from Entamoeba histolytica (Eh-CaBP)

  20. Enhanced functional and structural domain assignments using remote similarity detection procedures for proteins encoded in the genome of Mycobacterium tuberculosis H37Rv

    Indian Academy of Sciences (India)

    Seema Namboori; Natasha Mhatre; Sentivel Sujatha; Narayanaswamy Srinivasan; Shashi Bhushan Pandit

    2004-09-01

    The sequencing of the Mycobacterium tuberculosis (MTB) H37Rv genome has facilitated deeper insights into the biology of MTB, yet the functions of many MTB proteins are unknown. We have used sensitive profile-based search procedures to assign functional and structural domains to infer functions of gene products encoded in MTB. These domain assignments have been made using a compendium of sequence and structural domain families. Functions are predicted for 78% of the encoded gene products. For 69% of these, functions can be inferred by domain assignments. The functions for the rest are deduced from their homology to proteins of known function. Superfamily relationships between families of unknown and known structures have increased structural information by ∼ 11%. Remote similarity detection methods have enabled domain assignments for 1325 ‘hypothetical proteins’. The most populated families in MTB are involved in lipid metabolism, entry and survival of the bacillus in host. Interestingly, for 353 proteins, which we refer to as MTB-specific, no homologues have been identified. Numerous, previously unannotated, hypothetical proteins have been assigned domains and some of these could perhaps be the possible chemotherapeutic targets. MTB-specific proteins might include factors responsible for virulence. Importantly, these assignments could be valuable for experimental endeavors. The detailed results are publicly available at http://hodgkin.mbu.iisc.ernet.in/∼dots.

  1. Pseudo 5D HN(C)N Experiment to Facilitate the Assignment of Backbone Resonances in Proteins Exhibiting High Backbone Shift Degeneracy

    CERN Document Server

    Kumar, Dinesh; Shukla, Vaibhav Kumar; Pandey, Himanshu; Arora, Ashish; Guleria, Anupam

    2014-01-01

    Assignment of protein backbone resonances is most routinely carried out using triple resonance three dimensional NMR experiments involving amide 1H and 15N resonances. However for intrinsically unstructured proteins, alpha-helical proteins or proteins containing several disordered fragments, the assignment becomes problematic because of high degree of backbone shift degeneracy. In this backdrop, a novel reduced dimensionality (RD) experiment -(5,3)D-hNCO-CANH- is presented to facilitate (and/or to validate) the sequential backbone resonance assignment in such proteins. The proposed 3D NMR experiment makes use of the modulated amide 15N chemical shifts (resulting from the joint sampling along both its indirect dimensions) to resolve the ambiguity involved in connecting the neighboring amide resonances (i.e. HiNi and Hi-1Ni-1) for overlapping amide NH peaks. The experiment -encoding 5D spectral information- leads to a conventional 3D spectrum with significantly reduced spectral crowding and complexity. The impr...

  2. How do you assign persistent identifiers to extracts from large, complex, dynamic data sets that underpin scholarly publications?

    Science.gov (United States)

    Wyborn, Lesley; Car, Nicholas; Evans, Benjamin; Klump, Jens

    2016-04-01

    Persistent identifiers in the form of a Digital Object Identifier (DOI) are becoming more mainstream, assigned at both the collection and dataset level. For static datasets, this is a relatively straight-forward matter. However, many new data collections are dynamic, with new data being appended, models and derivative products being revised with new data, or the data itself revised as processing methods are improved. Further, because data collections are becoming accessible as services, researchers can log in and dynamically create user-defined subsets for specific research projects: they also can easily mix and match data from multiple collections, each of which can have a complex history. Inevitably extracts from such dynamic data sets underpin scholarly publications, and this presents new challenges. The National Computational Infrastructure (NCI) has been experiencing and making progress towards addressing these issues. The NCI is large node of the Research Data Services initiative (RDS) of the Australian Government's research infrastructure, which currently makes available over 10 PBytes of priority research collections, ranging from geosciences, geophysics, environment, and climate, through to astronomy, bioinformatics, and social sciences. Data are replicated to, or are produced at, NCI and then processed there to higher-level data products or directly analysed. Individual datasets range from multi-petabyte computational models and large volume raster arrays, down to gigabyte size, ultra-high resolution datasets. To facilitate access, maximise reuse and enable integration across the disciplines, datasets have been organized on a platform called the National Environmental Research Data Interoperability Platform (NERDIP). Combined, the NERDIP data collections form a rich and diverse asset for researchers: their co-location and standardization optimises the value of existing data, and forms a new resource to underpin data-intensive Science. New publication

  3. Analysis and application of large-scale protein-protein interaction data sets

    Institute of Scientific and Technical Information of China (English)

    SUN Jingchun; XU Jinlin; LI Yixue; SHI Tieliu

    2005-01-01

    Protein-protein interactions play key roles in cells. Lots of experimental approaches and in silico methods have been developed to identify and predict large-scale protein-protein interactions. However, compared with the traditionally experimental results, the high-throughput protein-protein interaction data often contain the false positives in high probability. In order to fully utilize the large-scale data, it is necessary to develop bioinformatic methods for systematically evaluating those data in order to further improve the data reliability and mine biological information. This review summarizes the methodologies of analysis and application of high-throughput protein-protein interaction data, including the evaluation methods, the relationship between protein-protein interaction data and other protein biological information, and their applications in biological study. In addition, this paper also suggests some interesting topics on mining high-throughput protein-protein interaction data.

  4. Proton-detected scalar coupling based assignment strategies in MAS solid-state NMR spectroscopy applied to perdeuterated proteins

    Science.gov (United States)

    Linser, Rasmus; Fink, Uwe; Reif, Bernd

    2008-07-01

    Assignment of proteins in MAS (magic angle spinning) solid-state NMR relies so far on correlations among heteronuclei. This strategy is based on well dispersed resonances in the 15N dimension. In many complex cases like membrane proteins or amyloid fibrils, an additional frequency dimension is desirable in order to spread the amide resonances. We show here that proton detected HNCO, HNCA, and HNCACB type experiments can successfully be implemented in the solid-state. Coherences are sufficiently long lived to allow pulse schemes of a duration greater than 70 ms before incrementation of the first indirect dimension. The achieved resolution is comparable to the resolution obtained in solution-state NMR experiments. We demonstrate the experiments using a triply labeled sample of the SH3 domain of chicken α-spectrin, which was re-crystallized in H 2O/D 2O using a ratio of 1/9. We employ paramagnetic relaxation enhancement (PRE) using EDTA chelated Cu II to enable rapid data acquisition.

  5. Disulfide assignment of the C-terminal cysteine knot of agouti-related protein (AGRP) by direct sequencing analysis.

    Science.gov (United States)

    Young, Y; Zeni, L; Rosenfeld, R D; Stark, K L; Rohde, M F; Haniu, M

    1999-12-01

    We have assigned the disulfide structure of Md-65 agouti-related protein (Md65-AGRP) using differential reduction and alkylation followed by direct sequencing analysis. The mature human AGRP is a single polypeptide chain of 112 amino acid residues, consisting of an N-terminal acidic region and a unique C-terminal cysteine-rich domain. The C-terminal domain, a 48 amino acid peptide named Md65-AGRP, was expressed in Escherichia coil cells and refolded under different conditions from the mature recombinant protein. The disulfide bonds in the cystine knot structure of Md65-AGRP were partially reduced using tris(2-carboxyethyl) phosphine (TCEP) under acidic conditions, followed by alkylation with N-ethylmaleimide (NEM). The procedure generated several isoforms with varying degrees of NEM alkylation. The multiple forms of Md65-AGRP generated by partial reduction and NEM modification were then completely reduced and carboxymethylated to identify unreactive disulfide bonds. Differentially labeled Md65-AGRP were directly sequenced and analyzed by MALDI mass spectrometry. The results confirmed that Md65-AGRP contained the same disulfide structure as that of Md5-AGRP reported previously [Bures, E. J., Hui, J. O., Young, Y. et al. (1998) Biochemistry 37, 12172-12177].

  6. Comparative 2D NMR studies of human insulin and des-pentapeptide insulin: Sequential resonance assignment and implications for protein dynamics and receptor recognition

    Energy Technology Data Exchange (ETDEWEB)

    Hua, Qingxin (Harvard Medical School, Boston, MA (United States)); Weiss, M.A. (Harvard Medical School, Boston, MA (United States) Massachusetts General Hospital, Boston, MA (United States))

    1991-06-04

    The solution structure and dynamics of human insulin are ivestigated by 2D {sup 1}H NMR spectroscopy in reference to a previously analyzed analogue, des-pentapeptide (B26-B30) insulin. This spectroscopic comparison is of interest since (i) the structure of the C-terminal region of the B-chain has not been determined in the monomeric state and (ii) the role of this region in binding to the insulin receptor has been the subject of long-standing speculation. The present NMR studies are conducted in the presence of an organic cosolvent (20% acetic acid), under which conditions both proteins are monomeric and stably folded. Complete sequential assignment of human insulin is obtained and leads to the following conclusions. (1) The secondary structure of the insulin monomer (three {alpha}-helices and B-chain {beta}-turn) is similar to that observed in the 2-Zn crustal state. (2) The folding of DPI is essentially the same as the corresponding portion of intact insulin, in accord with the similarities between their respective crystal structues. (3) residues B24-B28 adopt an extended configuration in the monomer and pack against the hydrophobic core as in crystallographic dimers; residues B29 and B30 are largely disordered. (4) The insulin fold is shown to provide a model for collective motions in a protein with implications for the mechanism of protein-protein recognition. To their knowledge, this paper describes the first detailed analysis of a protein NMR spectrum under conditions of extensive conformational broadening.

  7. Comparative 2D NMR studies of human insulin and des-pentapeptide insulin: Sequential resonance assignment and implications for protein dynamics and receptor recognition

    International Nuclear Information System (INIS)

    The solution structure and dynamics of human insulin are ivestigated by 2D 1H NMR spectroscopy in reference to a previously analyzed analogue, des-pentapeptide (B26-B30) insulin. This spectroscopic comparison is of interest since (i) the structure of the C-terminal region of the B-chain has not been determined in the monomeric state and (ii) the role of this region in binding to the insulin receptor has been the subject of long-standing speculation. The present NMR studies are conducted in the presence of an organic cosolvent (20% acetic acid), under which conditions both proteins are monomeric and stably folded. Complete sequential assignment of human insulin is obtained and leads to the following conclusions. (1) The secondary structure of the insulin monomer (three α-helices and B-chain β-turn) is similar to that observed in the 2-Zn crustal state. (2) The folding of DPI is essentially the same as the corresponding portion of intact insulin, in accord with the similarities between their respective crystal structues. (3) residues B24-B28 adopt an extended configuration in the monomer and pack against the hydrophobic core as in crystallographic dimers; residues B29 and B30 are largely disordered. (4) The insulin fold is shown to provide a model for collective motions in a protein with implications for the mechanism of protein-protein recognition. To their knowledge, this paper describes the first detailed analysis of a protein NMR spectrum under conditions of extensive conformational broadening

  8. Pseudo 5D HN(C)N experiment to facilitate the assignment of backbone resonances in proteins exhibiting high backbone shift degeneracy

    Energy Technology Data Exchange (ETDEWEB)

    Kumar, Dinesh, E-mail: dineshcbmr@gmail.com [Centre of Biomedical Research (CBMR), SGPGIMS Campus, Raibareli Road, Lucknow 226014 (India); Raikwal, Nisha [Centre of Biomedical Research (CBMR), SGPGIMS Campus, Raibareli Road, Lucknow 226014 (India); Shukla, Vaibhav Kumar; Pandey, Himanshu; Arora, Ashish [Molecular and Structural Biology Division, CSIR, Central Drug Research Institute, Lucknow 226031 (India); Guleria, Anupam, E-mail: anuguleriaphy@gmail.com [Centre of Biomedical Research (CBMR), SGPGIMS Campus, Raibareli Road, Lucknow 226014 (India)

    2014-09-30

    Graphical abstract: - Highlights: • A reduced dimensionality experiment – referred as pseudo 5D HN(C)N- is presented. • Encodes highly resolved 5D spectral information in a 3D spectrum. • Superior in terms of peak dispersion. • Facilitates assignment of crowded HSQC spectra of moderately sized proteins. • Modulated {sup 15}N chemical shifts are used to break the amide shift degeneracy. - Abstract: Assignment of protein backbone resonances is most routinely carried out using triple resonance three-dimensional NMR experiments involving amide {sup 1}H/{sup 15}N resonances. However for intrinsically unstructured proteins, alpha-helical proteins or proteins containing several disordered fragments, the assignment becomes problematic because of high-degree of backbone shift degeneracy. In this backdrop, a novel reduced-dimensionality (RD) experiment –(5, 3)D-hNCO-CANH- is presented to facilitate/validate the sequential backbone resonance assignment in such proteins. The proposed 3D NMR experiment makes use of the modulated amide {sup 15}N chemical shifts (resulting from the joint sampling along both its indirect dimensions) to resolve the ambiguity involved in connecting the neighboring amide resonances (i.e. H{sub i}N{sub i} and H{sub i−1}N{sub i−1}) for overlapping amide-NH peaks. The experiment -in combination with routine triple resonance 3D-NMR experiments involving backbone amide ({sup 1}H/{sup 15}N) and carbon ({sup 13}C{sup α}/{sup 13}C′) chemical shifts- will serve as a powerful complementary tool to achieve the nearly complete assignment of protein backbone resonances in a time efficient manner.

  9. Two- and three-dimensional sup 1 H NMR studies of a wheat phospholipid transfer protein: Sequential resonance assignments and secondary structure

    Energy Technology Data Exchange (ETDEWEB)

    Simorre, J.P.; Caille, A. (Centre National de la Recherche Scientifique, Orleans (France)); Marion, D. (Laboratoire de Resonance Magnetique en Biologie et Medecine, Grenoble (France)); Marion, D. (INRA, Nantes (France)); Ptak, M. (Centre National de la Recherche Scientifique, Orleans (France) Univ. d' Orleans (France))

    1991-12-10

    Two- and three-dimensional {sup 1}H NMR experiments have been used to sequentially assign nearly all proton resonances of the 90 residues of wheat phospholipid transfer protein. Only a few side-chain protons were not identified because of degeneracy or overlapping. The identification of spin systems and the sequential assignment were made at the same time by combining the data of the two- and three-dimensional experiments. The classical two-dimensional COSY, HOHAHA, and NOESY experiments benefit from both good resolution and high sensitivity, allowing the detection of long-range dipolar connectivities. The three-dimensional HOHAHA-NOESY experiment offers the advantage of a faster and unambiguous assignment. As a matter of fact, homonuclear three-dimensional NMR spectroscopy prove to be a very efficient method for resonance assignments of protein {sup 1}H NMR spectra which cannot be unraveled by 2D methods. An assignment strategy which overcomes most of the ambiguities has been proposed, in which each individual assignment toward the C-terminal end is supported by another in the opposite direction originating from a completely different part of the spectrum. Location of secondary structures of the phospholipid transfer protein was determined by using the method of analysis introduced here and was confirmed by {sup 3}J{sub {alpha}NH} coupling and NH exchange rates. Except for the C-terminal part, the polypeptide chain appears to be organized mainly as helical fragments connected by disulfide bridges. Further modeling will display the overall folding of the protein and should provide a better understanding of its interactions with lipids.

  10. Complex protein nanopatterns over large areas via colloidal lithography

    DEFF Research Database (Denmark)

    Kristensen, Stine H; Pedersen, Gitte Albinus; Ogaki, Ryosuke;

    2013-01-01

    matrix proteins (vitronectin) or cellular ligands (the extracellular domain of E-cadherin) in the nanopatterns, whereas the selective poly(l-lysine)–poly(ethylene glycol) functionalization of the SiO2 matrix renders it protein repellent. Cell studies, as a proof of principle, demonstrate the potential...... are used to generate complex protein nanopatterns over large areas. Simple circular patches or more complex ring structures are produced in addition to hierarchical patterns of smaller patches. The gold regions are modified through alkanethiol chemistry, which enables the preparation of extracellular...

  11. Protein resonance assignment at MAS frequencies approaching 100 kHz: a quantitative comparison of J-coupling and dipolar-coupling-based transfer methods

    Energy Technology Data Exchange (ETDEWEB)

    Penzel, Susanne; Smith, Albert A.; Agarwal, Vipin; Hunkeler, Andreas [ETH Zürich, Physical Chemistry (Switzerland); Org, Mai-Liis; Samoson, Ago, E-mail: ago.samoson@ttu.ee [Tallinn University of Technology, NMR Instituut, Tartu Teadus, Tehnomeedikum (Estonia); Böckmann, Anja, E-mail: a.bockmann@ibcp.fr [UMR 5086 CNRS/Université de Lyon 1, Institut de Biologie et Chimie des Protéines (France); Ernst, Matthias, E-mail: maer@ethz.ch; Meier, Beat H., E-mail: beme@ethz.ch [ETH Zürich, Physical Chemistry (Switzerland)

    2015-10-15

    We discuss the optimum experimental conditions to obtain assignment spectra for solid proteins at magic-angle spinning (MAS) frequencies around 100 kHz. We present a systematic examination of the MAS dependence of the amide proton T{sub 2}′ times and a site-specific comparison of T{sub 2}′ at 93 kHz versus 60 kHz MAS frequency. A quantitative analysis of transfer efficiencies of building blocks, as they are used for typical 3D experiments, was performed. To do this, we compared dipolar-coupling and J-coupling based transfer steps. The building blocks were then combined into 3D experiments for sequential resonance assignment, where we evaluated signal-to-noise ratio and information content of the different 3D spectra in order to identify the best assignment strategy. Based on this comparison, six experiments were selected to optimally assign the model protein ubiquitin, solely using spectra acquired at 93 kHz MAS. Within 3 days of instrument time, the required spectra were recorded from which the backbone resonances have been assigned to over 96 %.

  12. A large piece of a small pie: Minimum wages and unemployment benefits in an assignment model with search frictions

    NARCIS (Netherlands)

    P.A. Gautier (Pieter); C.N. Teulings (Coen)

    2003-01-01

    textabstractMost empirical studies on the minimum wage find a spike at the minimum wage, compression of wage differentials at a large interval above the minimum wage and small employment losses. This paper offers a search model which is consistent with these facts. We consider a continuum of worker

  13. Six- and seven-dimensional experiments by combination of sparse random sampling and projection spectroscopy dedicated for backbone resonance assignment of intrinsically disordered proteins

    Energy Technology Data Exchange (ETDEWEB)

    Żerko, Szymon; Koźmiński, Wiktor, E-mail: kozmin@chem.uw.edu.pl [University of Warsaw, Faculty of Chemistry, Biological and Chemical Research Centre (Poland)

    2015-11-15

    Two novel six- and seven-dimensional NMR experiments are proposed. The new experiments employ non-uniform sampling that enables achieving high resolution in four indirectly detected dimensions and synchronous sampling in the additional dimensions using projection spectroscopy principle. The resulted data sets could be processed as five-dimensional data using existing software. The experiments facilitate resonance assignment of intrinsically disordered proteins. The novel experiments were successfully tested using 1 mM sample of α-synuclein on 600 and 800 MHz NMR spectrometers equipped with standard room temperature probes. The experiments allowed backbone assignment from a 1-day acquisition.

  14. Secondary structural analysis of proteins based on 13C chemical shift assignments in unresolved solid-state NMR spectra enhanced by fragmented structure database

    International Nuclear Information System (INIS)

    Magic-angle-spinning solid-state 13C NMR spectroscopy is useful for structural analysis of non-crystalline proteins. However, the signal assignments and structural analysis are often hampered by the signal overlaps primarily due to minor structural heterogeneities, especially for uniformly-13C,15N labeled samples. To overcome this problem, we present a method for assigning 13C chemical shifts and secondary structures from unresolved two-dimensional 13C–13C MAS NMR spectra by spectral fitting, named reconstruction of spectra using protein local structures (RESPLS). The spectral fitting was conducted using databases of protein fragmented structures related to 13Cα, 13Cβ, and 13C′ chemical shifts and cross-peak intensities. The experimental 13C–13C inter- and intra-residue correlation spectra of uniformly isotope-labeled ubiquitin in the lyophilized state had a few broad peaks. The fitting analysis for these spectra provided sequence-specific Cα, Cβ, and C′ chemical shifts with an accuracy of about 1.5 ppm, which enabled the assignment of the secondary structures with an accuracy of 79 %. The structural heterogeneity of the lyophilized ubiquitin is revealed from the results. Test of RESPLS analysis for simulated spectra of five different types of proteins indicated that the method allowed the secondary structure determination with accuracy of about 80 % for the 50–200 residue proteins. These results demonstrate that the RESPLS approach expands the applicability of the NMR to non-crystalline proteins exhibiting unresolved 13C NMR spectra, such as lyophilized proteins, amyloids, membrane proteins and proteins in living cells.

  15. Backbone and Ile-δ1, Leu, Val Methyl 1H, 13C and 15N NMR chemical shift assignments for human interferon-stimulated gene 15 protein

    Energy Technology Data Exchange (ETDEWEB)

    Yin, Cuifeng; Aramini, James M.; Ma, LiChung; Cort, John R.; Swapna, G.V.T.; Krug, R. M.; Montelione, Gaetano

    2011-10-01

    Human interferon-stimulated gene 15 protein (ISG15), also called ubiquitin cross-reactive protein (UCRP), is the first identified ubiquitin-like protein containing two ubiquitin-like domains fused in tandem. The active form of ISG15 is conjugated to target proteins via the C-terminal glycine residue through an isopeptide bond in a manner similar to ubiquitin. The biological role of ISG15 is strongly associated with the modulation of cell immune function, and there is mounting evidence suggesting that many viral pathogens evade the host innate immune response by interfering with ISG15 conjugation to both host and viral proteins in a variety of ways. Here we report nearly complete backbone 1HN, 15N, 13CO, and 13Ca, as well as side chain 13Cb, methyl (Ile-d1, Leu, Val), amide (Asn, Gln), and indole NH (Trp) NMR resonance assignments for the 157-residue human ISG15 protein. These resonance assignments provide the basis for future structural and functional solution NMR studies of the biologically important human ISG15 protein.

  16. Insights into Hox protein function from a large scale combinatorial analysis of protein domains.

    Directory of Open Access Journals (Sweden)

    Samir Merabet

    2011-10-01

    Full Text Available Protein function is encoded within protein sequence and protein domains. However, how protein domains cooperate within a protein to modulate overall activity and how this impacts functional diversification at the molecular and organism levels remains largely unaddressed. Focusing on three domains of the central class Drosophila Hox transcription factor AbdominalA (AbdA, we used combinatorial domain mutations and most known AbdA developmental functions as biological readouts to investigate how protein domains collectively shape protein activity. The results uncover redundancy, interactivity, and multifunctionality of protein domains as salient features underlying overall AbdA protein activity, providing means to apprehend functional diversity and accounting for the robustness of Hox-controlled developmental programs. Importantly, the results highlight context-dependency in protein domain usage and interaction, allowing major modifications in domains to be tolerated without general functional loss. The non-pleoitropic effect of domain mutation suggests that protein modification may contribute more broadly to molecular changes underlying morphological diversification during evolution, so far thought to rely largely on modification in gene cis-regulatory sequences.

  17. Hepatitis B virus large surface protein: function and fame

    OpenAIRE

    Churin, Yuri; Roderfeld, Martin; Roeb, Elke

    2015-01-01

    Chronic infection with hepatitis B virus (HBV) is the leading cause of liver cirrhosis and hepatocellular carcinoma worldwide. HBV life cycle begins with viral attachment to hepatocytes, mediated by the large HBV surface protein (LHBs). Identification of the sodium-taurocholate cotransporting polypeptide (NTCP) as a HBV receptor has revealed a suitable target for viral entry inhibition. Analysis of serum hepatitis B surface antigen (HBsAg) level is a non-invasive diagnostic parameter that imp...

  18. Predicting protein functions from redundancies in large-scale protein interaction networks

    Science.gov (United States)

    Samanta, Manoj Pratim; Liang, Shoudan

    2003-01-01

    Interpreting data from large-scale protein interaction experiments has been a challenging task because of the widespread presence of random false positives. Here, we present a network-based statistical algorithm that overcomes this difficulty and allows us to derive functions of unannotated proteins from large-scale interaction data. Our algorithm uses the insight that if two proteins share significantly larger number of common interaction partners than random, they have close functional associations. Analysis of publicly available data from Saccharomyces cerevisiae reveals >2,800 reliable functional associations, 29% of which involve at least one unannotated protein. By further analyzing these associations, we derive tentative functions for 81 unannotated proteins with high certainty. Our method is not overly sensitive to the false positives present in the data. Even after adding 50% randomly generated interactions to the measured data set, we are able to recover almost all (approximately 89%) of the original associations.

  19. Spectral editing at ultra-fast magic-angle-spinning in solid-state NMR: facilitating protein sequential signal assignment by HIGHLIGHT approach.

    Science.gov (United States)

    Wang, Songlin; Matsuda, Isamu; Long, Fei; Ishii, Yoshitaka

    2016-02-01

    This study demonstrates a novel spectral editing technique for protein solid-state NMR (SSNMR) to simplify the spectrum drastically and to reduce the ambiguity for protein main-chain signal assignments in fast magic-angle-spinning (MAS) conditions at a wide frequency range of 40-80 kHz. The approach termed HIGHLIGHT (Wang et al., in Chem Comm 51:15055-15058, 2015) combines the reverse (13)C, (15)N-isotope labeling strategy and selective signal quenching using the frequency-selective REDOR pulse sequence under fast MAS. The scheme allows one to selectively observe the signals of "highlighted" labeled amino-acid residues that precede or follow unlabeled residues through selectively quenching (13)CO or (15)N signals for a pair of consecutively labeled residues by recoupling (13)CO-(15)N dipolar couplings. Our numerical simulation results showed that the scheme yielded only ~15% loss of signals for the highlighted residues while quenching as much as ~90% of signals for non-highlighted residues. For lysine-reverse-labeled micro-crystalline GB1 protein, the 2D (15)N/(13)Cα correlation and 2D (13)Cα/(13)CO correlation SSNMR spectra by the HIGHLIGHT approach yielded signals only for six residues following and preceding the unlabeled lysine residues, respectively. The experimental dephasing curves agreed reasonably well with the corresponding simulation results for highlighted and quenched residues at spinning speeds of 40 and 60 kHz. The compatibility of the HIGHLIGHT approach with fast MAS allows for sensitivity enhancement by paramagnetic assisted data collection (PACC) and (1)H detection. We also discuss how the HIGHLIGHT approach facilitates signal assignments using (13)C-detected 3D SSNMR by demonstrating full sequential assignments of lysine-reverse-labeled micro-crystalline GB1 protein (~300 nmol), for which data collection required only 11 h. The HIGHLIGHT approach offers valuable means of signal assignments especially for larger proteins through reducing the

  20. Spectral editing at ultra-fast magic-angle-spinning in solid-state NMR: facilitating protein sequential signal assignment by HIGHLIGHT approach

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Songlin; Matsuda, Isamu; Long, Fei; Ishii, Yoshitaka, E-mail: yishii@uic.edu [University of Illinois at Chicago, Department of Chemistry (United States)

    2016-02-15

    This study demonstrates a novel spectral editing technique for protein solid-state NMR (SSNMR) to simplify the spectrum drastically and to reduce the ambiguity for protein main-chain signal assignments in fast magic-angle-spinning (MAS) conditions at a wide frequency range of 40–80 kHz. The approach termed HIGHLIGHT (Wang et al., in Chem Comm 51:15055–15058, 2015) combines the reverse {sup 13}C, {sup 15}N-isotope labeling strategy and selective signal quenching using the frequency-selective REDOR pulse sequence under fast MAS. The scheme allows one to selectively observe the signals of “highlighted” labeled amino-acid residues that precede or follow unlabeled residues through selectively quenching {sup 13}CO or {sup 15}N signals for a pair of consecutively labeled residues by recoupling {sup 13}CO–{sup 15}N dipolar couplings. Our numerical simulation results showed that the scheme yielded only ∼15 % loss of signals for the highlighted residues while quenching as much as ∼90 % of signals for non-highlighted residues. For lysine-reverse-labeled micro-crystalline GB1 protein, the 2D {sup 15}N/{sup 13}C{sub α} correlation and 2D {sup 13}C{sub α}/{sup 13}CO correlation SSNMR spectra by the HIGHLIGHT approach yielded signals only for six residues following and preceding the unlabeled lysine residues, respectively. The experimental dephasing curves agreed reasonably well with the corresponding simulation results for highlighted and quenched residues at spinning speeds of 40 and 60 kHz. The compatibility of the HIGHLIGHT approach with fast MAS allows for sensitivity enhancement by paramagnetic assisted data collection (PACC) and {sup 1}H detection. We also discuss how the HIGHLIGHT approach facilitates signal assignments using {sup 13}C-detected 3D SSNMR by demonstrating full sequential assignments of lysine-reverse-labeled micro-crystalline GB1 protein (∼300 nmol), for which data collection required only 11 h. The HIGHLIGHT approach offers valuable

  1. Spectral editing at ultra-fast magic-angle-spinning in solid-state NMR: facilitating protein sequential signal assignment by HIGHLIGHT approach

    International Nuclear Information System (INIS)

    This study demonstrates a novel spectral editing technique for protein solid-state NMR (SSNMR) to simplify the spectrum drastically and to reduce the ambiguity for protein main-chain signal assignments in fast magic-angle-spinning (MAS) conditions at a wide frequency range of 40–80 kHz. The approach termed HIGHLIGHT (Wang et al., in Chem Comm 51:15055–15058, 2015) combines the reverse 13C, 15N-isotope labeling strategy and selective signal quenching using the frequency-selective REDOR pulse sequence under fast MAS. The scheme allows one to selectively observe the signals of “highlighted” labeled amino-acid residues that precede or follow unlabeled residues through selectively quenching 13CO or 15N signals for a pair of consecutively labeled residues by recoupling 13CO–15N dipolar couplings. Our numerical simulation results showed that the scheme yielded only ∼15 % loss of signals for the highlighted residues while quenching as much as ∼90 % of signals for non-highlighted residues. For lysine-reverse-labeled micro-crystalline GB1 protein, the 2D 15N/13Cα correlation and 2D 13Cα/13CO correlation SSNMR spectra by the HIGHLIGHT approach yielded signals only for six residues following and preceding the unlabeled lysine residues, respectively. The experimental dephasing curves agreed reasonably well with the corresponding simulation results for highlighted and quenched residues at spinning speeds of 40 and 60 kHz. The compatibility of the HIGHLIGHT approach with fast MAS allows for sensitivity enhancement by paramagnetic assisted data collection (PACC) and 1H detection. We also discuss how the HIGHLIGHT approach facilitates signal assignments using 13C-detected 3D SSNMR by demonstrating full sequential assignments of lysine-reverse-labeled micro-crystalline GB1 protein (∼300 nmol), for which data collection required only 11 h. The HIGHLIGHT approach offers valuable means of signal assignments especially for larger proteins through reducing the

  2. Spectral editing at ultra-fast magic-angle-spinning in solid-state NMR: facilitating protein sequential signal assignment by HIGHLIGHT approach.

    Science.gov (United States)

    Wang, Songlin; Matsuda, Isamu; Long, Fei; Ishii, Yoshitaka

    2016-02-01

    This study demonstrates a novel spectral editing technique for protein solid-state NMR (SSNMR) to simplify the spectrum drastically and to reduce the ambiguity for protein main-chain signal assignments in fast magic-angle-spinning (MAS) conditions at a wide frequency range of 40-80 kHz. The approach termed HIGHLIGHT (Wang et al., in Chem Comm 51:15055-15058, 2015) combines the reverse (13)C, (15)N-isotope labeling strategy and selective signal quenching using the frequency-selective REDOR pulse sequence under fast MAS. The scheme allows one to selectively observe the signals of "highlighted" labeled amino-acid residues that precede or follow unlabeled residues through selectively quenching (13)CO or (15)N signals for a pair of consecutively labeled residues by recoupling (13)CO-(15)N dipolar couplings. Our numerical simulation results showed that the scheme yielded only ~15% loss of signals for the highlighted residues while quenching as much as ~90% of signals for non-highlighted residues. For lysine-reverse-labeled micro-crystalline GB1 protein, the 2D (15)N/(13)Cα correlation and 2D (13)Cα/(13)CO correlation SSNMR spectra by the HIGHLIGHT approach yielded signals only for six residues following and preceding the unlabeled lysine residues, respectively. The experimental dephasing curves agreed reasonably well with the corresponding simulation results for highlighted and quenched residues at spinning speeds of 40 and 60 kHz. The compatibility of the HIGHLIGHT approach with fast MAS allows for sensitivity enhancement by paramagnetic assisted data collection (PACC) and (1)H detection. We also discuss how the HIGHLIGHT approach facilitates signal assignments using (13)C-detected 3D SSNMR by demonstrating full sequential assignments of lysine-reverse-labeled micro-crystalline GB1 protein (~300 nmol), for which data collection required only 11 h. The HIGHLIGHT approach offers valuable means of signal assignments especially for larger proteins through reducing the

  3. Proton-detected MAS NMR experiments based on dipolar transfers for backbone assignment of highly deuterated proteins

    Science.gov (United States)

    Chevelkov, Veniamin; Habenstein, Birgit; Loquet, Antoine; Giller, Karin; Becker, Stefan; Lange, Adam

    2014-05-01

    Proton-detected solid-state NMR was applied to a highly deuterated insoluble, non-crystalline biological assembly, the Salmonella typhimurium type iii secretion system (T3SS) needle. Spectra of very high resolution and sensitivity were obtained at a low protonation level of 10-20% at exchangeable amide positions. We developed efficient experimental protocols for resonance assignment tailored for this system and the employed experimental conditions. Using exclusively dipolar-based interspin magnetization transfers, we recorded two sets of 3D spectra allowing for an almost complete backbone resonance assignment of the needle subunit PrgI. The additional information provided by the well-resolved proton dimension revealed the presence of two sets of resonances in the N-terminal helix of PrgI, while in previous studies employing 13C detection only a single set of resonances was observed.

  4. Large-volume protein crystal growth for neutron macromolecular crystallography.

    Science.gov (United States)

    Ng, Joseph D; Baird, James K; Coates, Leighton; Garcia-Ruiz, Juan M; Hodge, Teresa A; Huang, Sijay

    2015-04-01

    Neutron macromolecular crystallography (NMC) is the prevailing method for the accurate determination of the positions of H atoms in macromolecules. As neutron sources are becoming more available to general users, finding means to optimize the growth of protein crystals to sizes suitable for NMC is extremely important. Historically, much has been learned about growing crystals for X-ray diffraction. However, owing to new-generation synchrotron X-ray facilities and sensitive detectors, protein crystal sizes as small as in the nano-range have become adequate for structure determination, lessening the necessity to grow large crystals. Here, some of the approaches, techniques and considerations for the growth of crystals to significant dimensions that are now relevant to NMC are revisited. These include experimental strategies utilizing solubility diagrams, ripening effects, classical crystallization techniques, microgravity and theoretical considerations.

  5. Large scale protein separations: engineering aspects of chromatography.

    Science.gov (United States)

    Chisti, Y; Moo-Young, M

    1990-01-01

    The engineering considerations common to large scale chromatographic purification of proteins are reviewed. A discussion of the industrial chromatography fundamentals is followed by aspects which affect the scale of separation. The separation column geometry, the effect of the main operational parameters on separation performance, and the physical characteristics of column packing are treated. Throughout, the emphasis is on ion exchange and size exclusion techniques which together constitute the major portion of commercial chromatographic protein purifications. In all cases, the state of current technology is examined and areas in need of further development are noted. The physico-chemical advances now underway in chromatographic separation of biopolymers would ensure a substantially enhanced role for these techniques in industrial production of products of new biotechnology.

  6. Purification, renaturation, and reconstituted protein kinase activity of the Sendai virus large (L) protein: L protein phosphorylates the NP and P proteins in vitro.

    OpenAIRE

    Einberger, H; Mertz, R; Hofschneider, P H; Neubert, W J

    1990-01-01

    Sodium dodecyl sulfate-solubilized Sendai virus large (L) protein was highly purified by a one-step procedure, using hydroxylapatite column chromatography. Monoclonal antibodies addressed to the carboxyl-terminal amino acid sequence of the L protein were used for monitoring L protein during purification. By removing sodium dodecyl sulfate from purified L protein, a protein kinase activity was successfully renatured. P and NP proteins served as its substrates. After immunoprecipitation with an...

  7. Reduced dimensionality tailored HN(C)N experiments for facile backbone resonance assignment of proteins through unambiguous identification of sequential HSQC peaks

    Science.gov (United States)

    Kumar, Dinesh

    2013-12-01

    Two novel reduced dimensionality (RD) tailored HN(C)N [S.C. Panchal, N.S. Bhavesh, R.V. Hosur, Improved 3D triple resonance experiments, HNN and HN(C)N, for HN and 15N sequential correlations in (13C, 15N) labeled proteins: application to unfolded proteins, J. Biomol. NMR 20 (2001) 135-147] experiments are proposed to facilitate the backbone resonance assignment of proteins both in terms of its accuracy and speed. These experiments - referred here as (4,3)D-hNCOcaNH and (4,3)D-hNcoCANH - exploit the linear combination of backbone 15N and 13C‧/13Cα chemical shifts simultaneously to achieve higher peak dispersion and randomness along their respective F1 dimensions. Simply, this has been achieved by modulating the backbone 15N(i) chemical shifts with that of 13C‧ (i - 1)/13Cα (i - 1) spins following the established reduced dimensionality NMR approach [T. Szyperski, D.C. Yeh, D.K. Sukumaran, H.N. Moseley, G.T. Montelione, Reduced-dimensionality NMR spectroscopy for high-throughput protein resonance assignment, Proc. Natl. Acad. Sci. USA 99 (2002) 8009-8014]. Though the modification is simple it has resulted an ingenious improvement of HN(C)N both in terms of peak dispersion and easiness of establishing the sequential connectivities. The increased dispersion along F1 dimension solves two purposes here: (i) resolves the ambiguities arising because of degenerate 15N chemical shifts and (ii) reduces the signal overlap in F2(15N)-F3(1H) planes (an important requisite in HN(C)N based assignment protocol for facile and unambiguous identification of sequentially connected HSQC peaks). The performance of both these experiments and the assignment protocol has been demonstrated using bovine apo Calbindin-d9k (75 aa) and urea denatured UNC60B (a 152 amino acid ADF/cofilin family protein of Caenorhabditis elegans), as representatives of folded and unfolded protein systems, respectively.

  8. A non-uniformly sampled 4D HCC(CO)NH-TOCSY experiment processed using maximum entropy for rapid protein sidechain assignment

    Science.gov (United States)

    Mobli, Mehdi; Stern, Alan S.; Bermel, Wolfgang; King, Glenn F.; Hoch, Jeffrey C.

    2010-05-01

    One of the stiffest challenges in structural studies of proteins using NMR is the assignment of sidechain resonances. Typically, a panel of lengthy 3D experiments are acquired in order to establish connectivities and resolve ambiguities due to overlap. We demonstrate that these experiments can be replaced by a single 4D experiment that is time-efficient, yields excellent resolution, and captures unique carbon-proton connectivity information. The approach is made practical by the use of non-uniform sampling in the three indirect time dimensions and maximum entropy reconstruction of the corresponding 3D frequency spectrum. This 4D method will facilitate automated resonance assignment procedures and it should be particularly beneficial for increasing throughput in NMR-based structural genomics initiatives.

  9. Two-dimensional NMR and photo-CIDNP studies of the insulin monomer: Assignment of aromatic resonances with application to protein folding, structure, and dynamics

    International Nuclear Information System (INIS)

    The aromatic 1H NMR resonances of the insulin monomer are assigned at 500 MHz by comparative studies of chemically modified and genetically altered variants, including a mutant insulin (PheB25 → Leu) associated with diabetes mellitus. The two histidines, three phenylalanines, and four tyrosines are observed to be in distinct local environments; their assignment provides sensitive markers for studies of tertiary structure, protein dynamics, and protein folding. The environments of the tyrosine residues have also been investigated by photochemically induced dynamic nuclear polarization (photo-CIDNP) and analyzed in relation to packing constrains in the crystal structures of insulin. Dimerization involving specific B-chain interactions is observed with increasing protein concentration and is shown to depend on temperature, pH, and solvent composition. The differences between proinsulin and mini-proinsulin suggest a structural mechanism for the observation that the fully reduced B29-A1 analogue folds more efficiently than proinsulin to form the correct pattern of disulfide bonds. These results are discussed in relation to molecular mechanics calculations of insulin based on the available crystal structures

  10. Two-dimensional NMR and photo-CIDNP studies of the insulin monomer: Assignment of aromatic resonances with application to protein folding, structure, and dynamics

    Energy Technology Data Exchange (ETDEWEB)

    Weiss, M.A.; Shoelson, S.E. (Harvard Medical School, Boston, MA (USA) Massachusetts General Hospital, Boston (USA)); Nguyen, D.T.; O' Shea, E.; Karplus, M. (Harvard Univ., Cambridge, MA (USA)); Khait, I.; Neuringer, L.J. (Massachusetts Institute of Technology, Cambridge (USA)); Inouye, K. (Shionogi and Co., Ltd., Osaka (Japan)); Frank, B.H.; Beckage, M. (Eli Lilly and Co., Indianapolis, IN (USA))

    1989-12-12

    The aromatic {sup 1}H NMR resonances of the insulin monomer are assigned at 500 MHz by comparative studies of chemically modified and genetically altered variants, including a mutant insulin (PheB25 {yields} Leu) associated with diabetes mellitus. The two histidines, three phenylalanines, and four tyrosines are observed to be in distinct local environments; their assignment provides sensitive markers for studies of tertiary structure, protein dynamics, and protein folding. The environments of the tyrosine residues have also been investigated by photochemically induced dynamic nuclear polarization (photo-CIDNP) and analyzed in relation to packing constrains in the crystal structures of insulin. Dimerization involving specific B-chain interactions is observed with increasing protein concentration and is shown to depend on temperature, pH, and solvent composition. The differences between proinsulin and mini-proinsulin suggest a structural mechanism for the observation that the fully reduced B29-A1 analogue folds more efficiently than proinsulin to form the correct pattern of disulfide bonds. These results are discussed in relation to molecular mechanics calculations of insulin based on the available crystal structures.

  11. Reduced Dimensionality tailored HN(C)N Pulse Sequences for Efficient Backbone Resonance Assignment of Proteins through Rapid Identification of Sequential HSQC peaks

    CERN Document Server

    Kumar, Dinesh

    2013-01-01

    Two novel reduced dimensionality (RD) experiments -(4,3)D-hNCOcaNH and (4,3)D-hNcoCANH- have been presented here to facilitate the backbone resonance assignment of proteins both in terms of its accuracy and speed. The experiments basically represent an improvisation of previously reported HN(C)N experiment [Panchal et. al., J. Biomol. NMR. (2002), 20 (2), 135-147] and exploit the simple reduced dimensionality NMR concept [Szyperski et. al. (2002), Proc. Natl. Acad. Sci. U.S.A. 99(12), 8009-8014] to achieve (a) higher dispersion and resolution along the co-evolved F1 dimension and (b) rapid identification of sequential HSQC peaks on its F2(15N)- F3(1H) planes. The current implementation is based on the fact that the linear combination of 15N and 13CO/13Ca chemical shifts offers relatively better dispersion and randomness compared to the individual chemical shifts; thus enables the assignment of crowded HSQC spectra by resolving the ambiguities generally encountered in HNCN based assignment protocol because of ...

  12. Hepatitis B virus large surface protein: function and fame.

    Science.gov (United States)

    Churin, Yuri; Roderfeld, Martin; Roeb, Elke

    2015-02-01

    Chronic infection with hepatitis B virus (HBV) is the leading cause of liver cirrhosis and hepatocellular carcinoma worldwide. HBV life cycle begins with viral attachment to hepatocytes, mediated by the large HBV surface protein (LHBs). Identification of the sodium-taurocholate cotransporting polypeptide (NTCP) as a HBV receptor has revealed a suitable target for viral entry inhibition. Analysis of serum hepatitis B surface antigen (HBsAg) level is a non-invasive diagnostic parameter that improves HBV treatment opportunities. Furthermore, HBsAg plays an important role in manipulation of host immune response by HBV. However, observations in patients with chronic hepatitis B under conditions of immune suppression and in transgenic mouse models of HBV infection suggest, that in absence of adaptive immune responses cellular mechanisms induced by HBV may also lead to the development of liver diseases. Thus, the multifaceted pathological aspects of HBsAg predetermine the design of new therapeutical options modulating associated biological implications. PMID:25713800

  13. Systemic transport of Alfalfa mosaic virus can be mediated by the movement proteins of several viruses assigned to five genera of the 30K family.

    Science.gov (United States)

    Fajardo, Thor V M; Peiró, Ana; Pallás, Vicente; Sánchez-Navarro, Jesús

    2013-03-01

    We previously showed that the movement protein (MP) gene of Alfalfa mosaic virus (AMV) is functionally exchangeable for the cell-to-cell transport of the corresponding genes of Tobacco mosaic virus (TMV), Brome mosaic virus, Prunus necrotic ringspot virus, Cucumber mosaic virus and Cowpea mosaic virus. We have analysed the capacity of the heterologous MPs to systemically transport the corresponding chimeric AMV genome. All MPs were competent in systemic transport but required the fusion at their C terminus of the coat protein-interacting C-terminal 44 aa (A44) of the AMV MP. Except for the TMV MP, the presence of the hybrid virus in upper leaves correlated with the capacity to move locally. These results suggest that all the MPs assigned to the 30K superfamily should be exchangeable not only for local virus movement but also for systemic transport when the A44 fragment is present. PMID:23136366

  14. 3D-TROSY-based backbone and ILV-methyl resonance assignments of a 319-residue homodimer from a single protein sample

    Energy Technology Data Exchange (ETDEWEB)

    Krejcirikova, Anna; Tugarinov, Vitali, E-mail: vitali@umd.edu [University of Maryland, Department of Chemistry and Biochemistry (United States)

    2012-10-15

    The feasibility of practically complete backbone and ILV methyl chemical shift assignments from a single [U-{sup 2}H,{sup 15}N,{sup 13}C; Ile{delta}1-{l_brace}{sup 13}CH{sub 3}{r_brace}; Leu,Val-{l_brace}{sup 13}CH{sub 3}/{sup 12}CD{sub 3}{r_brace}]-labeled protein sample of the truncated form of ligand-free Bst-Tyrosyl tRNA Synthetase (Bst-{Delta}YRS), a 319-residue predominantly helical homodimer, is established. Protonation of ILV residues at methyl positions does not appreciably detract from the quality of TROSY triple resonance data. The assignments are performed at 40 Degree-Sign C to improve the sensitivity of the measurements and alleviate the overlap of {sup 1}H-{sup 15}N correlations in the abundant {alpha}-helical segments of the protein. A number of auxiliary approaches are used to assist in the assignment process: (1) selection of {sup 1}H-{sup 15}N amide correlations of certain residue types (Ala, Thr/Ser) that simplifies 2D {sup 1}H-{sup 15}N TROSY spectra, (2) straightforward identification of ILV residue types from the methyl-detected 'out-and-back' HMCM(CG)CBCA experiment, and (3) strong sequential HN-HN NOE connectivities in the helical regions. The two subunits of Bst-YRS were predicted earlier to exist in two different conformations in the absence of ligands. In agreement with our earlier findings (Godoy-Ruiz in J Am Chem Soc 133:19578-195781, 2011), no evidence of dimer asymmetry has been observed in either amide- or methyl-detected experiments.

  15. Molecular characterization and chromosomal assignment of equine cartilage derived retinoic acid sensitive protein (CD-RAP)/melanoma inhibitory activity (MIA)

    DEFF Research Database (Denmark)

    Berg, Lise Charlotte; Mata, Xavier; Thomsen, Preben Dybdahl

    2008-01-01

    Cartilage-derived retinoic acid sensitive protein (CD-RAP) also known as melanoma inhibitory activity (MIA) has already been established as a marker for chondrocyte differentiation and a number of cancerous condition sin humans. Studies have also shown that CD-RAP/MIA is a potential marker of joi...

  16. The human urinary proteome contains more than 1500 proteins, including a large proportion of membrane proteins

    Science.gov (United States)

    Adachi, Jun; Kumar, Chanchal; Zhang, Yanling; Olsen, Jesper V; Mann, Matthias

    2006-01-01

    Background Urine is a desirable material for the diagnosis and classification of diseases because of the convenience of its collection in large amounts; however, all of the urinary proteome catalogs currently being generated have limitations in their depth and confidence of identification. Our laboratory has developed methods for the in-depth characterization of body fluids; these involve a linear ion trap-Fourier transform (LTQ-FT) and a linear ion trap-orbitrap (LTQ-Orbitrap) mass spectrometer. Here we applied these methods to the analysis of the human urinary proteome. Results We employed one-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis and reverse phase high-performance liquid chromatography for protein separation and fractionation. Fractionated proteins were digested in-gel or in-solution, and digests were analyzed with the LTQ-FT and LTQ-Orbitrap at parts per million accuracy and with two consecutive stages of mass spectrometric fragmentation. We identified 1543 proteins in urine obtained from ten healthy donors, while essentially eliminating false-positive identifications. Surprisingly, nearly half of the annotated proteins were membrane proteins according to Gene Ontology (GO) analysis. Furthermore, extracellular, lysosomal, and plasma membrane proteins were enriched in the urine compared with all GO entries. Plasma membrane proteins are probably present in urine by secretion in exosomes. Conclusion Our analysis provides a high-confidence set of proteins present in human urinary proteome and provides a useful reference for comparing datasets obtained using different methodologies. The urinary proteome is unexpectedly complex and may prove useful in biomarker discovery in the future. PMID:16948836

  17. Sample preparation of membrane proteins suitable for solid-state MAS NMR and development of assignment strategies

    OpenAIRE

    Hiller, Matthias

    2009-01-01

    Although the basic structure of biological membranes is provided by the lipid bilayer, most of the specific functions are carried out by membrane proteins (MPs) such as channels, ion-pumps and receptors. Additionally, it is known, that mutations in MPs are directly or indirectly involved in many diseases. Thus, structure determination of MPs is of major interest not only in structural biology but also in pharmacology, especially for drug development. Advances in structural biology of membrane...

  18. Investigating the Role of Large-Scale Domain Dynamics in Protein-Protein Interactions

    Science.gov (United States)

    Delaforge, Elise; Milles, Sigrid; Huang, Jie-rong; Bouvier, Denis; Jensen, Malene Ringkjøbing; Sattler, Michael; Hart, Darren J.; Blackledge, Martin

    2016-01-01

    Intrinsically disordered linkers provide multi-domain proteins with degrees of conformational freedom that are often essential for function. These highly dynamic assemblies represent a significant fraction of all proteomes, and deciphering the physical basis of their interactions represents a considerable challenge. Here we describe the difficulties associated with mapping the large-scale domain dynamics and describe two recent examples where solution state methods, in particular NMR spectroscopy, are used to investigate conformational exchange on very different timescales. PMID:27679800

  19. Redundancies in Large-scale Protein Interaction Networks

    CERN Document Server

    Samanta, M P; Samanta, Manoj Pratim; Liang, Shoudan

    2003-01-01

    Understanding functional associations among genes discovered in sequencing projects is a key issue in post-genomic biology. However, reliable interpretation of the protein interaction data has been difficult. In this work, we show that if two proteins share significantly larger number of common interaction partners than random, they have close functional associations. Analysis of publicly available data from Saccharomyces cerevisiae reveals more than 2800 reliable functional associations, 29% of which involve at least one unannotated protein. By further analyzing these associations, we derive tentative functions for 81 unannotated proteins with high certainty.

  20. A large-scale evaluation of computational protein function prediction

    NARCIS (Netherlands)

    Radivojac, P.; Clark, W.T.; Oron, T.R.; Schnoes, A.M.; Wittkop, T.; Kourmpetis, Y.A.I.; Dijk, van A.D.J.; Friedberg, I.

    2013-01-01

    Automated annotation of protein function is challenging. As the number of sequenced genomes rapidly grows, the overwhelming majority of protein products can only be annotated computationally. If computational predictions are to be relied upon, it is crucial that the accuracy of these methods be high

  1. Large-scale biophysical evaluation of protein PEGylation effects

    DEFF Research Database (Denmark)

    Vernet, Erik; Popa, Gina; Pozdnyakova, Irina;

    2016-01-01

    of PEGylation on the thermal stability of a protein based on data generated by circular dichroism (CD), differential scanning calorimetry (DSC), or differential scanning fluorimetry (DSF). In addition, DSF was validated as a fast and inexpensive screening method for thermal unfolding studies of PEGylated...... proteins. Multivariate data analysis revealed clear trends in biophysical properties upon PEGylation for a subset of proteins, although no universal trends were found. Taken together, these findings are important in the consideration of biophysical methods and evaluation of second......-generation biopharmaceutical drug candidates....

  2. Spectral editing of two-dimensional magic-angle-spinning solid-state NMR spectra for protein resonance assignment and structure determination

    International Nuclear Information System (INIS)

    Several techniques for spectral editing of 2D 13C–13C correlation NMR of proteins are introduced. They greatly reduce the spectral overlap for five common amino acid types, thus simplifying spectral assignment and conformational analysis. The carboxyl (COO) signals of glutamate and aspartate are selected by suppressing the overlapping amide N–CO peaks through 13C–15N dipolar dephasing. The sidechain methine (CH) signals of valine, lecuine, and isoleucine are separated from the overlapping methylene (CH2) signals of long-chain amino acids using a multiple-quantum dipolar transfer technique. Both the COO and CH selection methods take advantage of improved dipolar dephasing by asymmetric rotational-echo double resonance (REDOR), where every other π-pulse is shifted from the center of a rotor period tr by about 0.15 tr. This asymmetry produces a deeper minimum in the REDOR dephasing curve and enables complete suppression of the undesired signals of immobile segments. Residual signals of mobile sidechains are positively identified by dynamics editing using recoupled 13C–1H dipolar dephasing. In all three experiments, the signals of carbons within a three-bond distance from the selected carbons are detected in the second spectral dimension via 13C spin exchange. The efficiencies of these spectral editing techniques range from 60 % for the COO and dynamic selection experiments to 25 % for the CH selection experiment, and are demonstrated on well-characterized model proteins GB1 and ubiquitin.

  3. Variability in automated assignment of NOESY spectra and three-dimensional structure determination: A test case on three small disulfide-bonded proteins

    International Nuclear Information System (INIS)

    Three independent runs of automatic assignment and structure calculations were performed on three small proteins, calcicludine from the venom of the green mamba Dendroaspis angusticeps, κ-conotoxin PVIIA from the purple cone Conus purpurascens and HsTX1, a short scorpion toxin from the venom of Heterometrus spinnifer. At the end of all the runs, the number of cross peaks which remained unassigned (0.6%, 1.4% and 2% for calcicludine, κ-conotoxin and HsTX1, respectively), as well as the number of constraints which were rejected as producing systematic violations (2.7%, 1.0%, and 1.4% for calcicludine, κ-conotoxin and HsTX1, respectively) were low. The conformation of the initial model used in the procedure (linear model or constructed by homology) has no influence on the final structures. Mainly two parameters control the procedure: the chemical shift tolerance and the cut-off distance. Independent runs of structure calculations, using the same parameters, yield structures for which the rmsd between averaged structures and the rmsd around each averaged structure were of the same order of magnitude. A different cut-off distance and a different chemical shift tolerance yield rmsd values on final average structures which did not differ more than 0.5 A compared to the rmsd obtained around the averaged structure for each calculation. These results show that the procedure is robust when applied to such a small disulfide-bonded protein

  4. Oligomeric viral proteins: small in size, large in presence

    Science.gov (United States)

    Jayaraman, Bhargavi; Smith, Amber M.; Fernandes, Jason D.; Frankel, Alan D.

    2016-01-01

    Viruses are obligate parasites that rely heavily on host cellular processes for replication. The small number of proteins typically encoded by a virus is faced with selection pressures that lead to the evolution of distinctive structural properties, allowing each protein to maintain its function under constraints such as small genome size, high mutation rate, and rapidly changing fitness conditions. One common strategy for this evolution is to utilize small building blocks to generate protein oligomers that assemble in multiple ways, thereby diversifying protein function and regulation. In this review, we discuss specific cases that illustrate how oligomerization is used to generate a single defined functional state, to modulate activity via different oligomeric states, or to generate multiple functional forms via different oligomeric states. PMID:27685368

  5. 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 from...... of noncoding DNA sequence, identified peptides localize coding sequences (exons) in a confined region of the genome, which contains the cognate gene. The approach does not require prior information about putative ORFs as predicted by computerized gene finding algorithms. The method scales to the complete human...... genome and allows identification, mapping, cloning and assistance in gene prediction of any protein for which minimal mass spectrometric information can be obtained. Several novel proteins from Arabidopsis thaliana and human have been discovered in this way....

  6. Combined automated NOE assignment and structure calculation with CYANA

    Energy Technology Data Exchange (ETDEWEB)

    Güntert, Peter, E-mail: guentert@em.uni-frankfurt.de; Buchner, Lena [Goethe University Frankfurt am Main, Center for Biomolecular Magnetic Resonance, Institute of Biophysical Chemistry (Germany)

    2015-08-15

    The automated assignment of NOESY cross peaks has become a fundamental technique for NMR protein structure analysis. A widely used algorithm for this purpose is implemented in the program CYANA. It has been used for a large number of structure determinations of proteins in solution but was so far not described in full detail. In this paper we present a complete description of the CYANA implementation of automated NOESY assignment, which differs extensively from its predecessor CANDID by the use of a consistent probabilistic treatment, and we discuss its performance in the second round of the critical assessment of structure determination by NMR.

  7. Recent advances in large-scale protein interactome mapping

    OpenAIRE

    Virja Mehta; Laura Trinkle-Mulcahy

    2016-01-01

    Protein-protein interactions (PPIs) underlie most, if not all, cellular functions. The comprehensive mapping of these complex networks of stable and transient associations thus remains a key goal, both for systems biology-based initiatives (where it can be combined with other ‘omics’ data to gain a better understanding of functional pathways and networks) and for focused biological studies. Despite the significant challenges of such an undertaking, major strides have been made over the past f...

  8. Chaperonin Structure - The Large Multi-Subunit Protein Complex

    Directory of Open Access Journals (Sweden)

    Irena Roterman

    2009-03-01

    Full Text Available The multi sub-unit protein structure representing the chaperonins group is analyzed with respect to its hydrophobicity distribution. The proteins of this group assist protein folding supported by ATP. The specific axial symmetry GroEL structure (two rings of seven units stacked back to back - 524 aa each and the GroES (single ring of seven units - 97 aa each polypeptide chains are analyzed using the hydrophobicity distribution expressed as excess/deficiency all over the molecule to search for structure-to-function relationships. The empirically observed distribution of hydrophobic residues is confronted with the theoretical one representing the idealized hydrophobic core with hydrophilic residues exposure on the surface. The observed discrepancy between these two distributions seems to be aim-oriented, determining the structure-to-function relation. The hydrophobic force field structure generated by the chaperonin capsule is presented. Its possible influence on substrate folding is suggested.

  9. Recent advances in large-scale protein interactome mapping.

    Science.gov (United States)

    Mehta, Virja; Trinkle-Mulcahy, Laura

    2016-01-01

    Protein-protein interactions (PPIs) underlie most, if not all, cellular functions. The comprehensive mapping of these complex networks of stable and transient associations thus remains a key goal, both for systems biology-based initiatives (where it can be combined with other 'omics' data to gain a better understanding of functional pathways and networks) and for focused biological studies. Despite the significant challenges of such an undertaking, major strides have been made over the past few years. They include improvements in the computation prediction of PPIs and the literature curation of low-throughput studies of specific protein complexes, but also an increase in the deposition of high-quality data from non-biased high-throughput experimental PPI mapping strategies into publicly available databases. PMID:27158474

  10. NMR experiments for resonance assignments of 13C, 15N doubly-labeled flexible polypeptides: Application to the human prion protein hPrP(23-230)

    International Nuclear Information System (INIS)

    A combination of three heteronuclear three-dimensional NMR experiments tailored for sequential resonance assignments in uniformly 15N, 13C-labeled flexible polypeptide chains is described. The 3D (H)N(CO-TOCSY)NH, 3D (H)CA(CO-TOCSY)NH and 3D (H)CBCA(CO-TOCSY)NH schemes make use of the favorable 15N chemical shift dispersion in unfolded polypeptides, exploit the slow transverse 15N relaxation rates of unfolded polypeptides in high resolution constant-time [1H, 15N]-correlation experiments, and use carbonyl carbon homonuclear isotropic mixing to transfer magnetization sequentially along the amino acid sequence. Practical applications are demonstrated with the 100-residue flexible tail of the recombinant human prion protein, making use of spectral resolution up to 0.6 Hz in the 15N dimension, simultaneous correlation with the two adjacent amino acid residues to overcome problems associated with spectral overlap, and the potential of the presently described experiments to establish nearest-neighbor correlations across proline residues in the amino acid sequence

  11. Spectral editing of two-dimensional magic-angle-spinning solid-state NMR spectra for protein resonance assignment and structure determination

    Energy Technology Data Exchange (ETDEWEB)

    Schmidt-Rohr, K.; Fritzsching, K. J.; Liao, S. Y.; Hong Mei, E-mail: mhong@iastate.edu [Iowa State University, Department of Chemistry and Ames Laboratory (United States)

    2012-12-15

    Several techniques for spectral editing of 2D {sup 13}C-{sup 13}C correlation NMR of proteins are introduced. They greatly reduce the spectral overlap for five common amino acid types, thus simplifying spectral assignment and conformational analysis. The carboxyl (COO) signals of glutamate and aspartate are selected by suppressing the overlapping amide N-CO peaks through {sup 13}C-{sup 15}N dipolar dephasing. The sidechain methine (CH) signals of valine, lecuine, and isoleucine are separated from the overlapping methylene (CH{sub 2}) signals of long-chain amino acids using a multiple-quantum dipolar transfer technique. Both the COO and CH selection methods take advantage of improved dipolar dephasing by asymmetric rotational-echo double resonance (REDOR), where every other {pi}-pulse is shifted from the center of a rotor period t{sub r} by about 0.15 t{sub r}. This asymmetry produces a deeper minimum in the REDOR dephasing curve and enables complete suppression of the undesired signals of immobile segments. Residual signals of mobile sidechains are positively identified by dynamics editing using recoupled {sup 13}C-{sup 1}H dipolar dephasing. In all three experiments, the signals of carbons within a three-bond distance from the selected carbons are detected in the second spectral dimension via {sup 13}C spin exchange. The efficiencies of these spectral editing techniques range from 60 % for the COO and dynamic selection experiments to 25 % for the CH selection experiment, and are demonstrated on well-characterized model proteins GB1 and ubiquitin.

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

    Energy Technology Data Exchange (ETDEWEB)

    Hodsdon, M.E.; Toner, J.J.; Cistola, D.P. [Washington Univ. School of Medicine, St. Louis, MO (United States)

    1994-12-01

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

  13. Assignment of methyl NMR resonances of a 52 kDa protein with residue-specific 4D correlation maps

    International Nuclear Information System (INIS)

    Methyl groups have become key probes for structural and functional studies by nuclear magnetic resonance. However, their NMR signals cluster in a small spectral region and assigning their resonances can be a tedious process. Here, we present a method that facilitates assignment of methyl resonances from assigned amide groups. Calculating the covariance between sensitive methyl and amide 3D spectra, each providing correlations to Cα and Cβ separately, produces 4D correlation maps directly correlating methyl groups to amide groups. Optimal correlation maps are obtained by extracting residue-specific regions, applying derivative to the dimensions subject to covariance, and multiplying 4D maps stemming from different 3D spectra. The latter procedure rescues weak signals that may be missed in traditional assignment procedures. Using these covariance correlation maps, nearly all assigned isoleucine, leucine, and valine amide resonances of a 52 kDa nonribosomal peptide synthetase cyclization domain were paired with their corresponding methyl groups

  14. Assignment of methyl NMR resonances of a 52 kDa protein with residue-specific 4D correlation maps

    Energy Technology Data Exchange (ETDEWEB)

    Mishra, Subrata H.; Frueh, Dominique P., E-mail: dfrueh@jhmi.edu [Johns Hopkins University School of Medicine, Department of Biophysics and Biophysical Chemistry (United States)

    2015-07-15

    Methyl groups have become key probes for structural and functional studies by nuclear magnetic resonance. However, their NMR signals cluster in a small spectral region and assigning their resonances can be a tedious process. Here, we present a method that facilitates assignment of methyl resonances from assigned amide groups. Calculating the covariance between sensitive methyl and amide 3D spectra, each providing correlations to C{sup α} and C{sup β} separately, produces 4D correlation maps directly correlating methyl groups to amide groups. Optimal correlation maps are obtained by extracting residue-specific regions, applying derivative to the dimensions subject to covariance, and multiplying 4D maps stemming from different 3D spectra. The latter procedure rescues weak signals that may be missed in traditional assignment procedures. Using these covariance correlation maps, nearly all assigned isoleucine, leucine, and valine amide resonances of a 52 kDa nonribosomal peptide synthetase cyclization domain were paired with their corresponding methyl groups.

  15. 4D Non-uniformly sampled HCBCACON and {sup 1}J(NC{sup {alpha}})-selective HCBCANCO experiments for the sequential assignment and chemical shift analysis of intrinsically disordered proteins

    Energy Technology Data Exchange (ETDEWEB)

    Novacek, Jiri [Masaryk University, Faculty of Science, NCBR, and CEITEC (Czech Republic); Haba, Noam Y.; Chill, Jordan H. [Bar Ilan University, Department of Chemistry (Israel); Zidek, Lukas, E-mail: lzidek@chemi.muni.cz; Sklenar, Vladimir [Masaryk University, Faculty of Science, NCBR, and CEITEC (Czech Republic)

    2012-06-15

    A pair of 4D NMR experiments for the backbone assignment of disordered proteins is presented. The experiments exploit {sup 13}C direct detection and non-uniform sampling of the indirectly detected dimensions, and provide correlations of the aliphatic proton (H{sup {alpha}}, and H{sup {beta}}) and carbon (C{sup {alpha}}, C{sup {beta}}) resonance frequencies to the protein backbone. Thus, all the chemical shifts regularly used to map the transient secondary structure motifs in the intrinsically disordered proteins (H{sup {alpha}}, C{sup {alpha}}, C{sup {beta}}, C Prime , and N) can be extracted from each spectrum. Compared to the commonly used assignment strategy based on matching the C{sup {alpha}} and C{sup {beta}} chemical shifts, inclusion of the H{sup {alpha}} and H{sup {beta}} provides up to three extra resonance frequencies that decrease the chance of ambiguous assignment. The experiments were successfully applied to the original assignment of a 12.8 kDa intrinsically disordered protein having a high content of proline residues (26 %) in the sequence.

  16. CO{sub H}(N)CACB experiments for assigning backbone resonances in {sup 13}C/{sup 15}N-labeled proteins

    Energy Technology Data Exchange (ETDEWEB)

    Astrof, Nathan; Bracken, Clay; Cavanagh, John; Palmer, Arthur G

    1998-05-15

    A triple resonance NMR experiment, denoted CO{sub H}(N)CACB, correlates{sup 1}H{sup N} and {sup 13}CO spins with the{sup 13}C{sup {alpha}} and{sup 13}C{sup {beta}} spins of adjacent amino acids. The pulse sequence is an 'out-and-back' design that starts with{sup 1}H{sup N} magnetization and transfers coherence via the {sup 15}N spin simultaneously to the {sup 13}CO and{sup 13}C{sup {alpha}} spins, followed by transfer to the{sup 13}C{sup {beta}} spin. Two versions of the sequence are presented: one in which the {sup 13}CO spins are frequency labeled during an incremented t{sub 1} evolution period prior to transfer of magnetization from the {sup 13}C{sup {alpha}} to the{sup 13}C{sup {beta}} resonances, and one in which the{sup 13}CO spins are frequency labeled in a constant-time manner during the coherence transfer to and from the{sup 13}C{sup {beta}} resonances. Because {sup 13}COand {sup 15}N chemical shifts are largely uncorrelated, the technique will be especially useful when degeneracy in the{sup 1}H{sup N}-{sup 15}N chemical shifts hinders resonance assignment. The CO{sub H}(N)CACB experiment is demonstrated using uniformly {sup 13}C/{sup 15}N-labeled ubiquitin.

  17. Guaranteed Discrete Energy Optimization on Large Protein Design Problems.

    Science.gov (United States)

    Simoncini, David; Allouche, David; de Givry, Simon; Delmas, Céline; Barbe, Sophie; Schiex, Thomas

    2015-12-01

    In Computational Protein Design (CPD), assuming a rigid backbone and amino-acid rotamer library, the problem of finding a sequence with an optimal conformation is NP-hard. In this paper, using Dunbrack's rotamer library and Talaris2014 decomposable energy function, we use an exact deterministic method combining branch and bound, arc consistency, and tree-decomposition to provenly identify the global minimum energy sequence-conformation on full-redesign problems, defining search spaces of size up to 10(234). This is achieved on a single core of a standard computing server, requiring a maximum of 66GB RAM. A variant of the algorithm is able to exhaustively enumerate all sequence-conformations within an energy threshold of the optimum. These proven optimal solutions are then used to evaluate the frequencies and amplitudes, in energy and sequence, at which an existing CPD-dedicated simulated annealing implementation may miss the optimum on these full redesign problems. The probability of finding an optimum drops close to 0 very quickly. In the worst case, despite 1,000 repeats, the annealing algorithm remained more than 1 Rosetta unit away from the optimum, leading to design sequences that could differ from the optimal sequence by more than 30% of their amino acids. PMID:26610100

  18. Easy and unambiguous sequential assignments of intrinsically disordered proteins by correlating the backbone {sup 15}N or {sup 13}C′ chemical shifts of multiple contiguous residues in highly resolved 3D spectra

    Energy Technology Data Exchange (ETDEWEB)

    Yoshimura, Yuichi; Kulminskaya, Natalia V.; Mulder, Frans A. A., E-mail: fmulder@chem.au.dk [Aarhus University, Department of Chemistry and Interdisciplinary Nanoscience Center (iNANO) (Denmark)

    2015-02-15

    Sequential resonance assignment strategies are typically based on matching one or two chemical shifts of adjacent residues. However, resonance overlap often leads to ambiguity in resonance assignments in particular for intrinsically disordered proteins. We investigated the potential of establishing connectivity through the three-bond couplings between sequentially adjoining backbone carbonyl carbon nuclei, combined with semi-constant time chemical shift evolution, for resonance assignments of small folded and larger unfolded proteins. Extended sequential connectivity strongly lifts chemical shift degeneracy of the backbone nuclei in disordered proteins. We show here that 3D (H)N(COCO)NH and (HN)CO(CO)NH experiments with relaxation-optimized multiple pulse mixing correlate up to seven adjacent backbone amide nitrogen or carbonyl carbon nuclei, respectively, and connections across proline residues are also obtained straightforwardly. Multiple, recurrent long-range correlations with ultra-high resolution allow backbone {sup 1}H{sup N}, {sup 15}N{sup H}, and {sup 13}C′ resonance assignments to be completed from a single pair of 3D experiments.

  19. Large-scale protein-protein interaction analysis in Arabidopsis mesophyll protoplasts by split firefly luciferase complementation.

    Directory of Open Access Journals (Sweden)

    Jian-Feng Li

    Full Text Available Protein-protein interactions (PPIs constitute the regulatory network that coordinates diverse cellular functions. There are growing needs in plant research for creating protein interaction maps behind complex cellular processes and at a systems biology level. However, only a few approaches have been successfully used for large-scale surveys of PPIs in plants, each having advantages and disadvantages. Here we present split firefly luciferase complementation (SFLC as a highly sensitive and noninvasive technique for in planta PPI investigation. In this assay, the separate halves of a firefly luciferase can come into close proximity and transiently restore its catalytic activity only when their fusion partners, namely the two proteins of interest, interact with each other. This assay was conferred with quantitativeness and high throughput potential when the Arabidopsis mesophyll protoplast system and a microplate luminometer were employed for protein expression and luciferase measurement, respectively. Using the SFLC assay, we could monitor the dynamics of rapamycin-induced and ascomycin-disrupted interaction between Arabidopsis FRB and human FKBP proteins in a near real-time manner. As a proof of concept for large-scale PPI survey, we further applied the SFLC assay to testing 132 binary PPIs among 8 auxin response factors (ARFs and 12 Aux/IAA proteins from Arabidopsis. Our results demonstrated that the SFLC assay is ideal for in vivo quantitative PPI analysis in plant cells and is particularly powerful for large-scale binary PPI screens.

  20. Automated assignment of NMR chemical shifts based on a known structure and 4D spectra.

    Science.gov (United States)

    Trautwein, Matthias; Fredriksson, Kai; Möller, Heiko M; Exner, Thomas E

    2016-08-01

    Apart from their central role during 3D structure determination of proteins the backbone chemical shift assignment is the basis for a number of applications, like chemical shift perturbation mapping and studies on the dynamics of proteins. This assignment is not a trivial task even if a 3D protein structure is known and needs almost as much effort as the assignment for structure prediction if performed manually. We present here a new algorithm based solely on 4D [(1)H,(15)N]-HSQC-NOESY-[(1)H,(15)N]-HSQC spectra which is able to assign a large percentage of chemical shifts (73-82 %) unambiguously, demonstrated with proteins up to a size of 250 residues. For the remaining residues, a small number of possible assignments is filtered out. This is done by comparing distances in the 3D structure to restraints obtained from the peak volumes in the 4D spectrum. Using dead-end elimination, assignments are removed in which at least one of the restraints is violated. Including additional information from chemical shift predictions, a complete unambiguous assignment was obtained for Ubiquitin and 95 % of the residues were correctly assigned in the 251 residue-long N-terminal domain of enzyme I. The program including source code is available at https://github.com/thomasexner/4Dassign . PMID:27484442

  1. Nano-Mole Scale Side-Chain Signal Assignment by 1H-Detected Protein Solid-State NMR by Ultra-Fast Magic-Angle Spinning and Stereo-Array Isotope Labeling

    KAUST Repository

    Wang, Songlin

    2015-04-09

    We present a general approach in 1H-detected 13C solid-state NMR (SSNMR) for side-chain signal assignments of 10-50 nmol quantities of proteins using a combination of a high magnetic field, ultra-fast magic-angle spinning (MAS) at ~80 kHz, and stereo-array-isotope-labeled (SAIL) proteins [Kainosho M. et al., Nature 440, 52–57, 2006]. First, we demonstrate that 1H indirect detection improves the sensitivity and resolution of 13C SSNMR of SAIL proteins for side-chain assignments in the ultra-fast MAS condition. 1H-detected SSNMR was performed for micro-crystalline ubiquitin (~55 nmol or ~0.5mg) that was SAIL-labeled at seven isoleucine (Ile) residues. Sensitivity was dramatically improved by 1H-detected 2D 1H/13C SSNMR by factors of 5.4-9.7 and 2.1-5.0, respectively, over 13C-detected 2D 1H/13C SSNMR and 1D 13C CPMAS, demonstrating that 2D 1H-detected SSNMR offers not only additional resolution but also sensitivity advantage over 1D 13C detection for the first time. High 1H resolution for the SAIL-labeled side-chain residues offered reasonable resolution even in the 2D data. A 1H-detected 3D 13C/13C/1H experiment on SAIL-ubiquitin provided nearly complete 1H and 13C assignments for seven Ile residues only within ~2.5 h. The results demonstrate the feasibility of side-chain signal assignment in this approach for as little as 10 nmol of a protein sample within ~3 days. The approach is likely applicable to a variety of proteins of biological interest without any requirements of highly efficient protein expression systems.

  2. Large-scale analysis of phosphorylation site occupancy in eukaryotic proteins

    DEFF Research Database (Denmark)

    Rao, R Shyama Prasad; Møller, Ian Max

    2012-01-01

    in proteins is currently lacking. We have therefore analyzed the occurrence and occupancy of phosphorylated sites (~ 100,281) in a large set of eukaryotic proteins (~ 22,995). Phosphorylation probability was found to be much higher in both the  termini of protein sequences and this is much pronounced...... maximum randomness. An analysis of phosphorylation motifs indicated that just 40 motifs and a much lower number of associated kinases might account for nearly 50% of the known phosphorylations in eukaryotic proteins. Our results provide a broad picture of the phosphorylation sites in eukaryotic proteins....

  3. Core-Cone Structured Monodispersed Mesoporous Silica Nanoparticles with Ultra-large Cavity for Protein Delivery.

    Science.gov (United States)

    Xu, Chun; Yu, Meihua; Noonan, Owen; Zhang, Jun; Song, Hao; Zhang, Hongwei; Lei, Chang; Niu, Yuting; Huang, Xiaodan; Yang, Yannan; Yu, Chengzhong

    2015-11-25

    A new type of monodispersed mesoporous silica nanoparticles with a core-cone structure (MSN-CC) has been synthesized. The large cone-shaped pores are formed by silica lamellae closely packed encircling a spherical core, showing a structure similar to the flower dahlia. MSN-CC has a large pore size of 45 nm and a high pore volume of 2.59 cm(3) g(-1). MSN-CC demonstrates a high loading capacity of large proteins and successfully delivers active β-galactosidase into cells, showing their potential as efficient nanocarriers for the cellular delivery of proteins with large molecular weights. PMID:26426420

  4. A Large Scale Separation of Taxanes from the Bark Extract of Taxus yunnanesis and 1H- and 13C-NMR Assignments for 7-epi-10-Deacetyltaxol

    Institute of Scientific and Technical Information of China (English)

    薛军; 卜海山; 曹春阳; 吴厚铭; 陈建民

    2001-01-01

    A large-scale separation of paclitaxel from semi-purified bark extract of Taxus yunnanesis was investigated. The chromatographic behavior of paclitaxel and two close eluting analogues, cephalomannine and 7-epi-10-deacetyltaxol were sytematically studied on a C18 bonded phase column with different mobile phase in reverse phase mode. According to the notably different selectivity of the methanol and acetonitrile with water in the mobile phase and the most important requirement of capacity in preparative chromatography, the optimum suitably mobile phase used in a large-scale isolation of paclitaxel and 7-epi-10-deacetyltaxol on a preparative C18 column was given.Cephalomannine was eliminated by ozonolysis and after then separated throughout a normal phase silica column.The whole large-scale process for high purity paclitaxel from the bark extract of Taxus yunnanesis consisted of a preliminary purification with Biotage FLASH 150i systen based on a prepacked normal phase silica cartridge followed by using a C18 Nova-pakTM column in Waters PrepLCTM 4000 prepparative HPLC system. The structure of 7-epi-10-deacetyltaxol was elucidated by 2O NMR technologies of TOCSY, DQF-COSY,HMQC and HMBC, etc.

  5. Ribosome Profiling Provides Evidence that Large Noncoding RNAs Do Not Encode Proteins

    OpenAIRE

    Guttman, Mitchell; Russell, Pamela; Ingolia, Nicholas T.; Weissman, Jonathan S.; Lander, Eric S.; Lander, Eric S.

    2013-01-01

    Large noncoding RNAs are emerging as an important component in cellular regulation. Considerable evidence indicates that these transcripts act directly as functional RNAs rather than through an encoded protein product. However, a recent study of ribosome occupancy reported that many large intergenic ncRNAs (lincRNAs) are bound by ribosomes, raising the possibility that they are translated into proteins. Here, we show that classical noncoding RNAs and 5′ UTRs show the same ribosome occupancy a...

  6. Large-scale identification of membrane proteins with properties favorable for crystallization.

    Science.gov (United States)

    Kim, Jared; Kagawa, Allison; Kurasaki, Kellie; Ataie, Niloufar; Cho, Il Kyu; Li, Qing X; Ng, Ho Leung

    2015-11-01

    Membrane protein crystallography is notoriously difficult due to challenges in protein expression and issues of degradation and structural stability. We have developed a novel method for large-scale screening of native sources for integral membrane proteins that have intrinsic biochemical properties favorable for crystallization. Highly expressed membrane proteins that are thermally stable and nonaggregating in detergent solutions were identified by mass spectrometry from Escherichia coli, Saccharomyces cerevisiae, and Sus scrofa cerebrum. Many of the membrane proteins identified had been crystallized previously, supporting the promise of the approach. Most identified proteins have known functions and include high-value targets such as transporters and ATPases. To validate the method, we recombinantly expressed and purified the yeast protein, Yop1, which is responsible for endoplasmic reticulum curvature. We demonstrate that Yop1 can be purified with the detergent dodecylmaltoside without aggregating.

  7. “C.R.E.A.T.E.”-ing Unique Primary-Source Research Paper Assignments for a Pleasure and Pain Course Teaching Neuroscientific Principles in a Large General Education Undergraduate Course

    Science.gov (United States)

    Bodnar, Richard J.; Rotella, Francis M.; Loiacono, Ilyssa; Coke, Tricia; Olsson, Kerstin; Barrientos, Alicia; Blachorsky, Lauren; Warshaw, Deena; Buras, Agata; Sanchez, Ciara M.; Azad, Raihana; Stellar, James R.

    2016-01-01

    A large (250 registrants) General Education lecture course, Pleasure and Pain, presented basic neuroscience principles as they related to animal and human models of pleasure and pain by weaving basic findings related to food and drug addiction and analgesic states with human studies examining empathy, social neuroscience and neuroeconomics. In its first four years, the course grade was based on weighted scores from two multiple-choice exams and a five-page review of three unique peer-reviewed research articles. Although well-registered and well-received, 18% of the students received Incomplete grades, primarily due to failing to submit the paper that went largely unresolved and eventually resulted in a failing grade. To rectify this issue, a modified version of the C.R.E.A.T.E. (Consider, Read, Elucidate hypotheses, Analyze and interpret data, Think of the next Experiment) method replaced the paper with eight structured assignments focusing on an initial general-topic article, the introduction-methods, and results-discussion of each of three related peer-review neuroscience-related articles, and a final summary. Compliance in completing these assignments was very high, resulting in only 11 INC grades out of 228 students. Thus, use of the C.R.E.A.T.E. method reduced the percentage of problematic INC grades from 18% to 4.8%, a 73% decline, without changing the overall grade distribution. Other analyses suggested the students achieved a deeper understanding of the scientific process using the C.R.E.A.T.E. method relative to the original term paper assignment. PMID:27385918

  8. "C.R.E.A.T.E."-ing Unique Primary-Source Research Paper Assignments for a Pleasure and Pain Course Teaching Neuroscientific Principles in a Large General Education Undergraduate Course.

    Science.gov (United States)

    Bodnar, Richard J; Rotella, Francis M; Loiacono, Ilyssa; Coke, Tricia; Olsson, Kerstin; Barrientos, Alicia; Blachorsky, Lauren; Warshaw, Deena; Buras, Agata; Sanchez, Ciara M; Azad, Raihana; Stellar, James R

    2016-01-01

    A large (250 registrants) General Education lecture course, Pleasure and Pain, presented basic neuroscience principles as they related to animal and human models of pleasure and pain by weaving basic findings related to food and drug addiction and analgesic states with human studies examining empathy, social neuroscience and neuroeconomics. In its first four years, the course grade was based on weighted scores from two multiple-choice exams and a five-page review of three unique peer-reviewed research articles. Although well-registered and well-received, 18% of the students received Incomplete grades, primarily due to failing to submit the paper that went largely unresolved and eventually resulted in a failing grade. To rectify this issue, a modified version of the C.R.E.A.T.E. (Consider, Read, Elucidate hypotheses, Analyze and interpret data, Think of the next Experiment) method replaced the paper with eight structured assignments focusing on an initial general-topic article, the introduction-methods, and results-discussion of each of three related peer-review neuroscience-related articles, and a final summary. Compliance in completing these assignments was very high, resulting in only 11 INC grades out of 228 students. Thus, use of the C.R.E.A.T.E. method reduced the percentage of problematic INC grades from 18% to 4.8%, a 73% decline, without changing the overall grade distribution. Other analyses suggested the students achieved a deeper understanding of the scientific process using the C.R.E.A.T.E. method relative to the original term paper assignment.

  9. "C.R.E.A.T.E."-ing Unique Primary-Source Research Paper Assignments for a Pleasure and Pain Course Teaching Neuroscientific Principles in a Large General Education Undergraduate Course.

    Science.gov (United States)

    Bodnar, Richard J; Rotella, Francis M; Loiacono, Ilyssa; Coke, Tricia; Olsson, Kerstin; Barrientos, Alicia; Blachorsky, Lauren; Warshaw, Deena; Buras, Agata; Sanchez, Ciara M; Azad, Raihana; Stellar, James R

    2016-01-01

    A large (250 registrants) General Education lecture course, Pleasure and Pain, presented basic neuroscience principles as they related to animal and human models of pleasure and pain by weaving basic findings related to food and drug addiction and analgesic states with human studies examining empathy, social neuroscience and neuroeconomics. In its first four years, the course grade was based on weighted scores from two multiple-choice exams and a five-page review of three unique peer-reviewed research articles. Although well-registered and well-received, 18% of the students received Incomplete grades, primarily due to failing to submit the paper that went largely unresolved and eventually resulted in a failing grade. To rectify this issue, a modified version of the C.R.E.A.T.E. (Consider, Read, Elucidate hypotheses, Analyze and interpret data, Think of the next Experiment) method replaced the paper with eight structured assignments focusing on an initial general-topic article, the introduction-methods, and results-discussion of each of three related peer-review neuroscience-related articles, and a final summary. Compliance in completing these assignments was very high, resulting in only 11 INC grades out of 228 students. Thus, use of the C.R.E.A.T.E. method reduced the percentage of problematic INC grades from 18% to 4.8%, a 73% decline, without changing the overall grade distribution. Other analyses suggested the students achieved a deeper understanding of the scientific process using the C.R.E.A.T.E. method relative to the original term paper assignment. PMID:27385918

  10. Scaffolding students’ assignments

    DEFF Research Database (Denmark)

    Slot, Marie Falkesgaard

    2013-01-01

    , on the shoulders of linguistics research, results of “assignment-didactics” as a useful but also very tentative term in the context of the assessment of design principles for the design of assignments. Based on empirical studies, I discuss cognition, multi-modality and collaboration as crucial types of scaffolding...

  11. Combination of Bcl-2 and MYC protein expression improves high-risk stratification in diffuse large B-cell lymphoma

    Directory of Open Access Journals (Sweden)

    Wang J

    2015-09-01

    Full Text Available Jing Wang,* Min Zhou,* Jing-Yan Xu,* Bing Chen, Jian OuyangDepartment of Hematology, The Affiliated Drum Tower Hospital of Nanjing University Medical School, Nanjing, Jiangsu, People’s Republic of China*These authors contributed equally to this work and should be considered as cofirst authorsPurpose: To evaluate whether the addition of two biological markers (MYC and BCL-2 protein overexpression improves the stratification of high-risk patients with diffuse large B-cell lymphoma (DLBCL.Method: Seven risk factors were identified at diagnosis, and a maximum of 7 points were assigned to each patient. The patients were classified according to four risk groups: low (0–1, low-intermediate (2–3, high-intermediate (4, and high (5–7. Only high-risk patients with DLBCL were included in this analysis. We retrospectively examined 20 cases from 2008 to 2013 at the Nanjing Drum Tower Hospital.Results: The median expression of MYC protein was 60%, and 17 of 20 (65% evaluable cases overexpressed MYC. The median expression of BCL-2 protein was also 60%. Eighteen of 20 (90% evaluable cases showed BCL-2 overexpression. Additionally, 12 out of 20 cases (60% demonstrated coexpression of MYC and BCL-2 proteins. The percentages of overall survival and progression-free survival at the median follow-up time (36 months were 33.3%±16.1% and 16.9%±13.5%, respectively. By comparison, nine, four, and 20 patients were classified as high risk based on the International Prognostic Index (IPI, National Comprehensive Cancer Network(NCCN-IPI, and revised IPI criteria, respectively. According to the IPI and NCCN-IPI stratification, the risk groups demonstrated closely overlapping survival curves. In addition, four out of 20 cases were identified as low-intermediate risk according to the NCCN-IPI criteria.Conclusion: The addition of MYC and BCL-2 protein expression to the IPI could identify a subset of DLBCL patients with high-risk clinicopathological characteristics and

  12. STAQ: Static Traffic Assignment with Queing

    NARCIS (Netherlands)

    Brederode, L.; Bliemer, M.C.J.; Wismans, L.

    2010-01-01

    Because of computation time issues on large networks, most strategic regional and urban transport models today use static instead of dynamic traffic assignment procedures. Mathematical models of traffic assignment are usually based upon Wardrop’s principle. To solve this static traffic equilibrium p

  13. rRNA maturation in yeast cells depleted of large ribosomal subunit proteins.

    Directory of Open Access Journals (Sweden)

    Gisela Pöll

    Full Text Available The structural constituents of the large eukaryotic ribosomal subunit are 3 ribosomal RNAs, namely the 25S, 5.8S and 5S rRNA and about 46 ribosomal proteins (r-proteins. They assemble and mature in a highly dynamic process that involves more than 150 proteins and 70 small RNAs. Ribosome biogenesis starts in the nucleolus, continues in the nucleoplasm and is completed after nucleo-cytoplasmic translocation of the subunits in the cytoplasm. In this work we created 26 yeast strains, each of which conditionally expresses one of the large ribosomal subunit (LSU proteins. In vivo depletion of the analysed LSU r-proteins was lethal and led to destabilisation and degradation of the LSU and/or its precursors. Detailed steady state and metabolic pulse labelling analyses of rRNA precursors in these mutant strains showed that LSU r-proteins can be grouped according to their requirement for efficient progression of different steps of large ribosomal subunit maturation. Comparative analyses of the observed phenotypes and the nature of r-protein-rRNA interactions as predicted by current atomic LSU structure models led us to discuss working hypotheses on i how individual r-proteins control the productive processing of the major 5' end of 5.8S rRNA precursors by exonucleases Rat1p and Xrn1p, and ii the nature of structural characteristics of nascent LSUs that are required for cytoplasmic accumulation of nascent subunits but are nonessential for most of the nuclear LSU pre-rRNA processing events.

  14. Detection and assignment of proteins encoded by rice black streaked dwarf fijivirus S7, S8, S9 and S10.

    Science.gov (United States)

    Isogai, M; Uyeda, I; Lee, B C

    1998-06-01

    The proteins encoded by rice black streaked dwarf fijivirus (RBSDV) genomic segments 7-10 (S7-S10) were characterized. Open reading frames (ORFs) from these segments were expressed as fusion proteins in Escherichia coli. Antibodies raised against the expressed products were used as probes to determine whether the viral ORFs encode structural proteins. In Western blots, antibodies to the expressed S8 and S10 products reacted with a core capsid (65 kDa) and a major outer capsid (56 kDa) protein, respectively, while none of the antibodies to S7 and S9 products reacted with structural proteins. Antisera to RBSDV S7 ORF1 and S9 ORF1 each detected a single protein of the predicted size in total protein extracts from infected rice plants and viruliferous Laodelphax striatellus. Immunoelectron microscopy revealed that antibodies to RBSDV S7 ORF1 and RBSDV S9 ORF1 reacted with tubular structures and viroplasm, respectively, in sections of both infected maize plants and viruliferous L. striatellus. Antisera to ORF2 of S7 and S9 failed to detect any proteins in the infected tissue using either Western blotting or immuno-electron microscopic techniques.

  15. Unfolding the Role of Large Heat Shock Proteins: New Insights and Therapeutic Implications

    Directory of Open Access Journals (Sweden)

    Daming eZuo

    2016-03-01

    Full Text Available Heat shock proteins (HSPs of eukaryotes are evolutionarily conserved molecules present in all the major intracellular organelles. They mainly function as molecular chaperones and participate in maintenance of protein homeostasis in physiological state and under stressful conditions. Despite their relative abundance, the large HSPs, i.e., Hsp110 and glucose-regulated protein 170 (Grp170, have received less attention compared to other conventional HSPs. These proteins are distantly related to the Hsp70 and belong to the Hsp70 superfamily. Increased sizes of Hsp110 and Grp170, largely due to the presence of a loop structure, appears to result in their exceptional capability in binding to polypeptide substrates or non-protein ligands, such as pathogen-associated molecules. These interactions that occur in the extracellular environment during tissue injury or microbial infection may lead to amplification of an immune response engaging both innate and adaptive immune components. Here we review the current advances in understanding these large HSPs as molecular chaperones in proteostasis control and immune modulation as well as their therapeutic implications in treatment of cancer and neurodegeneration. Given their unique immunoregulatory activities, we also discuss the emerging evidence of their potential involvement in inflammatory and immune-related diseases.

  16. Historical WBAN ID Assignments

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — 4"x6" index cards represent the first written assignments of Weather Bureau Army Navy (WBAN) station identifier numbers by the National Climatic Data Center....

  17. MODERN NMR TECHNIQUES FOR THE STUDY OF LARGE PROTEINS IN SOLUTION

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    @@ A number of important methodological developments in high resolution NMR spectroscopy have led to significant increases in the size limitations that previously impeded solution structural studies of macromolecules. Specifically, isotope labeling and TROSY triple resonance spectroscopy has resulted in substantial sensitivity and resolution gain for applications to large molecular weight proteins.

  18. The DEAD box protein Mrh4 functions in the assembly of the mitochondrial large ribosomal subunit.

    Science.gov (United States)

    De Silva, Dasmanthie; Fontanesi, Flavia; Barrientos, Antoni

    2013-11-01

    Proteins in a cell are universally synthesized by ribosomes. Mitochondria contain their own ribosomes, which specialize in the synthesis of a handful of proteins required for oxidative phosphorylation. The pathway of mitoribosomal biogenesis and factors involved are poorly characterized. An example is the DEAD box proteins, widely known to participate in the biogenesis of bacterial and cytoplasmic eukaryotic ribosomes as either RNA helicases or RNA chaperones, whose mitochondrial counterparts remain completely unknown. Here, we have identified the Saccharomyces cerevisiae mitochondrial DEAD box protein Mrh4 as essential for large mitoribosome subunit biogenesis. Mrh4 interacts with the 21S rRNA, mitoribosome subassemblies, and fully assembled mitoribosomes. In the absence of Mrh4, the 21S rRNA is matured and forms part of a large on-pathway assembly intermediate missing proteins Mrpl16 and Mrpl39. We conclude that Mrh4 plays an essential role during the late stages of mitoribosome assembly by promoting remodeling of the 21S rRNA-protein interactions.

  19. Characterisation of silent and active genes for a variable large protein of Borrelia recurrentis

    Directory of Open Access Journals (Sweden)

    Scragg Ian G

    2002-10-01

    Full Text Available Abstract Background We report the characterisation of the variable large protein (vlp gene expressed by clinical isolate A1 of Borrelia recurrentis; the agent of the life-threatening disease louse-borne relapsing fever. Methods The major vlp protein of this isolate was characterised and a DNA probe created. Use of this together with standard molecular methods was used to determine the location of the vlp1B. recurrentis A1 gene in both this and other isolates. Results This isolate was found to carry silent and expressed copies of the vlp1B. recurrentis A1 gene on plasmids of 54 kbp and 24 kbp respectively, whereas a different isolate, A17, had only the silent vlp1B. recurrentis A17 on a 54 kbp plasmid. Silent and expressed vlp1 have identical mature protein coding regions but have different 5' regions, both containing different potential lipoprotein leader sequences. Only one form of vlp1 is transcribed in the A1 isolate of B. recurrentis, yet both 5' upstream sequences of this vlp1 gene possess features of bacterial promoters. Conclusion Taken together these results suggest that antigenic variation in B. recurrentis may result from recombination of variable large and small protein genes at the junction between lipoprotein leader sequence and mature protein coding region. However, this hypothetical model needs to be validated by further identification of expressed and silent variant protein genes in other B. recurrentis isolates.

  20. Detecting remote evolutionary relationships among proteins by large-scale semantic embedding.

    Directory of Open Access Journals (Sweden)

    Iain Melvin

    Full Text Available Virtually every molecular biologist has searched a protein or DNA sequence database to find sequences that are evolutionarily related to a given query. Pairwise sequence comparison methods--i.e., measures of similarity between query and target sequences--provide the engine for sequence database search and have been the subject of 30 years of computational research. For the difficult problem of detecting remote evolutionary relationships between protein sequences, the most successful pairwise comparison methods involve building local models (e.g., profile hidden Markov models of protein sequences. However, recent work in massive data domains like web search and natural language processing demonstrate the advantage of exploiting the global structure of the data space. Motivated by this work, we present a large-scale algorithm called ProtEmbed, which learns an embedding of protein sequences into a low-dimensional "semantic space." Evolutionarily related proteins are embedded in close proximity, and additional pieces of evidence, such as 3D structural similarity or class labels, can be incorporated into the learning process. We find that ProtEmbed achieves superior accuracy to widely used pairwise sequence methods like PSI-BLAST and HHSearch for remote homology detection; it also outperforms our previous RankProp algorithm, which incorporates global structure in the form of a protein similarity network. Finally, the ProtEmbed embedding space can be visualized, both at the global level and local to a given query, yielding intuition about the structure of protein sequence space.

  1. 42 CFR 433.146 - Rights assigned; assignment method.

    Science.gov (United States)

    2010-10-01

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

  2. PPS, a large multidomain protein, functions with sex-lethal to regulate alternative splicing in Drosophila.

    Directory of Open Access Journals (Sweden)

    Matthew L Johnson

    2010-03-01

    Full Text Available Alternative splicing controls the expression of many genes, including the Drosophila sex determination gene Sex-lethal (Sxl. Sxl expression is controlled via a negative regulatory mechanism where inclusion of the translation-terminating male exon is blocked in females. Previous studies have shown that the mechanism leading to exon skipping is autoregulatory and requires the SXL protein to antagonize exon inclusion by interacting with core spliceosomal proteins, including the U1 snRNP protein Sans-fille (SNF. In studies begun by screening for proteins that interact with SNF, we identified PPS, a previously uncharacterized protein, as a novel component of the machinery required for Sxl male exon skipping. PPS encodes a large protein with four signature motifs, PHD, BRK, TFS2M, and SPOC, typically found in proteins involved in transcription. We demonstrate that PPS has a direct role in Sxl male exon skipping by showing first that loss of function mutations have phenotypes indicative of Sxl misregulation and second that the PPS protein forms a complex with SXL and the unspliced Sxl RNA. In addition, we mapped the recruitment of PPS, SXL, and SNF along the Sxl gene using chromatin immunoprecipitation (ChIP, which revealed that, like many other splicing factors, these proteins bind their RNA targets while in close proximity to the DNA. Interestingly, while SNF and SXL are specifically recruited to their predicted binding sites, PPS has a distinct pattern of accumulation along the Sxl gene, associating with a region that includes, but is not limited to, the SxlPm promoter. Together, these data indicate that PPS is different from other splicing factors involved in male-exon skipping and suggest, for the first time, a functional link between transcription and SXL-mediated alternative splicing. Loss of zygotic PPS function, however, is lethal to both sexes, indicating that its role may be of broad significance.

  3. Accounting for Large Amplitude Protein Deformation during in Silico Macromolecular Docking

    Directory of Open Access Journals (Sweden)

    Chantal Prévost

    2011-02-01

    Full Text Available Rapid progress of theoretical methods and computer calculation resources has turned in silico methods into a conceivable tool to predict the 3D structure of macromolecular assemblages, starting from the structure of their separate elements. Still, some classes of complexes represent a real challenge for macromolecular docking methods. In these complexes, protein parts like loops or domains undergo large amplitude deformations upon association, thus remodeling the surface accessible to the partner protein or DNA.We discuss the problems linked with managing such rearrangements in docking methods and we review strategies that are presently being explored, as well as their limitations and success.

  4. Accounting for large amplitude protein deformation during in silico macromolecular docking.

    Science.gov (United States)

    Bastard, Karine; Saladin, Adrien; Prévost, Chantal

    2011-02-22

    Rapid progress of theoretical methods and computer calculation resources has turned in silico methods into a conceivable tool to predict the 3D structure of macromolecular assemblages, starting from the structure of their separate elements. Still, some classes of complexes represent a real challenge for macromolecular docking methods. In these complexes, protein parts like loops or domains undergo large amplitude deformations upon association, thus remodeling the surface accessible to the partner protein or DNA. We discuss the problems linked with managing such rearrangements in docking methods and we review strategies that are presently being explored, as well as their limitations and success.

  5. Human heterochromatin proteins form large domains containing KRAB-ZNF genes

    OpenAIRE

    Vogel, Maartje J.; Guelen, Lars; de Wit, Elzo; Hupkes, Daniel Peric; Lodén, Martin; Talhout, Wendy; Feenstra, Marike; Abbas, Ben; Classen, Anne-Kathrin; van Steensel, Bas

    2006-01-01

    Heterochromatin is important for gene regulation and chromosome structure, but the genes that are occupied by heterochromatin proteins in the mammalian genome are largely unknown. We have adapted the DamID method to systematically identify target genes of the heterochromatin proteins HP1 and SUV39H1 in human and mouse cells. Unexpectedly, we found that CBX1 (formerly HP1β) and SUV39H1 bind to genes encoding KRAB domain containing zinc finger (KRAB-ZNF) transcriptional repressors. These genes ...

  6. Establishment and characterization of human engineered cells stably expressing large extracellular matrix proteins.

    Science.gov (United States)

    Kwon, Daekee; Kang, Gwang-Sik; Han, Dong Keun; Park, Kwideok; Kim, Jae-Hwan; Lee, Soo-Hong

    2014-01-01

    Commercially available extracellular matrix (ECM) hydrogel-coated culture plates have been used to study the relationship between the ECM microenvironment and stem cell behavior. However, it is unclear whether ECM-coated dishes mimic the natural ECM microenvironment because the architecture of the ECM is constructed of randomly distributed fibers. The purpose of this study was the production and confirmation of human engineered cell lines stably expressing large ECM proteins such as collagen I/II and fibronectin. First, large (over 10 kb) ECM vectors encoding human collagen I/II and fibronectin were constructed and the circular vectors were linearized. Second, the linear ECM vectors were introduced into immortalized human embryonic kidney cells using various transfection methods. The polyethylenimine and liposome methods showed higher efficiencies than electroporation for transfection of these large vectors. Third, human ECM engineered cells were established by stable integration of the vector into the genomic DNA and resulted in stable overexpression of mRNA and proteins. In summary, human engineered cell lines stably expressing large ECM proteins such as human collagen I/II and fibronectin were successfully prepared, and secretion of the ECM components into the surrounding environment was confirmed by immunocytochemistry. Thus, human ECM engineered cells naturally secreting ECM components could be valuable for studying the relationship between the native ECM microenvironment and stem cell behavior.

  7. Effects of dietary restriction followed by high dietary energy or protein on compensatory growth of Ashanti Black × Large White crossbred weaner pigs.

    Science.gov (United States)

    Addah, Weseh; Dzewu, Reuben Rudolph Kafui; Alenyorege, Benjamin

    2016-01-01

    The study determined the effect of re-alimenting dietary protein or energy on compensatory growth. Eighteen Ashanti Black × Large White crossbred weaner pigs (7.5 ± 0.30 kg) were randomly assigned to one of three dietary treatments in a completely randomized design resulting in three replicate pens per treatment (n = 3) and two pigs per pen. In the first treatment, pigs were fed ad libitum a diet containing 12.0 MJ/kg of metabolizable energy (ME) and 14.4% crude protein (CP) (maintenance diet) for 56 days. In the second and third dietary treatments, pigs were fed the maintenance diet for the initial 28 days and then switched to a high protein (17.4% dry matter (DM) CP; protein) or high (14.0 MJ/kg DM; energy) diet for the rest of the 28-day period. Dry matter intake and growth performance were similar (P ≥ 0.52) among treatments during the first 28 days of restrictive feeding, but pigs re-alimented with the protein diet achieved superior (P = 0.004) DM intake, average daily gain (ADG), and feed efficiency than those fed the maintenance diet or re-alimented with the energy diet in the re-alimentation period. At the end of the entire 56-day period, pigs re-alimented with the protein diet had higher (P ≥ 0.01) live weight gains and ADG compared with those fed the maintenance diet or re-alimented with the energy diet, but DM intake was similar (P = 0.66) among treatments. It was concluded that re-alimentation with protein rather than energy can improve compensatory growth of Ashanti Black × Large White crossbred weaner pigs. PMID:26494544

  8. Random Cell Identifiers Assignment

    Directory of Open Access Journals (Sweden)

    Robert Bestak

    2012-01-01

    Full Text Available Despite integration of advanced functions that enable Femto Access Points (FAPs to be deployed in a plug-and-play manner, the femtocell concept still cause several opened issues to be resolved. One of them represents an assignment of Physical Cell Identifiers (PCIs to FAPs. This paper analyses a random based assignment algorithm in LTE systems operating in diverse femtocell scenarios. The performance of the algorithm is evaluated by comparing the number of confusions for various femtocell densities, PCI ranges and knowledge of vicinity. Simulation results show that better knowledge of vicinity can significantly reduce the number of confusions events.

  9. Exploring the composition of protein-ligand binding sites on a large scale.

    Directory of Open Access Journals (Sweden)

    Nickolay A Khazanov

    Full Text Available The residue composition of a ligand binding site determines the interactions available for diffusion-mediated ligand binding, and understanding general composition of these sites is of great importance if we are to gain insight into the functional diversity of the proteome. Many structure-based drug design methods utilize such heuristic information for improving prediction or characterization of ligand-binding sites in proteins of unknown function. The Binding MOAD database if one of the largest curated sets of protein-ligand complexes, and provides a source of diverse, high-quality data for establishing general trends of residue composition from currently available protein structures. We present an analysis of 3,295 non-redundant proteins with 9,114 non-redundant binding sites to identify residues over-represented in binding regions versus the rest of the protein surface. The Binding MOAD database delineates biologically-relevant "valid" ligands from "invalid" small-molecule ligands bound to the protein. Invalids are present in the crystallization medium and serve no known biological function. Contacts are found to differ between these classes of ligands, indicating that residue composition of biologically relevant binding sites is distinct not only from the rest of the protein surface, but also from surface regions capable of opportunistic binding of non-functional small molecules. To confirm these trends, we perform a rigorous analysis of the variation of residue propensity with respect to the size of the dataset and the content bias inherent in structure sets obtained from a large protein structure database. The optimal size of the dataset for establishing general trends of residue propensities, as well as strategies for assessing the significance of such trends, are suggested for future studies of binding-site composition.

  10. Large cryptic internal sequence repeats in protein structures from Homo sapiens

    Indian Academy of Sciences (India)

    R Sarani; N A Udayaprakash; R Subashini; P Mridula; T Yamane; K Sekar

    2009-03-01

    Amino acid sequences are known to constantly mutate and diverge unless there is a limiting condition that makes such a change deleterious. However, closer examination of the sequence and structure reveals that a few large, cryptic repeats are nevertheless sequentially conserved. This leads to the question of why only certain repeats are conserved at the sequence level. It would be interesting to find out if these sequences maintain their conservation at the three-dimensional structure level. They can play an active role in protein and nucleotide stability, thus not only ensuring proper functioning but also potentiating malfunction and disease. Therefore, insights into any aspect of the repeats – be it structure, function or evolution – would prove to be of some importance. This study aims to address the relationship between protein sequence and its three-dimensional structure, by examining if large cryptic sequence repeats have the same structure.

  11. Direct detection of x-rays for protein crystallography employing a thick, large area CCD

    Science.gov (United States)

    Atac, Muzaffer; McKay, Timothy

    1999-01-01

    An apparatus and method for directly determining the crystalline structure of a protein crystal. The crystal is irradiated by a finely collimated x-ray beam. The interaction of the x-ray beam with the crystal produces scattered x-rays. These scattered x-rays are detected by means of a large area, thick CCD which is capable of measuring a significant number of scattered x-rays which impact its surface. The CCD is capable of detecting the position of impact of the scattered x-ray on the surface of the CCD and the quantity of scattered x-rays which impact the same cell or pixel. This data is then processed in real-time and the processed data is outputted to produce a image of the structure of the crystal. If this crystal is a protein the molecular structure of the protein can be determined from the data received.

  12. Gene and protein expression proifling analysis of young spike development in large spike wheat germplasms

    Institute of Scientific and Technical Information of China (English)

    CHEN Dan; ZHANG Jin-peng; LIU Wei-hua; WU Xiao-yang; YANGXin-ming; LIXiu-quan; LUYu-qing; LI Li-hui

    2016-01-01

    The wheat grain number per spike (GNPS) is a major yield-limiting factor in wheat-breeding programs. Germplasms with a high GNPS are therefore valuable for increasing wheat yield potential. To investigate the molecular characteristics of young spike development in large-spike wheat germplasms with high GNPS, we performed gene and protein expression proifling analysis with three high-GNPS wheat lines (Pubing 3228, Pubing 3504 and 4844-12) and one low-GNPS control variety (Fukuho). The phenotypic data for the spikes in two growth seasons showed that the GNPS of the three large-spike wheat lines were signiifcantly higher than that of the Fukuho control line. The Affymetrix wheat chip and isobaric tags for relative and absolute quantitation-tandam mass spectrometry (iTRAQ-MS/MS) technology were employed for gene and protein expression proifling analyses of young spike development, respectively, at the lforet primordia differentiation stage. A total of 598 differentialy expressed transcripts (270 up-regulated and 328 down-regulated) and 280 proteins (122 up-regulated and 158 down-regulated) were identiifed in the three high-GNPS lines compared with the control line. We found that the expression of some lforal development-related genes, includingWknox1b, theAP2 domain protein kinase and the transcription factorHUA2, were up-regulated in the high-GNPS lines. The expression of theSHEPHERD (SHD) gene was up-regulated at both the transcript and protein levels. Overal, these results suggest that multiple regulatory pathways, including theCLAVATApathway and the meristem-maintaining KNOX protein pathway, take part in the development of the high-GNPS phenotype in our wheat germplasms.

  13. Accelerating large-scale protein structure alignments with graphics processing units

    Directory of Open Access Journals (Sweden)

    Pang Bin

    2012-02-01

    Full Text Available Abstract Background Large-scale protein structure alignment, an indispensable tool to structural bioinformatics, poses a tremendous challenge on computational resources. To ensure structure alignment accuracy and efficiency, efforts have been made to parallelize traditional alignment algorithms in grid environments. However, these solutions are costly and of limited accessibility. Others trade alignment quality for speedup by using high-level characteristics of structure fragments for structure comparisons. Findings We present ppsAlign, a parallel protein structure Alignment framework designed and optimized to exploit the parallelism of Graphics Processing Units (GPUs. As a general-purpose GPU platform, ppsAlign could take many concurrent methods, such as TM-align and Fr-TM-align, into the parallelized algorithm design. We evaluated ppsAlign on an NVIDIA Tesla C2050 GPU card, and compared it with existing software solutions running on an AMD dual-core CPU. We observed a 36-fold speedup over TM-align, a 65-fold speedup over Fr-TM-align, and a 40-fold speedup over MAMMOTH. Conclusions ppsAlign is a high-performance protein structure alignment tool designed to tackle the computational complexity issues from protein structural data. The solution presented in this paper allows large-scale structure comparisons to be performed using massive parallel computing power of GPU.

  14. Utilization of Methyl Proton Resonances in Cross-Saturation Measurement for Determining the Interfaces of Large Protein-Protein Complexes

    International Nuclear Information System (INIS)

    Cross-saturation experiments allow the identification of the contact residues of large protein complexes (MW>50 K) more rigorously than conventional NMR approaches which involve chemical shift perturbations and hydrogen-deuterium exchange experiments [Takahashi et al. (2000) Nat. Struct. Biol., 7, 220-223]. In the amide proton-based cross-saturation experiment, the combined use of high deuteration levels for non-exchangeable protons of the ligand protein and a solvent with a low concentration of 1H2Ogreatly enhanced the selectivity of the intermolecular cross-saturation phenomenon. Unfortunately, experimental limitations caused losses in sensitivity. Furthermore, since main chain amide protons are not generally exposed to solvent, the efficiency of the saturation transfer directed to the main chain amide protons is not very high. Here we propose an alternative cross-saturation experiment which utilizes the methyl protons of the side chains of the ligand protein. Owing to the fast internal rotation along the methyl axis, we theoretically and experimentally demonstrated the enhanced efficiency of this approach. The methyl-utilizing cross-saturation experiment has clear advantages in sensitivity and saturation transfer efficiency over the amide proton-based approach

  15. My Favorite Assignment.

    Science.gov (United States)

    Hebert, Margaret; And Others

    1991-01-01

    Contains seven brief articles which offer assignments designed to help students perform job searches, write job application letters, answer difficult questions, write letters of resignation, alleviate fears of public speaking, use the interview effectively in the business communication, and develop listening skills. (PRA)

  16. Large-scale identification of odorant-binding proteins and chemosensory proteins from expressed sequence tags in insects

    Directory of Open Access Journals (Sweden)

    Zhang Yong-Jun

    2009-12-01

    Full Text Available Abstract Background Insect odorant binding proteins (OBPs and chemosensory proteins (CSPs play an important role in chemical communication of insects. Gene discovery of these proteins is a time-consuming task. In recent years, expressed sequence tags (ESTs of many insect species have accumulated, thus providing a useful resource for gene discovery. Results We have developed a computational pipeline to identify OBP and CSP genes from insect ESTs. In total, 752,841 insect ESTs were examined from 54 species covering eight Orders of Insecta. From these ESTs, 142 OBPs and 177 CSPs were identified, of which 117 OBPs and 129 CSPs are new. The complete open reading frames (ORFs of 88 OBPs and 123 CSPs were obtained by electronic elongation. We randomly chose 26 OBPs from eight species of insects, and 21 CSPs from four species for RT-PCR validation. Twenty two OBPs and 16 CSPs were confirmed by RT-PCR, proving the efficiency and reliability of the algorithm. Together with all family members obtained from the NCBI (OBPs or the UniProtKB (CSPs, 850 OBPs and 237 CSPs were analyzed for their structural characteristics and evolutionary relationship. Conclusions A large number of new OBPs and CSPs were found, providing the basis for deeper understanding of these proteins. In addition, the conserved motif and evolutionary analysis provide some new insights into the evolution of insect OBPs and CSPs. Motif pattern fine-tune the functions of OBPs and CSPs, leading to the minor difference in binding sex pheromone or plant volatiles in different insect Orders.

  17. Rice Ribosomal Protein Large Subunit Genes and Their Spatio-temporal and Stress Regulation.

    Science.gov (United States)

    Moin, Mazahar; Bakshi, Achala; Saha, Anusree; Dutta, Mouboni; Madhav, Sheshu M; Kirti, P B

    2016-01-01

    Ribosomal proteins (RPs) are well-known for their role in mediating protein synthesis and maintaining the stability of the ribosomal complex, which includes small and large subunits. In the present investigation, in a genome-wide survey, we predicted that the large subunit of rice ribosomes is encoded by at least 123 genes including individual gene copies, distributed throughout the 12 chromosomes. We selected 34 candidate genes, each having 2-3 identical copies, for a detailed characterization of their gene structures, protein properties, cis-regulatory elements and comprehensive expression analysis. RPL proteins appear to be involved in interactions with other RP and non-RP proteins and their encoded RNAs have a higher content of alpha-helices in their predicted secondary structures. The majority of RPs have binding sites for metal and non-metal ligands. Native expression profiling of 34 ribosomal protein large (RPL) subunit genes in tissues covering the major stages of rice growth shows that they are predominantly expressed in vegetative tissues and seedlings followed by meiotically active tissues like flowers. The putative promoter regions of these genes also carry cis-elements that respond specifically to stress and signaling molecules. All the 34 genes responded differentially to the abiotic stress treatments. Phytohormone and cold treatments induced significant up-regulation of several RPL genes, while heat and H2O2 treatments down-regulated a majority of them. Furthermore, infection with a bacterial pathogen, Xanthomonas oryzae, which causes leaf blight also induced the expression of 80% of the RPL genes in leaves. Although the expression of RPL genes was detected in all the tissues studied, they are highly responsive to stress and signaling molecules indicating that their encoded proteins appear to have roles in stress amelioration besides house-keeping. This shows that the RPL gene family is a valuable resource for manipulation of stress tolerance in rice

  18. Human heterochromatin proteins form large domains containing KRAB-ZNF genes.

    Science.gov (United States)

    Vogel, Maartje J; Guelen, Lars; de Wit, Elzo; Peric-Hupkes, Daniel; Lodén, Martin; Talhout, Wendy; Feenstra, Marike; Abbas, Ben; Classen, Anne-Kathrin; van Steensel, Bas

    2006-12-01

    Heterochromatin is important for gene regulation and chromosome structure, but the genes that are occupied by heterochromatin proteins in the mammalian genome are largely unknown. We have adapted the DamID method to systematically identify target genes of the heterochromatin proteins HP1 and SUV39H1 in human and mouse cells. Unexpectedly, we found that CBX1 (formerly HP1beta) and SUV39H1 bind to genes encoding KRAB domain containing zinc finger (KRAB-ZNF) transcriptional repressors. These genes constitute one of the largest gene families and are organized in clusters in the human genome. Preference of CBX1 for this gene family was observed in both human and mouse cells. High-resolution mapping on human chromosome 19 revealed that CBX1 coats large domains 0.1-4 Mb in size, which coincide with the position of KRAB-ZNF gene clusters. These domains show an intricate CBX1 binding pattern: While CBX1 is globally elevated throughout the domains, it is absent from the promoters and binds more strongly to the 3' ends of KRAB-ZNF genes. KRAB-ZNF domains contain large numbers of LINE elements, which may contribute to CBX1 recruitment. These results uncover a surprising link between heterochromatin and a large family of regulatory genes in mammals. We suggest a role for heterochromatin in the evolution of the KRAB-ZNF gene family. PMID:17038565

  19. Large

    OpenAIRE

    Toufik Berri; Said Azizi

    2014-01-01

    Cystic lymphangioma (CL) of the parotid gland is an uncommon benign congenital tumor and very few cases have been reported in adults. We report and discuss the case of a large CL of the parotid gland in a 66-year-old woman. On computed tomography (CT) the tumor appeared as a large, well defined, unilocular cyst of the parotid region. The lesion was surgically removed and the diagnosis was confirmed on postoperative histology. CL of the parotid gland in adults presents difficult challenges ...

  20. Intracellular protein delivery by hollow mesoporous silica capsules with a large surface hole

    International Nuclear Information System (INIS)

    We prepared cell membrane-permeable hollow mesoporous silica capsules (HMSCs) by a simple new method. CTAB micellar assembly in cholesterol emulsion gave rise to a novel capsular morphology of the HMSC particles. The HMSCs consisted of mesostructured silica walls with a large surface hole (25–50 nm) and the average particle dimension was 100–300 nm. They exhibited high surface areas of up to 719.3 m2 g−1 and a mesoporous range of pores of 2.4–2.7 nm. The surface-functionalized HMSCs could also be prepared by a similar co-condensation method using tetraethoxysilane with various organoalkoxysilane precursors in the presence of cholesterol. These organically modified HMSCs could be further modified on demand. For example, a carboxy-functionalized HMSC could be surface-functionalized by a green fluorescent 5-aminofluorescein (AFL) through an amidation reaction to afford a fluorescent AFL–HMSC. The hollow capsular morphology of the HMSCs with a large surface hole enabled us to develop very efficient intracellular delivery systems for membrane-impermeable ions, molecules, and various functional proteins. Non-covalent sequestration and delivery of proteins as well as covalent linkage of fluorescent molecules on the silica surface are effective for this system. The highly negatively charged green fluorescent probe mag-fluo-4 could be intracellularly delivered into HeLa cells by HMSC without any difficulty. The HMSCs could also effectively transport large functional proteins such as antibodies into HeLa cells. The efficiency of protein delivery by HMSC seems to be 3–22-fold higher than that of mesoporous silica nanospheres (MSNs) based on confocal laser scanning microscopy (CLSM) analysis. (paper)

  1. (H)N(COCA)NH and HN(COCA)NH experiments for 1H-15N backbone assignments in 13C/15N-labeled proteins

    Energy Technology Data Exchange (ETDEWEB)

    Bracken, Clay; Palmer, Arthur G. III [Columbia University, Department of Biochemistry and Molecular Biophysics (United States); Cavanagh, John [New York State Department of Health, NMR Structural Biology Facility, Wadsworth Center (United States)

    1997-01-15

    Triple resonance HN(COCA)NH pulse sequences for correlating 1H(i), 15N(i),1H(i-1), and 15N(i-1) spins that utilize overlapping coherence transfer periods provide increased sensitivity relative to pulse sequences that utilize sequential coherence transfer periods. Although the overlapping sequence elements reduce the overall duration of the pulse sequences, the principal benefit derives from a reduction in the number of 180 deg. pulses. Two versions of the technique are presented: a 3D (H)N(COCA)NH experiment that correlates 15N(i),1H(i-1), and 15N(i-1) spins, and a 3D HN(COCA)NH experiment that correlates 1H(i), 15N(i),1H(i-1), and 15N(i-1) spins by simultaneously encoding the 1H(i) and 15N(i) chemical shifts during the t1 evolution period. The methods are demonstrated on a 13C/15N-enriched sample of the protein ubiquitin and are easily adapted for application to 2H/13C/15N-enriched proteins.

  2. Rapamycin-binding FKBP25 associates with diverse proteins that form large intracellular entities

    Energy Technology Data Exchange (ETDEWEB)

    Galat, Andrzej, E-mail: galat@dsvidf.cea.fr; Thai, Robert

    2014-08-08

    Highlights: • The hFKBP25 interacts with diverse components of macromolecular entities. • We show that the endogenous human FKBP25 is bound to polyribosomes. • The endogenous hFKBP25 co-immunoprecipitated with nucleosomal proteins. • FKBP25 could induce conformational switch in macromolecular complexes. - Abstract: In this paper, we show some evidence that a member of the FK506-binding proteins, FKBP25 is associated to diverse components that are part of several different intracellular large-molecular mass entities. The FKBP25 is a high-affinity rapamycin-binding immunophilin, which has nuclear translocation signals present in its PPIase domain but it was detected both in the cytoplasm compartment and in the nuclear proteome. Analyses of antiFKBP25-immunoprecipitated proteins have revealed that the endogenous FKBP25 is associated to the core histones of the nucleosome, and with several proteins forming spliceosomal complexes and ribosomal subunits. Using polyclonal antiFKBP25 we have detected FKBP25 associated with polyribosomes. Added RNAs or 0.5 M NaCl release FKBP25 that was associated with the polyribosomes indicating that the immunophilin has an intrinsic capacity to form complexes with polyribonucleotides via its charged surface patches. Rapamycin or FK506 treatments of the polyribosomes isolated from porcine brain, HeLa and K568 cells caused a residual release of the endogenous FKBP25, which suggests that the immunophilin also binds to some proteins via its PPIase cavity. Our proteomics study indicates that the nuclear pool of the FKBP25 targets various nuclear proteins that are crucial for packaging of DNA, chromatin remodeling and pre-mRNA splicing whereas the cytosolic pool of this immunophilin is bound to some components of the ribosome.

  3. Large-Area Protein Patterns Generated by Ordered Binary Colloidal Assemblies as Templates

    DEFF Research Database (Denmark)

    Singh, Gurvinder; Gohri, Vipul; Pillai, Saju;

    2011-01-01

    .10 to 0.50. This allows masks with variable morphology and thus production of chemical patterns of tunable geometry. Finally, the Au or SiO(2) regions of the pattern are surface modified with protein resistant oligoethyleneglycol self-assembled molecules, which facilitates site selective adsorption......We demonstrate the use of binary colloidal assemblies as lithographic masks to generate tunable Au patterns on SiO(2) substrates with dimensions ranging from micrometers to nanometers. Such patterns can be modified with different chemistries to create patterns with well-defined sites for selective...... adsorption of proteins, where the pattern size and spacing is adjustable depending on particle choice. In our system, the binary colloidal assemblies contain large and small particles of similar or different material and are self-assembled from dilute dispersions with particle size ratios ranging from 0...

  4. On the accuracy of protein determination in large biological samples by prompt gamma neutron activation analysis

    International Nuclear Information System (INIS)

    A prompt gamma neutron activation analysis (PGNAA) facility has been developed for the determination of nitrogen and thus total protein in large volume biological samples or the whole body of small animals. In the present work, the accuracy of nitrogen determination by PGNAA in phantoms of known composition as well as in four raw ground meat samples of about 1 kg mass was examined. Dumas combustion and Kjeldahl techniques were also used for the assessment of nitrogen concentration in the meat samples. No statistically significant differences were found between the concentrations assessed by the three techniques. The results of this work demonstrate the applicability of PGNAA for the assessment of total protein in biological samples of 0.25-1.5 kg mass, such as a meat sample or the body of small animal even in vivo with an equivalent radiation dose of about 40 mSv

  5. In various protein complexes, disordered protomers have large per-residue surface areas and area of protein-, DNA- and RNA-binding interfaces.

    Science.gov (United States)

    Wu, Zhonghua; Hu, Gang; Yang, Jianyi; Peng, Zhenling; Uversky, Vladimir N; Kurgan, Lukasz

    2015-09-14

    We provide first large scale analysis of the peculiarities of surface areas of 5658 dissimilar (below 50% sequence similarity) proteins with known 3D-structures that bind to proteins, DNA or RNAs. We show here that area of the protein surface is highly correlated with the protein length. The size of the interface surface is only modestly correlated with the protein size, except for RNA-binding proteins where larger proteins are characterized by larger interfaces. Disordered proteins with disordered interfaces are characterized by significantly larger per-residue areas of their surfaces and interfaces when compared to the structured proteins. These result are applicable for proteins involved in interaction with DNA, RNA, and proteins and suggest that disordered proteins and binding regions are less compact and more likely to assume extended shape. We demonstrate that disordered protein binding residues in the interfaces of disordered proteins drive the increase in the per residue area of these interfaces. Our results can be used to predict in silico whether a given protomer from the DNA, RNA or protein complex is likely to be disordered in its unbound form.

  6. Job Assignments under Moral Hazard

    DEFF Research Database (Denmark)

    Koch, Alexander; Nafziger, Julia

    Inefficient job assignments are usually explained with incomplete information about employees' abilities or contractual imperfections. We show that inefficient assignments arise even without uncertainty about the employee's ability and with complete contracts. Building on this result we provide...

  7. More General Optimal Offset Assignment

    Directory of Open Access Journals (Sweden)

    Sven Mallach

    2015-06-01

    Full Text Available This manuscript presents exact approaches to the general offset assignment problem arising in the address code generation phase of compilers for application-specific processors. First, integer programming models for architecture-dependent and theoretically motivated special cases of the problem are established. Then, these models are extended to provide the first widely applicable formulations for the most general problem setting, supporting processors with several address registers and complex addressing capabilities. Existing heuristics are similarly extended and practical applicability of the proposed methods is demonstrated by experimental evaluation using an established and large benchmark set. The experiments allow us to study the impact of exploiting more complex memory addressing capabilities on the address computation costs of real-world programs. We also show how to integrate operand reordering techniques for commutative instructions into existing solution approaches.

  8. HLA-G and MHC Class II Protein Expression in Diffuse Large B-Cell Lymphoma.

    Science.gov (United States)

    Jesionek-Kupnicka, Dorota; Bojo, Marcin; Prochorec-Sobieszek, Monika; Szumera-Ciećkiewicz, Anna; Jabłońska, Joanna; Kalinka-Warzocha, Ewa; Kordek, Radzisław; Młynarski, Wojciech; Robak, Tadeusz; Warzocha, Krzysztof; Lech-Maranda, Ewa

    2016-06-01

    The expression of human leukocyte antigen-G (HLA-G) and HLA class II protein was studied by immunohistochemical staining of lymph nodes from 148 patients with diffuse large B-cell lymphoma (DLBCL) and related to the clinical course of the disease. Negative HLA-G expression was associated with a lower probability of achieving a complete remission (p = 0.04). Patients with negative HLA-G expression tended towards a lower 3-year overall survival (OS) rate compared to those with positive expression of HLA-G (p = 0.08). When restricting the analysis to patients receiving chemotherapy with rituximab, the estimated 3-year OS rate of patients with positive HLA-G expression was 73.3 % compared with 47.5 % (p = 0.03) in those with negative expression. Patients with negative HLA class II expression presented a lower 3-year OS rate compared to subjects with positive expression (p = 0.04). The loss of HLA class II expression (p = 0.05) and belonging to the intermediate high/high IPI risk group (p = 0.001) independently increased the risk of death. HLA class II expression also retained its prognostic value in patients receiving rituximab; the 3-year OS rate was 65.3 % in patients with positive HLA class II expression versus 29.6 % (p = 0.04) in subjects that had loss of HLA class II expression. To our knowledge, for the first time, the expression of HLA-G protein in DLBCL and its association with the clinical course of the disease was demonstrated. Moreover, the link between losing HLA class II protein expression and poor survival of patients treated with immunochemotherapy was confirmed.

  9. Vertebrate Protein CTCF and its Multiple Roles in a Large-Scale Regulation of Genome Activity

    Science.gov (United States)

    Nikolaev, L.G; Akopov, S.B; Didych, D.A; Sverdlov, E.D

    2009-01-01

    The CTCF transcription factor is an 11 zinc fingers multifunctional protein that uses different zinc finger combinations to recognize and bind different sites within DNA. CTCF is thought to participate in various gene regulatory networks including transcription activation and repression, formation of independently functioning chromatin domains and regulation of imprinting. Sequencing of human and other genomes opened up a possibility to ascertain the genomic distribution of CTCF binding sites and to identify CTCF-dependent cis-regulatory elements, including insulators. In the review, we summarized recent data on genomic distribution of CTCF binding sites in the human and other genomes within a framework of the loop domain hypothesis of large-scale regulation of the genome activity. We also tried to formulate possible lines of studies on a variety of CTCF functions which probably depend on its ability to specifically bind DNA, interact with other proteins and form di- and multimers. These three fundamental properties allow CTCF to serve as a transcription factor, an insulator and a constitutive dispersed genome-wide demarcation tool able to recruit various factors that emerge in response to diverse external and internal signals, and thus to exert its signal-specific function(s). PMID:20119526

  10. Vertebrate Protein CTCF and its Multiple Roles in a Large-Scale Regulation of Genome Activity.

    Science.gov (United States)

    Nikolaev, L G; Akopov, S B; Didych, D A; Sverdlov, E D

    2009-08-01

    The CTCF transcription factor is an 11 zinc fingers multifunctional protein that uses different zinc finger combinations to recognize and bind different sites within DNA. CTCF is thought to participate in various gene regulatory networks including transcription activation and repression, formation of independently functioning chromatin domains and regulation of imprinting. Sequencing of human and other genomes opened up a possibility to ascertain the genomic distribution of CTCF binding sites and to identify CTCF-dependent cis-regulatory elements, including insulators. In the review, we summarized recent data on genomic distribution of CTCF binding sites in the human and other genomes within a framework of the loop domain hypothesis of large-scale regulation of the genome activity. We also tried to formulate possible lines of studies on a variety of CTCF functions which probably depend on its ability to specifically bind DNA, interact with other proteins and form di- and multimers. These three fundamental properties allow CTCF to serve as a transcription factor, an insulator and a constitutive dispersed genome-wide demarcation tool able to recruit various factors that emerge in response to diverse external and internal signals, and thus to exert its signal-specific function(s). PMID:20119526

  11. Survey of large protein complexes D. vulgaris reveals great structural diversity

    Energy Technology Data Exchange (ETDEWEB)

    Han, B.-G.; Dong, M.; Liu, H.; Camp, L.; Geller, J.; Singer, M.; Hazen, T. C.; Choi, M.; Witkowska, H. E.; Ball, D. A.; Typke, D.; Downing, K. H.; Shatsky, M.; Brenner, S. E.; Chandonia, J.-M.; Biggin, M. D.; Glaeser, R. M.

    2009-08-15

    An unbiased survey has been made of the stable, most abundant multi-protein complexes in Desulfovibrio vulgaris Hildenborough (DvH) that are larger than Mr {approx} 400 k. The quaternary structures for 8 of the 16 complexes purified during this work were determined by single-particle reconstruction of negatively stained specimens, a success rate {approx}10 times greater than that of previous 'proteomic' screens. In addition, the subunit compositions and stoichiometries of the remaining complexes were determined by biochemical methods. Our data show that the structures of only two of these large complexes, out of the 13 in this set that have recognizable functions, can be modeled with confidence based on the structures of known homologs. These results indicate that there is significantly greater variability in the way that homologous prokaryotic macromolecular complexes are assembled than has generally been appreciated. As a consequence, we suggest that relying solely on previously determined quaternary structures for homologous proteins may not be sufficient to properly understand their role in another cell of interest.

  12. Artificial membrane-binding proteins stimulate oxygenation of stem cells during engineering of large cartilage tissue

    Science.gov (United States)

    Armstrong, James P. K.; Shakur, Rameen; Horne, Joseph P.; Dickinson, Sally C.; Armstrong, Craig T.; Lau, Katherine; Kadiwala, Juned; Lowe, Robert; Seddon, Annela; Mann, Stephen; Anderson, J. L. Ross; Perriman, Adam W.; Hollander, Anthony P.

    2015-06-01

    Restricted oxygen diffusion can result in central cell necrosis in engineered tissue, a problem that is exacerbated when engineering large tissue constructs for clinical application. Here we show that pre-treating human mesenchymal stem cells (hMSCs) with synthetic membrane-active myoglobin-polymer-surfactant complexes can provide a reservoir of oxygen capable of alleviating necrosis at the centre of hyaline cartilage. This is achieved through the development of a new cell functionalization methodology based on polymer-surfactant conjugation, which allows the delivery of functional proteins to the hMSC membrane. This new approach circumvents the need for cell surface engineering using protein chimerization or genetic transfection, and we demonstrate that the surface-modified hMSCs retain their ability to proliferate and to undergo multilineage differentiation. The functionalization technology is facile, versatile and non-disruptive, and in addition to tissue oxygenation, it should have far-reaching application in a host of tissue engineering and cell-based therapies.

  13. Interactive Assignments for Online Students

    Directory of Open Access Journals (Sweden)

    Pam Lowry

    2009-04-01

    Full Text Available Students can experience first hand through interactive assignments what is involved in teaching an online course. Most students develop a whole new appreciation for the student learning process. Faculty are beginning to realize that online instruction is more than a series of readings posted to a course management system. This paper summarizes the faculty member's instructional strategies involved when creating student interaction assignments. The paper also summarizes the assignments, discussion board, and trends in education from the student's perspective. In summary, it concludes with the faculty's overall perspective concerning these assignments and how the assignments could be more effective for the student.

  14. Large-scale identification of odorant-binding proteins and chemosensory proteins from expressed sequence tags in insects

    OpenAIRE

    Zhang Yong-Jun; Dong Shuang-Lin; Fang Shao-Qing; Zhang Lan; He Peng; Xu Ya-Long; Li Fei

    2009-01-01

    Abstract Background Insect odorant binding proteins (OBPs) and chemosensory proteins (CSPs) play an important role in chemical communication of insects. Gene discovery of these proteins is a time-consuming task. In recent years, expressed sequence tags (ESTs) of many insect species have accumulated, thus providing a useful resource for gene discovery. Results We have developed a computational pipeline to identify OBP and CSP genes from insect ESTs. In total, 752,841 insect ESTs were examined ...

  15. Backbone and sidechain 1H, 13C and 15N resonance assignments of the human brain-type fatty acid binding protein (FABP7) in its apo form and the holo forms binding to DHA, oleic acid, linoleic acid and elaidic acid

    DEFF Research Database (Denmark)

    Oeemig, Jesper S; Jørgensen, Mathilde L; Hansen, Mikka S;

    2009-01-01

    In this manuscript, we present the backbone and side chain assignments of human brain-type fatty acid binding protein, also known as FABP7, in its apo form and in four different holo forms, bound to DHA, oleic acid, linoleic acid and elaidic acid.......In this manuscript, we present the backbone and side chain assignments of human brain-type fatty acid binding protein, also known as FABP7, in its apo form and in four different holo forms, bound to DHA, oleic acid, linoleic acid and elaidic acid....

  16. Incentivized optimal advert assignment via utility decomposition

    NARCIS (Netherlands)

    F. Kelly; P. Key; N. Walton

    2014-01-01

    We consider a large-scale Ad-auction where adverts are assigned over a potentially infinite number of searches. We capture the intrinsic asymmetries in information between advertisers, the advert platform and the space of searches: advertisers know and can optimize the average performance of their a

  17. Complete sequence-specific 1H NMR assignments for human insulin

    International Nuclear Information System (INIS)

    Solvent conditions where human insulin could be studied by high-resolution NMR were determined. Both low pH and addition of acetonitrile were required to overcome the protein's self-association and to obtain useful spectra. Two hundred eighty-six 1H resonances were located and assigned to specific sites on the protein by using two-dimensional NMR methods. The presence and position of numerous dNN sequential NOE's indicate that the insulin conformation seen in crystallographic studies is largely retained under these solution conditions. Slowly exchanging protons were observed for seven backbone amide protons and were assigned to positions A15 and A16 and to positions B15-B19. These amides all occur within helical regions of the protein

  18. Complete sequence-specific sup 1 H NMR assignments for human insulin

    Energy Technology Data Exchange (ETDEWEB)

    Kline, A.D.; Justice, R.M. Jr. (Eli Lilly and Co., Indianapolis, IN (USA))

    1990-03-27

    Solvent conditions where human insulin could be studied by high-resolution NMR were determined. Both low pH and addition of acetonitrile were required to overcome the protein's self-association and to obtain useful spectra. Two hundred eighty-six {sup 1}H resonances were located and assigned to specific sites on the protein by using two-dimensional NMR methods. The presence and position of numerous d{sub NN} sequential NOE's indicate that the insulin conformation seen in crystallographic studies is largely retained under these solution conditions. Slowly exchanging protons were observed for seven backbone amide protons and were assigned to positions A15 and A16 and to positions B15-B19. These amides all occur within helical regions of the protein.

  19. Balanced input-output assignment

    Science.gov (United States)

    Gawronski, W.; Hadaegh, F. Y.

    1989-01-01

    Actuator/sensor locations and balanced representations of linear systems are considered for a given set of controllability and observability grammians. The case of equally controlled and observed states is given special attention. The assignability of grammians is examined, and the conditions for their existence are presented, along with several algorithms for their determination. Although an arbitrary positive semidefinite matrix is not always assignable, the identity grammian is shown to be always assignable. The results are extended to the case of flexible structures.

  20. Systematic evaluation of combined automated NOE assignment and structure calculation with CYANA

    Energy Technology Data Exchange (ETDEWEB)

    Buchner, Lena; Güntert, Peter, E-mail: guentert@em.uni-frankfurt.de [Goethe University Frankfurt am Main, Institute of Biophysical Chemistry, Center for Biomolecular Magnetic Resonance, and Frankfurt Institute of Advanced Studies (Germany)

    2015-05-15

    The automated assignment of NOESY cross peaks has become a fundamental technique for NMR protein structure analysis. A widely used algorithm for this purpose is implemented in the program CYANA. It has been used for a large number of structure determinations of proteins in solution but a systematic evaluation of its performance has not yet been reported. In this paper we systematically analyze the reliability of combined automated NOESY assignment and structure calculation with CYANA under a variety of conditions on the basis of the experimental NMR data sets of ten proteins. To evaluate the robustness of the algorithm, the original high-quality experimental data sets were modified in different ways to simulate the effect of data imperfections, i.e. incomplete or erroneous chemical shift assignments, missing NOESY cross peaks, inaccurate peak positions, inaccurate peak intensities, lower dimensionality NOESY spectra, and higher tolerances for the matching of chemical shifts and peak positions. The results show that the algorithm is remarkably robust with regard to imperfections of the NOESY peak lists and the chemical shift tolerances but susceptible to lacking or erroneous resonance assignments, in particular for nuclei that are involved in many NOESY cross peaks.

  1. Folding 19 proteins to their native state and stability of large proteins from a coarse-grained model.

    Science.gov (United States)

    Kapoor, Abhijeet; Travesset, Alex

    2014-03-01

    We develop an intermediate resolution model, where the backbone is modeled with atomic resolution but the side chain with a single bead, by extending our previous model (Proteins (2013) DOI: 10.1002/prot.24269) to properly include proline, preproline residues and backbone rigidity. Starting from random configurations, the model properly folds 19 proteins (including a mutant 2A3D sequence) into native states containing β sheet, α helix, and mixed α/β. As a further test, the stability of H-RAS (a 169 residue protein, critical in many signaling pathways) is investigated: The protein is stable, with excellent agreement with experimental B-factors. Despite that proteins containing only α helices fold to their native state at lower backbone rigidity, and other limitations, which we discuss thoroughly, the model provides a reliable description of the dynamics as compared with all atom simulations, but does not constrain secondary structures as it is typically the case in more coarse-grained models. Further implications are described.

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

    Science.gov (United States)

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

    2014-08-01

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

  3. Debottlenecking recombinant protein production in Bacillus megaterium under large-scale conditions--targeted precursor feeding designed from metabolomics.

    Science.gov (United States)

    Korneli, Claudia; Bolten, Christoph Josef; Godard, Thibault; Franco-Lara, Ezequiel; Wittmann, Christoph

    2012-06-01

    In the present work the impact of large production scale was investigated for Bacillus megaterium expressing green fluorescent protein (GFP). Specifically designed scale-down studies, mimicking the intermittent and continuous nutrient supply of large- and small-scale processes, were carried out for this purpose. The recombinant strain revealed a 40% reduced GFP yield for the large-scale conditions. In line with extended carbon loss via formation of acetate and carbon dioxide, this indicated obvious limitations in the underlying metabolism of B. megaterium under the large-scale conditions. Quantitative analysis of intracellular amino acids via validated fast filtration protocols revealed that their level strongly differed between the two scenarios. During cultivation in large-scale set-up, the availability of most amino acids, serving as key building blocks of the recombinant protein, was substantially reduced. This was most pronounced for tryptophan, aspartate, histidine, glutamine, and lysine. In contrast alanine was increased, probably related to a bottleneck at the level of pyruvate which also triggered acetate overflow metabolism. The pre-cursor quantifications could then be exploited to verify the presumed bottlenecks and improve recombinant protein production under large-scale conditions. Addition of only 5 mM tryptophan, aspartate, histidine, glutamine, and lysine to the feed solution increased the GFP yield by 100%. This rational concept of driving the lab scale productivity of recombinant microorganisms under suboptimal feeding conditions emulating large scale can easily be extended to other processes and production hosts. PMID:22252649

  4. MYC protein expression and genetic alterations have prognostic impact in diffuse large B-cell lymphoma treated with immunochemotherapy

    OpenAIRE

    Valera Barros, Alexandra; López Guillermo, Armando; Cardesa Salzmann, Antonio; Climent, Fina; González Barca, Eva; Mercadal, Santiago; Espinosa, Iñigo; Novelli, Silvana; Briones, Javier; Mate, José L.; Salamero, Olga; Sancho, Juan M.; Arenillas, Leonor; Serrano, Sergi; Erill, Nadina

    2013-01-01

    MYC alterations influence the survival of patients with diffuse large B-cell lymphoma. Most studies have focused on MYC translocations but there is little information regarding the impact of numerical alterations and protein expression. We analyzed the genetic alterations and protein expression of MYC, BCL2, BCL6, and MALT1 in 219 cases of diffuse large B-cell lymphoma. MYC rearrangement occurred as the sole abnormality (MYC single-hit) in 3% of cases, MYC and concurrent BCL2 and/or BCL6 rear...

  5. 76 FR 55880 - Recording Assignments

    Science.gov (United States)

    2011-09-09

    ..., depending on the date they were recorded. The public may also search patent and trademark assignment... United States Patent and Trademark Office Recording Assignments ACTION: Proposed collection; comment request. SUMMARY: The United States Patent and Trademark Office (USPTO), as part of its continuing...

  6. Assigning agents to a line

    DEFF Research Database (Denmark)

    Hougaard, Jens Leth; Moreno-Ternero, Juan D.; Østerdal, Lars Peter Raahave

    2014-01-01

    We consider the problem of assigning agents to slots on a line, where only one agent can be served at a slot and each agent prefers to be served as close as possible to his target. Our focus is on aggregate gap minimizing methods, i.e., those that minimize the total gap between targets and assigned...

  7. An approach to large scale identification of non-obvious structural similarities between proteins

    Directory of Open Access Journals (Sweden)

    Cherkasov Artem

    2004-05-01

    Full Text Available Abstract Background A new sequence independent bioinformatics approach allowing genome-wide search for proteins with similar three dimensional structures has been developed. By utilizing the numerical output of the sequence threading it establishes putative non-obvious structural similarities between proteins. When applied to the testing set of proteins with known three dimensional structures the developed approach was able to recognize structurally similar proteins with high accuracy. Results The method has been developed to identify pathogenic proteins with low sequence identity and high structural similarity to host analogues. Such protein structure relationships would be hypothesized to arise through convergent evolution or through ancient horizontal gene transfer events, now undetectable using current sequence alignment techniques. The pathogen proteins, which could mimic or interfere with host activities, would represent candidate virulence factors. The developed approach utilizes the numerical outputs from the sequence-structure threading. It identifies the potential structural similarity between a pair of proteins by correlating the threading scores of the corresponding two primary sequences against the library of the standard folds. This approach allowed up to 64% sensitivity and 99.9% specificity in distinguishing protein pairs with high structural similarity. Conclusion Preliminary results obtained by comparison of the genomes of Homo sapiens and several strains of Chlamydia trachomatis have demonstrated the potential usefulness of the method in the identification of bacterial proteins with known or potential roles in virulence.

  8. Fast and Accurate Protein False Discovery Rates on Large-Scale Proteomics Data Sets with Percolator 3.0

    Science.gov (United States)

    The, Matthew; MacCoss, Michael J.; Noble, William S.; Käll, Lukas

    2016-08-01

    Percolator is a widely used software tool that increases yield in shotgun proteomics experiments and assigns reliable statistical confidence measures, such as q values and posterior error probabilities, to peptides and peptide-spectrum matches (PSMs) from such experiments. Percolator's processing speed has been sufficient for typical data sets consisting of hundreds of thousands of PSMs. With our new scalable approach, we can now also analyze millions of PSMs in a matter of minutes on a commodity computer. Furthermore, with the increasing awareness for the need for reliable statistics on the protein level, we compared several easy-to-understand protein inference methods and implemented the best-performing method—grouping proteins by their corresponding sets of theoretical peptides and then considering only the best-scoring peptide for each protein—in the Percolator package. We used Percolator 3.0 to analyze the data from a recent study of the draft human proteome containing 25 million spectra (PM:24870542). The source code and Ubuntu, Windows, MacOS, and Fedora binary packages are available from http://percolator.ms/ under an Apache 2.0 license.

  9. Clean SEA-TROSY Experiments to Map Solvent Exposed Amides in Large Proteins

    Institute of Scientific and Technical Information of China (English)

    林东海

    2004-01-01

    It is well known that the SEA-TROSY experiment could alleviate some of the problems of resonance overlap in 15N/2H labeled proteins as it was designed to selectively map solvent exposed amide protons. However, SEATROSY spectra may be contaminated with exchange-relayed NOE contributions from fast exchanged hydroxyl or amine protons and contributions from longitudinal relaxation. Also, perdeuteration of the protein sample is a prerequisite for this experiment. In this communication, a modified version, clean SEA-TROSY, was proposed to eliminate these artifacts and to allow the experiment to be applied to protonated or partially deuterated proteins and protein complexes.

  10. An Assignment Sequence for Underprepared Writers.

    Science.gov (United States)

    Nimmo, Kristi

    2000-01-01

    Presents a sequenced writing assignment on shopping to aid basic writers. Describes a writing assignment focused around online and mail-order shopping. Notes steps in preparing for the assignment, the sequence, and discusses responses to the assignments. (SC)

  11. FragAnchor: A Large-Scale Predictor of Glycosylphosphatidylinositol Anchors in Eukaryote Protein Sequences by Qualitative Scoring

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    A glycosylphosphatidylinositol (GPI) anchor is a common but complex C-terminal post-translational modification of extracellular proteins in eukaryotes. Here we investigate the problem of correctly annotating GPI-anchored proteins for the growing number of sequences in public databases. We developed a computational system, called FragAnchor, based on the tandem use of a neural network (NN) and a hidden Markov model (HMM). Firstly, NN selects potential GPI-anchored proteins in a dataset, then HMM parses these potential GPI signals and refines the prediction by qualitative scoring. FragAnchor correctly predicted 91% of all the GPI-anchored proteins annotated in the Swiss-Prot database.In a large-scale analysis of 29 eukaryote proteomes, FragAnchor predicted that the percentage of highly probable GPI-anchored proteins is between 0.21% and 2.01%. The distinctive feature of FragAnchor, compared with other systems,is that it targets only the C-terminus of a protein, making it less sensitive to the background noise found in databases and possible incomplete protein sequences. Moreover, FragAnchor can be used to predict GPI-anchored proteins in all eukaryotes. Finally, by using qualitative scoring, the predictions combine both sensitivity and information content. The predictor is publicly available at http: // navet. ics. hawaii.edu/~fraganchor/NNHMM/NNHMM.html.

  12. Machine Learning-based Classification of Diffuse Large B-cell Lymphoma Patients by Their Protein Expression Profiles.

    Science.gov (United States)

    Deeb, Sally J; Tyanova, Stefka; Hummel, Michael; Schmidt-Supprian, Marc; Cox, Juergen; Mann, Matthias

    2015-11-01

    Characterization of tumors at the molecular level has improved our knowledge of cancer causation and progression. Proteomic analysis of their signaling pathways promises to enhance our understanding of cancer aberrations at the functional level, but this requires accurate and robust tools. Here, we develop a state of the art quantitative mass spectrometric pipeline to characterize formalin-fixed paraffin-embedded tissues of patients with closely related subtypes of diffuse large B-cell lymphoma. We combined a super-SILAC approach with label-free quantification (hybrid LFQ) to address situations where the protein is absent in the super-SILAC standard but present in the patient samples. Shotgun proteomic analysis on a quadrupole Orbitrap quantified almost 9,000 tumor proteins in 20 patients. The quantitative accuracy of our approach allowed the segregation of diffuse large B-cell lymphoma patients according to their cell of origin using both their global protein expression patterns and the 55-protein signature obtained previously from patient-derived cell lines (Deeb, S. J., D'Souza, R. C., Cox, J., Schmidt-Supprian, M., and Mann, M. (2012) Mol. Cell. Proteomics 11, 77-89). Expression levels of individual segregation-driving proteins as well as categories such as extracellular matrix proteins behaved consistently with known trends between the subtypes. We used machine learning (support vector machines) to extract candidate proteins with the highest segregating power. A panel of four proteins (PALD1, MME, TNFAIP8, and TBC1D4) is predicted to classify patients with low error rates. Highly ranked proteins from the support vector analysis revealed differential expression of core signaling molecules between the subtypes, elucidating aspects of their pathobiology. PMID:26311899

  13. A general Monte Carlo/simulated annealing algorithm for resonance assignment in NMR of uniformly labeled biopolymers

    Science.gov (United States)

    Hu, Kan-Nian; Qiang, Wei; Tycko, Robert

    2011-01-01

    We describe a general computational approach to site-specific resonance assignments in multidimensional NMR studies of uniformly 15N,13C-labeled biopolymers, based on a simple Monte Carlo/simulated annealing (MCSA) algorithm contained in the program MCASSIGN2. Input to MCASSIGN2 includes lists of multidimensional signals in the NMR spectra with their possible residue-type assignments (which need not be unique), the biopolymer sequence, and a table that describes the connections that relate one signal list to another. As output, MCASSIGN2 produces a high-scoring sequential assignment of the multidimensional signals, using a score function that rewards good connections (i.e., agreement between relevant sets of chemical shifts in different signal lists) and penalizes bad connections, unassigned signals, and assignment gaps. Examination of a set of high-scoring assignments from a large number of independent runs allows one to determine whether a unique assignment exists for the entire sequence or parts thereof. We demonstrate the MCSA algorithm using two-dimensional (2D) and three-dimensional (3D) solid state NMR spectra of several model protein samples (α-spectrin SH3 domain and protein G/B1 microcrystals, HET-s218–289 fibrils), obtained with magic-angle spinning and standard polarization transfer techniques. The MCSA algorithm and MCASSIGN2 program can accommodate arbitrary combinations of NMR spectra with arbitrary dimensionality, and can therefore be applied in many areas of solid state and solution NMR. PMID:21710190

  14. Online Assignments in Economics: A Test of Their Effectiveness

    Science.gov (United States)

    Kennelly, Brendan; Considine, John; Flannery, Darragh

    2011-01-01

    This article compares the effectiveness of online and paper-based assignments and tutorials using summative assessment results. All of the students in a large managerial economics course at National University of Ireland, Galway were asked to do six assignments online using Aplia and to do two on paper. The authors examined whether a student's…

  15. Large-scale analysis of in Vivo phosphorylated membrane proteins by immobilized metal ion affinity chromatography and mass spectrometry

    DEFF Research Database (Denmark)

    Nühse, Thomas S; Stensballe, Allan; Jensen, Ole N;

    2003-01-01

    Global analyses of protein phosphorylation require specific enrichment methods because of the typically low abundance of phosphoproteins. To date, immobilized metal ion affinity chromatography (IMAC) for phosphopeptides has shown great promise for large-scale studies, but has a reputation for poo...

  16. Graph-based methods for large-scale protein classification and orthology inference

    NARCIS (Netherlands)

    Kuzniar, A.

    2009-01-01

    The quest for understanding how proteins evolve and function has been a prominent and costly human endeavor. With advances in genomics and use of bioinformatics tools, the diversity of proteins in present day genomes can now be studied more efficiently than ever before. This thesis describes computa

  17. Incentivized optimal advert assignment via utility decomposition

    OpenAIRE

    Kelly, F.; Key, P.; Walton, N.

    2014-01-01

    We consider a large-scale Ad-auction where adverts are assigned over a potentially infinite number of searches. We capture the intrinsic asymmetries in information between advertisers, the advert platform and the space of searches: advertisers know and can optimize the average performance of their advertisement campaign; the platform knows and can optimize on each search instance; and, neither party knows the distribution of the infinite number of searches that can occur. We look at maximizin...

  18. Exploring Symmetry as an Avenue to the Computational Design of Large Protein Domains

    Energy Technology Data Exchange (ETDEWEB)

    Fortenberry, Carie; Bowman, Elizabeth Anne; Proffitt, Will; Dorr, Brent; Combs, Steven; Harp, Joel; Mizoue, Laura; Meiler, Jens (Vanderbilt)

    2012-03-15

    It has been demonstrated previously that symmetric, homodimeric proteins are energetically favored, which explains their abundance in nature. It has been proposed that such symmetric homodimers underwent gene duplication and fusion to evolve into protein topologies that have a symmetric arrangement of secondary structure elements - 'symmetric superfolds'. Here, the ROSETTA protein design software was used to computationally engineer a perfectly symmetric variant of imidazole glycerol phosphate synthase and its corresponding symmetric homodimer. The new protein, termed FLR, adopts the symmetric ({beta}{alpha}){sub 8} TIM-barrel superfold. The protein is soluble and monomeric and exhibits two-fold symmetry not only in the arrangement of secondary structure elements but also in sequence and at atomic detail, as verified by crystallography. When cut in half, FLR dimerizes readily to form the symmetric homodimer. The successful computational design of FLR demonstrates progress in our understanding of the underlying principles of protein stability and presents an attractive strategy for the in silico construction of larger protein domains from smaller pieces.

  19. Proteomic analysis of pure human airway gland mucus reveals a large component of protective proteins.

    Directory of Open Access Journals (Sweden)

    Nam Soo Joo

    Full Text Available Airway submucosal glands contribute to innate immunity and protect the lungs by secreting mucus, which is required for mucociliary clearance and which also contains antimicrobial, anti-inflammatory, anti-proteolytic and anti-oxidant proteins. We stimulated glands in tracheal trimmings from three lung donors and collected droplets of uncontaminated mucus as they formed at the gland orifices under an oil layer. We analyzed the mucus using liquid chromatography-tandem mass spectrometry (LC-MS/MS. Analysis identified 5486 peptides and 441 proteins from across the 3 samples (269-319 proteins per subject. We focused on 269 proteins common to at least 2 0f 3 subjects, of which 102 (38% had protective or innate immunity functions. While many of these have long been known to play such roles, for many others their cellular protective functions have only recently been appreciated in addition to their well-studied biologic functions (e.g. annexins, apolipoproteins, gelsolin, hemoglobin, histones, keratins, and lumican. A minority of the identified proteins are known to be secreted via conventional exocytosis, suggesting that glandular secretion occurs via multiple mechanisms. Two of the observed protective proteins, major vault protein and prohibitin, have not been observed in fluid from human epithelial cultures or in fluid from nasal or bronchoalveolar lavage. Further proteomic analysis of pure gland mucus may help clarify how healthy airways maintain a sterile environment.

  20. Job Assignment with Multivariate Skills

    OpenAIRE

    Brilon, Stefanie

    2010-01-01

    This paper analyzes the job assignment problem faced by a firm when workers' skills are distributed along several dimensions and jobs require different skills to varying extent. I derive optimal assignment rules with and without slot constraints, and show that under certain circumstances workers may get promoted although in their new job they are expected to be less productive than in their old job. This can be interpreted as a version of the Peter Principle which states that workers get prom...

  1. Protein-ligand binding affinities from large-scale quantum mechanical simulations

    OpenAIRE

    Fox, Stephen J.

    2012-01-01

    The accurate prediction of protein-drug binding affinities is a major aim of computational drug optimisation and development. A quantitative measure of binding affinity is provided by the free energy of binding, and such calculations typically require extensive configurational sampling of entities such as proteins with thousands of atoms. Current binding free energy methods use force fields to perform the configurational sampling and to compute interaction energies. Due to the empirical natur...

  2. The J Domain of Simian Virus 40 Large T Antigen Is Required To Functionally Inactivate RB Family Proteins

    OpenAIRE

    Zalvide, Juan; Stubdal, Hilde; DeCaprio, James A.

    1998-01-01

    Transformation by simian virus 40 large T antigen (TAg) is dependent on the inactivation of cellular tumor suppressors. Transformation minimally requires the following three domains: (i) a C-terminal domain that mediates binding to p53; (ii) the LXCXE domain (residues 103 to 107), necessary for binding to the retinoblastoma tumor suppressor protein, pRB, and the related p107 and p130; and (iii) an N-terminal domain that is homologous to the J domain of DnaJ molecular chaperone proteins. We ha...

  3. Robust classification of protein variation using structural modelling and large-scale data integration.

    Science.gov (United States)

    Baugh, Evan H; Simmons-Edler, Riley; Müller, Christian L; Alford, Rebecca F; Volfovsky, Natalia; Lash, Alex E; Bonneau, Richard

    2016-04-01

    Existing methods for interpreting protein variation focus on annotating mutation pathogenicity rather than detailed interpretation of variant deleteriousness and frequently use only sequence-based or structure-based information. We present VIPUR, a computational framework that seamlessly integrates sequence analysis and structural modelling (using the Rosetta protein modelling suite) to identify and interpret deleterious protein variants. To train VIPUR, we collected 9477 protein variants with known effects on protein function from multiple organisms and curated structural models for each variant from crystal structures and homology models. VIPUR can be applied to mutations in any organism's proteome with improved generalized accuracy (AUROC .83) and interpretability (AUPR .87) compared to other methods. We demonstrate that VIPUR's predictions of deleteriousness match the biological phenotypes in ClinVar and provide a clear ranking of prediction confidence. We use VIPUR to interpret known mutations associated with inflammation and diabetes, demonstrating the structural diversity of disrupted functional sites and improved interpretation of mutations associated with human diseases. Lastly, we demonstrate VIPUR's ability to highlight candidate variants associated with human diseases by applying VIPUR to de novo variants associated with autism spectrum disorders. PMID:26926108

  4. Single-Molecule Analysis of Protein Large-Amplitude Conformational Transitions

    Science.gov (United States)

    Yang, Haw

    2011-03-01

    Proteins have evolved to harness thermal fluctuations, rather than frustrated by them, to carry out chemical transformations and mechanical work. What are, then, the operation and design principles of protein machines? To frame the problem in a tractable way, several basic questions have been formulated to guide the experimental design: (a) How many conformational states can a protein sample on the functionally important timescale? (b) What are the inter-conversion rates between states? (c) How do ligand binding or interactions with other proteins modulate the motions? (d) What are the structural basis of flexibility and its underlying molecular mechanics? Guided by this framework, we have studied protein tyrosine phosphatase B, PtpB, from M. tuberculosis (a virulence factor of tuberculosis and a potential drug target) and adenylate kinase, AK, from E. coli (a ubiquitous energy-balancing enzyme in cells). These domain movements have been followed in real time on their respective catalytic timescales using high-resolution single-molecule Förster resonance energy transfer (FRET) spectroscopy. It is shown quantitatively that both PtpB and AK are capable of dynamically sampling two distinct states that correlate well with those observed by x-ray crystallography. Integrating these microscopic dynamics into macroscopic kinetics allows us to place the experimentally measured free-energy landscape in the context of enzymatic turnovers.

  5. THE REQUIREMENTS OF CRUDE PROTEIN BY LARGE WHITE BREEDING SOWS AND PIGLETS IN GHANA

    Directory of Open Access Journals (Sweden)

    S.W.A. RHULE

    2012-07-01

    Full Text Available Eighteen Large White gilts at an average initial live weight of 100.39kg were distributed by completely randomized block design over three treatments. There were three cereals-based diets with three different calculated levels of crude protein (CP namely control breeder diet, Diet 1 (13.16% CP, Diet 2 (12.56% CP and Diet 3 (12.01% CP with corresponding lactation diets namely control lactation Diet 1 (LD1 (16.00% CP, Diet 2 (LD2 (14.50%CP and Diet 3 (LD3 (13.20%CP. Additionally there were three creep diets comprising control Diet 1 (CD1 with 23.01%CP, Diet 2 (CD2 with 21.46% CP and Diet 3 (CD3 with 20.00% CP. The average live litter size of sows were 8.50, 8.40 and 6.67 on Diets 1, 2, and 3 respectively (P>0.05. The number of piglets weaned decreased with decreasing CP in the diet being 7.83. 7.80 and 5.33 on Diets 1, 2 and 3 respectively but were not found to be significantly (P>0.05 different. The average birth weights of the piglets were similar at 1.45, 1.34 and 1.40 kg on Diets 1, 2 and 3 respectively (P>0.05. The milk yield by the sows at 28 days of lactation were found to be 3.06, 3.03 and 5.44kg/day on LD1, 2 and 3 and significantly (P<0.05 different being higher on the lowest CP diet. There was inconsistency with the milk yield at 42 days of lactation with significantly (P<0.05 different values of 4.87, 8.33 and 3.60kg/day on lactation with lactation diet 1, 2, and 3 respectively. It was observed that live weight gains by the sows during gestation significantly (P<0.05 increased with decreasing CP levels in the diets and were 30.3, 36.3 and 34.0 kg on Diets 1, 2 and 3 respectively. The ADGs of the piglets on the creep diets were 0.22, 0.17 and 0.19 kg/day on CD1, CD2 and CD3 respectively, and found to be significantly (P<0.05 different. The study indicated that decreasing levels of CP in the diet at gestation could affect the reproductive performance of the Sow. Decreasing CP in the lactation diet significantly affected sow performance

  6. Recent advances in large-scale protein interactome mapping [version 1; referees: 3 approved

    Directory of Open Access Journals (Sweden)

    Virja Mehta

    2016-04-01

    Full Text Available Protein-protein interactions (PPIs underlie most, if not all, cellular functions. The comprehensive mapping of these complex networks of stable and transient associations thus remains a key goal, both for systems biology-based initiatives (where it can be combined with other ‘omics’ data to gain a better understanding of functional pathways and networks and for focused biological studies. Despite the significant challenges of such an undertaking, major strides have been made over the past few years. They include improvements in the computation prediction of PPIs and the literature curation of low-throughput studies of specific protein complexes, but also an increase in the deposition of high-quality data from non-biased high-throughput experimental PPI mapping strategies into publicly available databases.

  7. Genus-specific protein binding to the large clusters of DNA repeats (short regularly spaced repeats) present in Sulfolobus genomes

    DEFF Research Database (Denmark)

    Peng, Xu; Brügger, Kim; Shen, Biao;

    2003-01-01

    Short regularly spaced repeats (SRSRs) occur in multiple large clusters in archaeal chromosomes and as smaller clusters in some archaeal conjugative plasmids and bacterial chromosomes. The sequence, size, and spacing of the repeats are generally constant within a cluster but vary between clusters....... For the crenarchaeon Sulfolobus solfataricus P2, the repeats in the genome fall mainly into two closely related sequence families that are arranged in seven clusters containing a total of 441 repeats which constitute ca. 1% of the genome. The Sulfolobus conjugative plasmid pNOB8 contains a small cluster of six repeats...... that are identical in sequence to one of the repeat variants in the S. solfataricus chromosome. Repeats from the pNOB8 cluster were amplified and tested for protein binding with cell extracts from S. solfataricus. A 17.5-kDa SRSR-binding protein was purified from the cell extracts and sequenced. The protein is N...

  8. Fast large-scale clustering of protein structures using Gauss integrals

    DEFF Research Database (Denmark)

    Harder, Tim; Borg, Mikael; Boomsma, Wouter;

    2011-01-01

    trajectories. Results: We present Pleiades, a novel approach to clustering protein structures with a rigorous mathematical underpinning. The method approximates clustering based on the root mean square deviation by rst mapping structures to Gauss integral vectors – which were introduced by Røgen and co...

  9. Large scale identification and categorization of protein sequences using structured logistic regression

    DEFF Research Database (Denmark)

    Pedersen, Bjørn Panella; Ifrim, Georgiana; Liboriussen, Poul;

    2014-01-01

    Abstract Background Structured Logistic Regression (SLR) is a newly developed machine learning tool first proposed in the context of text categorization. Current availability of extensive protein sequence databases calls for an automated method to reliably classify sequences and SLR seems well...

  10. Morphological and protein profile comparison of large vessel and microvascular endothelial cells in culture

    Energy Technology Data Exchange (ETDEWEB)

    Beer, D.M.; Kim, J.S.; Carson, M.P.; Haudeuschild, C.C.; Patton, W.F.; Jacobson, B.S.

    1986-05-01

    Bovine adrenal medulla (AmMEC) and brain (BrMEC) microvessel endothelial cells, and bovine aortic (BAE) endothelial cells were isolated and cultured under identical conditions using a modification of a technique previously described for BrMEC. The cells were isolated and passaged under conditions minimizing cell surface alterations. Primary cultures were confluent in 4-6 days at a plating density in the region of 10/sup 4/ cells/cm/sup 2/. BAEs maintained a cobblestone morphology and a denser monolayer than MECs in primary and passaged cells whether the cells were passaged using Pancreatin, Trypsin-EDTA, or Collagenase-EDTA. MECs were initially elongate and became more like BAEs with passaging. BAEs and AmMECs were examined for differences in whole cell, Triton extracted cytoskeleton and plasma membrane (PM) protein profiles by two-dimensional gel electrophoresis. Cells were labeled with /sup 35/S-methionine and PM by lactoperoxidase catalyzed iodination. Though for the most part protein patterns were similar, several proteins in the PM and cytoskeletal preparations differed. A significant difference in the isoelectric forms of proteins with the same molecular weight was observed in the PM.

  11. Protein crystal growth in microgravity review of large scale temperature induction method: bovine insulin, human insulin and human alpha interferon

    Science.gov (United States)

    Long, Marianna M.; Bishop, John Bradford; Nagabhushan, Tattanahalli L.; Reichert, Paul; Smith, G. David; DeLucas, Lawrence J.

    1996-10-01

    The protein crystal growth facility (PCF) is space-flight hardware that accommodates large scale protein crystal growth experiments using temperature change as the inductive step. Recent modifications include specialized instrumentation for monitoring crystal nucleation with laser light scattering. This paper reviews results from the PCF's first seven flights on the Space Shuttle, the last with laser light scattering instrumentation. The PCF's objective is twofold: (1) production of high quality protein crystals for X-ray analysis and subsequent structure based drug design and (2) preparation of a large quantity of relatively contaminant free crystals for use as time-release protein pharmaceuticals. The first three Shuttle flights with bovine insulin constituted the PCF's proof of concept, demonstrating that the space-grown crystals were larger and diffracted to higher resolution than their earth-grown counterparts. The later four PCF missions were used to grow recombinant human insulin crystals for X-ray analysis and to continue productions trials aimed at the development of a processing facility for crystalline recombinant alpha interferon.

  12. Assignment of phycocyanobilin in HMPT using triple resonance experiments.

    Science.gov (United States)

    Röben, Marco; Schmieder, Peter

    2011-09-01

    A complete assignment of all resonances of a small organic molecule is a prerequisite for a structure determination using NMR spectroscopy. This is conventionally obtained using a well-established strategy based on COSY, HMQC and HMBC spectra. In case of phycocyanobilin (PCB) in HMPT this strategy was unsuccessful due to the symmetry of the molecule and extreme signal overlap. Since (13)C and (15)N labeled material was available, an alternative strategy for resonance assignment was used. Triple resonance experiments derived from experiments conventionally performed for proteins are sensitive and easy to analyze. Their application led to a complete and unambiguous assignment using three types of experiments. PMID:21815209

  13. Large-scale prediction of drug-target interactions using protein sequences and drug topological structures

    Energy Technology Data Exchange (ETDEWEB)

    Cao Dongsheng [Research Center of Modernization of Traditional Chinese Medicines, Central South University, Changsha 410083 (China); Liu Shao [Xiangya Hospital, Central South University, Changsha 410008 (China); Xu Qingsong [School of Mathematical Sciences and Computing Technology, Central South University, Changsha 410083 (China); Lu Hongmei; Huang Jianhua [Research Center of Modernization of Traditional Chinese Medicines, Central South University, Changsha 410083 (China); Hu Qiannan [Key Laboratory of Combinatorial Biosynthesis and Drug Discovery (Wuhan University), Ministry of Education, and Wuhan University School of Pharmaceutical Sciences, Wuhan 430071 (China); Liang Yizeng, E-mail: yizeng_liang@263.net [Research Center of Modernization of Traditional Chinese Medicines, Central South University, Changsha 410083 (China)

    2012-11-08

    Highlights: Black-Right-Pointing-Pointer Drug-target interactions are predicted using an extended SAR methodology. Black-Right-Pointing-Pointer A drug-target interaction is regarded as an event triggered by many factors. Black-Right-Pointing-Pointer Molecular fingerprint and CTD descriptors are used to represent drugs and proteins. Black-Right-Pointing-Pointer Our approach shows compatibility between the new scheme and current SAR methodology. - Abstract: The identification of interactions between drugs and target proteins plays a key role in the process of genomic drug discovery. It is both consuming and costly to determine drug-target interactions by experiments alone. Therefore, there is an urgent need to develop new in silico prediction approaches capable of identifying these potential drug-target interactions in a timely manner. In this article, we aim at extending current structure-activity relationship (SAR) methodology to fulfill such requirements. In some sense, a drug-target interaction can be regarded as an event or property triggered by many influence factors from drugs and target proteins. Thus, each interaction pair can be represented theoretically by using these factors which are based on the structural and physicochemical properties simultaneously from drugs and proteins. To realize this, drug molecules are encoded with MACCS substructure fingerings representing existence of certain functional groups or fragments; and proteins are encoded with some biochemical and physicochemical properties. Four classes of drug-target interaction networks in humans involving enzymes, ion channels, G-protein-coupled receptors (GPCRs) and nuclear receptors, are independently used for establishing predictive models with support vector machines (SVMs). The SVM models gave prediction accuracy of 90.31%, 88.91%, 84.68% and 83.74% for four datasets, respectively. In conclusion, the results demonstrate the ability of our proposed method to predict the drug

  14. Plant Cell Wall Proteins: A Large Body of Data, but What about Runaways?

    OpenAIRE

    Cécile Albenne; Hervé Canut; Laurent Hoffmann; Elisabeth Jamet

    2014-01-01

    Plant cell wall proteomics has been a very dynamic field of research for about fifteen years. A full range of strategies has been proposed to increase the number of identified proteins and to characterize their post-translational modifications. The protocols are still improving to enlarge the coverage of cell wall proteomes. Comparisons between these proteomes have been done based on various working strategies or different physiological stages. In this review, two points are highlighted. The ...

  15. A clustering method for robust and reliable large scale functional and structural protein sequence annotation

    OpenAIRE

    Piovesan, Damiano

    2013-01-01

    Bioinformatics, in the last few decades, has played a fundamental role to give sense to the huge amount of data produced. Obtained the complete sequence of a genome, the major problem of knowing as much as possible of its coding regions, is crucial. Protein sequence annotation is challenging and, due to the size of the problem, only computational approaches can provide a feasible solution. As it has been recently pointed out by the Critical Assessment of Function Annotations (CAFA), most accu...

  16. Large surface proteins of hepatitis B virus containing the pre-s sequence.

    Science.gov (United States)

    Heermann, K H; Goldmann, U; Schwartz, W; Seyffarth, T; Baumgarten, H; Gerlich, W H

    1984-11-01

    The sequence of hepatitis B virus DNA contains an open reading frame which codes for a not-yet-identified protein of at least 389 amino acids. Only the products starting at the third (GP33/GP36) or the fourth (P24/GP27) initiation signal have been characterized as components of the viral surface antigen. We found a larger protein, P39, and its glycosylated form, GP42, in hepatitis B virus particles and viral surface antigen filaments. Immunological cross-reactions showed that P39/GP42 is partially homologous to P24/GP27 and GP33/GP36. The unique portion of its sequence bound monoclonal antibodies which had been induced by immunization with hepatitis B virus particles. Proteolytic cleavage patterns and subtype-specific size differences suggested that the sequence of P39 starts with the first initiation signal of the open reading frame. Its amino-terminal part (pre-s coded) is exposed at the viral surface and, probably, is highly immunogenic. A model is presented of how the open reading frame for the viral envelope leads to defined amounts of three different proteins. PMID:6492255

  17. Pseudocontact Shift-Driven Iterative Resampling for 3D Structure Determinations of Large Proteins.

    Science.gov (United States)

    Pilla, Kala Bharath; Otting, Gottfried; Huber, Thomas

    2016-01-29

    Pseudocontact shifts (PCSs) induced by paramagnetic lanthanides produce pronounced effects in nuclear magnetic resonance spectra, which are easily measured and which deliver valuable long-range structure restraints. Even sparse PCS data greatly enhance the success rate of 3D (3-dimensional) structure predictions of proteins by the modeling program Rosetta. The present work extends this approach to 3D structures of larger proteins, comprising more than 200 residues, which are difficult to model by Rosetta without additional experimental restraints. The new algorithm improves the fragment assembly method of Rosetta by utilizing PCSs generated from paramagnetic lanthanide ions attached at four different sites as the only experimental restraints. The sparse PCS data are utilized at multiple stages, to identify native-like local structures, to rank the best structural models and to rebuild the fragment libraries. The fragment libraries are refined iteratively until convergence. The PCS-driven iterative resampling algorithm is strictly data dependent and shown to generate accurate models for a benchmark set of eight different proteins, ranging from 100 to 220 residues, using solely PCSs of backbone amide protons.

  18. Probabilistic analysis of power assignments

    NARCIS (Netherlands)

    Graaf, de Maurits; Manthey, Bodo

    2014-01-01

    A fundamental problem for wireless ad hoc networks is the assignment of suitable transmission powers to the wireless devices such that the resulting communication graph is connected. The goal is to minimize the total transmit power in order to maximize the life-time of the network. Our aim is a prob

  19. Who Benefits from Homework Assignments?

    Science.gov (United States)

    Ronning, Marte

    2011-01-01

    Using Dutch data on pupils in elementary school this paper is the first empirical study to analyze whether assigning homework has a heterogeneous impact on pupil achievement. Addressing potential biases by using a difference-in-difference approach, I find that the test score gap is larger in classes where everybody gets homework than in classes…

  20. Understanding Protein Synthesis: A Role-Play Approach in Large Undergraduate Human Anatomy and Physiology Classes

    Science.gov (United States)

    Sturges, Diana; Maurer, Trent W.; Cole, Oladipo

    2009-01-01

    This study investigated the effectiveness of role play in a large undergraduate science class. The targeted population consisted of 298 students enrolled in 2 sections of an undergraduate Human Anatomy and Physiology course taught by the same instructor. The section engaged in the role-play activity served as the study group, whereas the section…

  1. Classifying proteins into functional groups based on all-versus-all BLAST of 10 million proteins.

    Science.gov (United States)

    Kolker, Natali; Higdon, Roger; Broomall, William; Stanberry, Larissa; Welch, Dean; Lu, Wei; Haynes, Winston; Barga, Roger; Kolker, Eugene

    2011-01-01

    To address the monumental challenge of assigning function to millions of sequenced proteins, we completed the first of a kind all-versus-all sequence alignments using BLAST for 9.9 million proteins in the UniRef100 database. Microsoft Windows Azure produced over 3 billion filtered records in 6 days using 475 eight-core virtual machines. Protein classification into functional groups was then performed using Hive and custom jars implemented on top of Apache Hadoop utilizing the MapReduce paradigm. First, using the Clusters of Orthologous Genes (COG) database, a length normalized bit score (LNBS) was determined to be the best similarity measure for classification of proteins. LNBS achieved sensitivity and specificity of 98% each. Second, out of 5.1 million bacterial proteins, about two-thirds were assigned to significantly extended COG groups, encompassing 30 times more assigned proteins. Third, the remaining proteins were classified into protein functional groups using an innovative implementation of a single-linkage algorithm on an in-house Hadoop compute cluster. This implementation significantly reduces the run time for nonindexed queries and optimizes efficient clustering on a large scale. The performance was also verified on Amazon Elastic MapReduce. This clustering assigned nearly 2 million proteins to approximately half a million different functional groups. A similar approach was applied to classify 2.8 million eukaryotic sequences resulting in over 1 million proteins being assign to existing KOG groups and the remainder clustered into 100,000 functional groups. PMID:21809957

  2. Protein Biomarkers for Insulin Resistance and Type 2 Diabetes Risk in Two Large Community Cohorts.

    Science.gov (United States)

    Nowak, Christoph; Sundström, Johan; Gustafsson, Stefan; Giedraitis, Vilmantas; Lind, Lars; Ingelsson, Erik; Fall, Tove

    2016-01-01

    Insulin resistance (IR) is a precursor of type 2 diabetes (T2D), and improved risk prediction and understanding of the pathogenesis are needed. We used a novel high-throughput 92-protein assay to identify circulating biomarkers for HOMA of IR in two cohorts of community residents without diabetes (n = 1,367) (mean age 73 ± 3.6 years). Adjusted linear regression identified cathepsin D and confirmed six proteins (leptin, renin, interleukin-1 receptor antagonist [IL-1ra], hepatocyte growth factor, fatty acid-binding protein 4, and tissue plasminogen activator [t-PA]) as IR biomarkers. Mendelian randomization analysis indicated a positive causal effect of IR on t-PA concentrations. Two biomarkers, IL-1ra (hazard ratio [HR] 1.28, 95% CI 1.03-1.59) and t-PA (HR 1.30, 1.02-1.65) were associated with incident T2D, and t-PA predicted 5-year transition to hyperglycemia (odds ratio 1.30, 95% CI 1.02-1.65). Additional adjustment for fasting glucose rendered both coefficients insignificant and revealed an association between renin and T2D (HR 0.79, 0.62-0.99). LASSO regression suggested a risk model including IL-1ra, t-PA, and the Framingham Offspring Study T2D score, but prediction improvement was nonsignificant (difference in C-index 0.02, 95% CI -0.08 to 0.12) over the T2D score only. In conclusion, proteomic blood profiling indicated cathepsin D as a new IR biomarker and suggested a causal effect of IR on t-PA. PMID:26420861

  3. Low HIP1R mRNA and protein expression are associated with worse survival in diffuse large B-cell lymphoma patients treated with R-CHOP

    DEFF Research Database (Denmark)

    Wong, K. K.; Ch'ng, E. S.; Loo, S. K.;

    2015-01-01

    Huntingtin-interacting protein 1-related (HIP1R) is an endocytic protein involved in receptor trafficking, including regulating cell surface expression of receptor tyrosine kinases. We have previously shown that low HIP1R protein expression was associated with poorer survival in diffuse large B-c...

  4. Muted protein is involved in the targeting of CD63 to large dense-core vesicles of chromaffin cells.

    Science.gov (United States)

    Zhenhua, Hao; Wei, Li

    2016-08-01

    Large dense-core vesicles (LDCVs) are characterized as a class of lysosome-related organelles (LROs), which undergo regulated release and play important roles in development, metabolism and homeostasis. The Muted protein is a subunit of the biogenesis of lysosome-related organelles complex-1 (BLOC-1), which functions in the biogenesis of lysosomes and LROs. CD63 is a membrane component of lysosomes and LROs. Whether and how CD63 is sorted into LDCVs is largely unknown. In this study, we aim to identify the localization of CD63 in chromaffin cells by colocalization, living cell imaging and cell fractionation. We found that a proportion of CD63-YFP colocalized with NPY-dsRed labeled LDCVs. By sucrose density gradient fractionation, a proportion of CD63 was found to be highly enriched in LDCVs fractions. The Muted mutant mouse is a model of Hermansky-Pudlak syndrome (HPS). We also found that the level of CD63 was significantly decreased in Muted-deficient adrenal glands, suggesting that the Muted protein is important for the steady-state level of CD63. Our results suggest that CD63 is a membrane component of LDCVs and the stability of CD63 is dependent on the Muted protein, which provides a clue to the pathogenesis of LRO defects in HPS. PMID:27531610

  5. NPM/ALK binds and phosphorylates the RNA/DNA-binding protein PSF in anaplastic large-cell lymphoma.

    Science.gov (United States)

    Galietta, Annamaria; Gunby, Rosalind H; Redaelli, Sara; Stano, Paola; Carniti, Cristiana; Bachi, Angela; Tucker, Philip W; Tartari, Carmen J; Huang, Ching-Jung; Colombo, Emanuela; Pulford, Karen; Puttini, Miriam; Piazza, Rocco G; Ruchatz, Holger; Villa, Antonello; Donella-Deana, Arianna; Marin, Oriano; Perrotti, Danilo; Gambacorti-Passerini, Carlo

    2007-10-01

    The oncogenic fusion tyrosine kinase nucleophosmin/anaplastic lymphoma kinase (NPM/ALK) induces cellular transformation in anaplastic large-cell lymphomas (ALCLs) carrying the t(2;5) chromosomal translocation. Protein-protein interactions involving NPM/ALK are important for the activation of downstream signaling pathways. This study was aimed at identifying novel NPM/ALK-binding proteins that might contribute to its oncogenic transformation. Using a proteomic approach, several RNA/DNA-binding proteins were found to coimmunoprecipitate with NPM/ALK, including the multifunctional polypyrimidine tract binding proteinassociated splicing factor (PSF). The interaction between NPM/ALK and PSF was dependent on an active ALK kinase domain and PSF was found to be tyrosine-phosphorylated in NPM/ALK-expressing cell lines and in primary ALK(+) ALCL samples. Furthermore, PSF was shown to be a direct substrate of purified ALK kinase domain in vitro, and PSF Tyr293 was identified as the site of phosphorylation. Y293F PSF was not phosphorylated by NPM/ALK and was not delocalized in NPM/ALK(+) cells. The expression of ALK fusion proteins induced delocalization of PSF from the nucleus to the cytoplasm and forced overexpression of PSF-inhibited proliferation and induced apoptosis in cells expressing NPM/ALK. PSF phosphorylation also increased its binding to RNA and decreased the PSF-mediated suppression of GAGE6 expression. These results identify PSF as a novel NPM/ALK-binding protein and substrate, and suggest that PSF function may be perturbed in NPM/ALK-transformed cells.

  6. Calibrated peer review assignments for the earth sciences

    Science.gov (United States)

    Rudd, J.A., II; Wang, V.Z.; Cervato, C.; Ridky, R.W.

    2009-01-01

    Calibrated Peer Review ??? (CPR), a web-based instructional tool developed as part of the National Science Foundation reform initiatives in undergraduate science education, allows instructors to incorporate multiple writing assignments in large courses without overwhelming the instructor. This study reports successful implementation of CPR in a large, introductory geology course and student learning of geoscience content. For each CPR assignment in this study, students studied web-based and paper resources, wrote an essay, and reviewed seven essays (three from the instructor, three from peers, and their own) on the topic. Although many students expressed negative attitudes and concerns, particularly about the peer review process of this innovative instructional approach, they also recognized the learning potential of completing CPR assignments. Comparing instruction on earthquakes and plate boundaries using a CPR assignment vs. an instructional video lecture and homework essay with extensive instructor feedback, students mastered more content via CPR instruction.

  7. Annotation and analysis of a large cuticular protein family with the R&R Consensus in Anopheles gambiae

    Directory of Open Access Journals (Sweden)

    He Ningjia

    2008-01-01

    Full Text Available Abstract Background The most abundant family of insect cuticular proteins, the CPR family, is recognized by the R&R Consensus, a domain of about 64 amino acids that binds to chitin and is present throughout arthropods. Several species have now been shown to have more than 100 CPR genes, inviting speculation as to the functional importance of this large number and diversity. Results We have identified 156 genes in Anopheles gambiae that code for putative cuticular proteins in this CPR family, over 1% of the total number of predicted genes in this species. Annotation was verified using several criteria including identification of TATA boxes, INRs, and DPEs plus support from proteomic and gene expression analyses. Two previously recognized CPR classes, RR-1 and RR-2, form separate, well-supported clades with the exception of a small set of genes with long branches whose relationships are poorly resolved. Several of these outliers have clear orthologs in other species. Although both clades are under purifying selection, the RR-1 variant of the R&R Consensus is evolving at twice the rate of the RR-2 variant and is structurally more labile. In contrast, the regions flanking the R&R Consensus have diversified in amino-acid composition to a much greater extent in RR-2 genes compared with RR-1 genes. Many genes are found in compact tandem arrays that may include similar or dissimilar genes but always include just one of the two classes. Tandem arrays of RR-2 genes frequently contain subsets of genes coding for highly similar proteins (sequence clusters. Properties of the proteins indicated that each cluster may serve a distinct function in the cuticle. Conclusion The complete annotation of this large gene family provides insight on the mechanisms of gene family evolution and clues about the need for so many CPR genes. These data also should assist annotation of other Anopheles genes.

  8. The Crystal Structure of Bacteriophage HK97 gp6: Defining a Large Family of Head-Tail Connector Proteins

    Energy Technology Data Exchange (ETDEWEB)

    Cardarelli, Lia; Lam, Robert; Tuite, Ashleigh; Baker, Lindsay A; Sadowski, Paul D; Radford, Devon R; Rubinstein, John L; Battaile, Kevin P; Chirgadze, Nickolay; Maxwell, Karen L; Davidson, Alan R [UHN; (Toronto); (HWMRI)

    2011-11-23

    The final step in the morphogenesis of long-tailed double-stranded DNA bacteriophages is the joining of the DNA-filled head to the tail. The connector is a specialized structure of the head that serves as the interface for tail attachment and the point of egress for DNA from the head during infection. Here, we report the determination of a 2.1 Å crystal structure of gp6 of bacteriophage HK97. Through structural comparisons, functional studies, and bioinformatic analysis, gp6 has been determined to be a component of the connector of phage HK97 that is evolutionarily related to gp15, a well-characterized connector component of bacteriophage SPP1. Whereas the structure of gp15 was solved in a monomeric form, gp6 crystallized as an oligomeric ring with the dimensions expected for a connector protein. Although this ring is composed of 13 subunits, which does not match the symmetry of the connector within the phage, sequence conservation and modeling of this structure into the cryo-electron microscopy density of the SPP1 connector indicate that this oligomeric structure represents the arrangement of gp6 subunits within the mature phage particle. Through sequence searches and genomic position analysis, we determined that gp6 is a member of a large family of connector proteins that are present in long-tailed phages. We have also identified gp7 of HK97 as a homologue of gp16 of phage SPP1, which is the second component of the connector of this phage. These proteins are members of another large protein family involved in connector assembly.

  9. Fleet Assignment Using Collective Intelligence

    Science.gov (United States)

    Antoine, Nicolas E.; Bieniawski, Stefan R.; Kroo, Ilan M.; Wolpert, David H.

    2004-01-01

    Product distribution theory is a new collective intelligence-based framework for analyzing and controlling distributed systems. Its usefulness in distributed stochastic optimization is illustrated here through an airline fleet assignment problem. This problem involves the allocation of aircraft to a set of flights legs in order to meet passenger demand, while satisfying a variety of linear and non-linear constraints. Over the course of the day, the routing of each aircraft is determined in order to minimize the number of required flights for a given fleet. The associated flow continuity and aircraft count constraints have led researchers to focus on obtaining quasi-optimal solutions, especially at larger scales. In this paper, the authors propose the application of this new stochastic optimization algorithm to a non-linear objective cold start fleet assignment problem. Results show that the optimizer can successfully solve such highly-constrained problems (130 variables, 184 constraints).

  10. Who benefits from homework assignments?

    OpenAIRE

    Rønning, Marte

    2008-01-01

    Abstract: Using Dutch data on pupils in elementary school this paper is the first empirical study that analyzes whether assigning homework has an heterogeneous impact on pupil achievement. Addressing potential biases that arise from unobserved school quality, pupil selection by exploiting different methods, I find that the test score gap is larger in classes where everybody gets homework than in classes where nobody gets homework. More precisely pupils belonging to the upper part of the so...

  11. Mining Large Scale Tandem Mass Spectrometry Data for Protein Modifications Using Spectral Libraries.

    Science.gov (United States)

    Horlacher, Oliver; Lisacek, Frederique; Müller, Markus

    2016-03-01

    Experimental improvements in post-translational modification (PTM) detection by tandem mass spectrometry (MS/MS) has allowed the identification of vast numbers of PTMs. Open modification searches (OMSs) of MS/MS data, which do not require prior knowledge of the modifications present in the sample, further increased the diversity of detected PTMs. Despite much effort, there is still a lack of functional annotation of PTMs. One possibility to narrow the annotation gap is to mine MS/MS data deposited in public repositories and to correlate the PTM presence with biological meta-information attached to the data. Since the data volume can be quite substantial and contain tens of millions of MS/MS spectra, the data mining tools must be able to cope with big data. Here, we present two tools, Liberator and MzMod, which are built using the MzJava class library and the Apache Spark large scale computing framework. Liberator builds large MS/MS spectrum libraries, and MzMod searches them in an OMS mode. We applied these tools to a recently published set of 25 million spectra from 30 human tissues and present tissue specific PTMs. We also compared the results to the ones obtained with the OMS tool MODa and the search engine X!Tandem.

  12. Mining Large Scale Tandem Mass Spectrometry Data for Protein Modifications Using Spectral Libraries.

    Science.gov (United States)

    Horlacher, Oliver; Lisacek, Frederique; Müller, Markus

    2016-03-01

    Experimental improvements in post-translational modification (PTM) detection by tandem mass spectrometry (MS/MS) has allowed the identification of vast numbers of PTMs. Open modification searches (OMSs) of MS/MS data, which do not require prior knowledge of the modifications present in the sample, further increased the diversity of detected PTMs. Despite much effort, there is still a lack of functional annotation of PTMs. One possibility to narrow the annotation gap is to mine MS/MS data deposited in public repositories and to correlate the PTM presence with biological meta-information attached to the data. Since the data volume can be quite substantial and contain tens of millions of MS/MS spectra, the data mining tools must be able to cope with big data. Here, we present two tools, Liberator and MzMod, which are built using the MzJava class library and the Apache Spark large scale computing framework. Liberator builds large MS/MS spectrum libraries, and MzMod searches them in an OMS mode. We applied these tools to a recently published set of 25 million spectra from 30 human tissues and present tissue specific PTMs. We also compared the results to the ones obtained with the OMS tool MODa and the search engine X!Tandem. PMID:26653734

  13. Role of pRb-related proteins in simian virus 40 large-T-antigen-mediated transformation.

    OpenAIRE

    Zalvide, J; DeCaprio, J A

    1995-01-01

    Simian virus 40 large T-antigen (TAg) transformation is thought to be mediated, at least in part, by binding to and modulating the function of certain cellular proteins, including the retinoblastoma tumor suppressor gene product, pRb. TAg can disrupt the inhibitory complexes formed by pRb with the oncogenic transcription factor E2F, and this mechanism has been suggested to be important for TAg-mediated transformation. Residues 102 to 114 of TAg (including the LXCXE motif) are required for bin...

  14. biGBac enables rapid gene assembly for the expression of large multisubunit protein complexes.

    Science.gov (United States)

    Weissmann, Florian; Petzold, Georg; VanderLinden, Ryan; Huis In 't Veld, Pim J; Brown, Nicholas G; Lampert, Fabienne; Westermann, Stefan; Stark, Holger; Schulman, Brenda A; Peters, Jan-Michael

    2016-05-10

    Analyses of protein complexes are facilitated by methods that enable the generation of recombinant complexes via coexpression of their subunits from multigene DNA constructs. However, low experimental throughput limits the generation of such constructs in parallel. Here we describe a method that allows up to 25 cDNAs to be assembled into a single baculoviral expression vector in only two steps. This method, called biGBac, uses computationally optimized DNA linker sequences that enable the efficient assembly of linear DNA fragments, using reactions developed by Gibson for the generation of synthetic genomes. The biGBac method uses a flexible and modular "mix and match" approach and enables the generation of baculoviruses from DNA constructs at any assembly stage. Importantly, it is simple, efficient, and fast enough to allow the manual generation of many multigene expression constructs in parallel. We have used this method to generate and characterize recombinant forms of the anaphase-promoting complex/cyclosome, cohesin, and kinetochore complexes. PMID:27114506

  15. Identification of antithrombin-modulating genes. Role of LARGE, a gene encoding a bifunctional glycosyltransferase, in the secretion of proteins?

    Directory of Open Access Journals (Sweden)

    María Eugenia de la Morena-Barrio

    Full Text Available The haemostatic relevance of antithrombin together with the low genetic variability of SERPINC1, and the high heritability of plasma levels encourage the search for modulating genes. We used a hypothesis-free approach to identify these genes, evaluating associations between plasma antithrombin and 307,984 polymorphisms in the GAIT study (352 individuals from 21 Spanish families. Despite no SNP reaching the genome wide significance threshold, we verified milder positive associations in 307 blood donors from a different cohort. This validation study suggested LARGE, a gene encoding a protein with xylosyltransferase and glucuronyltransferase activities that forms heparin-like linear polysaccharides, as a potential modulator of antithrombin based on the significant association of one SNPs, rs762057, with anti-FXa activity, particularly after adjustment for age, sex and SERPINC1 rs2227589 genotype, all factors influencing antithrombin levels (p = 0.02. Additional results sustained this association. LARGE silencing inHepG2 and HEK-EBNA cells did not affect SERPINC1 mRNA levels but significantly reduced the secretion of antithrombin with moderate intracellular retention. Milder effects were observed on α1-antitrypsin, prothrombin and transferrin. Our study suggests LARGE as the first known modifier of plasma antithrombin, and proposes a new role for LARGE in modulating extracellular secretion of certain glycoproteins.

  16. Identification of Antithrombin-Modulating Genes. Role of LARGE, a Gene Encoding a Bifunctional Glycosyltransferase, in the Secretion of Proteins?

    Science.gov (United States)

    de la Morena-Barrio, María Eugenia; Buil, Alfonso; Antón, Ana Isabel; Martínez-Martínez, Irene; Miñano, Antonia; Gutiérrez-Gallego, Ricardo; Navarro-Fernández, José; Aguila, Sonia; Souto, Juan Carlos; Vicente, Vicente; Soria, José Manuel; Corral, Javier

    2013-01-01

    The haemostatic relevance of antithrombin together with the low genetic variability of SERPINC1, and the high heritability of plasma levels encourage the search for modulating genes. We used a hypothesis-free approach to identify these genes, evaluating associations between plasma antithrombin and 307,984 polymorphisms in the GAIT study (352 individuals from 21 Spanish families). Despite no SNP reaching the genome wide significance threshold, we verified milder positive associations in 307 blood donors from a different cohort. This validation study suggested LARGE, a gene encoding a protein with xylosyltransferase and glucuronyltransferase activities that forms heparin-like linear polysaccharides, as a potential modulator of antithrombin based on the significant association of one SNPs, rs762057, with anti-FXa activity, particularly after adjustment for age, sex and SERPINC1 rs2227589 genotype, all factors influencing antithrombin levels (p = 0.02). Additional results sustained this association. LARGE silencing inHepG2 and HEK-EBNA cells did not affect SERPINC1 mRNA levels but significantly reduced the secretion of antithrombin with moderate intracellular retention. Milder effects were observed on α1-antitrypsin, prothrombin and transferrin. Our study suggests LARGE as the first known modifier of plasma antithrombin, and proposes a new role for LARGE in modulating extracellular secretion of certain glycoproteins. PMID:23705025

  17. Effects of fish protein hydrolysate on growth performance and humoral immune response in large yellow croaker (Pseudosciaena crocea R.)

    Institute of Scientific and Technical Information of China (English)

    Hong-gang TANG; Tian-xing WU; Zhan-yu ZHAO; Xiao-dong PAN

    2008-01-01

    We investigated the effects of fish protein hydrolysate (FPH) on growth performance and humoral immune response of the large yellow croaker (Pseudosciaena crocea R.). One thousand and two hundred large yellow croakers [initial average weight: (162.75±23.85) g] were divided into four groups and reared in floating sea cages (3 m×3 m×3 m). The animals were fed with 4 diets: basal diet only (control) or diets supplemented with 5%, 10% and 15% (w/w) FPH. The results show that dietary FPH levels significantly influenced the growth and immunity of the large yellow croaker. Compared with the control group, total weight gain (TWG) in all treatment groups, relative weight gain (RWG) and specific growth rate (SGR) in fish fed with diets supplemerited with 10% and 15% FPH were significantly increased (P<0.05). Similar results were observed in immune parameters [lysozyme activity, serum complements, immunoglobulin M (IgM)]. Lysozyme activity, complement C4 and IgM were also significantly increased (P<0.05) in fish fed with diets supplemented with 10% and 15% FPH, while complement C3 level was significantly increased (P<0.05) in all treatment groups. In general, with the supplementation of FPH, particularly at dose of 10%,the growth performance and immunity of the large yellow croaker can be improved effectively.

  18. Large-Scale Quantitative Assessment of Binding Preferences in Protein-Nucleic Acid Complexes.

    Science.gov (United States)

    Jakubec, Dávid; Hostas, Jirí; Laskowski, Roman A; Hobza, Pavel; Vondrásek, Jirí

    2015-04-14

    The growing number of high-quality experimental (X-ray, NMR) structures of protein–DNA complexes has sufficient enough information to assess whether universal rules governing the DNA sequence recognition process apply. While previous studies have investigated the relative abundance of various modes of amino acid–base contacts (van der Waals contacts, hydrogen bonds), relatively little is known about the energetics of these noncovalent interactions. In the present study, we have performed the first large-scale quantitative assessment of binding preferences in protein–DNA complexes by calculating the interaction energies in all 80 possible amino acid–DNA base combinations. We found that several mutual amino acid–base orientations featuring bidentate hydrogen bonds capable of unambiguous one-to-one recognition correspond to unique minima in the potential energy space of the amino acid–base pairs. A clustering algorithm revealed that these contacts form a spatially well-defined group offering relatively little conformational freedom. Various molecular mechanics force field and DFT-D ab initio calculations were performed, yielding similar results. PMID:26894243

  19. Genomic organization and reproductive regulation of a large lipid transfer protein in the varroa mite, Varroa destructor (Anderson & Trueman).

    Science.gov (United States)

    Cabrera, A R; Shirk, P D; Duehl, A J; Donohue, K V; Grozinger, C M; Evans, J D; Teal, P E A

    2013-10-01

    The complete genomic region and corresponding transcript of the most abundant protein in phoretic varroa mites, Varroa destructor (Anderson & Trueman), were sequenced and have homology with acarine hemelipoglycoproteins and the large lipid transfer protein (LLTP) super family. The genomic sequence of VdLLTP included 14 introns and the mature transcript coded for a predicted polypeptide of 1575 amino acid residues. VdLLTP shared a minimum of 25% sequence identity with acarine LLTPs. Phylogenetic assessment showed VdLLTP was most closely related to Metaseiulus occidentalis vitellogenin and LLTP proteins of ticks; however, no heme binding by VdLLTP was detected. Analysis of lipids associated with VdLLTP showed that it was a carrier for free and esterified C12 -C22 fatty acids from triglycerides, diacylglycerides and monoacylglycerides. Additionally, cholesterol and β-sitosterol were found as cholesterol esters linked to common fatty acids. Transcript levels of VdLLTP were 42 and 310 times higher in phoretic female mites when compared with males and quiescent deutonymphs, respectively. Coincident with initiation of the reproductive phase, VdLLTP transcript levels declined to a third of those in phoretic female mites. VdLLTP functions as an important lipid transporter and should provide a significant RNA interference target for assessing the control of varroa mites.

  20. Relationship between C-reactive protein and stroke: a large prospective community based study.

    Directory of Open Access Journals (Sweden)

    Yanfang Liu

    Full Text Available Previous studies have suggested that C-reactive protein (CRP was associated with risk of stroke. There were few studies in Asian population, or on stroke subtypes other than ischemic stroke. We thus investigated the relationship between CRP and the risks of all stroke and its subtypes in a Chinese adult population.In the current study, we included 90,517 Chinese adults free of stroke and myocardial infarction at baseline (June 2006 to October 2007 in analyses. Strokes were classified as ischemic stroke (IS, intracranial heamorrhage (ICH and subarachnoid heamorrhage (SAH. High-sensitivity CRP (hs-CRP were categorized into three groups: 3 mg/L. Cox proportional hazards regression was used to calculate the association between hs-CRP concentrations and all stroke, as well as its subtypes.During a median follow-up time of 49 months, we documented 1,472 incident stroke cases. Of which 1,049 (71.3% were IS, 383 (26.0% were ICH, and 40 (2.7% were SAH. After multivariate adjustment, hs-CRP concentrations ≥1 mg/L were associated with increased risks of all stroke (hs-CRP 1-3 mg/L: hazard ratio (HR 1.17, 95% confidential interval (CI 1.03-1.33; hs-CRP>3 mg/L: HR 1.25, 95% CI 1.07-1.46 and IS (hs-CRP 1-3 mg/L: HR 1.17, 95% CI 1.01-1.36; hs-CRP>3 mg/L: HR 1.33, 95% CI 1.11-1.60, but not with ICH and SAH. Subgroup analyses showed that higher hs-CRP concentration was more prone to be a risk factor for all stroke and IS in non-fatal stroke, male and hypertensive participants.We found that higher hs-CRP concentrations were associated with a higher risk of IS, particularly for non-fatal stroke, male and hypertensive subjects. In contrast, we did not observe significant associations between hs-CRP and ICH/SAH.

  1. Large-format imaging plate and weissenberg camera for accurate protein crystallographic data collection using synchrotron radiation.

    Science.gov (United States)

    Sakabe, K; Sasaki, K; Watanabe, N; Suzuki, M; Wang, Z G; Miyahara, J; Sakabe, N

    1997-05-01

    Off-line and on-line protein data-collection systems using an imaging plate as a detector are described and their components reported. The off-line scanner IPR4080 was developed for a large-format imaging plate ;BASIII' of dimensions 400 x 400 mm and 400 x 800 mm. The characteristics of this scanner are a dynamic range of 10(5) photons pixel(-1), low background noise and high sensitivity. A means of reducing electronic noise and a method for finding the origin of the noise are discussed in detail. A dedicated screenless Weissenberg camera matching IPR4080 with synchrotron radiation was developed and installed on beamline BL6B at the Photon Factory. This camera can attach one or two sheets of 400 x 800 mm large-format imaging plate inside the film cassette by evacuation. The positional reproducibility of the imaging plate on the cassette is so good that the data can be processed by batch job. Data of 93% completeness up to 1.6 A resolution were collected on a single axis rotation and the value of R(merge) becomes 4% from a tetragonal lysozyme crystal using a set of two imaging-plate sheets. Comparing two types of imaging plates, the signal-to-noise ratio of the ST-VIP-type imaging plate is 25% better than that of the BASIII-type imaging plate for protein data collection using 1.0 and 0.7 A X-rays. A new on-line protein data-collection system with imaging plates is specially designed to use synchrotron radiation X-rays at maximum efficiency.

  2. A Statistical Programme Assignment Model

    DEFF Research Database (Denmark)

    Rosholm, Michael; Staghøj, Jonas; Svarer, Michael

    When treatment effects of active labour market programmes are heterogeneous in an observable way  across the population, the allocation of the unemployed into different programmes becomes a particularly  important issue. In this paper, we present a statistical model designed to improve the present...... duration of unemployment spells may result if a statistical programme assignment model is introduced. We discuss several issues regarding the  plementation of such a system, especially the interplay between the statistical model and  case workers....

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

    KAUST Repository

    Jang, Richard

    2012-03-21

    Background: Chemical shift mapping is an important technique in NMR-based drug screening for identifying the atoms of a target protein that potentially bind to a drug molecule upon the molecule\\'s introduction in increasing concentrations. The goal is to obtain a mapping of peaks with known residue assignment from the reference spectrum of the unbound protein to peaks with unknown assignment in the target spectrum of the bound protein. Although a series of perturbed spectra help to trace a path from reference peaks to target peaks, a one-to-one mapping generally is not possible, especially for large proteins, due to errors, such as noise peaks, missing peaks, missing but then reappearing, overlapped, and new peaks not associated with any peaks in the reference. Due to these difficulties, the mapping is typically done manually or semi-automatically, which is not efficient for high-throughput drug screening.Results: We present PeakWalker, a novel peak walking algorithm for fast-exchange systems that models the errors explicitly and performs many-to-one mapping. On the proteins: hBclXL, UbcH5B, and histone H1, it achieves an average accuracy of over 95% with less than 1.5 residues predicted per target peak. Given these mappings as input, we present PeakAssigner, a novel combined structure-based backbone resonance and NOE assignment algorithm that uses just 15N-NOESY, while avoiding TOCSY experiments and 13C-labeling, to resolve the ambiguities for a one-to-one mapping. On the three proteins, it achieves an average accuracy of 94% or better.Conclusions: Our mathematical programming approach for modeling chemical shift mapping as a graph problem, while modeling the errors directly, is potentially a time- and cost-effective first step for high-throughput drug screening based on limited NMR data and homologous 3D structures. 2012 Jang et al.; licensee BioMed Central Ltd.

  4. Combining ambiguous chemical shift mapping with structure-based backbone and NOE assignment from 15N-NOESY

    KAUST Repository

    Jang, Richard

    2011-01-01

    Chemical shift mapping is an important technique in NMRbased drug screening for identifying the atoms of a target protein that potentially bind to a drug molecule upon the molecule\\'s introduction in increasing concentrations. The goal is to obtain a mapping of peaks with known residue assignment from the reference spectrum of the unbound protein to peaks with unknown assignment in the target spectrum of the bound protein. Although a series of perturbed spectra help to trace a path from reference peaks to target peaks, a one-to-one mapping generally is not possible, especially for large proteins, due to errors, such as noise peaks, missing peaks, missing but then reappearing, overlapped, and new peaks not associated with any peaks in the reference. Due to these difficulties, the mapping is typically done manually or semi-automatically. However, automated methods are necessary for high-throughput drug screening. We present PeakWalker, a novel peak walking algorithm for fast-exchange systems that models the errors explicitly and performs many-to-one mapping. On the proteins: hBclXL, UbcH5B, and histone H1, it achieves an average accuracy of over 95% with less than 1.5 residues predicted per target peak. Given these mappings as input, we present PeakAssigner, a novel combined structure-based backbone resonance and NOE assignment algorithm that uses just 15N-NOESY, while avoiding TOCSY experiments and 13C- labeling, to resolve the ambiguities for a one-toone mapping. On the three proteins, it achieves an average accuracy of 94% or better. Copyright © 2011 ACM.

  5. Two-sided matching in hierarchical organizations: an application for the assignment of military personnel

    OpenAIRE

    Robards, Paul Anthony

    2011-01-01

    Many large organizations rely on manual assignment processes, despite the theory of bounded rationality indicating that time and cognitive constraints would limit the quality of assignments. This research used participant experiments to explore the effect of information load on assignment quality: participants, motivated by induced value theory, performed the role of decision makers and information load was identified by the number of personnel requiring assignment and the number of attrib...

  6. Out-and-back {sup 13}C-{sup 13}C scalar transfers in protein resonance assignment by proton-detected solid-state NMR under ultra-fast MAS

    Energy Technology Data Exchange (ETDEWEB)

    Barbet-Massin, Emeline; Pell, Andrew J. [University of Lyon, CNRS/ENS Lyon/UCB Lyon 1, Centre de RMN a Tres Hauts Champs (France); Jaudzems, Kristaps [Latvian Institute of Organic Synthesis (Latvia); Franks, W. Trent; Retel, Joren S. [Leibniz-Institut fuer Molekulare Pharmakologie (Germany); Kotelovica, Svetlana; Akopjana, Inara; Tars, Kaspars [Biomedical Research and Study Center (Latvia); Emsley, Lyndon [University of Lyon, CNRS/ENS Lyon/UCB Lyon 1, Centre de RMN a Tres Hauts Champs (France); Oschkinat, Hartmut [Leibniz-Institut fuer Molekulare Pharmakologie (Germany); Lesage, Anne; Pintacuda, Guido, E-mail: guido.pintacuda@ens-lyon.fr [University of Lyon, CNRS/ENS Lyon/UCB Lyon 1, Centre de RMN a Tres Hauts Champs (France)

    2013-08-15

    We present here {sup 1}H-detected triple-resonance H/N/C experiments that incorporate CO-CA and CA-CB out-and-back scalar-transfer blocks optimized for robust resonance assignment in biosolids under ultra-fast magic-angle spinning (MAS). The first experiment, (H)(CO)CA(CO)NH, yields {sup 1}H-detected inter-residue correlations, in which we record the chemical shifts of the CA spins in the first indirect dimension while during the scalar-transfer delays the coherences are present only on the longer-lived CO spins. The second experiment, (H)(CA)CB(CA)NH, correlates the side-chain CB chemical shifts with the NH of the same residue. These high sensitivity experiments are demonstrated on both fully-protonated and 100 %-H{sup N} back-protonated perdeuterated microcrystalline samples of Acinetobacter phage 205 (AP205) capsids at 60 kHz MAS.

  7. Assignment Choice, Effort, and Assignment Completion: Does Work Ethic Predict Those Who Choose Higher-Effort Assignments?

    Science.gov (United States)

    Parkhurst, John T.; Fleisher, Matthew S.; Skinner, Christopher H.; Woehr, David J.; Hawthorn-Embree, Meredith L.

    2011-01-01

    After completing the Multidimensional Work-Ethic Profile (MWEP), 98 college students were given a 20-problem math computation assignment and instructed to stop working on the assignment after completing 10 problems. Next, they were allowed to choose to finish either the partially completed assignment that had 10 problems remaining or a new…

  8. Reciprocal expression of the endocytic protein HIP1R and its repressor FOXP1 predicts outcome in R-CHOP-treated diffuse large B-cell lymphoma patients

    DEFF Research Database (Denmark)

    Wong, K K; Gascoyne, D M; Brown, P J;

    2014-01-01

    We previously identified autoantibodies to the endocytic-associated protein Huntingtin-interacting protein 1-related (HIP1R) in diffuse large B-cell lymphoma (DLBCL) patients. HIP1R regulates internalization of cell surface receptors via endocytosis, a process relevant to many therapeutic strateg...

  9. Ensiled and dry cassava leaves, and sweet potato vines as a protein source in diets for growing Vietnamese large white Mong Cai pigs

    NARCIS (Netherlands)

    Nguyen, T.H.L.; Le, N.G.; Verstegen, M.W.A.; Hendriks, W.H.

    2010-01-01

    The aim of the present study was to evaluate the effects of replacing 70% of the protein from fish meal by protein from ensiled or dry cassava leaves and sweet potato vines on the performance and carcass characters of growing F1 (Large White¿Mong Cai) pigs in Central Vietnam. Twenty-five crossbred p

  10. Structural and functional studies of a large winged Z-DNA-binding domain of Danio rerio protein kinase PKZ.

    Science.gov (United States)

    Subramani, Vinod Kumar; Kim, Doyoun; Yun, Kyunghee; Kim, Kyeong Kyu

    2016-07-01

    The Z-DNA-binding domain of PKZ from zebrafish (Danio rerio; drZαPKZ ) contains the largest β-wing among known Z-DNA-binding domains. To elucidate the functional implication of the β-wing, we solved the crystal structure of apo-drZαPKZ . Structural comparison with its Z-DNA-bound form revealed a large conformational change within the β-wing during Z-DNA binding. Biochemical studies of protein mutants revealed that two basic residues in the β-wing are responsible for Z-DNA recognition as well as fast B-Z transition. Therefore, the extra basic residues in the β-wing of drZαPKZ are necessary for the fast B-Z transition activity. PMID:27265117

  11. Prevention of hepatitis B virus infection in vivo by entry inhibitors derived from the large envelope protein.

    Science.gov (United States)

    Petersen, Joerg; Dandri, Maura; Mier, Walter; Lütgehetmann, Marc; Volz, Tassilo; von Weizsäcker, Fritz; Haberkorn, Uwe; Fischer, Lutz; Pollok, Joerg-Matthias; Erbes, Berit; Seitz, Stefan; Urban, Stephan

    2008-03-01

    360 million people are chronically infected with the human hepatitis B virus (HBV) and are consequently prone to develop liver cirrhosis and hepatocellular carcinoma. As approved therapeutic regimens-which modulate patients' antiviral defenses or inhibit the viral reverse transcriptase-are generally noncurative, strategies interfering with other HBV replication steps are required. Expanding on our demonstration that acylated peptides derived from the large HBV envelope protein block virus entry in vitro, we show their applicability to prevent HBV or woolly monkey hepatitis B virus infection in vivo, using immunodeficient urokinase-type plasminogen activator (uPA) mice repopulated with primary human or Tupaia belangeri hepatocytes. Accumulation of the peptides in the liver, their extraordinary inhibitory potency and specific mode of action permit subcutaneous delivery at very low doses. Inhibition of hepadnavirus entry thus constitutes a therapeutic approach to prevent primary HBV infection, such as after liver transplantation, and might also restrain virus spread in chronically infected patients. PMID:18297057

  12. NUP-1 Is a large coiled-coil nucleoskeletal protein in trypanosomes with lamin-like functions.

    Directory of Open Access Journals (Sweden)

    Kelly N DuBois

    Full Text Available A unifying feature of eukaryotic nuclear organization is genome segregation into transcriptionally active euchromatin and transcriptionally repressed heterochromatin. In metazoa, lamin proteins preserve nuclear integrity and higher order heterochromatin organization at the nuclear periphery, but no non-metazoan lamin orthologues have been identified, despite the likely presence of nucleoskeletal elements in many lineages. This suggests a metazoan-specific origin for lamins, and therefore that distinct protein elements must compose the nucleoskeleton in other lineages. The trypanosomatids are highly divergent organisms and possess well-documented but remarkably distinct mechanisms for control of gene expression, including polycistronic transcription and trans-splicing. NUP-1 is a large protein localizing to the nuclear periphery of Trypanosoma brucei and a candidate nucleoskeletal component. We sought to determine if NUP-1 mediates heterochromatin organization and gene regulation at the nuclear periphery by examining the influence of NUP-1 knockdown on morphology, chromatin positioning, and transcription. We demonstrate that NUP-1 is essential and part of a stable network at the inner face of the trypanosome nuclear envelope, since knockdown cells have abnormally shaped nuclei with compromised structural integrity. NUP-1 knockdown also disrupts organization of nuclear pore complexes and chromosomes. Most significantly, we find that NUP-1 is required to maintain the silenced state of developmentally regulated genes at the nuclear periphery; NUP-1 knockdown results in highly specific mis-regulation of telomere-proximal silenced variant surface glycoprotein (VSG expression sites and procyclin loci, indicating a disruption to normal chromatin organization essential to life-cycle progression. Further, NUP-1 depletion leads to increased VSG switching and therefore appears to have a role in control of antigenic variation. Thus, analogous to vertebrate

  13. Protein kinase C inhibitor sotrastaurin selectively inhibits the growth of CD79 mutant diffuse large B-cell lymphomas.

    Science.gov (United States)

    Naylor, Tara L; Tang, Huaping; Ratsch, Boris A; Enns, Andreas; Loo, Alice; Chen, Liqing; Lenz, Peter; Waters, Nigel J; Schuler, Walter; Dörken, Bernd; Yao, Yung-Mae; Warmuth, Markus; Lenz, Georg; Stegmeier, Frank

    2011-04-01

    The activated B-cell-like (ABC) subtype of diffuse large B-cell lymphoma (DLBCL) correlates with poor prognosis. The ABC subtype of DLBCL is associated with constitutive activation of the NF-κB pathway, and oncogenic lesions have been identified in its regulators, including CARD11/CARMA1 (caspase recruitment domain-containing protein 11), A20/TNFAIP3, and CD79A/B. In this study, we offer evidence of therapeutic potential for the selective PKC (protein kinase C) inhibitor sotrastaurin (STN) in preclinical models of DLBCL. A significant fraction of ABC DLBCL cell lines exhibited strong sensitivity to STN, and we found that the molecular nature of NF-κB pathway lesions predicted responsiveness. CD79A/B mutations correlated with STN sensitivity, whereas CARD11 mutations rendered ABC DLBCL cell lines insensitive. Growth inhibitory effects of PKC inhibition correlated with NF-κB pathway inhibition and were mediated by induction of G₁-phase cell-cycle arrest and/or cell death. We found that STN produced significant antitumor effects in a mouse xenograft model of CD79A/B-mutated DLBCL. Collectively, our findings offer a strong rationale for the clinical evaluation of STN in ABC DLBCL patients who harbor CD79 mutations also illustrating the necessity to stratify DLBCL patients according to their genetic abnormalities.

  14. Managing voluntary turnover through challenging assignments

    NARCIS (Netherlands)

    Preenen, P.T.Y.; Pater, I.E. de; Vianen, A.E.M. van; Keijzer, L.

    2011-01-01

    This study examines employees' challenging assignments as manageable means to reduce turnover intentions, job search behaviors, and voluntary turnover. Results indicate that challenging assignments are negatively related to turnover intentions and job search behaviors and that these relationships ar

  15. Capacity constrained assignment in spatial databases

    DEFF Research Database (Denmark)

    U, Leong Hou; Yiu, Man Lung; Mouratidis, Kyriakos;

    2008-01-01

    Given a point set P of customers (e.g., WiFi receivers) and a point set Q of service providers (e.g., wireless access points), where each q 2 Q has a capacity q.k, the capacity constrained assignment (CCA) is a matching M Q × P such that (i) each point q 2 Q (p 2 P) appears at most k times (at most......, the quality of q's service to p in a given (q, p) pair is anti-proportional to their distance. Although max-flow algorithms are applicable to this problem, they require the complete distance-based bipartite graph between Q and P. For large spatial datasets, this graph is expensive to compute and it may be too...

  16. The Development of Gel Media and Columns for Large-Scale Chromatography of Proteins,a Historical Review

    Institute of Scientific and Technical Information of China (English)

    Jan-Christer; Janson

    2002-01-01

    Thr first dedicated protein chromatography media were introduced during the 1950s and 1960s.There was an early awareness of the possibility of using these for production applications within the biopharmaceutical industry.However,the crucial limitation was the fact that those media that were most compatible with proteins lent themselves less favourably to scaling-up.The problems were primarily physical.Thus the fibrous cellulose media showed bed cracking tendencies and the bead shaped polyacrylamide.dextran,and agarose gel media,then available, were too soft to stand the hydrodynamic forces acting in large columns,leading to bed compaction and increased pressure drop.At the time,the best solution to the latter problem,after a number of intermediary solutions were tried,was the introductionof the stacked column concept in which several short column segments were connected by small bore tubing,thus reducing the force acting on the particles in each bed com partment,However,the ultimate remedy,the introduction of chromatographic matrices that combine the desired features of adequate rigidity,macroporosity,biocompatibility,chemical stability(for CIP and SIP0and derivatizability,did not occur until the middle of the 1980s when adequately cross-linked agarose gel media such as Sepharose Fast Flow were made available.The paper also recognizes the many attempts made during the past 50 years to develop continous chromatography columns.Most of the designs are based on an annular bed or on an array of annularly arranged parallel columns continuously fed with samples in a cyclic manner.The introduction of media and columns for expanded bed adsorption followed a demand for rewer pruification steps and shorter process times.In recent years,columns have been ntroduced that allow packing and repacking without needing to open the column.The review provides an historical account of the developments that have led to the present state-of-the-art both regarding large diameter columns

  17. 5 CFR 870.901 - Assignments permitted.

    Science.gov (United States)

    2010-01-01

    ... (CONTINUED) FEDERAL EMPLOYEES' GROUP LIFE INSURANCE PROGRAM Assignments of Life Insurance § 870.901..., and compensationers. (2) An individual may assign ownership of all life insurance under this part... Benefit before assigning the remainder of his/her insurance, the amount of Basic insurance does...

  18. 7 CFR 1437.104 - Assigned production.

    Science.gov (United States)

    2010-01-01

    ... 7 Agriculture 10 2010-01-01 2010-01-01 false Assigned production. 1437.104 Section 1437.104... Determining Yield Coverage Using Actual Production History § 1437.104 Assigned production. (a) When determining losses under this section, assigned production will be used to offset the loss of production...

  19. Lexical Stress Assignment in Italian Developmental Dyslexia

    Science.gov (United States)

    Paizi, Despina; Zoccolotti, Pierluigi; Burani, Cristina

    2011-01-01

    Stress assignment to Italian polysyllabic words is unpredictable, because stress is neither marked nor predicted by rule. Stress assignment, especially to low frequency words, has been reported to be a function of stress dominance and stress neighbourhood. Two experiments investigate stress assignment in sixth-grade, skilled and dyslexic, readers.…

  20. Integrated assignment and path planning

    Science.gov (United States)

    Murphey, Robert A.

    2005-11-01

    A surge of interest in unmanned systems has exposed many new and challenging research problems across many fields of engineering and mathematics. These systems have the potential of transforming our society by replacing dangerous and dirty jobs with networks of moving machines. This vision is fundamentally separate from the modern view of robotics in that sophisticated behavior is realizable not by increasing individual vehicle complexity, but instead through collaborative teaming that relies on collective perception, abstraction, decision making, and manipulation. Obvious examples where collective robotics will make an impact include planetary exploration, space structure assembly, remote and undersea mining, hazardous material handling and clean-up, and search and rescue. Nonetheless, the phenomenon driving this technology trend is the increasing reliance of the US military on unmanned vehicles, specifically, aircraft. Only a few years ago, following years of resistance to the use of unmanned systems, the military and civilian leadership in the United States reversed itself and have recently demonstrated surprisingly broad acceptance of increasingly pervasive use of unmanned platforms in defense surveillance, and even attack. However, as rapidly as unmanned systems have gained acceptance, the defense research community has discovered the technical pitfalls that lie ahead, especially for operating collective groups of unmanned platforms. A great deal of talent and energy has been devoted to solving these technical problems, which tend to fall into two categories: resource allocation of vehicles to objectives, and path planning of vehicle trajectories. An extensive amount of research has been conducted in each direction, yet, surprisingly, very little work has considered the integrated problem of assignment and path planning. This dissertation presents a framework for studying integrated assignment and path planning and then moves on to suggest an exact

  1. Optimisation of timetable-based, stochastic transit assignment models based on MSA

    DEFF Research Database (Denmark)

    Nielsen, Otto Anker; Frederiksen, Rasmus Dyhr

    2006-01-01

    (CRM), such a large-scale transit assignment model was developed and estimated. The Stochastic User Equilibrium problem was solved by the Method of Successive Averages (MSA). However, the model suffered from very large calculation times. The paper focuses on how to optimise transit assignment models...

  2. An optimal query assignment for wireless sensor networks

    NARCIS (Netherlands)

    Mitici, Mihaela; Onderwater, Martijn; Graaf, de Maurits; Ommeren, van Jan-Kees; Dijk, van Nico; Goseling, Jasper

    2012-01-01

    With the increased use of large-scale real-time embedded sensor networks, new control mechanisms are needed to avoid congestion and meet required Quality of Service (QoS) levels. In this paper, we propose a Markov Decision Problem (MDP) to prescribe an optimal query assignment strategy that achieves

  3. Semiclassical Assignment of the Vibrational Spectrum of N2O

    NARCIS (Netherlands)

    Waalkens, Holger; Jung, Christof; Taylor, Howard S.

    2002-01-01

    The vibrational spectrum of N2O as given by an effective spectroscopic Hamiltonian based on the existence of a superpolyad number is analyzed and assigned in terms of classical motions. The effective Hamiltonian includes a large number of resonances of which only one is dominant for low and intermed

  4. Using Clouds for MapReduce Measurement Assignments

    Science.gov (United States)

    Rabkin, Ariel; Reiss, Charles; Katz, Randy; Patterson, David

    2013-01-01

    We describe our experiences teaching MapReduce in a large undergraduate lecture course using public cloud services and the standard Hadoop API. Using the standard API, students directly experienced the quality of industrial big-data tools. Using the cloud, every student could carry out scalability benchmarking assignments on realistic hardware,…

  5. The domain architecture of large guanine nucleotide exchange factors for the small GTP-binding protein Arf

    Directory of Open Access Journals (Sweden)

    Geldner Niko

    2005-02-01

    Full Text Available Abstract Background Small G proteins, which are essential regulators of multiple cellular functions, are activated by guanine nucleotide exchange factors (GEFs that stimulate the exchange of the tightly bound GDP nucleotide by GTP. The catalytic domain responsible for nucleotide exchange is in general associated with non-catalytic domains that define the spatio-temporal conditions of activation. In the case of small G proteins of the Arf subfamily, which are major regulators of membrane trafficking, GEFs form a heterogeneous family whose only common characteristic is the well-characterized Sec7 catalytic domain. In contrast, the function of non-catalytic domains and how they regulate/cooperate with the catalytic domain is essentially unknown. Results Based on Sec7-containing sequences from fully-annotated eukaryotic genomes, including our annotation of these sequences from Paramecium, we have investigated the domain architecture of large ArfGEFs of the BIG and GBF subfamilies, which are involved in Golgi traffic. Multiple sequence alignments combined with the analysis of predicted secondary structures, non-structured regions and splicing patterns, identifies five novel non-catalytic structural domains which are common to both subfamilies, revealing that they share a conserved modular organization. We also report a novel ArfGEF subfamily with a domain organization so far unique to alveolates, which we name TBS (TBC-Sec7. Conclusion Our analysis unifies the BIG and GBF subfamilies into a higher order subfamily, which, together with their being the only subfamilies common to all eukaryotes, suggests that they descend from a common ancestor from which species-specific ArfGEFs have subsequently evolved. Our identification of a conserved modular architecture provides a background for future functional investigation of non-catalytic domains.

  6. An Investigation of the Partial-Assignment Completion Effect on Students' Assignment Choice Behavior

    Science.gov (United States)

    Hawthorn-Embree, Meredith L.; Skinner, Christopher H.; Parkhurst, John; Conley, Elisha

    2011-01-01

    This study was designed to investigate the partial assignment completion effect. Seventh-grade students were given a math assignment. After working for 5 min, they were interrupted and their partially completed assignments were collected. About 20 min later, students were given their partially completed assignment and a new, control assignment…

  7. Heteronuclear 2D NMR studies on an engineered insulin monomer: Assignments and characterization of the receptor-binding surface by selective 2H and 13C labeling with application to protein design

    International Nuclear Information System (INIS)

    Insulin provides an important model for the application of genetic engineering to rational protein design and has been well characterized in the crystal state. However, self-association of insulin in solution has precluded complementary 2D NMR study under physiological conditions. The authors demonstrate here that such limitations may be circumvented by the use of a monomeric analogue that contains three amino acid substitutions on the protein surface (HisB10 → Asp, ProB28 → Lys, and LysB29 → Pro); this analogue (designated DKP-insulin) retains native receptor-binding potency. Comparative 1H NMR studies of native human insulin and a series of three related analogues-(i) the singly substituted analogue [HisB10→Asp], (ii) the doubly substituted analogue [ProB28→Lys; LysB29→Pro], and (iii) DKP-insulin-demonstrate progressive reduction in concentration-dependent line-broadening in accord with the results of analytical ultracentrifugation. Extensive nonlocal interactions are observed in the NOESY spectrum of DKP-insulin, indicating that this analogue adopts a compact and stably folded structure as a monomer in overall accord with crystal models. Site-specific 2H and 13C isotopic labels are introduced by semisynthesis as probes for the structure and dynamics of the receptor-binding surface. These studies confirm and extend under physiological conditions the results of a previous 2D NMR analysis of native insulin in 20% acetic acid. Implications for the role of protein flexibility in receptor recognition are discussed with application to the design of novel insulin analogues

  8. Sequence analysis of the large polymerase (L) protein of the US strain of avian metapneumovirus indicates a close resemblance to that of the human metapneumovirus.

    Science.gov (United States)

    Govindarajan, Dhanasekaran; Samal, Siba K

    2004-09-15

    The complete nucleotide sequence of the large polymerase (L) protein of the avian metapneumovirus subgroup C strain Colorado was determined. The L protein gene of avian pneumovirus Colorado isolate (APV-C) was 6173 nucleotides in length from the gene-start to the gene-end and encoded a polypeptide of 2005 amino acids in length. The length of the L protein of APV-C was exactly the same as that of human metapneumovirus (hMPV) and one amino acid longer than the L protein of APV subgroup A. The L protein of APV-C showed 80% amino acid identity with the L protein of hMPV, but only 64% amino acid identity with the L protein of APV-A. The nucleotide and deduced amino acid sequences were compared with the corresponding sequences of eleven other paramyxoviruses. All six domains characteristic of paramyxovirus L proteins were also observed in the L protein of APV-C. All the polymerase core motifs in domain III were conserved to nearly 100% in the metapneumoviruses. Similarly, the putative ATP-binding motif in domain VI was completely conserved among the metapneumoviruses and differed in length, by one intermediate residue, from other paramyxoviruses. Phylogenetic analysis of the different L proteins also revealed a closer relationship between APV-C and hMPV.

  9. Job Assignments, Intrinsic Motivation and Explicit Incentives

    OpenAIRE

    Nafziger, Julia

    2008-01-01

    This paper considers the interplay of job assignments with the intrinsic and extrinsic motivation of an agent. Job assignments influence the self confidence of the agent, and thereby his intrinsic motivation. Monetary reward allow the principal to complement intrinsic motivation with extrinsic incentives. The main result is that the principal chooses an inefficient job assignment rule to enhance the agent's intrinsic motivation even though she can motivate him with monetary rewards. This show...

  10. MYC protein expression and genetic alterations have prognostic impact in patients with diffuse large B-cell lymphoma treated with immunochemotherapy

    OpenAIRE

    Valera, Alexandra; López-Guillermo, Armando; Cardesa-Salzmann, Teresa; Climent, Fina; González-Barca, Eva; Mercadal, Santiago; Espinosa, Íñigo; Novelli, Silvana; Briones, Javier; Mate, José L.; Salamero, Olga; Sancho, Juan M.; Arenillas, Leonor; Serrano, Sergi; Erill, Nadina

    2013-01-01

    MYC alterations influence the survival of patients with diffuse large B-cell lymphoma. Most studies have focused on MYC translocations but there is little information regarding the impact of numerical alterations and protein expression. We analyzed the genetic alterations and protein expression of MYC, BCL2, BCL6, and MALT1 in 219 cases of diffuse large B-cell lymphoma. MYC rearrangement occurred as the sole abnormality (MYC single-hit) in 3% of cases, MYC and concurrent BCL2 and/or BCL6 rear...

  11. Wildlife forensic science: A review of genetic geographic origin assignment.

    Science.gov (United States)

    Ogden, Rob; Linacre, Adrian

    2015-09-01

    Wildlife forensic science has become a key means of enforcing legislation surrounding the illegal trade in protected and endangered species. A relatively new dimension to this area of forensic science is to determine the geographic origin of a seized sample. This review focuses on DNA testing, which relies on assignment of an unknown sample to its genetic population of origin. Key examples of this are the trade in timber, fish and ivory and these are used only to illustrate the large number of species for which this type of testing is potentially available. The role of mitochondrial and nuclear DNA markers is discussed, alongside a comparison of neutral markers with those exhibiting signatures of selection, which potentially offer much higher levels of assignment power to address specific questions. A review of assignment tests is presented along with detailed methods for evaluating error rates and considerations for marker selection. The availability and quality of reference data are of paramount importance to support assignment applications and ensure reliability of any conclusions drawn. The genetic methods discussed have been developed initially as investigative tools but comment is made regarding their use in courts. The potential to compliment DNA markers with elemental assays for greater assignment power is considered and finally recommendations are made for the future of this type of testing.

  12. A Kelch Motif-Containing Serine/Threonine Protein Phosphatase Determines the Large Grain QTL Trait in Rice

    Institute of Scientific and Technical Information of China (English)

    Zejun Hu; Haohua He; Shiyong Zhang; Fan Sun; Xiaoyun Xin; Wenxiang Wang; Xi Qian; Jingshui Yang; Xiaojin Luo

    2012-01-01

    A thorough understanding of the genetic basis of rice grain traits is critical for the improvement of rice (Oryza sativa L.) varieties.In this study,we generated an F2 population by crossing the large-grain japonica cultivar CW23 with Peiai 64 (PA64),an elite indica small-grain cultivar.Using QTL analysis,17 QTLs for five grain traits were detected on four different chromosomes.Eight of the QTLs were newly-identified in this study.In particular,qGL3-1,a newly-identified grain length QTL with the highest LOD value and largest phenotypic variation,was fine-mapped to the 17 kb region of chromosome 3.A serine/threonine protein phosphatase gene encoding a repeat domain containing two Kelch motifs was identified as the unique candidate gene corresponding to this QTL.A comparison of PA64 and CW23 sequences revealed a single nucleotide substitution (C→A) at position 1092 in exon 10,resulting in replacement of Asp (D) in PA64 with Glu (E) in CW23 for the 364th amino acid.This variation is located at the D position of the conserved sequence motif AVLDT of the Kelch repeat.Genetic analysis of a near-isogenic line (NIL) for qGL3-1 revealed that the allele qGL3-1 from CW23 has an additive or partly dominant effect,and is suitable for use in molecular marker-assisted selection.

  13. Large scale interaction analysis predicts that the Gerbera hybrida floral E function is provided both by general and specialized proteins

    Directory of Open Access Journals (Sweden)

    Elomaa Paula

    2010-06-01

    Full Text Available Abstract Background The ornamental plant Gerbera hybrida bears complex inflorescences with morphologically distinct floral morphs that are specific to the sunflower family Asteraceae. We have previously characterized several MADS box genes that regulate floral development in Gerbera. To study further their behavior in higher order complex formation according to the quartet model, we performed yeast two- and three-hybrid analysis with fourteen Gerbera MADS domain proteins to analyze their protein-protein interaction potential. Results The exhaustive pairwise interaction analysis showed significant differences in the interaction capacity of different Gerbera MADS domain proteins compared to other model plants. Of particular interest in these assays was the behavior of SEP-like proteins, known as GRCDs in Gerbera. The previously described GRCD1 and GRCD2 proteins, which are specific regulators involved in stamen and carpel development, respectively, showed very limited pairwise interactions, whereas the related GRCD4 and GRCD5 factors displayed hub-like positions in the interaction map. We propose GRCD4 and GRCD5 to provide a redundant and general E function in Gerbera, comparable to the SEP proteins in Arabidopsis. Based on the pairwise interaction data, combinations of MADS domain proteins were further subjected to yeast three-hybrid assays. Gerbera B function proteins showed active behavior in ternary complexes. All Gerbera SEP-like proteins with the exception of GRCD1 were excellent partners for B function proteins, further implicating the unique role of GRCD1 as a whorl- and flower-type specific C function partner. Conclusions Gerbera MADS domain proteins exhibit both conserved and derived behavior in higher order protein complex formation. This protein-protein interaction data can be used to classify and compare Gerbera MADS domain proteins to those of Arabidopsis and Petunia. Combined with our reverse genetic studies of Gerbera, these

  14. Large-scale inference of protein tissue origin in gram-positive sepsis plasma using quantitative targeted proteomics.

    Science.gov (United States)

    Malmström, Erik; Kilsgård, Ola; Hauri, Simon; Smeds, Emanuel; Herwald, Heiko; Malmström, Lars; Malmström, Johan

    2016-01-06

    The plasma proteome is highly dynamic and variable, composed of proteins derived from surrounding tissues and cells. To investigate the complex processes that control the composition of the plasma proteome, we developed a mass spectrometry-based proteomics strategy to infer the origin of proteins detected in murine plasma. The strategy relies on the construction of a comprehensive protein tissue atlas from cells and highly vascularized organs using shotgun mass spectrometry. The protein tissue atlas was transformed to a spectral library for highly reproducible quantification of tissue-specific proteins directly in plasma using SWATH-like data-independent mass spectrometry analysis. We show that the method can determine drastic changes of tissue-specific protein profiles in blood plasma from mouse animal models with sepsis. The strategy can be extended to several other species advancing our understanding of the complex processes that contribute to the plasma proteome dynamics.

  15. NMR assignments of mitochondrial cyclophilin Cpr3 from Saccharomyces cerevisiae.

    Science.gov (United States)

    Shukla, Vaibhav Kumar; Singh, Jai Shankar; Trivedi, Dipesh; Hosur, Ramakrishna V; Kumar, Ashutosh

    2016-04-01

    Cyclophilins regulate protein folding, transport and signalling through catalysis of proline isomerization, and are ubiquitously expressed in both prokaryotes and eukaryotes. Cpr3 is the yeast mitochondrial cyclophilin and it is structurally and biophysically uncharacterized so far. Yeast cyclophilin gene cpr3 is essential for the lactate metabolism. Here, we report (1)H, (13)C, and (15)N chemical shift assignments of Cpr3 protein determined by various 2D and 3D heteronuclear NMR experiments at pH 6.5, and temperature 298 K. PMID:26897529

  16. 48 CFR 208.7002 - Assignment authority.

    Science.gov (United States)

    2010-10-01

    ... 48 Federal Acquisition Regulations System 3 2010-10-01 2010-10-01 false Assignment authority. 208..., DEPARTMENT OF DEFENSE ACQUISITION PLANNING REQUIRED SOURCES OF SUPPLIES AND SERVICES Coordinated Acquisition 208.7002 Assignment authority. (a) Under the DoD Coordinated Acquisition Program,...

  17. Individualized Assignments in an Experimental Psychology Course.

    Science.gov (United States)

    Hovancik, John R.

    1984-01-01

    A computer is used to individualize student assignments in statistics. The principal benefit of individualized activities is that they emphasize decision-making processes rather than correct answers. Students in an individualized assignment group scored higher on an examination than those in a comparison group. (RM)

  18. Detecting Plagiarism in MS Access Assignments

    Science.gov (United States)

    Singh, Anil

    2013-01-01

    Assurance of individual effort from students in computer-based assignments is a challenge. Due to digitization, students can easily use a copy of their friend's work and submit it as their own. Plagiarism in assignments puts students who cheat at par with those who work honestly and this compromises the learning evaluation process. Using a…

  19. A Role-Based Fuzzy Assignment Model

    Institute of Scientific and Technical Information of China (English)

    ZUO Bao-he; FENG Shan

    2002-01-01

    It's very important to dynamically assign the tasks to corresponding actors in workflow management system, especially in complex applications. This improves the flexibility of workflow systems.In this paper, a role-based workflow model with fuzzy optimized intelligent assignment is proposed and applied in the investment management system. A groupware-based software model is also proposed.

  20. Stress Assignment in Reading Italian Polysyllabic Pseudowords

    Science.gov (United States)

    Sulpizio, Simone; Arduino, Lisa S.; Paizi, Despina; Burani, Cristina

    2013-01-01

    In 4 naming experiments we investigated how Italian readers assign stress to pseudowords. We assessed whether participants assign stress following distributional information such as stress neighborhood (the proportion and number of existent words sharing orthographic ending and stress pattern) and whether such distributional information affects…

  1. Online Discussion Assignments Improve Students' Class Preparation

    Science.gov (United States)

    Lineweaver, Tara T.

    2010-01-01

    To increase the number of students who read the text before class and to promote student interaction centering on text material, I developed an online discussion assignment as a required component of a cognitive psychology course. Across 2 studies, this assignment had a limited effect on examination performance, but students completing online…

  2. A Model of Hierarchical Key Assignment Scheme

    Institute of Scientific and Technical Information of China (English)

    ZHANG Zhigang; ZHAO Jing; XU Maozhi

    2006-01-01

    A model of the hierarchical key assignment scheme is approached in this paper, which can be used with any cryptography algorithm. Besides, the optimal dynamic control property of a hierarchical key assignment scheme will be defined in this paper. Also, our scheme model will meet this property.

  3. Nitrogen-detected CAN and CON experiments as alternative experiments for main chain NMR resonance assignments.

    Science.gov (United States)

    Takeuchi, Koh; Heffron, Gregory; Sun, Zhen-Yu J; Frueh, Dominique P; Wagner, Gerhard

    2010-08-01

    Heteronuclear direct-detection experiments, which utilize the slower relaxation properties of low gamma nuclei, such as (13)C have recently been proposed for sequence-specific assignment and structural analyses of large, unstructured, and/or paramagnetic proteins. Here we present two novel (15)N direct-detection experiments. The CAN experiment sequentially connects amide (15)N resonances using (13)C(alpha) chemical shift matching, and the CON experiment connects the preceding (13)C' nuclei. When starting from the same carbon polarization, the intensities of nitrogen signals detected in the CAN or CON experiments would be expected four times lower than those of carbon resonances observed in the corresponding (13)C-detecting experiment, NCA-DIPAP or NCO-IPAP (Bermel et al. 2006b; Takeuchi et al. 2008). However, the disadvantage due to the lower gamma is counteracted by the slower (15)N transverse relaxation during detection, the possibility for more efficient decoupling in both dimensions, and relaxation optimized properties of the pulse sequences. As a result, the median S/N in the (15)N observe CAN experiment is 16% higher than in the (13)C observe NCA-DIPAP experiment. In addition, significantly higher sensitivity was observed for those residues that are hard to detect in the NCA-DIPAP experiment, such as Gly, Ser and residues with high-field C(alpha) resonances. Both CAN and CON experiments are able to detect Pro resonances that would not be observed in conventional proton-detected experiments. In addition, those experiments are free from problems of incomplete deuterium-to-proton back exchange in amide positions of perdeuterated proteins expressed in D(2)O. Thus, these features and the superior resolution of (15)N-detected experiments provide an attractive alternative for main chain assignments. The experiments are demonstrated with the small model protein GB1 at conditions simulating a 150 kDa protein, and the 52 kDa glutathione S-transferase dimer, GST.

  4. Airport Gate Assignment: New Model and Implementation

    CERN Document Server

    Li, Chendong

    2008-01-01

    Airport gate assignment is of great importance in airport operations. In this paper, we study the Airport Gate Assignment Problem (AGAP), propose a new model and implement the model with Optimization Programming language (OPL). With the objective to minimize the number of conflicts of any two adjacent aircrafts assigned to the same gate, we build a mathematical model with logical constraints and the binary constraints, which can provide an efficient evaluation criterion for the Airlines to estimate the current gate assignment. To illustrate the feasibility of the model we construct experiments with the data obtained from Continental Airlines, Houston Gorge Bush Intercontinental Airport IAH, which indicate that our model is both energetic and effective. Moreover, we interpret experimental results, which further demonstrate that our proposed model can provide a powerful tool for airline companies to estimate the efficiency of their current work of gate assignment.

  5. Efficient Credit Assignment through Evaluation Function Decomposition

    Science.gov (United States)

    Agogino, Adrian; Turner, Kagan; Mikkulainen, Risto

    2005-01-01

    Evolutionary methods are powerful tools in discovering solutions for difficult continuous tasks. When such a solution is encoded over multiple genes, a genetic algorithm faces the difficult credit assignment problem of evaluating how a single gene in a chromosome contributes to the full solution. Typically a single evaluation function is used for the entire chromosome, implicitly giving each gene in the chromosome the same evaluation. This method is inefficient because a gene will get credit for the contribution of all the other genes as well. Accurately measuring the fitness of individual genes in such a large search space requires many trials. This paper instead proposes turning this single complex search problem into a multi-agent search problem, where each agent has the simpler task of discovering a suitable gene. Gene-specific evaluation functions can then be created that have better theoretical properties than a single evaluation function over all genes. This method is tested in the difficult double-pole balancing problem, showing that agents using gene-specific evaluation functions can create a successful control policy in 20 percent fewer trials than the best existing genetic algorithms. The method is extended to more distributed problems, achieving 95 percent performance gains over tradition methods in the multi-rover domain.

  6. Potential protein post-translational modifications and find potential single amino acid substitutions in hepatitis B large envelope protein Posibles modificaciones postranslacionales y sustituciones de un solo aminoácido en la proteína grande de la superficie del virus de la hepatitis B

    OpenAIRE

    V Wiwanitkit

    2008-01-01

    Post-translational modifications of proteins control many biological processes. This is also important process in virus including hepatitis B. However, there is no in-depth study on the whole hepatitis B virus large envelope protein. In this work, potential protein post-translational modifications in hepatitis B virus large envelope protein were determined by a standard bioinformatics technique. Furthermore, potential single amino acid substitutions in hepatitis B large envelope protein were ...

  7. Inferring Protein Associations Using Protein Pulldown Assays

    Energy Technology Data Exchange (ETDEWEB)

    Sharp, Julia L.; Anderson, Kevin K.; Daly, Don S.; Auberry, Deanna L.; Borkowski, John J.; Cannon, William R.

    2007-02-01

    Background: One method to infer protein-protein associations is through a “bait-prey pulldown” assay using a protein affinity agent and an LC-MS (liquid chromatography-mass spectrometry)-based protein identification method. False positive and negative protein identifications are not uncommon, however, leading to incorrect inferences. Methods: A pulldown experiment generates a protein association matrix wherein each column represents a sample from one bait protein, each row represents one prey protein and each cell contains a presence/absence association indicator. Our method evaluates the presence/absence pattern across a prey protein (row) with a Likelihood Ratio Test (LRT), computing its p-value with simulated LRT test statistic distributions after a check with simulated binomial random variates disqualified the large sample 2 test. A pulldown experiment often involves hundreds of tests so we apply the false discovery rate method to control the false positive rate. Based on the p-value, each prey protein is assigned a category (specific association, non-specific association, or not associated) and appraised with respect to the pulldown experiment’s goal and design. The method is illustrated using a pulldown experiment investigating the protein complexes of Shewanella oneidensis MR-1. Results: The Monte Carlo simulated LRT p-values objectively reveal specific and ubiquitous prey, as well as potential systematic errors. The example analysis shows the results to be biologically sensible and more realistic than the ad hoc screening methods previously utilized. Conclusions: The method presented appears to be informative for screening for protein-protein associations.

  8. Ran GTPase-activating protein 1 is a therapeutic target in diffuse large B-cell lymphoma.

    Directory of Open Access Journals (Sweden)

    Kung-Chao Chang

    Full Text Available Lymphoma-specific biomarkers contribute to therapeutic strategies and the study of tumorigenesis. Diffuse large B-cell lymphoma (DLBCL is the most common type of malignant lymphoma. However, only 50% of patients experience long-term survival after current treatment; therefore, developing novel therapeutic strategies is warranted. Comparative proteomic analysis of two DLBCL lines with a B-lymphoblastoid cell line (LCL showed differential expression of Ran GTPase-activating protein 1 (RanGAP1 between them, which was confirmed using immunoblotting. Immunostaining showed that the majority of DLBCLs (92%, 46/50 were RanGAP1(+, while reactive lymphoid hyperplasia (n = 12 was RanGAP1(+ predominantly in germinal centers. RanGAP1 was also highly expressed in other B-cell lymphomas (BCL, n = 180 with brisk mitotic activity (B-lymphoblastic lymphoma/leukemia: 93%, and Burkitt lymphoma: 95% or cell-cycle dysregulation (mantle cell lymphoma: 83%, and Hodgkin's lymphoma 91%. Interestingly, serum RanGAP1 level was higher in patients with high-grade BCL (1.71 ± 2.28 ng/mL, n = 62 than in low-grade BCL (0.75 ± 2.12 ng/mL, n = 52 and healthy controls (0.55 ± 1.58 ng/mL, n = 75 (high-grade BCL vs. low-grade BCL, p = 0.002; high-grade BCL vs. control, p < 0.001, Mann-Whitney U test. In vitro, RNA interference of RanGAP1 showed no effect on LCL but enhanced DLBCL cell death (41% vs. 60%; p = 0.035 and cell-cycle arrest (G0/G1: 39% vs. 49%, G2/M: 19.0% vs. 7.5%; p = 0.030 along with decreased expression of TPX2 and Aurora kinases, the central regulators of mitotic cell division. Furthermore, ON 01910.Na (Estybon, a multikinase inhibitor induced cell death, mitotic cell arrest, and hyperphosphorylation of RanGAP1 in DLBCL cell lines but no effects in normal B and T cells. Therefore, RanGAP1 is a promising marker and therapeutic target for aggressive B-cell lymphoma, especially DLBCL.

  9. Sifting through genomes with iterative-sequence clustering produces a large, phylogenetically diverse protein-family resource

    Directory of Open Access Journals (Sweden)

    Sharpton Thomas J

    2012-10-01

    Full Text Available Abstract Background New computational resources are needed to manage the increasing volume of biological data from genome sequencing projects. One fundamental challenge is the ability to maintain a complete and current catalog of protein diversity. We developed a new approach for the identification of protein families that focuses on the rapid discovery of homologous protein sequences. Results We implemented fully automated and high-throughput procedures to de novo cluster proteins into families based upon global alignment similarity. Our approach employs an iterative clustering strategy in which homologs of known families are sifted out of the search for new families. The resulting reduction in computational complexity enables us to rapidly identify novel protein families found in new genomes and to perform efficient, automated updates that keep pace with genome sequencing. We refer to protein families identified through this approach as “Sifting Families,” or SFams. Our analysis of ~10.5 million protein sequences from 2,928 genomes identified 436,360 SFams, many of which are not represented in other protein family databases. We validated the quality of SFam clustering through statistical as well as network topology–based analyses. Conclusions We describe the rapid identification of SFams and demonstrate how they can be used to annotate genomes and metagenomes. The SFam database catalogs protein-family quality metrics, multiple sequence alignments, hidden Markov models, and phylogenetic trees. Our source code and database are publicly available and will be subject to frequent updates (http://edhar.genomecenter.ucdavis.edu/sifting_families/.

  10. Dietary protein intake and coronary heart disease in a large community based cohort: results from the Atherosclerosis Risk in Communities (ARIC study [corrected].

    Directory of Open Access Journals (Sweden)

    Bernhard Haring

    Full Text Available BACKGROUND: Prospective data examining the relationship between dietary protein intake and incident coronary heart disease (CHD are inconclusive. Most evidence is derived from homogenous populations such as health professionals. Large community-based analyses in more diverse samples are lacking. METHODS: We studied the association of protein type and major dietary protein sources and risk for incident CHD in 12,066 middle-aged adults (aged 45-64 at baseline, 1987-1989 from four U.S. communities enrolled in the Atherosclerosis Risk in Communities (ARIC Study who were free of diabetes mellitus and cardiovascular disease at baseline. Dietary protein intake was assessed at baseline and after 6 years of follow-up by food frequency questionnaire. Our primary outcome was adjudicated coronary heart disease events or deaths with following up through December 31, 2010. Cox proportional hazard models with multivariable adjustment were used for statistical analyses. RESULTS: During a median follow-up of 22 years, there were 1,147 CHD events. In multivariable analyses total, animal and vegetable protein were not associated with an increased risk for CHD before or after adjustment. In food group analyses of major dietary protein sources, protein intake from red and processed meat, dairy products, fish, nuts, eggs, and legumes were not significantly associated with CHD risk. The hazard ratios [with 95% confidence intervals] for risk of CHD across quintiles of protein from poultry were 1.00 [ref], 0.83 [0.70-0.99], 0.93 [0.75-1.15], 0.88 [0.73-1.06], 0.79 [0.64-0.98], P for trend  = 0.16. Replacement analyses evaluating the association of substituting one source of dietary protein for another or of decreasing protein intake at the expense of carbohydrates or total fats did not show any statistically significant association with CHD risk. CONCLUSION: Based on a large community cohort we found no overall relationship between protein type and major dietary

  11. 5 CFR 870.902 - Making an assignment.

    Science.gov (United States)

    2010-01-01

    ... assignment. (a) To assign insurance, an insured individual must complete an approved assignment form. Only the insured individual can make an assignment; no one can assign on behalf of an insured individual... 5 Administrative Personnel 2 2010-01-01 2010-01-01 false Making an assignment. 870.902 Section...

  12. The Assignment Game : The Reduced Game

    OpenAIRE

    Owen, Guillermo

    1992-01-01

    Let v be an assignment game. For a given reference payoff vector (x; y), and a coalition S, bargaining within the coalition can be represented by either the reduced game or the derived game. It is known that the reduced game need not be an assignment game (in fact, it need not be super additive) while the derived game is another assignment game, with modified reservation prices. We prove that, when the reference vector is in the core of the game, the derived game is the super additive cover o...

  13. MYC protein expression and genetic alterations have prognostic impact in patients with diffuse large B-cell lymphoma treated with immunochemotherapy

    Science.gov (United States)

    Valera, Alexandra; López-Guillermo, Armando; Cardesa-Salzmann, Teresa; Climent, Fina; González-Barca, Eva; Mercadal, Santiago; Espinosa, Íñigo; Novelli, Silvana; Briones, Javier; Mate, José L.; Salamero, Olga; Sancho, Juan M.; Arenillas, Leonor; Serrano, Sergi; Erill, Nadina; Martínez, Daniel; Castillo, Paola; Rovira, Jordina; Martínez, Antonio; Campo, Elias; Colomo, Luis

    2013-01-01

    MYC alterations influence the survival of patients with diffuse large B-cell lymphoma. Most studies have focused on MYC translocations but there is little information regarding the impact of numerical alterations and protein expression. We analyzed the genetic alterations and protein expression of MYC, BCL2, BCL6, and MALT1 in 219 cases of diffuse large B-cell lymphoma. MYC rearrangement occurred as the sole abnormality (MYC single-hit) in 3% of cases, MYC and concurrent BCL2 and/or BCL6 rearrangements (MYC double/triple-hit) in 4%, MYC amplifications in 2% and MYC gains in 19%. MYC single-hit, MYC double/triple-hit and MYC amplifications, but not MYC gains or other gene rearrangements, were associated with unfavorable progression-free survival and overall survival. MYC protein expression, evaluated using computerized image analysis, captured the unfavorable prognosis of MYC translocations/amplifications and identified an additional subset of patients without gene alterations but with similar poor prognosis. Patients with tumors expressing both MYC/BCL2 had the worst prognosis, whereas those with double-negative tumors had the best outcome. High MYC expression was associated with shorter overall survival irrespectively of the International Prognostic Index and BCL2 expression. In conclusion, MYC protein expression identifies a subset of diffuse large B-cell lymphoma with very poor prognosis independently of gene alterations and other prognostic parameters. PMID:23716551

  14. Use of the myosin motor domain as large-affinity tag for the expression and purification of proteins in Dictyostelium discoideum.

    Science.gov (United States)

    Kollmar, Martin

    2006-08-15

    The cellular slime mold Dictyostelium discoideum is increasingly be used for the overexpression of proteins. Dictyostelium is amenable to classical and molecular genetic approaches and can easily be grown in large quantities. It contains a variety of chaperones and folding enzymes, and is able to perform all kinds of post-translational protein modifications. Here, new expression vectors are presented that have been designed for the production of proteins in large quantities for biochemical and structural studies. The expression cassettes of the most successful vectors are based on a tandem affinity purification tag consisting of an octahistidine tag followed by the myosin motor domain tag. The myosin motor domain not only strongly enhances the production of fused proteins but is also used for a fast affinity purification step through its ATP-dependent binding to actin. The applicability of the new system has been demonstrated for the expression and purification of subunits of the dynein-dynactin motor protein complex from different species. PMID:16516959

  15. Studies on the Coordination of Ribosomal Protein Assembly Events Involved in Processing and Stabilization of Yeast Early Large Ribosomal Subunit Precursors.

    Directory of Open Access Journals (Sweden)

    Uli Ohmayer

    Full Text Available Cellular production of ribosomes involves the formation of highly defined interactions between ribosomal proteins (r-proteins and ribosomal RNAs (rRNAs. Moreover in eukaryotic cells, efficient ribosome maturation requires the transient association of a large number of ribosome biogenesis factors (RBFs with newly forming ribosomal subunits. Here, we investigated how r-protein assembly events in the large ribosomal subunit (LSU rRNA domain II are coordinated with each other and with the association of RBFs in early LSU precursors of the yeast Saccharomyces cerevisiae. Specific effects on the pre-ribosomal association of RBFs could be observed in yeast mutants blocked in LSU rRNA domain II assembly. Moreover, formation of a cluster of r-proteins was identified as a downstream event in LSU rRNA domain II assembly. We analyzed in more detail the functional relevance of eukaryote specific bridges established by this r-protein cluster between LSU rRNA domain II and VI and discuss how they can support the stabilization and efficient processing of yeast early LSU precursor RNAs.

  16. Single-Molecule Methods for the Large-Scale Characterization of Expression Levels and Protein-Protein Interactions in Shewanella Oneidensis MR-1

    Energy Technology Data Exchange (ETDEWEB)

    Weiss, Shimon; Michalet, Xavier

    2008-10-01

    This project has demonstrated a new approach to localize binding sites of proteins regulating gene expression (also known as transcription factors) on the genome of bacteria. Knowledge of the precise binding site(s) of a specific transcription factor helps determining its role in the cell cycle and by extension provides further understanding of the mechanisms at play in the organism. The approach entails labeling transcription factors (or any other DNA-binding protein of interest) with quantum dots, a new class of very bright fluorescent probes, which allow detection of individual molecules with a simple microscope. Detection is then followed with very accurate localization of the probe (with nanometer resolution) with respect to specific parts of the DNA or other proteins bound to the DNA. We have confirmed the precision of our measurement using another technique based on atomic force microscopy, which provides a nanometer-resolution topographic picture of a sample. Quantum dots and DNA are readily observable (and distinguishable) in the atomic force microscope, and can be simultaneously observed by fluorescence microscopy, allowing a direct comparison of the two methods. Precise nanometer-localization of protein binding sites using fluorescent quantum dots is thus a direct and visual method for physical mapping of transcription factor binding sites on whole genomes.

  17. A new routing assignment model for Internet

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    This paper builds a new traffic assignment model for Internet. The routing policy based on this model can realize load splitting in a network automatically, which overcomes the shortcoming of OSPF and satisfies the load balancing requirement of IETF.

  18. Crosstalk-Aware Routing Resource Assignment

    Institute of Scientific and Technical Information of China (English)

    Hai-Long Yao; Yi-Ci Cai; Qiang Zhou; Xian-Long Hong

    2005-01-01

    Crosstalk noise is one of the emerging issues in deep sub-micrometer technology which causes many undesired effects on the circuit performance. In this paper, a Crosstalk-Aware Routing Resource Assignment (CARRA) algorithm is proposed, which integrates the routing layers and tracks to address the crosstalk noise issue during the track/layer assignment stage. The CARRA problem is formulated as a weighted bipartite matching problem and solved using the linear assignment algorithm. The crosstalk risks between nets are represented by an undirected graph and the maximum number of the concurrent crosstalk risking nets is computed as the max clique of the graph. Then the nets in each max clique are assigned to disadjacent tracks. Thus the crosstalk noise can be avoided based on the clique concept. The algorithm is tested on IBM benchmarks and the experimental results show that it can improve the final routing layout a lot with little loss of the completion rate.

  19. Inactivation of pRB-related proteins p130 and p107 mediated by the J domain of simian virus 40 large T antigen.

    OpenAIRE

    Stubdal, H; Zalvide, J; Campbell, K S; Schweitzer, C; Roberts, T.M.; DeCaprio, J A

    1997-01-01

    Inactivation of the retinoblastoma tumor suppressor protein (pRB) contributes to tumorigenesis in a wide variety of cancers. In contrast, the role of the two pRB-related proteins, p130 and p107, in oncogenic transformation is unclear. The LXCXE domain of simian virus 40 large T antigen (TAg) specifically binds to pRB, p107, and p130. We have previously shown that the N terminus and the LXCXE domain of TAg cooperate to alter the phosphorylation state of p130 and p107. Here, we demonstrate that...

  20. A gene pair from the human major histocompatibility complex encodes large proline-rich proteins with multiple repeated motifs and a single ubiquitin-like domain.

    OpenAIRE

    Banerji, J.; Sands, J; Strominger, J.L.; Spies, T

    1990-01-01

    A large number of genes has been identified previously between the class I and class II gene families within the class III region of the human major histocompatibility complex. The complete sequences of two of these genes, BAT2 and BAT3 (where BAT is HLA-B-associated transcript), which are closely linked, were determined from cDNA clones. The putative BAT2 and BAT3 proteins are 228 and 110 kDa, respectively, and do not appear to be members of any known family of proteins. However, BAT3 contai...

  1. Pd(0)-Catalyzed Direct C-H Functionalization of 2-H-4-Benzylidene Imidazolones: Friendly and Large-Scale Access to GFP and Kaede Protein Fluorophores.

    Science.gov (United States)

    Muselli, Mickaël; Baudequin, Christine; Perrio, Cécile; Hoarau, Christophe; Bischoff, Laurent

    2016-04-11

    The first one-pot synthesis of N-substituted 2-H-4-benzylidene imidazolones and their subsequent palladium-catalyzed and copper-assisted direct C2-H arylation and alkenylation with aryl- and alkenylhalides are described. This innovative synthesis is step-economical, azide-free, high yielding, highly flexible in the introduction of a variety of electronically different groups, and can be operated on large-scale. Moreover, the method allows direct access to C2-arylated or alkenylated imidazolone-based green fluorescent protein (GFP) and Kaede protein fluorophores, including ortho-hydroxylated models. PMID:26960963

  2. Hierarchical method of task assignment for multiple cooperating UAV teams

    Institute of Scientific and Technical Information of China (English)

    Xiaoxuan Hu; Huawei Ma; Qingsong Ye; He Luo

    2015-01-01

    The problem of task assignment for multiple cooperat-ing unmanned aerial vehicle (UAV) teams is considered. Multiple UAVs forming several smal teams are needed to perform attack tasks on a set of predetermined ground targets. A hierarchical task assignment method is presented to address the problem. It breaks the original problem down to three levels of sub-problems: tar-get clustering, cluster al ocation and target assignment. The first two sub-problems are central y solved by using clustering algo-rithms and integer linear programming, respectively, and the third sub-problem is solved in a distributed and paral el manner, using a mixed integer linear programming model and an improved ant colony algorithm. The proposed hierarchical method can reduce the computational complexity of the task assignment problem con-siderably, especial y when the number of tasks or the number of UAVs is large. Experimental results show that this method is feasi-ble and more efficient than non-hierarchical methods.

  3. Using Magnets and Classroom Flipping to Promote Student Engagement and Learning about Protein Translation in a Large Microbiology Class

    Directory of Open Access Journals (Sweden)

    Jennifer Lynn McLean

    2016-05-01

    Full Text Available It is generally accepted within the education community that active learning is superior to traditional lecturing alone. Many science educators, however, are reluctant to give up classroom time for activities because they fear that they will not have time to cover as much content. Classroom flipping has been gaining momentum in higher education as one way to engage students in the classroom while still exposing students to the same volume of course content. The activity presented here demonstrates how flipping one lecture period can be used in conjunction with an engaging in-class activity to teach a concept that is often difficult for students to learn through lecture alone. Specifically, we asked students to view a lecture video on bacterial protein translation before coming to class. We then used the classroom period to conduct a hands-on activity that allowed students to interact with magnetic pieces representing the components of protein translation to generate a protein from a given piece of DNA. Survey data showed that students liked the flipped classroom format associated with this activity, but they would not want every class flipped, and they perceived that the hands-on protein translation activity helped them to learn the material. Preliminary summative assessment data showed that this activity may have been useful in helping students to achieve the fundamental learning outcome that students will be able to translate a protein from a given piece of bacterial DNA.

  4. Extending the eNOE data set of large proteins by evaluation of NOEs with unresolved diagonals

    Energy Technology Data Exchange (ETDEWEB)

    Chi, Celestine N.; Strotz, Dean; Riek, Roland, E-mail: roland.riek@phys.chem.ethz.ch; Vögeli, Beat, E-mail: beat.voegeli@phys.chem.ethz.ch [Swiss Federal Institute of Technology, Laboratory of Physical Chemistry, Vladimir-Prelog Weg 2 (Switzerland)

    2015-05-15

    The representation of a protein’s spatial sampling at atomic resolution is fundamental for understanding its function. NMR has been established as the best-suited technique toward this goal for small proteins. However, the accessible information content rapidly deteriorates with increasing protein size. We have recently demonstrated that for small proteins distance restraints with an accuracy smaller than 0.1 Å can be obtained by replacing traditional semi-quantitative Nuclear Overhauser Effects (NOEs) with exact NOEs (eNOE). The high quality of the data allowed us to calculate structural ensembles of the small model protein GB3 consisting of multiple rather than a single state. The analysis has been limited to small proteins because NOEs of spins with unresolved diagonal peaks cannot be used. Here we propose a simple approach to translate such NOEs into correct upper distance restraints, which opens access to larger biomolecules. We demonstrate that for 16 kDa cyclophilin A the collection of such restraints extends the original 1254 eNOEs to 3471.

  5. Competitive Traffic Assignment in Road Networks

    Directory of Open Access Journals (Sweden)

    Krylatov Alexander Y.

    2016-09-01

    Full Text Available Recently in-vehicle route guidance and information systems are rapidly developing. Such systems are expected to reduce congestion in an urban traffic area. This social benefit is believed to be reached by imposing the route choices on the network users that lead to the system optimum traffic assignment. However, guidance service could be offered by different competitive business companies. Then route choices of different mutually independent groups of users may reject traffic assignment from the system optimum state. In this paper, a game theoretic approach is shown to be very efficient to formalize competitive traffic assignment problem with various groups of users in the form of non-cooperative network game with the Nash equilibrium search. The relationships between the Wardrop’s system optimum associated with the traffic assignment problem and the Nash equilibrium associated with the competitive traffic assignment problem are investigated. Moreover, some related aspects of the Nash equilibrium and the Wardrop’s user equilibrium assignments are also discussed.

  6. Large-scale structure of RecA protein from Deinococcus radiodurance and its complexes in solution

    Science.gov (United States)

    Karelov, D. V.; Lebedev, D. V.; Suslov, A. V.; Shalguev, V. I.; Kuklin, A. I.; Islamov, A. Kh; Lauter, H.; Lanzov, V. A.; Isaev-Ivanov, V. V.

    2008-03-01

    Different conformational states of the filaments formed by RecA protein from a radiation resistant strain Deinococcus radiodurance (RecADr) in solution were investigated using small angle neutron scattering. Scattering by the protein self-polymer was consistent with a long helix model, with the pitch of the helix being lower than that in the crystal structure. Compared to those of RecA proteins from Escherichia coli and Pseudomonas aeruginosa, helical filaments of RecA from D. radiodurance exhibited a lower helical pitch and lower stability at low Mg2+ concentrations or under conditions of elevated ionic strength in the absence of ATP (adenosine triphosphate). Formation of an active filament upon binding of ATPγS and either single- or double-stranded DNA brought about a significant increase in the helix pitch and a moderate decrease in the cross-sectional gyration radius, but resulted in little change in the number of monomers per helix turn. The helix pitch value of the RecADr presynaptic complex was conservative and close to that found for other RecA proteins and their analogs.

  7. Flexible taxonomic assignment of ambiguous sequencing reads

    Directory of Open Access Journals (Sweden)

    Jansson Jesper

    2011-01-01

    Full Text Available Abstract Background To characterize the diversity of bacterial populations in metagenomic studies, sequencing reads need to be accurately assigned to taxonomic units in a given reference taxonomy. Reads that cannot be reliably assigned to a unique leaf in the taxonomy (ambiguous reads are typically assigned to the lowest common ancestor of the set of species that match it. This introduces a potentially severe error in the estimation of bacteria present in the sample due to false positives, since all species in the subtree rooted at the ancestor are implicitly assigned to the read even though many of them may not match it. Results We present a method that maps each read to a node in the taxonomy that minimizes a penalty score while balancing the relevance of precision and recall in the assignment through a parameter q. This mapping can be obtained in time linear in the number of matching sequences, because LCA queries to the reference taxonomy take constant time. When applied to six different metagenomic datasets, our algorithm produces different taxonomic distributions depending on whether coverage or precision is maximized. Including information on the quality of the reads reduces the number of unassigned reads but increases the number of ambiguous reads, stressing the relevance of our method. Finally, two measures of performance are described and results with a set of artificially generated datasets are discussed. Conclusions The assignment strategy of sequencing reads introduced in this paper is a versatile and a quick method to study bacterial communities. The bacterial composition of the analyzed samples can vary significantly depending on how ambiguous reads are assigned depending on the value of the q parameter. Validation of our results in an artificial dataset confirm that a combination of values of q produces the most accurate results.

  8. Protein crystal growth in microgravity review of large scale temperature induction method: Bovine insulin, human insulin and human α-interferon

    International Nuclear Information System (INIS)

    The Protein Crystal Growth Facility (PCF) is space-flight hardware that accommodates large scale protein crystal growth experiments using temperature change as the inductive step. Recent modifications include specialized instrumentation for monitoring crystal nucleation with laser light scattering. This paper reviews results from its first seven flights on the Space Shuttle, the last with laser light scattering instrumentation in place. The PCF's objective is twofold: (1) the production of high quality protein crystals for x-ray analysis and subsequent structure-based drug design and (2) preparation of a large quantity of relatively contaminant free crystals for use as time-release protein pharmaceuticals. The first three Shuttle flights with bovine insulin constituted the PCF's proof of concept, demonstrating that the space-grown crystals were larger and diffracted to higher resolution than their earth-grown counterparts. The later four PCF missions were used to grow recombinant human insulin crystals for x-ray analysis and continue productions trials aimed at the development of a processing facility for crystalline recombinant a-interferon

  9. Protein crystal growth in microgravity review of large scale temperature induction method: Bovine insulin, human insulin and human α-interferon

    Science.gov (United States)

    Long, Marianna M.; Bishop, John Bradford; Delucas, Lawrence J.; Nagabhushan, Tattanhalli L.; Reichert, Paul; Smith, G. David

    1997-01-01

    The Protein Crystal Growth Facility (PCF) is space-flight hardware that accommodates large scale protein crystal growth experiments using temperature change as the inductive step. Recent modifications include specialized instrumentation for monitoring crystal nucleation with laser light scattering. This paper reviews results from its first seven flights on the Space Shuttle, the last with laser light scattering instrumentation in place. The PCF's objective is twofold: (1) the production of high quality protein crystals for x-ray analysis and subsequent structure-based drug design and (2) preparation of a large quantity of relatively contaminant free crystals for use as time-release protein pharmaceuticals. The first three Shuttle flights with bovine insulin constituted the PCF's proof of concept, demonstrating that the space-grown crystals were larger and diffracted to higher resolution than their earth-grown counterparts. The later four PCF missions were used to grow recombinant human insulin crystals for x-ray analysis and continue productions trials aimed at the development of a processing facility for crystalline recombinant a-interferon.

  10. Application of a sensitive collection heuristic for very large protein families: Evolutionary relationship between adipose triglyceride lipase (ATGL and classic mammalian lipases

    Directory of Open Access Journals (Sweden)

    Berezovsky Igor

    2006-03-01

    Full Text Available Abstract Background Manually finding subtle yet statistically significant links to distantly related homologues becomes practically impossible for very populated protein families due to the sheer number of similarity searches to be invoked and analyzed. The unclear evolutionary relationship between classical mammalian lipases and the recently discovered human adipose triglyceride lipase (ATGL; a patatin family member is an exemplary case for such a problem. Results We describe an unsupervised, sensitive sequence segment collection heuristic suitable for assembling very large protein families. It is based on fan-like expanding, iterative database searches. To prevent inclusion of unrelated hits, additional criteria are introduced: minimal alignment length and overlap with starting sequence segments, finding starting sequences in reciprocal searches, automated filtering for compositional bias and repetitive patterns. This heuristic was implemented as FAMILYSEARCHER in the ANNIE sequence analysis environment and applied to search for protein links between the classical lipase family and the patatin-like group. Conclusion The FAMILYSEARCHER is an efficient tool for tracing distant evolutionary relationships involving large protein families. Although classical lipases and ATGL have no obvious sequence similarity and differ with regard to fold and catalytic mechanism, homology links detected with FAMILYSEARCHER show that they are evolutionarily related. The conserved sequence parts can be narrowed down to an ancestral core module consisting of three β-strands, one α-helix and a turn containing the typical nucleophilic serine. Moreover, this ancestral module also appears in numerous enzymes with various substrate specificities, but that critically rely on nucleophilic attack mechanisms.

  11. Superfamily assignments for the yeast proteome through integration of structure prediction with the gene ontology.

    Directory of Open Access Journals (Sweden)

    Lars Malmström

    2007-04-01

    Full Text Available Saccharomyces cerevisiae is one of the best-studied model organisms, yet the three-dimensional structure and molecular function of many yeast proteins remain unknown. Yeast proteins were parsed into 14,934 domains, and those lacking sequence similarity to proteins of known structure were folded using the Rosetta de novo structure prediction method on the World Community Grid. This structural data was integrated with process, component, and function annotations from the Saccharomyces Genome Database to assign yeast protein domains to SCOP superfamilies using a simple Bayesian approach. We have predicted the structure of 3,338 putative domains and assigned SCOP superfamily annotations to 581 of them. We have also assigned structural annotations to 7,094 predicted domains based on fold recognition and homology modeling methods. The domain predictions and structural information are available in an online database at http://rd.plos.org/10.1371_journal.pbio.0050076_01.

  12. Unifying Temporal and Structural Credit Assignment Problems

    Science.gov (United States)

    Agogino, Adrian K.; Tumer, Kagan

    2004-01-01

    Single-agent reinforcement learners in time-extended domains and multi-agent systems share a common dilemma known as the credit assignment problem. Multi-agent systems have the structural credit assignment problem of determining the contributions of a particular agent to a common task. Instead, time-extended single-agent systems have the temporal credit assignment problem of determining the contribution of a particular action to the quality of the full sequence of actions. Traditionally these two problems are considered different and are handled in separate ways. In this article we show how these two forms of the credit assignment problem are equivalent. In this unified frame-work, a single-agent Markov decision process can be broken down into a single-time-step multi-agent process. Furthermore we show that Monte-Carlo estimation or Q-learning (depending on whether the values of resulting actions in the episode are known at the time of learning) are equivalent to different agent utility functions in a multi-agent system. This equivalence shows how an often neglected issue in multi-agent systems is equivalent to a well-known deficiency in multi-time-step learning and lays the basis for solving time-extended multi-agent problems, where both credit assignment problems are present.

  13. Definition of MYC genetic heteroclonality in diffuse large B-cell lymphoma with 8q24 rearrangement and its impact on protein expression.

    Science.gov (United States)

    Valera, Alexandra; Epistolio, Samantha; Colomo, Lluis; Riva, Alice; Balagué, Olga; Dlouhy, Ivan; Tzankov, Alexandar; Bühler, Marco; Haralambieva, Eugenia; Campo, Elias; Soldini, Davide; Mazzucchelli, Luca; Martin, Vittoria

    2016-08-01

    MYC rearrangement can be detected in a subgroup of diffuse large B-cell lymphoma characterized by unfavorable prognosis. In contrast to Burkitt lymphoma, the correlation between MYC rearrangement and MYC protein expression in diffuse large B-cell lymphoma is less clear, as approximately one-third of rearranged cases show negative or low expression by immunohistochemistry. To better understand whether specific characteristics of the MYC rearrangement may influence its protein expression, we investigated 43 de novo diffuse large B-cell lymphoma positive for 8q24 rearrangement by FISH, using 14 Burkitt lymphoma for comparison. Different cell populations (clones), breakpoints (classical vs non-classical FISH patterns), partner genes (IGH vs non-IGH) and immunostaining were detected and analyzed using computerized image systems. In a subgroup of diffuse large B-cell lymphoma, we observed different clones within the same tumor distinguishing the founder clone with MYC rearrangement alone from other subclones, carrying MYC rearrangement coupled with loss/extra copies of derivatives/normal alleles. This picture, which we defined MYC genetic heteroclonality, was found in 42% of cases and correlated to negative MYC expression (P=0.026). Non-classical FISH breakpoints were detected in 16% of diffuse large B-cell lymphoma without affecting expression (P=0.040). Non-IGH gene was the preferential partner of rearrangement in those diffuse large B-cell lymphoma showing MYC heteroclonality (P=0.016) and/or non-classical FISH breakpoints (P=0.058). MYC heteroclonality was not observed in Burkitt lymphoma and all cases had positive MYC expression. Non-classical FISH MYC breakpoint and non-IGH partner were found in 29 and 20% of Burkitt lymphoma, respectively. In conclusion, MYC genetic heteroclonality is a frequent event in diffuse large B-cell lymphoma and may have a relevant role in modulating MYC expression. PMID:27125356

  14. Large-scale analysis of protein expression changes in human keratinocytes immortalized by human papilloma virus type 16 E6 and E7 oncogenes

    Directory of Open Access Journals (Sweden)

    Arnouk Hilal

    2009-08-01

    Full Text Available Abstract Background Infection with high-risk type human papilloma viruses (HPVs is associated with cervical carcinomas and with a subset of head and neck squamous cell carcinomas. Viral E6 and E7 oncogenes cooperate to achieve cell immortalization by a mechanism that is not yet fully understood. Here, human keratinocytes were immortalized by long-term expression of HPV type 16 E6 or E7 oncoproteins, or both. Proteomic profiling was used to compare expression levels for 741 discrete protein features. Results Six replicate measurements were performed for each group using two-dimensional difference gel electrophoresis (2D-DIGE. The median within-group coefficient of variation was 19–21%. Significance of between-group differences was tested based on Significance Analysis of Microarray and fold change. Expression of 170 (23% of the protein features changed significantly in immortalized cells compared to primary keratinocytes. Most of these changes were qualitatively similar in cells immortalized by E6, E7, or E6/7 expression, indicating convergence on a common phenotype, but fifteen proteins (~2% were outliers in this regulatory pattern. Ten demonstrated opposite regulation in E6- and E7-expressing cells, including the cell cycle regulator p16INK4a; the carbohydrate binding protein Galectin-7; two differentially migrating forms of the intermediate filament protein Cytokeratin-7; HSPA1A (Hsp70-1; and five unidentified proteins. Five others had a pattern of expression that suggested cooperativity between the co-expressed oncoproteins. Two of these were identified as forms of the small heat shock protein HSPB1 (Hsp27. Conclusion This large-scale analysis provides a framework for understanding the cooperation between E6 and E7 oncoproteins in HPV-driven carcinogenesis.

  15. Competition assignment problem algorithm based on Hungarian method

    Institute of Scientific and Technical Information of China (English)

    KONG Chao; REN Yongtai; GE Huiling; DENG Hualing

    2007-01-01

    Traditional Hungarian method can only solve standard assignment problems, while can not solve competition assignment problems. This article emphatically discussed the difference between standard assignment problems and competition assignment problems. The kinds of competition assignment problem algorithms based on Hungarian method and the solutions of them were studied.

  16. Assignment of fields from particles to mesh

    CERN Document Server

    Duque, Daniel

    2016-01-01

    In Computational Fluid Dynamics there have been many attempts to combine the power of a fixed mesh on which to carry out spatial calculations with that of a set of particles that moves following the velocity field. These ideas indeed go back to Particle-in-Cell methods, proposed about 60 years ago. Of course, some procedure is needed to transfer field information between particles and mesh. There are many possible choices for this "assignment", or "projection". Several requirements may guide this choice. Two well-known ones are conservativity and stability, which apply to volume integrals of the fields. An additional one is here considered: preservation of information. This means that mesh interpolation, followed by mesh assignment, should leave the field values invariant. The resulting methods are termed "mass" assignments due to their strong similarities with the Finite Element Method. We test several procedures, including the well-known FLIP, on three scenarios: simple 1D convection, 2D convection of Zales...

  17. Medicare Accountable Care Organizations: Beneficiary Assignment Update.

    Science.gov (United States)

    Vaughn, Thomas; MacKinney, A Clinton; Mueller, Keith J; Ullrich, Fred; Zhu, Xi

    2016-06-01

    This brief updates Brief No. 2014-3 and explains changes in the Centers for Medicare & Medicaid Services (CMS) Accountable Care Organization (ACO) regulations issued in June 2015 pertaining to beneficiary assignment for Medicare Shared Savings Program ACOs. Overall, the regulatory changes are intended to (1) encourage ACOs to participate in two-sided risk contracts, (2) increase the likelihood that beneficiaries are assigned to the physician (and ACO) from whom they receive most of their primary care services, and (3) make it easier for Federally Qualified Health Centers (FQHCs) and Rural Health Clinics (RHCs) to participate in ACOs. Understanding ACO beneficiary assignment policies is critical for ACO in managing their panel of ACO providers and beneficiaries. PMID:27416650

  18. From course assignment paper to publishable manuscript.

    Science.gov (United States)

    Rew, Lynn

    2012-12-01

    Both undergraduate and graduate nursing students are expected to write numerous papers in their educational programs; however, most of these papers are never published. Many students and faculty lack the skills needed to convert a course assignment paper to a publishable manuscript. The purpose of this article is to describe 10 steps that can transform a course assignment paper into a publishable manuscript. These steps include outlining, clarifying the topic, clearly stating the purpose, identifying an appropriate audience, revising with faculty's feedback, querying journal editors, revising to conform to journal's author guidelines, requesting and responding to peer feedback, and finally editing and proofreading prior to submitting the manuscript. Faculty members are encouraged to make writing assignments that students can then convert to publishable manuscripts. Such publications form an essential cornerstone of professional holistic nursing.

  19. Medicare Accountable Care Organizations: Beneficiary Assignment Update.

    Science.gov (United States)

    Vaughn, Thomas; MacKinney, A Clinton; Mueller, Keith J; Ullrich, Fred; Zhu, Xi

    2016-06-01

    This brief updates Brief No. 2014-3 and explains changes in the Centers for Medicare & Medicaid Services (CMS) Accountable Care Organization (ACO) regulations issued in June 2015 pertaining to beneficiary assignment for Medicare Shared Savings Program ACOs. Overall, the regulatory changes are intended to (1) encourage ACOs to participate in two-sided risk contracts, (2) increase the likelihood that beneficiaries are assigned to the physician (and ACO) from whom they receive most of their primary care services, and (3) make it easier for Federally Qualified Health Centers (FQHCs) and Rural Health Clinics (RHCs) to participate in ACOs. Understanding ACO beneficiary assignment policies is critical for ACO in managing their panel of ACO providers and beneficiaries.

  20. Writing Assignments that Promote Active Learning

    Science.gov (United States)

    Narayanan, M.

    2014-12-01

    Encourage students to write a detailed, analytical report correlating classroom discussions to an important historical event or a current event. Motivate students interview an expert from industry on a topic that was discussed in class. Ask the students to submit a report with supporting sketches, drawings, circuit diagrams and graphs. Propose that the students generate a complete a set of reading responses pertaining to an assigned topic. Require each student to bring in one comment or one question about an assigned reading. The assignment should be a recent publication in an appropriate journal. Have the students conduct a web search on an assigned topic. Ask them to generate a set of ideas that can relate to classroom discussions. Provide the students with a study guide. The study guide should provide about 10 or 15 short topics. Quiz the students on one or two of the topics. Encourage the students to design or develop some creative real-world examples based on a chapter discussed or a topic of interest. Require that students originate, develop, support and defend a viewpoint using a specifically assigned material. Make the students practice using or utilizing a set of new technical terms they have encountered in an assigned chapter. Have students develop original examples explaining the different terms. Ask the students to select one important terminology from the previous classroom discussions. Encourage the students to explain why they selected that particular word. Ask them to talk about the importance of the terminology from the point of view of their educational objectives and future career. Angelo, T. A. (1991). Ten easy pieces: Assessing higher learning in four dimensions. In T. A. Angelo (Ed.), Classroom research: Early lessons from success (pp. 17-31). New Directions for Teaching and Learning, No. 46. San Francisco: Jossey-Bass.

  1. Expressions of tight junction proteins Occludin and Claudin-1 are under the circadian control in the mouse large intestine: implications in intestinal permeability and susceptibility to colitis.

    Directory of Open Access Journals (Sweden)

    Oh-oka Kyoko

    Full Text Available BACKGROUND & AIMS: The circadian clock drives daily rhythms in behavior and physiology. A recent study suggests that intestinal permeability is also under control of the circadian clock. However, the precise mechanisms remain largely unknown. Because intestinal permeability depends on tight junction (TJ that regulates the epithelial paracellular pathway, this study investigated whether the circadian clock regulates the expression levels of TJ proteins in the intestine. METHODS: The expression levels of TJ proteins in the large intestinal epithelium and colonic permeability were analyzed every 4, 6, or 12 hours between wild-type mice and mice with a mutation of a key clock gene Period2 (Per2; mPer2(m/m. In addition, the susceptibility to dextran sodium sulfate (DSS-induced colitis was compared between wild-type mice and mPer2(m/m mice. RESULTS: The mRNA and protein expression levels of Occludin and Claudin-1 exhibited daily variations in the colonic epithelium in wild-type mice, whereas they were constitutively high in mPer2(m/m mice. Colonic permeability in wild-type mice exhibited daily variations, which was inversely associated with the expression levels of Occludin and Claudin-1 proteins, whereas it was constitutively low in mPer2(m/m mice. mPer2(m/m mice were more resistant to the colonic injury induced by DSS than wild-type mice. CONCLUSIONS: Occludin and Claudin-1 expressions in the large intestine are under the circadian control, which is associated with temporal regulation of colonic permeability and also susceptibility to colitis.

  2. Tableaux for Dynamic Logic of Propositional Assignments

    OpenAIRE

    Lima, Tiago; Herzig, Andreas

    2014-01-01

    The Dynamic Logic for Propositional Assignments (DL-PA) has recently been studied as an alternative to Propositional Dynamic Logic (PDL). In DL-PA, the abstract atomic programs of PDL are replaced by assignments of propositional variables to truth values. This makes DL-PA enjoy some interesting meta-logical properties that PDL does not, such as eliminability of the Kleene star, compactness and interpolation. We define and analytic tableaux calculus for DL-PA and show that it matches the known...

  3. Evolving Networks with Nonlinear Assignment of Weight

    Institute of Scientific and Technical Information of China (English)

    TANG Chao; TANG Yi

    2006-01-01

    We propose a weighted evolving network model in which the underlying topological structure is still driven by the degree according to the preferential attachment rule while the weight assigned to the newly established edges is dependent on the degree in a nonlinear form. By varying the parameter α that controls the function determining the assignment of weight, a wide variety of power-law behaviours of the total weight distributions as well as the diversity of the weight distributions of edges are displayed. Variation of correlation and heterogeneity in the network is illustrated as well.

  4. Experimenting with Request Assignment Simulator (RAS

    Directory of Open Access Journals (Sweden)

    R. Arokia Paul Rajan

    2014-11-01

    Full Text Available There is no existence of dedicated simulators on the Internet that studies the impact of load balancing principles of the cloud architectures. Request Assignment Simulator (RAS is a customizable, visual tool that helps to understand the request assignment to the resources based on the load balancing principles. We have designed this simulator to fit into Infrastructure as a Service (IaaS cloud model. In this paper, we present a working manual useful for the conduct of experiment with RAS. The objective of this paper is to instill the user to understand the pertinent parameters in the cloud, their metrics, load balancing principles, and their impact on the performance.

  5. 48 CFR 211.274-5 - Policy for assignment of Government-assigned serial numbers.

    Science.gov (United States)

    2010-10-01

    ... 48 Federal Acquisition Regulations System 3 2010-10-01 2010-10-01 false Policy for assignment of... DEFENSE ACQUISITION REGULATIONS SYSTEM, DEPARTMENT OF DEFENSE ACQUISITION PLANNING DESCRIBING AGENCY NEEDS Using and Maintaining Requirements Documents 211.274-5 Policy for assignment of...

  6. 75 FR 55352 - Delegation of Authorities and Assignment of Responsibilities

    Science.gov (United States)

    2010-09-10

    ... of the Secretary Delegation of Authorities and Assignment of Responsibilities Secretary's Order 5-2010 Subject: Delegation of Authorities and Assignment of Responsibilities to the Administrator, Wage and Hour Division. 1. Purpose. To delegate authorities and assign responsibilities to...

  7. Reliable and inexpensive expression of large, tagged, exogenous proteins in murine bone marrow-derived macrophages using a second generation lentiviral system

    Directory of Open Access Journals (Sweden)

    Matthew R. Miller

    2015-08-01

    Full Text Available Over the past two decades, researchers have struggled to efficiently express foreign DNA in primary macrophages, impeding research progress. The applications of lipofection, electroporation, microinjection, and viral-mediated transfer typically result in disruptions in macrophage differentiation and function, low expression levels of exogenous proteins, limited efficiency and high cell mortality. In this report, after extensive optimization, we present a method of expressing large tagged proteins at high efficiency, consistency, and low cost using lentiviral infection. This method utilizes laboratory-propagated second generation plasmids to produce efficient virus that can be stored for later use. The expression of proteins up to 150 kDa in size is achieved in 30–70% of cells while maintaining normal macrophage differentiation and morphology as determined by fluorescence microscopy and Western blot analysis. This manuscript delineates the reagents and methods used to produce lentivirus to express exogenous DNA in murine bone marrow-derived macrophages sufficient for single cell microscopy as well as functional assays requiring large numbers of murine bone marrow-derived macrophages.

  8. A Literature Assignment with a Civic Emphasis.

    Science.gov (United States)

    Brough, Barbara L.

    1989-01-01

    Describes how students wrestled with an assignment that asked them to name their heroes or heroines, compare them to "Beowulf," and defend their choices in writing and class discussion. Tells how students gained insight into their community and the ways in which people make it better. (MG)

  9. Writing Assignments Based on Literary Works.

    Science.gov (United States)

    Matthews, Dorothy, Ed.

    1985-01-01

    The literature selections serving as the basis for writing assignments in the articles in this journal issue range from time-honored English classics ("Beowulf,""Sir Gawain and the Green Knight") and American standards ("A Farewell to Arms,""The Scarlet Letter") to contemporary fiction. The articles deal with works by women writers (Shirley…

  10. Tabu search for target-radar assignment

    DEFF Research Database (Denmark)

    Hindsberger, Magnus; Vidal, Rene Victor Valqui

    2000-01-01

    In the paper the problem of assigning air-defense illumination radars to enemy targets is presented. A tabu search metaheuristic solution is described and the results achieved are compared to those of other heuristic approaches, implementation and experimental aspects are discussed. It is argued ...

  11. Test-assignment: a quadratic coloring problem

    NARCIS (Netherlands)

    Duives, Jelle; Lodi, Andrea; Malaguti, Enrico

    2013-01-01

    We consider the problem of assigning the test variants of a written exam to the desks of a classroom in such a way that desks that are close-by receive different variants. The problem is a generalization of the Vertex Coloring and we model it as a binary quadratic problem. Exact solution methods bas

  12. Politics, Internet Assignments, and Civic Knowledge.

    Science.gov (United States)

    May, Vaughn

    2000-01-01

    Describes how one professor of American government fights the apathy and cynicism of college students toward politics by using the Internet to help students more fairly appraise the workings of the American political system. One assignment has students research and manage a particular public policy initiative through visits to Web sites…

  13. Teaching Historical Analysis through Creative Writing Assignments

    Science.gov (United States)

    Peterson, Janine Larmon; Graham, Lea

    2015-01-01

    Incorporating creative writing exercises in history courses can heighten students' critical reading and analytical skills in an active learning model. We identify and define two types of possible assignments that use model texts as their locus: centripetal, which focuses on specific context and disciplinary terms, and centrifugal, which address…

  14. On Online Assignments in a Calculus Class

    Science.gov (United States)

    Jungic, Veselin; Kent, Deborah; Menz, Petra

    2012-01-01

    In this paper, we describe our experience with the creation and utilization of online assignments for several calculus classes at Simon Fraser University (SFU). We present our findings regarding available software by considering the needs and perspectives of the instructors, students, and administrators. We provide a list of questions that guide…

  15. Olfactory receptor signaling is regulated by the post-synaptic density 95, Drosophila discs large, zona-occludens 1 (PDZ) scaffold multi-PDZ domain protein 1.

    LENUS (Irish Health Repository)

    Dooley, Ruth

    2009-12-01

    The unique ability of mammals to detect and discriminate between thousands of different odorant molecules is governed by the diverse array of olfactory receptors expressed by olfactory sensory neurons in the nasal epithelium. Olfactory receptors consist of seven transmembrane domain G protein-coupled receptors and comprise the largest gene superfamily in the mammalian genome. We found that approximately 30% of olfactory receptors possess a classical post-synaptic density 95, Drosophila discs large, zona-occludens 1 (PDZ) domain binding motif in their C-termini. PDZ domains have been established as sites for protein-protein interaction and play a central role in organizing diverse cell signaling assemblies. In the present study, we show that multi-PDZ domain protein 1 (MUPP1) is expressed in the apical compartment of olfactory sensory neurons. Furthermore, on heterologous co-expression with olfactory sensory neurons, MUPP1 was shown to translocate to the plasma membrane. We found direct interaction of PDZ domains 1 + 2 of MUPP1 with the C-terminus of olfactory receptors in vitro. Moreover, the odorant-elicited calcium response of OR2AG1 showed a prolonged decay in MUPP1 small interfering RNA-treated cells. We have therefore elucidated the first building blocks of the putative \\'olfactosome\\

  16. Effortless assignment with 4D covariance sequential correlation maps.

    Science.gov (United States)

    Harden, Bradley J; Mishra, Subrata H; Frueh, Dominique P

    2015-11-01

    Traditional Nuclear Magnetic Resonance (NMR) assignment procedures for proteins rely on preliminary peak-picking to identify and label NMR signals. However, such an approach has severe limitations when signals are erroneously labeled or completely neglected. The consequences are especially grave for proteins with substantial peak overlap, and mistakes can often thwart entire projects. To overcome these limitations, we previously introduced an assignment technique that bypasses traditional pick peaking altogether. Covariance Sequential Correlation Maps (COSCOMs) transform the indirect connectivity information provided by multiple 3D backbone spectra into direct (H, N) to (H, N) correlations. Here, we present an updated method that utilizes a single four-dimensional spectrum rather than a suite of three-dimensional spectra. We demonstrate the advantages of 4D-COSCOMs relative to their 3D counterparts. We introduce improvements accelerating their calculation. We discuss practical considerations affecting their quality. And finally we showcase their utility in the context of a 52 kDa cyclization domain from a non-ribosomal peptide synthetase.

  17. Assignment and Correspondence Tracking System - Tactical / Operational Reporting

    Data.gov (United States)

    Social Security Administration — Reporting data store for the Assignment and Correspondence Tracking System (ACT). ACT automates the assignment and tracking of correspondence processing within the...

  18. An optimal query assignment for wireless sensor networks

    OpenAIRE

    Mitici, Mihaela; Onderwater, Martijn; Graaf, de, G.; Ommeren, van, Jos; Dijk, van, G.; Goseling, Jasper

    2012-01-01

    With the increased use of large-scale real-time embedded sensor networks, new control mechanisms are needed to avoid congestion and meet required Quality of Service (QoS) levels. In this paper, we propose a Markov Decision Problem (MDP) to prescribe an optimal query assignment strategy that achieves a trade-off between two QoS requirements: query response time and data validity. Query response time is the time that queries spend in the sensor network until they are solved. Data validity (fres...

  19. Simian virus 40 large T antigen alters the phosphorylation state of the RB-related proteins p130 and p107.

    OpenAIRE

    Stubdal, H; Zalvide, J; DeCaprio, J A

    1996-01-01

    p130 and p107 are nuclear phosphoproteins related to the retinoblastoma gene product (pRb). pRb, p107, and p130 each undergo cell cycle-dependent phosphorylation, form complexes with the E2F family of transcription factors, and associate with oncoproteins of DNA tumor viruses, including simian virus 40 (SV40) large T antigen (TAg) and adenovirus ElA protein. The results of recent studies with mouse embryo fibroblasts (MEFs) lacking the retinoblastoma gene (Rb-1) have suggested that p130 and p...

  20. Wild-type, but not mutant, human p53 proteins inhibit the replication activities of simian virus 40 large tumor antigen.

    OpenAIRE

    Friedman, P N; Kern, S. E.; Vogelstein, B; Prives, C

    1990-01-01

    Murine p53 blocks many of the replication activities of simian virus 40 (SV40) large tumor antigen (T antigen) in vitro. As murine cells do not replicate SV40 DNA, it was of interest to determine how p53 from permissive human cells functions. Recombinant baculoviruses encoding either the wild-type form of human p53 or a mutant p53 cloned from a human tumor cell line were constructed, and p53 proteins were purified from infected insect cells. Surprisingly, we found that wild-type human p53 was...

  1. Eradication of large tumors expressing human papillomavirus E7 protein by therapeutic vaccination with E7 fused to the extra domain a from fibronectin.

    Science.gov (United States)

    Mansilla, Cristina; Berraondo, Pedro; Durantez, Maika; Martínez, Marta; Casares, Noelia; Arribillaga, Laura; Rudilla, Francesc; Fioravanti, Jessica; Lozano, Teresa; Villanueva, Lorea; Sarobe, Pablo; Borrás, Francisco; Leclerc, Claude; Prieto, Jesús; Lasarte, Juan José

    2012-08-01

    Cervical carcinoma is one of the most common cancers in women worldwide. It is well established that chronic infection of the genital tract by various mucosatropic human papillomavirus (HPV) types causes cervical cancer. Cellular immunity to E7 protein from HPV (HPVE7) has been associated with clinical and cytologic resolution of HPV-induced lesions. Thus, we decided to test if targeting of HPVE7 to dendritic cells using a fusion protein containing the extra domain A (EDA) from fibronectin, a natural ligand for TLR4, and HPVE7 (EDA-HPVE7) might be an efficient vaccine for the treatment of cervical carcinoma. We found that EDA-HPVE7 fusion protein was efficiently captured by bone marrow derived dendritic cells in vitro and induced their maturation, with the upregulation of maturation markers and the production of IL-12. Immunization of mice with EDA-HPVE7 fusion protein induced antitumor CD8(+) T cell responses in the absence of additional adjuvants. Repeated intratumoral administration of EDA-HPVE7 in saline was able to cure established TC-1 tumors of 5-7 mm in diameter. More importantly, intravenous injection with EDA-HPVE7 in combination with the TLR ligand polyinosinic-polycytidylic acid (pIC), or with low doses of cyclophosphamide and the TLR9 ligand CpG-B complexed in cationic lipids, were able to eradicate large established TC-1 tumors (1.2 cm in diameter). Thus, therapeutic vaccination with EDA-HPVE7 fusion protein may be effective in the treatment of human cervical carcinoma. PMID:21898393

  2. High levels of nuclear MYC protein predict the presence of MYC rearrangement in diffuse large B-cell lymphoma

    DEFF Research Database (Denmark)

    Green, Tina Marie; Nielsen, Ole; de Stricker, Karin;

    2012-01-01

    Determining the presence of MYC gene rearrangements is becoming an increasingly important part of the diagnostic workup in aggressive lymphoma. Cytogenetic MYC alterations aid in differentiating diffuse large B-cell lymphoma (DLBCL) from Burkitt lymphoma. In addition, MYC aberrations are associated...... with poor prognosis in DLBCL. Fluorescence in situ hybridization and karyotyping are standard tests for detecting MYC aberrations, but these techniques are laborious and expensive. Here, we studied MYC status of 219 DLBCLs and Burkitt lymphomas using fluorescence in situ hybridization, immunohistochemistry......, and quantitative real-time polymerase chain reaction (QRT-PCR). Overall, 15% of the cases had an MYC break. QRT-PCR analysis of MYC expression showed that 72% of DLBCLs with an MYC break had aberrantly high or low levels of MYC transcript. Excluding the cases with aberrantly low MYC expression, we found...

  3. Weekly Fleet Assignment Model and Algorithm

    Institute of Scientific and Technical Information of China (English)

    ZHU Xing-hui; ZHU Jin-fu; GONG Zai-wu

    2007-01-01

    A 0-1 integer programming model for weekly fleet assignment was put forward based on linear network and weekly flight scheduling in China. In this model, the objective function is to maximize the total profit of fleet assignment, subject to the constraints of coverage, aircraft flow balance, fleet size, aircraft availability, aircraft usage, flight restriction, aircraft seat capacity,and stopover. Then the branch-and-bound algorithm based on special ordered set was applied to solve the model. At last, a realworld case study on an airline with 5 fleets, 48 aircrafts and 1 786 flight legs indicated that the profit increase was $1591276 one week and the running time was no more than 4 min, which shows that the model and algorithm are fairly good for domestic airline.

  4. Phylogenetic analysis of four nuclear protein-encoding genes largely corroborates the traditional classification of Bivalvia (Mollusca).

    Science.gov (United States)

    Sharma, Prashant P; González, Vanessa L; Kawauchi, Gisele Y; Andrade, Sónia C S; Guzmán, Alejandra; Collins, Timothy M; Glover, Emily A; Harper, Elizabeth M; Healy, John M; Mikkelsen, Paula M; Taylor, John D; Bieler, Rüdiger; Giribet, Gonzalo

    2012-10-01

    Revived interest in molluscan phylogeny has resulted in a torrent of molecular sequence data from phylogenetic, mitogenomic, and phylogenomic studies. Despite recent progress, basal relationships of the class Bivalvia remain contentious, owing to conflicting morphological and molecular hypotheses. Marked incongruity of phylogenetic signal in datasets heavily represented by nuclear ribosomal genes versus mitochondrial genes has also impeded consensus on the type of molecular data best suited for investigating bivalve relationships. To arbitrate conflicting phylogenetic hypotheses, we evaluated the utility of four nuclear protein-encoding genes-ATP synthase β, elongation factor-1α, myosin heavy chain type II, and RNA polymerase II-for resolving the basal relationships of Bivalvia. We sampled all five major lineages of bivalves (Archiheterodonta, Euheterodonta [including Anomalodesmata], Palaeoheterodonta, Protobranchia, and Pteriomorphia) and inferred relationships using maximum likelihood and Bayesian approaches. To investigate the robustness of the phylogenetic signal embedded in the data, we implemented additional datasets wherein length variability and/or third codon positions were eliminated. Results obtained include (a) the clade (Nuculanida+Opponobranchia), i.e., the traditionally defined Protobranchia; (b) the monophyly of Pteriomorphia; (c) the clade (Archiheterodonta+Palaeoheterodonta); (d) the monophyly of the traditionally defined Euheterodonta (including Anomalodesmata); and (e) the monophyly of Heteroconchia, i.e., (Palaeoheterodonta+Archiheterodonta+Euheterodonta). The stability of the basal tree topology to dataset manipulation is indicative of signal robustness in these four genes. The inferred tree topology corresponds closely to those obtained by datasets dominated by nuclear ribosomal genes (18S rRNA and 28S rRNA), controverting recent taxonomic actions based solely upon mitochondrial gene phylogenies.

  5. Adaptive Task Assignment in Online Learning Environments

    OpenAIRE

    Andersen, Per-Arne; Kråkevik, Christian; Goodwin, Morten; Yazidi, Anis

    2016-01-01

    With the increasing popularity of online learning, intelligent tutoring systems are regaining increased attention. In this paper, we introduce adaptive algorithms for personalized assignment of learning tasks to student so that to improve his performance in online learning environments. As main contribution of this paper, we propose a a novel Skill-Based Task Selector (SBTS) algorithm which is able to approximate a student's skill level based on his performance and consequently suggest adequa...

  6. Online Ad Assignment with an Ad Exchange

    OpenAIRE

    Dvořák, Wolfgang; Henzinger, Monika

    2016-01-01

    Ad exchanges are becoming an increasingly popular way to sell advertisement slots on the internet. An ad exchange is basically a spot market for ad impressions. A publisher who has already signed contracts reserving advertisement impressions on his pages can choose between assigning a new ad impression for a new page view to a contracted advertiser or to sell it at an ad exchange. This leads to an online revenue maximization problem for the publisher. Given a new impression to sell decide whe...

  7. Structural Encoding of Static Single Assignment Form

    DEFF Research Database (Denmark)

    Gal, Andreas; Probst, Christian; Franz, Michael

    2005-01-01

    Static Single Assignment (SSA) form is often used as an intermediate representation during code optimization in Java Virtual Machines. Recently, SSA has successfully been used for bytecode verification. However, constructing SSA at the code consumer is costly. SSAbased mobile code transport formats...... Java bytecode. While the resulting bytecode sequence can still be directly executed by traditional Virtual Machines, our novel VM can infer SSA form and confirm its safety with virtually no overhead....

  8. Automated feedback generation for introductory programming assignments

    OpenAIRE

    Singh, Rishabh; Gulwani, Sumit; Solar-Lezama, Armando

    2013-01-01

    We present a new method for automatically providing feedback for introductory programming problems. In order to use this method, we need a reference implementation of the assignment, and an error model consisting of potential corrections to errors that students might make. Using this information, the system automatically derives minimal corrections to student's incorrect solutions, providing them with a measure of exactly how incorrect a given solution was, as well as feedback about what they...

  9. Assignation of limited resources in pediatrics

    Directory of Open Access Journals (Sweden)

    Garduño Espinosa Armando

    2014-07-01

    Full Text Available Limited resources assignation is fundamental in the development of health services and since they will never be enough, justice is- sues arise. Many distributive justice theories are discussed: liberal, equilable and utilitarian, as well as ethic principles and cost-benefit relation, that is, the consequences. Palliative medicine is suggested as a strategy to reduce the cost of hospitable care and to enhance its designation.

  10. File Assignment Policy in Network Storage System

    Institute of Scientific and Technical Information of China (English)

    CaoQiang; XieChang-sheng

    2003-01-01

    Network storage increase capacity and scalability of storage system, data availability and enables the sharing of data among clients. When the developing network technology reduce performance gap between disk and network, however,mismatched policies and access pattern can significantly reduce network storage performance. So the strategy of data place ment in system is an important factor that impacts the performance of overall system. In this paper, the two algorithms of file assignment are presented. One is Greed partition that aims at the load balance across all NADs (Network Attached Disk). The other is Sort partition that tries to minimize variance of service time in each NAD. Moreover, we also compare the performance of our two algorithms in practical environment. Our experimental results show that when the size distribution (load characters) of all assigning files is closer and larger, Sort partition provides consistently better response times than Greedy algorithm. However, when the range of all assigning files is wider, there are more small files and access rate is higher, the Greedy algorithm has superior performance in compared with the Sort partition in off-line.

  11. File Assignment Policy in Network Storage System

    Institute of Scientific and Technical Information of China (English)

    Cao Qiang; Xie Chang-sheng

    2003-01-01

    Network storage increase capacity and scalability of storage system, data availability and enables the sharing of data among clients. When the developing network technology reduce performance gap between disk and network, however, mismatched policies and access pattern can significantly reduce network storage performance. So the strategy of data placement in system is an important factor that impacts the performance of overall system. In this paper, the two algorithms of file assignment are presented. One is Greed partition that aims at the load balance across all NADs (Network Attached Disk). The other is Sort partition that tries to minimize variance of service time in each NAD. Moreover, we also compare the performance of our two algorithms in practical environment. Our experimental results show that when the size distribution (load characters) of all assigning files is closer and larger, Sort partition provides consistently better response times than Greedy algorithm. However, when the range of all assigning files is wider, there are more small files and access rate is higher, the Greedy algorithm has superior performance in compared with the Sort partition in off-line.

  12. Linking activity-based travel demand models and traffic assignment: A Flemish case study

    OpenAIRE

    Ramaekers, Katrien; Kochan, Bruno; BELLEMANS, Tom; JANSSENS, Davy; Wets, Geert

    2008-01-01

    A custom agent-based simulation framework is developed that combines the fields of traffic demand modeling and traffic assignment, applied to the region of Flanders (Belgium). The framework uses an activity-based approach to model traffic demand and an assignment module that is linked to the traffic demand module. Activity data for the framework is provided by a large scale survey, conducted on 2500 households in the study area. The agent-based simulation model consists of over six million ag...

  13. Computational Study of a Heterostructural Model of Type I Collagen and Implementation of an Amino Acid Potential Method Applicable to Large Proteins

    Directory of Open Access Journals (Sweden)

    Jay Eifler

    2014-02-01

    Full Text Available Collagen molecules are the primary structural proteins of many biological systems. Much progress has been made in the study of the structure and function of collagen, but fundamental understanding of its electronic structures at the atomic level is still lacking. We present the results of electronic structure and bonding calculations of a specific model of type I collagen using the density functional theory-based method. Information on density of states (DOS, partial DOS, effective charges, bond order values, and intra- and inter-molecular H-bonding are obtained and discussed. We further devised an amino-acid-based potential method (AAPM to circumvent the full self-consistent field (SCF calculation that can be applied to large proteins. The AAPM is validated by comparing the results with the full SCF calculation of the whole type I collagen model with three strands. The calculated effective charges on each atom in the model retained at least 95% accuracy. This technique provides a viable and efficient way to study the electronic structure of large complex biomaterials at the ab initio level.

  14. Blockade of oncogenic IκB kinase activity in diffuse large B-cell lymphoma by bromodomain and extraterminal domain protein inhibitors.

    Science.gov (United States)

    Ceribelli, Michele; Kelly, Priscilla N; Shaffer, Arthur L; Wright, George W; Xiao, Wenming; Yang, Yibin; Mathews Griner, Lesley A; Guha, Rajarshi; Shinn, Paul; Keller, Jonathan M; Liu, Dongbo; Patel, Paresma R; Ferrer, Marc; Joshi, Shivangi; Nerle, Sujata; Sandy, Peter; Normant, Emmanuel; Thomas, Craig J; Staudt, Louis M

    2014-08-01

    In the activated B-cell-like (ABC) subtype of diffuse large B-cell lymphoma (DLBCL), NF-κB activity is essential for viability of the malignant cells and is sustained by constitutive activity of IκB kinase (IKK) in the cytoplasm. Here, we report an unexpected role for the bromodomain and extraterminal domain (BET) proteins BRD2 and BRD4 in maintaining oncogenic IKK activity in ABC DLBCL. IKK activity was reduced by small molecules targeting BET proteins as well as by genetic knockdown of BRD2 and BRD4 expression, thereby inhibiting downstream NF-κB-driven transcriptional programs and killing ABC DLBCL cells. Using a high-throughput platform to screen for drug-drug synergy, we observed that the BET inhibitor JQ1 combined favorably with multiple drugs targeting B-cell receptor signaling, one pathway that activates IKK in ABC DLBCL. The BTK kinase inhibitor ibrutinib, which is in clinical development for the treatment of ABC DLBCL, synergized strongly with BET inhibitors in killing ABC DLBCL cells in vitro and in a xenograft mouse model. These findings provide a mechanistic basis for the clinical development of BET protein inhibitors in ABC DLBCL, particularly in combination with other modulators of oncogenic IKK signaling.

  15. Arabidopsis AtMORC4 and AtMORC7 Form Nuclear Bodies and Repress a Large Number of Protein-Coding Genes.

    Science.gov (United States)

    Harris, C Jake; Husmann, Dylan; Liu, Wanlu; Kasmi, Farid El; Wang, Haifeng; Papikian, Ashot; Pastor, William A; Moissiard, Guillaume; Vashisht, Ajay A; Dangl, Jeffery L; Wohlschlegel, James A; Jacobsen, Steven E

    2016-05-01

    The MORC family of GHKL ATPases are an enigmatic class of proteins with diverse chromatin related functions. In Arabidopsis, AtMORC1, AtMORC2, and AtMORC6 act together in heterodimeric complexes to mediate transcriptional silencing of methylated DNA elements. Here, we studied Arabidopsis AtMORC4 and AtMORC7. We found that, in contrast to AtMORC1,2,6, they act to suppress a wide set of non-methylated protein-coding genes that are enriched for those involved in pathogen response. Furthermore, atmorc4 atmorc7 double mutants show a pathogen response phenotype. We found that AtMORC4 and AtMORC7 form homomeric complexes in vivo and are concentrated in discrete nuclear bodies adjacent to chromocenters. Analysis of an atmorc1,2,4,5,6,7 hextuple mutant demonstrates that transcriptional de-repression is largely uncoupled from changes in DNA methylation in plants devoid of MORC function. However, we also uncover a requirement for MORC in both DNA methylation and silencing at a small but distinct subset of RNA-directed DNA methylation target loci. These regions are characterized by poised transcriptional potential and a low density of sites for symmetric cytosine methylation. These results provide insight into the biological function of MORC proteins in higher eukaryotes. PMID:27171361

  16. 28 CFR 301.103 - Inmate work assignments.

    Science.gov (United States)

    2010-07-01

    ... COMPENSATION General § 301.103 Inmate work assignments. The unit team of each inmate, which ordinarily designates work assignments, or whoever makes work assignments, shall review appropriate medical records... 28 Judicial Administration 2 2010-07-01 2010-07-01 false Inmate work assignments. 301.103...

  17. 38 CFR 19.3 - Assignment of proceedings.

    Science.gov (United States)

    2010-07-01

    ... 38 Pensions, Bonuses, and Veterans' Relief 2 2010-07-01 2010-07-01 false Assignment of proceedings... Assignment of proceedings. (a) Assignment. The Chairman may assign a proceeding instituted before the Board, including any motion, to an individual Member or to a panel of three or more Members for adjudication...

  18. 28 CFR 524.72 - CIM assignment categories.

    Science.gov (United States)

    2010-07-01

    .... Factors to consider in classifying an individual to this assignment include, but are not limited to... 28 Judicial Administration 2 2010-07-01 2010-07-01 false CIM assignment categories. 524.72 Section... assignment categories. CIM cases are classified according to the following assignments: (a) Witness...

  19. 49 CFR 821.35 - Assignment, duties and powers.

    Science.gov (United States)

    2010-10-01

    ... SAFETY BOARD RULES OF PRACTICE IN AIR SAFETY PROCEEDINGS Law Judges § 821.35 Assignment, duties and powers. (a) Assignment of law judge and duration of assignment. The chief law judge shall assign a law... addressed to the Case Manager for handling by the chief law judge, who may handle these matters...

  20. Sequence specific resonance assignment via Multicanonical Monte Carlo search using an ABACUS approach.

    Science.gov (United States)

    Lemak, Alexander; Steren, Carlos A; Arrowsmith, Cheryl H; Llinás, Miguel

    2008-05-01

    ABACUS [Grishaev et al. (2005) Proteins 61:36-43] is a novel protocol for automated protein structure determination via NMR. ABACUS starts from molecular fragments defined by unassigned J-coupled spin-systems and involves a Monte Carlo stochastic search in assignment space, probabilistic sequence selection, and assembly of fragments into structures that are used to guide the stochastic search. Here, we report further development of the two main algorithms that increase the flexibility and robustness of the method. Performance of the BACUS [Grishaev and Llinás (2004) J Biomol NMR 28:1-101] algorithm was significantly improved through use of sequential connectivities available from through-bond correlated 3D-NMR experiments, and a new set of likelihood probabilities derived from a database of 56 ultra high resolution X-ray structures. A Multicanonical Monte Carlo procedure, Fragment Monte Carlo (FMC), was developed for sequence-specific assignment of spin-systems. It relies on an enhanced assignment sampling and provides the uncertainty of assignments in a quantitative manner. The efficiency of the protocol was validated on data from four proteins of between 68-116 residues, yielding 100% accuracy in sequence specific assignment of backbone and side chain resonances.

  1. Replicating and Extending Research on the Partial Assignment Completion Effect: Is Sunk Cost Related to Partial Assignment Completion Strength?

    Science.gov (United States)

    Hawthorn-Embree, Meredith L.; Taylor, Emily P.; Skinner, Christopher H.; Parkhurst, John; Nalls, Meagan L.

    2014-01-01

    After students acquire a skill, mastery often requires them to choose to engage in assigned academic activities (e.g., independent seatwork, and homework). Although students may be more likely to choose to work on partially completed assignments than on new assignments, the partial assignment completion (PAC) effect may not be very powerful. The…

  2. Assignment procedure biases in randomized policy experiments

    DEFF Research Database (Denmark)

    Kirchsteiger, Georg; Sebald, Alexander Christopher; Aldashev, Gani

    2016-01-01

    Randomized controlled trials (RCT) have gained ground as the dominant tool for studying policy interventions in many fields of applied economics. We analyse theoretically encouragement and resentful demoralization in RCTs and show that these might be rooted in the same behavioural trait - people......’s propensity to act reciprocally. When people are motivated by reciprocity, the choice of assignment procedure influences the RCTs’ findings. We show that even credible and explicit randomization procedures do not guarantee an unbiased prediction of the impact of policy interventions; however, they minimize...

  3. Colony location algorithm for assignment problems

    Institute of Scientific and Technical Information of China (English)

    Dingwei WANG

    2004-01-01

    A novel algorithm called Colony Location Algorithm (CLA) is proposed. It mimics the phenomena in biotic conmunity that colonies of species could be located in the places most suitable to their growth. The factors working on the species location such as the nutrient of soil, resource competition between species, growth and decline process, and effect on environment were considered in CLA via the nutrient function, growth and decline rates, environment evaluation and fertilization strategy.CLA was applied to solve the classical assignment problems. The computation results show that CLA can achieve the optimal solution with higher possibility and shorter running time.

  4. 基于大体积循环进样的低丰度蛋白质富集%Protein enrichment based on large volume recycling injection

    Institute of Scientific and Technical Information of China (English)

    张权青; 张磊; 高小迪; 张维冰; 张庆合

    2014-01-01

    发展了一种大体积循环进样方法,用于富集低丰度蛋白质。在优化的色谱分离条件下,通过增加蛋白质样品的上样体积提高低丰度蛋白质的绝对含量;进一步采用增加样品进样循环次数的方法提高蛋白质的富集效率。以猪肝提取蛋白质为样品,每次上样量500μL 的大体积11次循环进样。根据色谱峰的信号强弱,选择了在原始谱图中看不到色谱峰、有较少小峰和有较多小峰出现的时间段等有代表性的馏分进行研究。在中等极性的组分中,保留时间为11.38 min 和12.58 min 组分的富集效率分别提高了52倍和61倍,实验结果与理论富集效率相近。所发展的方法为生物蛋白质样品研究提供了一种新的富集制备及检测方法。%Biological proteins have great differences in size,hydrophobicity,pH and relative abundance. Especially,the relative contents of high-abundance proteins and low abundance proteins can reach 11 orders of magnitude or more in blood. However,the contents of proteins which have an important impact on the biological function and have related to major diseases are usually very low in body fluids or tissues. So,it is very meaningful to detect the low abun-dance proteins in biological samples. A method of large volume recycling injection was devel-oped as a new sample enrichment technology in this study. The low absolute contents of pro-teins in a sample were improved by increasing the volume of sample,and the enrichment effi-ciency was also increased with the addition of injection times. With the pork liver protein as an example,under the optimized conditions,the representative contents which were no peaks or little peaks and more peaks with low signals were studied. The contents of peaks at the reten-tion times of 11. 38 min and 12. 58 min were enriched by 11 time injections of 500 μL each time. The enrichment factors respectively reached 52 and 61 which were closed to

  5. Assigning functional meaning to digital circuits

    Energy Technology Data Exchange (ETDEWEB)

    Eckmann, S.T.; Chisholm, G.H. [Argonne National Lab., IL (United States). Decision and Information Sciences Div.

    1997-07-01

    During computer-aided design, the problem of how to determine the logical function of a digital circuit arises in many contexts. For example, assigning functional meaning to a circuit is a fundamental operation in both reverse engineering and implementation validation. This report describes such a determination by discussing how a higher-level functional representation is constructed from a detailed circuit description (i.e., a gate-level netlist, which is a list of logic gates and their interconnections). The approach used involves transforming parts of the netlist into a functional representation and then manipulating this representation. Two types of functional representations are described: (1) a mathematical representation based on the logical operators ``exor`` and ``and`` and (2) a directed acyclic graph representation based on binary decision trees. Each representation provides a canonical form of the logical function being implemented (i.e., a form that is independent of implementation details). Such forms, however, have a well-known problem associated with the ordering of inputs: for each order, a unique form exists. A solution to this problem is given for both representations. Experimental results that demonstrate the use of these representations in the process of assigning functional meaning to a circuit are provided. The report also identifies and discusses issues critical to the performance required of this fundamental operation.

  6. A Silicon Nanomembrane Detector for Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS of Large Proteins

    Directory of Open Access Journals (Sweden)

    Jonghoo Park

    2013-10-01

    Full Text Available We describe a MALDI-TOF ion detector based on freestanding silicon nanomembrane technology. The detector is tested in a commercial MALDI-TOF mass spectrometer with equimolar mixtures of proteins. The operating principle of the nanomembrane detector is based on phonon-assisted field emission from these silicon nanomembranes, in which impinging ion packets excite electrons in the nanomembrane to higher energy states. Thereby the electrons can overcome the vacuum barrier and escape from the surface of the nanomembrane via field emission. Ion detection is demonstrated of apomyoglobin (16,952 Da, aldolase (39,212 Da, bovine serum albumin (66,430 Da, and their equimolar mixtures. In addition to the three intact ions, a large number of fragment ions are also revealed by the silicon nanomembrane detector, which are not observable with conventional detectors.

  7. A large multi-centre European study validates high-sensitivity C-reactive protein (hsCRP) as a clinical biomarker for the diagnosis of diabetes subtypes

    DEFF Research Database (Denmark)

    Thanabalasingham, G.; Shah, N.; Vaxillaire, M.;

    2011-01-01

    An accurate molecular diagnosis of diabetes subtype confers clinical benefits; however, many individuals with monogenic diabetes remain undiagnosed. Biomarkers could help to prioritise patients for genetic investigation. We recently demonstrated that high-sensitivity C-reactive protein (hs......CRP) levels are lower in UK patients with hepatocyte nuclear factor 1 alpha (HNF1A)-MODY than in other diabetes subtypes. In this large multi-centre study we aimed to assess the clinical validity of hsCRP as a diagnostic biomarker, examine the genotype-phenotype relationship and compare different hsCRP assays....... High-sensitivity CRP levels were analysed in individuals with HNF1A-MODY (n = 457), glucokinase (GCK)-MODY (n = 404), hepatocyte nuclear factor 4 alpha (HNF4A)-MODY (n = 54) and type 2 diabetes (n = 582) from seven European centres. Three common assays for hsCRP analysis were evaluated. We excluded 121...

  8. Characterization of p16 and E6 HPV-related proteins in uterine cervix high-grade lesions of patients treated by conization with large loop excision

    Science.gov (United States)

    RONCAGLIA, MARIA TERESA; FREGNANI, JOSÉ HUMBERTO T.G.; TACLA, MARICY; DE CAMPOS, SILVANA GISELE PEGORIN; CAIAFFA, HÉLIO HEHL; AB’SABER, ALEXANDRE; DA MOTTA, EDUARDO VIEIRA; ALVES, VENÂNCIO AVANCINI FERREIRA; BARACAT, EDMUND C.; LONGATTO FILHO, ADHEMAR

    2013-01-01

    Cervical cancer and its precursor lesions represent a significant public health problem for developing and less-developed countries. Cervical carcinogenesis is strongly correlated with persistent high-risk human papillomavirus (HPV) infection, which is mostly associated with expression of the p16 and E6 HPV-related proteins. The aim of this present study was to determine the expression of the p16 and E6 proteins in females with high-grade lesions treated with conization, and to discuss the role of these proteins as prognostic markers following treatment. In total, 114 females were treated for high-grade cervical intraepithelial neoplasia (CIN, grades 2/3) by conization with large loop excision of the transformation zone (LLETZ). Following surgery, the patients returned within 30–45 days for post-operative evaluation. A follow-up was conducted every 6 months for 2 years. At each follow-up appointment, a Pap smear, colposcopy and HPV DNA test were performed. E6 and p16 immunohistochemical tests were conducted on the surgical specimens. The positive expression of p16 was correlated with the presence of lesions with increased severity in the surgical specimens (P= 0.0001). The expression of E6 did not demonstrate the same correlation (P=0.131). The HPV DNA hybrid, collected in the first post-operative consultation as a predictor of the cytological abnormalities identified at the 24-month follow-up assessment, presented a sensitivity of 55.6%, a specificity of 84.8%, a positive predictive value of 33.3% and a negative predictive value of 93.3%. The role of p16INK4A as a marker of CIN was also demonstrated; the expression of p16 and E6, however, did not appear to be of any prognostic value in predicting the clearance of high-risk HPV following conization. A negative hybrid capture test was correlated with a disease-free outcome. PMID:23946778

  9. Simultaneously Optimizing Storage Location Assignment at Forward Area and Reserve Area - a Decomposition Based Heuristic

    DEFF Research Database (Denmark)

    Bloch, Søren; Christiansen, Christian Holk

    This paper addresses the Two-level Storage Location Assignment Problem (TSLAP) for two level low-level picker-to-part warehouses. The first level consists of a forward area from which all picking is done. The second level consists of a reserve storage area, from which the forward area...... is replenished. A large part of the cost of operating the warehouse is a function of the assignment of items to locations at both the reserve storage and the forward area. The cost of the storage location assignment at the forward area is dependent on the storage location assignment at the reserve area and visa...... versa. This, however, is often neglected in the existing literature. We solve the TSLAP simultaneously for the reserve area and the forward area. Based on randomly generated test instances we show that the solutions of TSLAP compare favorably to solutions found by other algorithms proposed...

  10. Involvement of Grb2 adaptor protein in nucleophosmin-anaplastic lymphoma kinase (NPM-ALK)-mediated signaling and anaplastic large cell lymphoma growth.

    Science.gov (United States)

    Riera, Ludovica; Lasorsa, Elena; Ambrogio, Chiara; Surrenti, Nadia; Voena, Claudia; Chiarle, Roberto

    2010-08-20

    Most anaplastic large cell lymphomas (ALCL) express oncogenic fusion proteins derived from chromosomal translocations or inversions of the anaplastic lymphoma kinase (ALK) gene. Frequently ALCL carry the t(2;5) translocation, which fuses the ALK gene to the nucleophosmin (NPM1) gene. The transforming activity mediated by NPM-ALK fusion induces different pathways that control proliferation and survival of lymphoma cells. Grb2 is an adaptor protein thought to play an important role in ALK-mediated transformation, but its interaction with NPM-ALK, as well as its function in regulating ALCL signaling pathways and cell growth, has never been elucidated. Here we show that active NPM-ALK, but not a kinase-dead mutant, bound and induced Grb2 phosphorylation in tyrosine 160. An intact SH3 domain at the C terminus of Grb2 was required for Tyr(160) phosphorylation. Furthermore, Grb2 did not bind to a single region but rather to different regions of NPM-ALK, mainly Tyr(152-156), Tyr(567), and a proline-rich region, Pro(415-417). Finally, shRNA knockdown experiments showed that Grb2 regulates primarily the NPM-ALK-mediated phosphorylation of SHP2 and plays a key role in ALCL cell growth.

  11. Crizotinib (PF-2341066) induces apoptosis due to downregulation of pSTAT3 and BCL-2 family proteins in NPM-ALK(+) anaplastic large cell lymphoma.

    Science.gov (United States)

    Hamedani, Farid Saei; Cinar, Munevver; Mo, Zhicheng; Cervania, Melissa A; Amin, Hesham M; Alkan, Serhan

    2014-04-01

    Nucleophosmin-anaplastic lymphoma kinase (NPM-ALK) is an aberrant fusion gene product with tyrosine kinase activity and is expressed in substantial subset of anaplastic large cell lymphomas (ALCL). It has been shown that NPM-ALK binds to and activates signal transducer and activator of transcription 3 (STAT3). Although NPM-ALK(+) ALCL overall shows a better prognosis, there is a sub-group of patients who relapses and is resistant to conventional chemotherapeutic regimens. NPM-ALK is a potential target for small molecule kinase inhibitors. Crizotinib (PF-2341066) is a small, orally bioavailable molecule that inhibits growth of tumors with ALK activity as shown in a subgroup of non-small lung cancer patients with EML4-ALK expression. In this study, we have investigated the in vitro effects of Crizotinib in ALCL cell line with NPM-ALK fusion. Crizotinib induced marked downregulation of STAT3 phosphorylation, which was associated with significant apoptotic cell death. Apoptosis induction was attributed to caspase-3 cleavage and marked downregulation of the Bcl-2 family of proteins including MCL-1. These findings implicate that Crizotinib has excellent potential to treat patients with NPM-ALK(+) ALCL through induction of apoptotic cell death and downregulation of major oncogenic proteins in this aggressive lymphoma.

  12. Channel Assignment Algorithms for MRMC Wireless Mesh Networks

    Directory of Open Access Journals (Sweden)

    Mohammad A Hoque

    2011-11-01

    Full Text Available The wireless mesh networksare considered as one of the vital elements in today’s converged networks,providing high bandwidth and connectivity over large geographical areas. Mesh routers equipped with multiple radios can significantly overcome the capacity problem and increase the aggregate throughput of the network where single radio nodessuffer from performancedegradation. Moreover, the market availability of cheap radios or network interfaces also makes multi-radio solutions more feasible.A key issue in such networks is how to efficiently design a channel assignmentscheme that utilizes the available channels as well as increases overall performance of the network. This paper provides an overall review on the issues pertaining to the channel assignment in WMNs and the most relevant approaches and solutions developed in the area. They include design challenges, goals and criteria; routing considerations, graph based solutions and challenges of partially overlapped channels. We conclude that the assignment of channels to the radio interfaces continuously poses significant challenges. Many research issues remain open for further investigation.

  13. A Large Repetitive RTX-Like Protein Mediates Water-Soaked Lesion Development, Leakage of Plant Cell Content and Host Colonization in the Pantoea stewartii subsp. stewartii Pathosystem.

    Science.gov (United States)

    Roper, M Caroline; Burbank, Lindsey P; Williams, Kayla; Viravathana, Polrit; Tien, Hsin-Yu; von Bodman, Susanne

    2015-12-01

    Pantoea stewartii subsp. stewartii is the etiological agent of Stewart's wilt and is a serious bacterial pathogen affecting sweet corn. During the leaf blight phase, P. stewartii colonizes the leaf apoplast and causes a characteristic water-soaked lesion. The Hrp type III secretion system has been implicated in the water-soaking phenotype, and the goal of this study was to investigate other potential factors that contribute to the plant cellular disruption associated with these lesions. The P. stewartii genome contains a gene encoding a large repetitive RTX toxin, designated rtx2. RTX toxins comprise a large family of pore-forming proteins, which are widely distributed among gram-negative bacteria. These cytotoxins usually lyse their target host cells and cause significant tissue damage as a consequence. We hypothesized that this RTX-like toxin plays a role in the water-soaking phase of infection due to its predicted cytolytic properties. Based on the data reported here, we conclude that RTX2 contributes significantly to the development of water-soaked lesions and leakage of plant cellular contents and is an important pathogenicity factor for P. stewartii. PMID:26284907

  14. Solving multiconstraint assignment problems using learning automata.

    Science.gov (United States)

    Horn, Geir; Oommen, B John

    2010-02-01

    This paper considers the NP-hard problem of object assignment with respect to multiple constraints: assigning a set of elements (or objects) into mutually exclusive classes (or groups), where the elements which are "similar" to each other are hopefully located in the same class. The literature reports solutions in which the similarity constraint consists of a single index that is inappropriate for the type of multiconstraint problems considered here and where the constraints could simultaneously be contradictory. This feature, where we permit possibly contradictory constraints, distinguishes this paper from the state of the art. Indeed, we are aware of no learning automata (or other heuristic) solutions which solve this problem in its most general setting. Such a scenario is illustrated with the static mapping problem, which consists of distributing the processes of a parallel application onto a set of computing nodes. This is a classical and yet very important problem within the areas of parallel computing, grid computing, and cloud computing. We have developed four learning-automata (LA)-based algorithms to solve this problem: First, a fixed-structure stochastic automata algorithm is presented, where the processes try to form pairs to go onto the same node. This algorithm solves the problem, although it requires some centralized coordination. As it is desirable to avoid centralized control, we subsequently present three different variable-structure stochastic automata (VSSA) algorithms, which have superior partitioning properties in certain settings, although they forfeit some of the scalability features of the fixed-structure algorithm. All three VSSA algorithms model the processes as automata having first the hosting nodes as possible actions; second, the processes as possible actions; and, third, attempting to estimate the process communication digraph prior to probabilistically mapping the processes. This paper, which, we believe, comprehensively reports the

  15. Improving load balance with flexibly assignable tasks

    Energy Technology Data Exchange (ETDEWEB)

    Pinar, Ali; Hendrickson, Bruce

    2003-09-09

    In many applications of parallel computing, distribution ofthe data unambiguously implies distribution of work among processors. Butthere are exceptions where some tasks can be assigned to one of severalprocessors without altering the total volume of communication. In thispaper, we study the problem of exploiting this flexibility in assignmentof tasks to improve load balance. We first model the problem in terms ofnetwork flow and use combinatorial techniques for its solution. Ourparametric search algorithms use maximum flow algorithms for probing on acandidate optimal solution value. We describe two algorithms to solve theassignment problem with \\logW_T and vbar P vbar probe calls, w here W_Tand vbar P vbar, respectively, denote the total workload and number ofproce ssors. We also define augmenting paths and cuts for this problem,and show that anyalgorithm based on augmenting paths can be used to findan optimal solution for the task assignment problem. We then consideracontinuous version of the problem, and formulate it as a linearlyconstrained optimization problem, i.e., \\min\\|Ax\\|_\\infty,\\; {\\rms.t.}\\;Bx=d. To avoid solving an intractable \\infty-norm optimization problem,we show that in this case minimizing the 2-norm is sufficient to minimizethe \\infty-norm, which reduces the problem to the well-studiedlinearly-constrained least squares problem. The continuous version of theproblem has the advantage of being easily amenable to parallelization.Our experiments with molecular dynamics and overlapped domaindecomposition applications proved the effectiveness of our methods withsignificant improvements in load balance. We also discuss how ourtechniques can be enhanced for heterogeneous systems.

  16. 7 CFR 1404.3 - Payments which may be assigned.

    Science.gov (United States)

    2010-01-01

    ... OF AGRICULTURE GENERAL REGULATIONS AND POLICIES ASSIGNMENT OF PAYMENTS § 1404.3 Payments which may be assigned. Except as otherwise provided in this part or in individual program regulations, contracts...

  17. Assignment Choice: Do Students Choose Briefer Assignments or Finishing What They Started?

    Science.gov (United States)

    Hawthorn-Embree, Meredith L.; Skinner, Christopher H.; Parkhurst, John; O'Neil, Michael; Conley, Elisha

    2010-01-01

    Academic skill development requires engagement in effortful academic behaviors. Although students may be more likely to choose to engage in behaviors that require less effort, they also may be motivated to complete assignments that they have already begun. Seventh-grade students (N = 88) began a mathematics computation worksheet, but were stopped…

  18. Efficient scheduling algorithm for demand-assigned TDMA satellite systems

    Science.gov (United States)

    Durrani, Sajjad H.; Jo, Kyung Y.

    1989-01-01

    An efficient scheduling algorithm (stations burst plan) for demand-assigned time-division multiple-access (TDMA) satellite network systems is introduced. The total demand for transmitting data through a transponder may exceed the available bit-rate capacity, and a scheduler of the system wishes to utilize the system with minimum changes of slot allocations while maximizing throughputs. By implementing such a burst-plan algorithm, transmission of all demanded data traffic can be completed with minimum unused resources (idle slots). The underlying ideas adopted for the algorithm are that jobs with shorter remaining processing times should have higher priorities and that as many jobs are processed at a time as possible. The algorithm is particularly useful for deriving smooth burst plans for a satellite system with a large number of ground stations.

  19. Learning through Writing: Teaching Critical Thinking Skills in Writing Assignments

    Science.gov (United States)

    Cavdar, Gamze; Doe, Sue

    2012-01-01

    Traditional writing assignments often fall short in addressing problems in college students' writing as too often these assignments fail to help students develop critical thinking skills and comprehension of course content. This article reports the use of a two-part (staged) writing assignment with postscript as a strategy for improving critical…

  20. 75 FR 55354 - Delegation of Authority and Assignment of Responsibilities

    Science.gov (United States)

    2010-09-10

    ... of the Secretary Delegation of Authority and Assignment of Responsibilities Secretary's Order 3-2010 Subject: Delegation of Authority and Assignment of Responsibilities to the Employee Benefits Security Administration. 1. Purpose. To delegate authority and assign responsibilities for the administration of...

  1. 7 CFR 762.160 - Assignment of guarantee.

    Science.gov (United States)

    2010-01-01

    ... of the loan. The holder must assign the guaranteed portion back to the original lender if requested... AGRICULTURE SPECIAL PROGRAMS GUARANTEED FARM LOANS § 762.160 Assignment of guarantee. (a) The following general requirements apply to assigning guaranteed loans: (1) Subject to Agency concurrence, the...

  2. 24 CFR 255.2 - GNMA right to assignment.

    Science.gov (United States)

    2010-04-01

    ... will have the right to perfect an assignment of the mortgage to itself. However, before exercising this right, GNMA will attempt to have the Mortgage assigned to another eligible coinsuring lender (unless... defaulting lender-issuer, GNMA will have the right to perfect an assignment of the Coinsured...

  3. 24 CFR 252.2 - GNMA right to assignment.

    Science.gov (United States)

    2010-04-01

    ....2 GNMA right to assignment. If the lender-issuer defaults on its obligations under the GNMA Mortgage... defaulting lender-issuer, GNMA will have the right to perfect an assignment of the mortgage to itself. However, before exercising this right, GNMA will attempt to have the Mortgage assigned to another...

  4. 24 CFR 251.2 - GNMA right to assignment.

    Science.gov (United States)

    2010-04-01

    ... right to assignment. If the lender-issuer defaults on its obligations under the GNMA Mortgage-Backed...-issuer, GNMA will have the right to perfect an assignment of the mortgage to itself. However, before exercising this right, GNMA will attempt to have the Mortgage assigned to another eligible coinsuring...

  5. Solving the k-cardinality assignment problem by transformation

    NARCIS (Netherlands)

    A. Volgenant

    2004-01-01

    The k-cardinality Linear Assignment Problem (k-LAP) with k a given integer is a generalization of the linear assignment problem: one wants to assign k rows (a free choice out of more rows) to k columns (a free choice out of more columns) minimizing the sum of the corresponding costs. For this polyno

  6. 42 CFR 60.38 - Assignment of a HEAL loan.

    Science.gov (United States)

    2010-10-01

    ... 42 Public Health 1 2010-10-01 2010-10-01 false Assignment of a HEAL loan. 60.38 Section 60.38... ASSISTANCE LOAN PROGRAM The Lender and Holder § 60.38 Assignment of a HEAL loan. A HEAL note may not be assigned except to another HEAL lender, the Student Loan Marketing Association (popularly known as...

  7. Evolutionary algorithm based index assignment algorithm for noisy channel

    Institute of Scientific and Technical Information of China (English)

    李天昊; 余松煜

    2004-01-01

    A globally optimal solution to vector quantization (VQ) index assignment on noisy channel, the evolutionary algorithm based index assignment algorithm (EAIAA), is presented. The algorithm yields a significant reduction in average distortion due to channel errors, over conventional arbitrary index assignment, as confirmed by experimental results over the memoryless binary symmetric channel (BSC) for any bit error.

  8. Probabilistic protein function prediction from heterogeneous genome-wide data.

    Directory of Open Access Journals (Sweden)

    Naoki Nariai

    Full Text Available Dramatic improvements in high throughput sequencing technologies have led to a staggering growth in the number of predicted genes. However, a large fraction of these newly discovered genes do not have a functional assignment. Fortunately, a variety of novel high-throughput genome-wide functional screening technologies provide important clues that shed light on gene function. The integration of heterogeneous data to predict protein function has been shown to improve the accuracy of automated gene annotation systems. In this paper, we propose and evaluate a probabilistic approach for protein function prediction that integrates protein-protein interaction (PPI data, gene expression data, protein motif information, mutant phenotype data, and protein localization data. First, functional linkage graphs are constructed from PPI data and gene expression data, in which an edge between nodes (proteins represents evidence for functional similarity. The assumption here is that graph neighbors are more likely to share protein function, compared to proteins that are not neighbors. The functional linkage graph model is then used in concert with protein domain, mutant phenotype and protein localization data to produce a functional prediction. Our method is applied to the functional prediction of Saccharomyces cerevisiae genes, using Gene Ontology (GO terms as the basis of our annotation. In a cross validation study we show that the integrated model increases recall by 18%, compared to using PPI data alone at the 50% precision. We also show that the integrated predictor is significantly better than each individual predictor. However, the observed improvement vs. PPI depends on both the new source of data and the functional category to be predicted. Surprisingly, in some contexts integration hurts overall prediction accuracy. Lastly, we provide a comprehensive assignment of putative GO terms to 463 proteins that currently have no assigned function.

  9. Assignment of amide proton signals by combined evaluation of HN, NN and HNCA MAS-NMR correlation spectra

    Energy Technology Data Exchange (ETDEWEB)

    Rossum, Barth-Jan van; Castellani, Federica [Forschungsinstitut fuer Molekulare Pharmakologie (FMP) (Germany); Pauli, Jutta [BAM (Germany); Rehbein, Kristina [Forschungsinstitut fuer Molekulare Pharmakologie (FMP) (Germany); Hollander, J.; Groot, Huub J.M. de [BAM (Germany); Oschkinat, Hartmut [Forschungsinstitut fuer Molekulare Pharmakologie (FMP) (Germany)], E-mail: Oschkinat@fmp-berlin.de

    2003-03-15

    In this paper, we present a strategy for the {sup 1}H{sup N} resonance assignment in solid-state magic-angle spinning (MAS) NMR, using the {alpha}-spectrin SH3 domain as an example. A novel 3D triple resonance experiment is presented that yields intraresidue H{sup N}-N-C{sup {alpha}} correlations, which was essential for the proton assignment. For the observable residues, 52 out of the 54 amide proton resonances were assigned from 2D ({sup 1}H-{sup 15}N) and 3D ({sup 1}H-{sup 15}N-{sup 13}C) heteronuclear correlation spectra. It is demonstrated that proton-driven spin diffusion (PDSD) experiments recorded with long mixing times (4 s) are helpful for confirming the assignment of the protein backbone {sup 15}N resonances and as an aid in the amide proton assignment.

  10. Induction and Persistence of Large γH2AX Foci by High Linear Energy Transfer Radiation in DNA-Dependent protein kinase–Deficient Cells

    Energy Technology Data Exchange (ETDEWEB)

    Bracalente, Candelaria; Ibañez, Irene L. [Departamento de Micro y Nanotecnología, Comisión Nacional de Energía Atómica, San Martín, Buenos Aires (Argentina); Consejo Nacional de Investigaciones Científicas y Técnicas, Buenos Aires (Argentina); Molinari, Beatriz [Departamento de Radiobiología, Comisión Nacional de Energía Atómica, San Martín, Buenos Aires (Argentina); Consejo Nacional de Investigaciones Científicas y Técnicas, Buenos Aires (Argentina); Palmieri, Mónica [Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires (Argentina); Kreiner, Andrés [Consejo Nacional de Investigaciones Científicas y Técnicas, Buenos Aires (Argentina); Gerencia de Investigación y Aplicaciones, Comisión Nacional de Energía Atómica, San Martín, Buenos Aires (Argentina); Escuela de Ciencia y Tecnología, Universidad Nacional de San Martín, San Martín, Buenos Aires (Argentina); Valda, Alejandro [Escuela de Ciencia y Tecnología, Universidad Nacional de San Martín, San Martín, Buenos Aires (Argentina); and others

    2013-11-15

    Purpose: To evaluate the cell response to DNA double-strand breaks induced by low and high linear energy transfer (LET) radiations when the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs), an essential protein of the nonhomologous end-joining repair pathway, lacks kinase activity. Methods and Materials: CHO10B2, a Chinese hamster ovary cell line, and its derived radiosensitive mutant cell line, irs-20, lacking DNA-PKcs activity, were evaluated after 0 to 3 Gy of γ-rays, plateau and Bragg peak protons, and lithium beams by clonogenic assay, and as a measurement of double-strand breaks, phosphorylated H2AX (γH2AX) foci number and size were quantified by immunocytofluorescence. Results: Irs-20 exhibited greater radiosensitivity and a higher amount of γH2AX foci than CHO10B2 at 6 hours after irradiation for all types of radiations. Remarkably, CHO10B2 and irs-20 maintained their difference in radiosensitivity after high-LET radiation. Six hours after low-LET radiations, irs-20 did not reach basal levels of γH2AX at high doses, whereas CHO10B2 recovered basal levels for all doses. After high-LET radiation, only CHO10B2 exhibited a reduction in γH2AX foci, but it never reached basal levels. Persistent foci in irs-20 confirmed a repair deficiency. Interestingly, after 30 minutes of high-LET radiation both cell lines exhibited large foci (size >0.9 μm{sup 2}) related to the damage nature, whereas at 6 hours irs-20 showed a higher amount of large foci than CHO10B2, with a 7-fold increase at 3 Gy, that could also be associated to radiosensitivity. Conclusions: We demonstrated, for the first time, an association between deficient DNA-PKcs activity and not only high levels of H2AX phosphorylation but also persistence and size increase of γH2AX foci after high-LET irradiation.

  11. An Interactive Introduction to Protein Structure

    Science.gov (United States)

    Lee, W. Theodore

    2004-01-01

    To improve student understanding of protein structure and the significance of noncovalent interactions in protein structure and function, students are assigned a project to write a paper complemented with computer-generated images. The assignment provides an opportunity for students to select a protein structure that is of interest and detail…

  12. A multicast dynamic wavelength assignment algorithm based on matching degree

    Institute of Scientific and Technical Information of China (English)

    WU Qi-wu; ZHOU Xian-wei; WANG Jian-ping; YIN Zhi-hong; ZHANG Long

    2009-01-01

    The wavelength assignment with multiple multicast requests in fixed routing WDM network is studied. A new multicast dynamic wavelength assignment algorithm is presented based on matching degree. First, the wavelength matching degree between available wavelengths and multicast routing trees is introduced into the algorithm. Then, the wavelength assign-ment is translated into the maximum weight matching in bipartite graph, and this matching problem is solved by using an extended Kuhn-Munkres algorithm. The simulation results prove that the overall optimal wavelength assignment scheme is obtained in polynomial time. At the same time, the proposed algorithm can reduce the connecting blocking probability and improve the system resource utilization.

  13. A Bayesian approach to simultaneously quantify assignments and linguistic uncertainty

    Energy Technology Data Exchange (ETDEWEB)

    Chavez, Gregory M [Los Alamos National Laboratory; Booker, Jane M [BOOKER SCIENTIFIC FREDERICKSBURG; Ross, Timothy J [UNM

    2010-10-07

    Subject matter expert assessments can include both assignment and linguistic uncertainty. This paper examines assessments containing linguistic uncertainty associated with a qualitative description of a specific state of interest and the assignment uncertainty associated with assigning a qualitative value to that state. A Bayesian approach is examined to simultaneously quantify both assignment and linguistic uncertainty in the posterior probability. The approach is applied to a simplified damage assessment model involving both assignment and linguistic uncertainty. The utility of the approach and the conditions under which the approach is feasible are examined and identified.

  14. Filtering high-throughput protein-protein interaction data using a combination of genomic features

    Directory of Open Access Journals (Sweden)

    Patil Ashwini

    2005-04-01

    Full Text Available Abstract Background Protein-protein interaction data used in the creation or prediction of molecular networks is usually obtained from large scale or high-throughput experiments. This experimental data is liable to contain a large number of spurious interactions. Hence, there is a need to validate the interactions and filter out the incorrect data before using them in prediction studies. Results In this study, we use a combination of 3 genomic features – structurally known interacting Pfam domains, Gene Ontology annotations and sequence homology – as a means to assign reliability to the protein-protein interactions in Saccharomyces cerevisiae determined by high-throughput experiments. Using Bayesian network approaches, we show that protein-protein interactions from high-throughput data supported by one or more genomic features have a higher likelihood ratio and hence are more likely to be real interactions. Our method has a high sensitivity (90% and good specificity (63%. We show that 56% of the interactions from high-throughput experiments in Saccharomyces cerevisiae have high reliability. We use the method to estimate the number of true interactions in the high-throughput protein-protein interaction data sets in Caenorhabditis elegans, Drosophila melanogaster and Homo sapiens to be 27%, 18% and 68% respectively. Our results are available for searching and downloading at http://helix.protein.osaka-u.ac.jp/htp/. Conclusion A combination of genomic features that include sequence, structure and annotation information is a good predictor of true interactions in large and noisy high-throughput data sets. The method has a very high sensitivity and good specificity and can be used to assign a likelihood ratio, corresponding to the reliability, to each interaction.

  15. Regulatory focus and the assignment of punishment

    Directory of Open Access Journals (Sweden)

    Chloe Carmichael

    2007-06-01

    Full Text Available Regulatory Focus has been demonstrated to influence human behavior in a number of domains, such as object valuation and readiness to commit time or money to social projects. It has also been demonstrated to influence an individual’s approach to mistakes; and a person’s preference for global or local processing of information. The present work seeks to consider how regulatory focus might interact with punitive behaviors, specifically, the assignment of legal punishment. In this study, 240 undergraduates completed a series of written instruments that assessed their regulatory focus. They read a vignette that described a target that commits a crime, is detected by the police, and is arrested due to a careless mistake. Participants were asked what level of legal punishment they deemed appropriate. Participants’ punitive evaluations show that there are significant interactions a between the regulatory focus of the participant and the regulatory focus of the target and b between the regulatory focus of the participant and the level of detail used to describe the target and her behavior. In each case, when the regulatory foci matched, causing ‘fit,’ the participant was more lenient than in the non-fit condition.

  16. Reference databases for taxonomic assignment in metagenomics.

    Science.gov (United States)

    Santamaria, Monica; Fosso, Bruno; Consiglio, Arianna; De Caro, Giorgio; Grillo, Giorgio; Licciulli, Flavio; Liuni, Sabino; Marzano, Marinella; Alonso-Alemany, Daniel; Valiente, Gabriel; Pesole, Graziano

    2012-11-01

    Metagenomics is providing an unprecedented access to the environmental microbial diversity. The amplicon-based metagenomics approach involves the PCR-targeted sequencing of a genetic locus fitting different features. Namely, it must be ubiquitous in the taxonomic range of interest, variable enough to discriminate between different species but flanked by highly conserved sequences, and of suitable size to be sequenced through next-generation platforms. The internal transcribed spacers 1 and 2 (ITS1 and ITS2) of the ribosomal DNA operon and one or more hyper-variable regions of 16S ribosomal RNA gene are typically used to identify fungal and bacterial species, respectively. In this context, reliable reference databases and taxonomies are crucial to assign amplicon sequence reads to the correct phylogenetic ranks. Several resources provide consistent phylogenetic classification of publicly available 16S ribosomal DNA sequences, whereas the state of ribosomal internal transcribed spacers reference databases is notably less advanced. In this review, we aim to give an overview of existing reference resources for both types of markers, highlighting strengths and possible shortcomings of their use for metagenomics purposes. Moreover, we present a new database, ITSoneDB, of well annotated and phylogenetically classified ITS1 sequences to be used as a reference collection in metagenomic studies of environmental fungal communities. ITSoneDB is available for download and browsing at http://itsonedb.ba.itb.cnr.it/.

  17. A Review on Eigenstructure Assignment Methods and Orthogonal Eigenstructure Control of Structural Vibrations

    Directory of Open Access Journals (Sweden)

    Mohammad Rastgaar

    2009-01-01

    Full Text Available This paper provides a state-of-the-art review of eigenstructure assignment methods for vibration cancellation. Eigenstructure assignment techniques have been widely used during the past three decades for vibration suppression in structures, especially in large space structures. These methods work similar to mode localization in which global vibrations are managed such that they remain localized within the structure. Such localization would help reducing vibrations more effectively than other methods of vibration cancellation, by virtue of confining the vibrations close to the source of disturbance. The common objective of different methods of eigenstructure assignment is to provide controller design freedom beyond pole placement, and define appropriate shapes for the eigenvectors of the systems. These methods; however, offer a large and complex design space of options that can often overwhelm the control designer. Recent developments in orthogonal eigenstructure control offers a significant simplification of the design task while allowing some experience-based design freedom. The majority of the papers from the past three decades in structural vibration cancellation using eigenstructure assignment methods are reviewed, along with recent studies that introduce new developments in eigenstructure assignment techniques.

  18. From Story to Analysis: Reflection and Uptake in the Literacy Narrative Assignment

    Science.gov (United States)

    Alexander, Kara Poe

    2015-01-01

    The literacy narrative assignment is popular with composition instructors because of the reflection it encourages in students. Previously, scholars have claimed that students demonstrate reflection in literacy narratives when they critique dominant ideologies. Largely absent, however, is research on what other elements might indicate reflection…

  19. Nitrogen-detected TROSY yields comparable sensitivity to proton-detected TROSY for non-deuterated, large proteins under physiological salt conditions

    Energy Technology Data Exchange (ETDEWEB)

    Takeuchi, Koh [National Institute for Advanced Industrial Science and Technology, Molecular Profiling Research Center for Drug Discovery (Japan); Arthanari, Haribabu [Harvard Medical School, Department of Biochemistry and Molecular Pharmacology (United States); Imai, Misaki [Japan Biological Informatics Consortium, Research and Development Department (Japan); Wagner, Gerhard, E-mail: gerhard-wagner@hms.harvard.edu [Harvard Medical School, Department of Biochemistry and Molecular Pharmacology (United States); Shimada, Ichio, E-mail: shimada@iw-nmr.f.u-tokyo.ac.jp [National Institute for Advanced Industrial Science and Technology, Molecular Profiling Research Center for Drug Discovery (Japan)

    2016-02-15

    Direct detection of the TROSY component of proton-attached {sup 15}N nuclei ({sup 15}N-detected TROSY) yields high quality spectra with high field magnets, by taking advantage of the slow {sup 15}N transverse relaxation. The slow transverse relaxation and narrow line width of the {sup 15}N-detected TROSY resonances are expected to compensate for the inherently low {sup 15}N sensitivity. However, the sensitivity of {sup 15}N-detected TROSY in a previous report was one-order of magnitude lower than in the conventional {sup 1}H-detected version. This could be due to the fact that the previous experiments were performed at low salt (0–50 mM), which is advantageous for {sup 1}H-detected experiments. Here, we show that the sensitivity gap between {sup 15}N and {sup 1}H becomes marginal for a non-deuterated, large protein (τ{sub c} = 35 ns) at a physiological salt concentration (200 mM). This effect is due to the high salt tolerance of the {sup 15}N-detected TROSY. Together with the previously reported benefits of the {sup 15}N-detected TROSY, our results provide further support for the significance of this experiment for structural studies of macromolecules when using high field magnets near and above 1 GHz.

  20. Y-box-binding protein-1 (YB-1) promotes cell proliferation, adhesion and drug resistance in diffuse large B-cell lymphoma.

    Science.gov (United States)

    Miao, Xiaobing; Wu, Yaxun; Wang, Yuchan; Zhu, Xinghua; Yin, Haibing; He, Yunhua; Li, Chunsun; Liu, Yushan; Lu, Xiaoyun; Chen, Yali; Shen, Rong; Xu, Xiaohong; He, Song

    2016-08-15

    YB-1 is a multifunctional protein, which has been shown to correlate with resistance to treatment of various tumor types. This study investigated the expression and biologic function of YB-1 in diffuse large B-cell lymphoma (DLBCL). Immunohistochemical analysis showed that the expression statuses of YB-1 and pYB-1(S102) were reversely correlated with the clinical outcomes of DLBCL patients. In addition, we found that YB-1 could promote the proliferation of DLBCL cells by accelerating the G1/S transition. Ectopic expression of YB-1 could markedly increase the expression of cell cycle regulators cyclin D1 and cyclin E. Furthermore, we found that adhesion of DLBCL cells to fibronectin (FN) could increase YB-1 phosphorylation at Ser102 and pYB-1(S102) nuclear translocation. In addition, overexpression of YB-1 could increase the adhesion of DLBCL cells to FN. Intriguingly, we found that YB-1 overexpression could confer drug resistance through cell-adhesion dependent and independent mechanisms in DLBCL. Silencing of YB-1 could sensitize DLBCL cells to mitoxantrone and overcome cell adhesion-mediated drug resistance (CAM-DR) phenotype in an AKT-dependent manner. PMID:27397581

  1. Protein kinase CK2 is widely expressed in follicular, Burkitt and diffuse large B-cell lymphomas and propels malignant B-cell growth.

    Science.gov (United States)

    Pizzi, Marco; Piazza, Francesco; Agostinelli, Claudio; Fuligni, Fabio; Benvenuti, Pietro; Mandato, Elisa; Casellato, Alessandro; Rugge, Massimo; Semenzato, Gianpietro; Pileri, Stefano A

    2015-03-30

    Serine-threonine kinase CK2 is highly expressed and pivotal for survival and proliferation in multiple myeloma, chronic lymphocytic leukemia and mantle cell lymphoma. Here, we investigated the expression of α catalytic and β regulatory CK2 subunits by immunohistochemistry in 57 follicular (FL), 18 Burkitt (BL), 52 diffuse large B-cell (DLBCL) non-Hodgkin lymphomas (NHL) and in normal reactive follicles. In silico evaluation of available Gene Expression Profile (GEP) data sets from patients and Western blot (WB) analysis in NHL cell-lines were also performed. Moreover, the novel, clinical-grade, ATP-competitive CK2-inhibitor CX-4945 (Silmitasertib) was assayed on lymphoma cells. CK2 was detected in 98.4% of cases with a trend towards a stronger CK2α immunostain in BL compared to FL and DLBCL. No significant differences were observed between Germinal Center B (GCB) and non-GCB DLBCL types. GEP data and WB confirmed elevated CK2 mRNA and protein levels as well as active phosphorylation of specific targets in NHL cells. CX-4945 caused a dose-dependent growth-arresting effect on GCB, non-GCB DLBCL and BL cell-lines and it efficiently shut off phosphorylation of NF-κB RelA and CDC37 on CK2 target sites. Thus, CK2 is highly expressed and could represent a suitable therapeutic target in BL, FL and DLBCL NHL.

  2. Large genomic fragment deletion and functional gene cassette knock-in via Cas9 protein mediated genome editing in one-cell rodent embryos.

    Science.gov (United States)

    Wang, Liren; Shao, Yanjiao; Guan, Yuting; Li, Liang; Wu, Lijuan; Chen, Fangrui; Liu, Meizhen; Chen, Huaqing; Ma, Yanlin; Ma, Xueyun; Liu, Mingyao; Li, Dali

    2015-01-01

    The CRISPR-Cas RNA-guided system has versatile uses in many organisms and allows modification of multiple target sites simultaneously. Generating novel genetically modified mouse and rat models is one valuable application of this system. Through the injection of Cas9 protein instead of mRNA into embryos, we observed fewer off-target effects of Cas9 and increased point mutation knock-in efficiency. Large genomic DNA fragment (up to 95 kb) deletion mice were generated for in vivo study of lncRNAs and gene clusters. Site-specific insertion of a 2.7 kb CreERT2 cassette into the mouse Nfatc1 locus allowed labeling and tracing of hair follicle stem cells. In addition, we combined the Cre-Loxp system with a gene-trap strategy to insert a GFP reporter in the reverse orientation into the rat Lgr5 locus, which was later inverted by Cre-mediated recombination, yielding a conditional knockout/reporter strategy suitable for mosaic mutation analysis. PMID:26620761

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

    KAUST Repository

    Abbas, Ahmed

    2014-04-19

    Despite significant advances in automated nuclear magnetic resonance-based protein structure determination, the high numbers of false positives and false negatives among the peaks selected by fully automated methods remain a problem. These false positives and negatives impair the performance of resonance assignment methods. One of the main reasons for this problem is that the computational research community often considers peak picking and resonance assignment to be two separate problems, whereas spectroscopists use expert knowledge to pick peaks and assign their resonances at the same time. We propose a novel framework that simultaneously conducts slice picking and spin system forming, an essential step in resonance assignment. Our framework then employs a genetic algorithm, directed by both connectivity information and amino acid typing information from the spin systems, to assign the spin systems to residues. The inputs to our framework can be as few as two commonly used spectra, i.e., CBCA(CO)NH and HNCACB. Different from the existing peak picking and resonance assignment methods that treat peaks as the units, our method is based on \\'slices\\', which are one-dimensional vectors in three-dimensional spectra that correspond to certain (N, H) values. Experimental results on both benchmark simulated data sets and four real protein data sets demonstrate that our method significantly outperforms the state-of-the-art methods while using a less number of spectra than those methods. Our method is freely available at http://sfb.kaust.edu.sa/Pages/Software.aspx. © 2014 Springer Science+Business Media.

  4. Isolation of cross-linked peptides by diagonal strong cation exchange chromatography for protein complex topology studies by peptide fragment fingerprinting from large sequence databases

    NARCIS (Netherlands)

    H. Buncherd; W. Roseboom; B. Ghavim; W. Du; L.J. de Koning; C.G. de Koster; L. de Jong

    2014-01-01

    Knowledge of spatial proximity of amino acid residues obtained by chemical cross-linking and mass spectrometric analysis provides information about protein folding, protein-protein interactions and topology of macromolecular assemblies. We show that the use of bis(succinimidyl)-3-azidomethyl glutara

  5. Introducing chemical biology applications to introductory organic chemistry students using series of weekly assignments.

    Science.gov (United States)

    Kanin, Maralee R; Pontrello, Jason K

    2016-03-01

    Calls to bring interdisciplinary content and examples into introductory science courses have increased, yet strategies that involve course restructuring often suffer from the need for a significant faculty commitment to motivate change. Minimizing the need for dramatic course reorganization, the structure, reactivity, and chemical biology applications of classes of biological monomers and polymers have been integrated into introductory organic chemistry courses through three series of semester-long weekly assignments that explored (a) Carbohydrates and Oligosaccharides, (b) Amino Acids, Peptides, and Proteins, and (c) Nucleosides, Nucleotides, and Nucleic Acids. Comparisons of unannounced pre- and post tests revealed improved understanding of a reaction introduced in the assignments, and course examinations evaluated cumulative assignment topics. Course surveys revealed that demonstrating biologically relevant applications consistently throughout the semesters enhanced student interest in the connection between basic organic chemistry content and its application to new and unfamiliar bio-related examples. Covering basic material related to these classes of molecules outside of the classroom opened lecture time to allow the instructor to further build on information developed through the weekly assignments, teaching advanced topics and applications typically not covered in an introductory organic chemistry lecture course. Assignments were implemented as homework, either with or without accompanying discussion, in both laboratory and lecture organic courses within the context of the existing course structures. © 2015 by The International Union of Biochemistry and Molecular Biology, 44:168-178, 2016. PMID:26560414

  6. Graphical analysis of NMR structural quality and interactive contact map of NOE assignments in ARIA

    Directory of Open Access Journals (Sweden)

    Malliavin Thérèse E

    2008-06-01

    Full Text Available Abstract Background The Ambiguous Restraints for Iterative Assignment (ARIA approach is widely used for NMR structure determination. It is based on simultaneously calculating structures and assigning NOE through an iterative protocol. The final solution consists of a set of conformers and a list of most probable assignments for the input NOE peak list. Results ARIA was extended with a series of graphical tools to facilitate a detailed analysis of the intermediate and final results of the ARIA protocol. These additional features provide (i an interactive contact map, serving as a tool for the analysis of assignments, and (ii graphical representations of structure quality scores and restraint statistics. The interactive contact map between residues can be clicked to obtain information about the restraints and their contributions. Profiles of quality scores are plotted along the protein sequence, and contact maps provide information of the agreement with the data on a residue pair level. Conclusion The graphical tools and outputs described here significantly extend the validation and analysis possibilities of NOE assignments given by ARIA as well as the analysis of the quality of the final structure ensemble. These tools are included in the latest version of ARIA, which is available at http://aria.pasteur.fr. The Web site also contains an installation guide, a user manual and example calculations.

  7. An Eigenstructure Assignment for a Static Synchronous Compensator

    Directory of Open Access Journals (Sweden)

    Ahmad N. Al-Husban

    2009-01-01

    Full Text Available Problem statement: Power flow through an AC transmission line is influenced by three basic electrical parameters, which are line impedance, magnitudes and phase-shift angle between the sending and receiving voltages. Therefore, the change in any of the three basic parameters means a change in the power flow through the transmission line. The aims of this research paper are: increase the power transfer capability of transmission systems, minimize the transmission losses, support a good voltage profile and retain system stability under large disturbances. Study the use of eigenstructure techniques for state feedback control of the power system static compensator. Therefore, the mathematical analysis was performed for eigenvector assignment, power flow transmission line and for the static compensator analysis based on the transformation of the three-phase into d-q frame. Approach: A novel control method for regulating the power system in case of abnormal conditions was carried out. The system considered is a static synchronous compensator. The study includes a detailed mathematical analysis of the impact of the shunt compensator on the power flow; investigation of the system constraints and their effects on the static compensator control; in addition simulation of static compensator to control a transmitted active power flow on the transmission line. The conducted method provides a way of constructing the state feedback gain matrix to satisfy a certain prescribed performance. Results: The solutions of the obtained equation were conducted using the computer simulation method for both open-loop and static compensator techniques. The result shows fast tracking of the power flow transient response when using the static compensator technique comparing with open-loop technique. However, the same trend of the behavior was observed for all cases. Conclusion: A new method for developing a parameterized feedback matrix that assigns a closed-loop prespecified

  8. The prognostic significance of STAT3 in invasive breast cancer: analysis of protein and mRNA expressions in large cohorts.

    Science.gov (United States)

    Aleskandarany, Mohammed A; Agarwal, Devika; Negm, Ola H; Ball, Graham; Elmouna, Ahmed; Ashankyty, Ibraheem; Nuglozeh, Edem; Fazaludeen, Mohammad F; Diez-Rodriguez, Maria; Nolan, Christopher C; Tighe, Patrick J; Green, Andrew R; Ellis, Ian O; Rakha, Emad A

    2016-02-01

    Signal transducer and activator of transcription (STAT) transcription factors family are involved in diverse cellular biological functions. Reports regarding the prognostic impact of STAT3 expression in breast cancer (BC) are variable whether being a factor of poor or good prognosis. Immunohistochemical expression of phospho-STAT3 (pSTAT3) was studied in large series of invasive BC (n = 1270). pSTAT3 and STAT3 were quantified using reverse phase protein array (RPPA) on proteins extracted from macro-dissected FFPE tissues (n = 49 cases). STAT3 gene expression in the METABRIC cohort was also investigated. STAT3 gene expression prognostic impact was externally validated using the online BC gene expression data (n = 26 datasets, 4.177 patients). pSTAT3 was expressed in the nuclei and cytoplasm of invasive BC cells. Nuclear pSTAT3 overexpression was positively associated with smaller tumour size, lower grade, good NPI, negative lymphovascular invasion (LVI), ER+, PgR+, p53-, HER2-, and low Ki67LI and an improved breast cancer-specific survival (BCSS), independently of other factors. On RPPA, the mean pSTAT3 and STAT3 expressions were higher in ER+, PgR+, and smaller size tumours. Higher STAT3 transcripts in the METABRIC cohort were observed in cases with favourable prognostic criteria and as well as improved BCSS within the whole cohort, ER+ cohort with and without hormonal therapy, and ER- cohort including those who did not receive adjuvant chemotherapy. Pooled STAT3 gene expression data in the external validation cohort showed an association with improved patients' outcome (P < 0.001, HR = 0.84, 95 % CI 0.79-0.90). Results of this study suggest nuclear localisation of pSTAT3 as favourable prognostic marker in invasive BC, results re-enforced by analysis of STAT3 gene expression data. This good prognostic advantage was maintained in patients who received and who did not receive adjuvant therapy. Therefore, STAT3 could have context-dependent molecular roles of in BC

  9. Understanding protein–protein interactions by genetic suppression

    Indian Academy of Sciences (India)

    Sitaraman Sujatha; Dipankar Chatterji

    2000-01-01

    Protein–protein interactions influence many cellular processes and it is increasingly being felt that even a weak and remote interplay between two subunits of a protein or between two proteins in a complex may govern the fate of a particular biochemical pathway. In a bacterial system where the complete genome sequence is available, it is an arduous task to assign function to a large number of proteins. It is possible that many of them are peripherally associated with a cellular event and it is very difficult to probe such interaction. However, mutations in the genes that encode such proteins (primary mutations) are useful in these studies. Isolation of a suppressor or a second-site mutation that restores the phenotype abolished by the primary mutation could be an elegant yet simple way to follow a set of interacting proteins. Such a reversion site need not necessarily be geometrically close to the primary mutation site.

  10. Cationic lipid-formulated DNA vaccine against hepatitis B virus: immunogenicity of MIDGE-Th1 vectors encoding small and large surface antigen in comparison to a licensed protein vaccine.

    Directory of Open Access Journals (Sweden)

    Anne Endmann

    Full Text Available Currently marketed vaccines against hepatitis B virus (HBV based on the small (S hepatitis B surface antigen (HBsAg fail to induce a protective immune response in about 10% of vaccinees. DNA vaccination and the inclusion of PreS1 and PreS2 domains of HBsAg have been reported to represent feasible strategies to improve the efficacy of HBV vaccines. Here, we evaluated the immunogenicity of SAINT-18-formulated MIDGE-Th1 vectors encoding the S or the large (L protein of HBsAg in mice and pigs. In both animal models, vectors encoding the secretion-competent S protein induced stronger humoral responses than vectors encoding the L protein, which was shown to be retained mainly intracellularly despite the presence of a heterologous secretion signal. In pigs, SAINT-18-formulated MIDGE-Th1 vectors encoding the S protein elicited an immune response of the same magnitude as the licensed protein vaccine Engerix-B, with S protein-specific antibody levels significantly higher than those considered protective in humans, and lasting for at least six months after the third immunization. Thus, our results provide not only the proof of concept for the SAINT-18-formulated MIDGE-Th1 vector approach but also confirm that with a cationic-lipid formulation, a DNA vaccine at a relatively low dose can elicit an immune response similar to a human dose of an aluminum hydroxide-adjuvanted protein vaccine in large animals.

  11. Backbone and sidechain 1H, 15N and 13C assignments of Tyrosine Phosphatase related to Biofilm formation A (TpbA) of Pseudomonas aeruginosa

    OpenAIRE

    Koveal, Dorothy; Jayasundera, Thusitha B.; Wood, Thomas K.; Peti, Wolfgang; Page, Rebecca

    2012-01-01

    The backbone and side chain resonance assignments of the Tyrosine Phosphatase related to Biofilm formation A (TpbA) of Pseudomonas aeruginosa have been determined based on triple-resonance experiments using uniformly [13C,15N]-labeled protein. This assignment is the first step towards the determination of the 3-dimensional structure of TpbA.

  12. Complete 1H, 15N and 13C assignment of trappin-2 and 1H assignment of its two domains, elafin and cementoin.

    Science.gov (United States)

    Loth, Karine; Alami, Soha Abou Ibrahim; Habès, Chahrazed; Garrido, Solène; Aucagne, Vincent; Delmas, Agnès F; Moreau, Thierry; Zani, Marie-Louise; Landon, Céline

    2016-04-01

    Trappin-2 is a serine protease inhibitor with a very narrow inhibitory spectrum and has significant anti-microbial activities. It is a 10 kDa cationic protein composed of two distinct domains. The N-terminal domain (38 residues) named cementoin is known to be intrinsically disordered when it is not linked to the elafin. The C-terminal domain (57 residues), corresponding to elafin, is a cysteine-rich domain stabilized by four disulfide bridges and is characterized by a flat core and a flexible N-terminal part. To our knowledge, there is no structural data available on trappin-2. We report here the complete (1)H, (15)N and (13)C resonance assignment of the recombinant trappin-2 and the (1)H assignments of cementoin and elafin, under the same experimental conditions. This is the first step towards the 3D structure determination of the trappin-2.

  13. Why the Rhetoric of CS Programming Assignments Matters

    Science.gov (United States)

    Wolfe, Joanna

    2004-06-01

    Despite the multiple potential benefits of asking students working on programming tasks to consider human factors, most programming assignments narrowly focus on technical details and requirements. Female students in particular may be attracted to assignments that emphasize human as well as technical factors. To assess how students respond to changes in the rhetorical presentation of programming instructions, 81 students completed questionnaires evaluating different assignment instructions. Students generally perceived assignments emphasizing real-world contexts and users as more motivating and enjoyable to program than those that did not emphasize human factors. Moreover, when asked what makes a "good" programming assignment, over half of the students volunteered that they looked for assignments stressing a real-world purpose, use or application.

  14. Using codon optimization, chaperone co-expression, and rational mutagenesis for production and NMR assignments of human eIF2α

    International Nuclear Information System (INIS)

    Producing a well behaved sample at high concentration is one of the main hurdles when starting a new project on an interesting protein. Especially when one attempts to overexpress a eukaryotic protein in bacteria, some difficulties are encountered, such as low expression level, low solubility, or even lack of folded structure. Overexpression in prokaryotic systems is highly desirable for cost-effective production of different isotope-labeled samples needed for NMR studies. Here we describe generally applicable methods for obtaining highly concentrated protein samples efficiently. This approach was developed as we tried to produce a NMR-suitable sample of the 35 kDa human translation initiation factor eIF2α, a protein that expresses poorly in E. coli and has very low solubility. First, an E. coli codon-optimized gene was synthesized on a thermal cycler, which increased the expression level by a factor of two. Second, we used co-expression of bacterial chaperone proteins, which largely increased the fraction of correctly folded protein found in the soluble phase. Third, we used rational mutagenesis guided by both the sequence alignment among homologues and the homology of one domain to a known fold for improving solubility and stability of the target protein by tenfold. Combining all these methods made it possible to produce from a one-liter preparation a 0.5 mM sample of human eIF2α that showed well-resolved NMR spectra and enabled nearly complete assignment of the protein. These methods may be generally useful for studies of other eukaryotic proteins that are otherwise difficult to express and exhibit poor solubility

  15. Electronic marking of mathematics assignments using Microsoft Word 2007

    OpenAIRE

    Lowe, Tim; Mestel, Ben; Arrowsmith, Gaynor

    2008-01-01

    This paper describes on-going work within the Department of Mathematics and Statistics at The Open University to enable distance learning students to electronically submit assignments rich in mathematical notation and diagrams, and for those assignments to be marked and returned electronically by their tutor. A trial is currently underway of a prototype system that enables students to submit assignments in a range of electronic formats, which are then converted to Microsoft Word 2007 format t...

  16. Strategies for Effective Comments on Students’ Written Assignments

    Institute of Scientific and Technical Information of China (English)

    任永东

    2014-01-01

    Comments on students’ assignments are important for English course. The loopholes in teachers’ working schemes-com-ments of their students’ written assignments are the clues around which the paper spreads. In the process of teaching, some English teach-ers pay so much attention to the contents taught in class but ignore the comments on students’ written assignments. Consequently, it is not applicable for students to correct their mistakes in time and they probably make the same mistakes repeatedly.

  17. Factors of Success of Female Expatriates During International Assignments

    OpenAIRE

    Lapinoja, Anu

    2012-01-01

    The aim of this bachelor’s thesis was to study what the factors of success of female expatriates are during international assignments. Also, the objective was to find out how companies could support and enhance these factors in the future and what kind of barriers of success there exist for female expatriates. The theoretical study examined international assignments in general, expatriates, success and failure of international assignments and the barriers of success. Also, female expatriates ...

  18. Integrated Project Scheduling and Staff Assignment with Controllable Processing Times

    OpenAIRE

    Victor Fernandez-Viagas; Framinan, Jose M.

    2014-01-01

    This paper addresses a decision problem related to simultaneously scheduling the tasks in a project and assigning the staff to these tasks, taking into account that a task can be performed only by employees with certain skills, and that the length of each task depends on the number of employees assigned. This type of problems usually appears in service companies, where both tasks scheduling and staff assignment are closely related. An integer programming model for the problem is proposed, tog...

  19. An Improved Point-track Optimal Assignment Algorithm

    OpenAIRE

    Zhonglei Zhang; Weihua Zhang; Li Zhou

    2013-01-01

    In order to improve the accuracy of data association of the Optimal Assignment (OA) algorithm based on dynamic information, an improved Point-Track Optimal Assignment (IPTOA) algorithm based on multi-source information is proposed. The improved algorithm gets valid 3-tuple of measurement set by solving 3-Dimensional (3-D) assignment problem which is based on dynamic information. Then fuses multi-source information by combination rule of D-S evidence theory and constructs the point-track corre...

  20. CYCLE TIMES ASSIGNMENT OF NONLINEAR DISCRETE EVENT DYNAMIC SYSTEMS

    Institute of Scientific and Technical Information of China (English)

    CHEN Wende

    2000-01-01

    In this paper, nonautonomous models of Discrete Event Dynamic Systems (DEDS) are established by min-max function, reachability and observability are defined,the problem on cycle times assignment of DEDS, which corresponds with the important problem on poles assignment of linear systems, is studied. By Gunawardena et al.'Duality Theorem following results are obtained: Cycle times of system can be assigned under state feedback(or output feedback) if and only if system is reachable (or reachable and obserbable).

  1. An efficient and impartial online algorithm for kidney assignment network

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    An online algorithm balancing the efficiency and equity principles is proposed for the kidney resource assignment when only the current patient and resource information is known to the assignment network. In the algorithm, the assignment is made according to the priority, which is calculated according to the efficiency principle and the equity principle. The efficiency principle is concerned with the post-transplantation immunity spending caused by the possible post-operation immunity rejection and patient’...

  2. Impact of English-language Proficiency on Chinese Expatriates’ Adjustment to Overseas Assignments

    Directory of Open Access Journals (Sweden)

    Xu, Xiaofei

    2012-01-01

    Full Text Available With the emergence of Chinese multinationals, increasingly large number of Chinese expatriates work overseas. However, little research has been conducted to investigate Chinese expatriates’ adjustment to overseas assignments. Drawing on prior studies in the areas of linguistics, communication, and business, the current study examines the impact of the English-language proficiency on the Chinese expatriates’ adjustment to overseas assignments.A Chinese multinational in the information technology industry, Company ABC, was selected as a subject company. Quantitative research method was adopted. The quantitative data were collected through a survey of 190 Chinese expatriates from Company ABC. Pearson’s Correlation Coefficient was performed to examine the relationships between English-language proficiency and variables related to the Chinese Expatriates’ Adjustment to Overseas Assignments. The findings show that English-language proficiency was found to enhance Chinese expatriates’ adjustment moderately. Moreover, further analysis indicates that different type of English-language proficiency affects different aspect of Chinese expatriates’ adjustment.

  3. Ant Colony Algorithm and Simulation for Robust Airport Gate Assignment

    Directory of Open Access Journals (Sweden)

    Hui Zhao

    2014-01-01

    Full Text Available Airport gate assignment is core task for airport ground operations. Due to the fact that the departure and arrival time of flights may be influenced by many random factors, the airport gate assignment scheme may encounter gate conflict and many other problems. This paper aims at finding a robust solution for airport gate assignment problem. A mixed integer model is proposed to formulate the problem, and colony algorithm is designed to solve this model. Simulation result shows that, in consideration of robustness, the ability of antidisturbance for airport gate assignment scheme has much improved.

  4. Using interviews to understand the assignment mechanism in a nonexperimental study: the case of eighth grade algebra.

    Science.gov (United States)

    Rickles, Jordan H

    2011-10-01

    Many inquiries regarding the causal effects of policies or programs are based on research designs where the treatment assignment process is unknown, and thus valid inferences depend on tenuous assumptions about the assignment mechanism. This article draws attention to the importance of understanding the assignment mechanism in policy and program evaluation studies, and illustrates how information collected through interviews can develop a richer understanding of the assignment mechanism. Focusing on the issue of student assignment to algebra in 8th grade, I show how a preliminary data collection effort aimed at understanding the assignment mechanism is particularly beneficial in multisite observational studies in education. The findings, based on ten interviews and administrative data from a large school district, draw attention to the often ignored heterogeneity in the assignment mechanism across schools. These findings likely extend beyond the current research project in question to related educational policy issues such as ability grouping, tracking, differential course taking, and curricular intensity, as well as other social programs in which the assignment mechanism can differ across sites.

  5. Fast and Accurate Identification of Cross-Linked Peptides for the Structural Analysis of Large Protein Complexes and Elucidation of Interaction Networks. / Tahir, Salman; Bukowski-Wills, Jimi-Carlo; Rasmussen, Morten; Rappsilber, Juri

    DEFF Research Database (Denmark)

    Rasmussen, Morten

    Fast and Accurate Identification of Cross-Linked Peptides for the structural analysis of large protein complexes and to elucidate interaction networks. Salman Tahir Jimi-Carlo Bukowski-Wills; Morten Rasmussen; Juri RappsilberWellcome Trust Centre for Cell Biology, Edinburgh , United Kingdom   Novel...

  6. Development of enrichment methods for cross-linked peptides to study the dynamic topology of large protein complexes by mass spectrometry

    NARCIS (Netherlands)

    H. Buncherd

    2013-01-01

    In the cell, most proteins carry out their functions as subunits of larger assemblies. Knowledge about the folding and mutual interactions of composing subunits is crucial to comprehend the dynamics and structures of such protein complexes and the molecular mechanisms underlying biological processes

  7. Active learning in pre-class assignments: Exploring the use of interactive simulations to enhance reading assignments

    CERN Document Server

    Stang, Jared B; Perez, Sarah; Ives, Joss; Roll, Ido

    2016-01-01

    Pre-class reading assignments help prepare students for active classes by providing a first exposure to the terms and concepts to be used during class. We investigate if the use of inquiry-oriented PhET-based activities in conjunction with pre-class reading assignments can improve both the preparation of students for in-class learning and student attitudes towards and engagement with pre-class assignments. Over three course modules covering different topics, students were assigned randomly to complete either a textbook-only pre-class assignment or both a textbook pre-class assignment and a PhET-based activity. The assignments helped prepare students for class, as measured by performance on the pre-class quiz relative to a beginning-of-semester pre-test, but no evidence for increased learning due the PhET activity was observed. Students rated the assignments which included PhET as more enjoyable and, for the topic latest in the semester, reported engaging more with the assignments when PhET was included.

  8. Effects of NMR spectral resolution on protein structure calculation.

    Directory of Open Access Journals (Sweden)

    Suhas Tikole

    Full Text Available Adequate digital resolution and signal sensitivity are two critical factors for protein structure determinations by solution NMR spectroscopy. The prime objective for obtaining high digital resolution is to resolve peak overlap, especially in NOESY spectra with thousands of signals where the signal analysis needs to be performed on a large scale. Achieving maximum digital resolution is usually limited by the practically available measurement time. We developed a method utilizing non-uniform sampling for balancing digital resolution and signal sensitivity, and performed a large-scale analysis of the effect of the digital resolution on the accuracy of the resulting protein structures. Structure calculations were performed as a function of digital resolution for about 400 proteins with molecular sizes ranging between 5 and 33 kDa. The structural accuracy was assessed by atomic coordinate RMSD values from the reference structures of the proteins. In addition, we monitored also the number of assigned NOESY cross peaks, the average signal sensitivity, and the chemical shift spectral overlap. We show that high resolution is equally important for proteins of every molecular size. The chemical shift spectral overlap depends strongly on the corresponding spectral digital resolution. Thus, knowing the extent of overlap can be a predictor of the resulting structural accuracy. Our results show that for every molecular size a minimal digital resolution, corresponding to the natural linewidth, needs to be achieved for obtaining the highest accuracy possible for the given protein size using state-of-the-art automated NOESY assignment and structure calculation methods.

  9. 一种可用于大规模单克隆抗体纯化的新型Protein A填料%A novel Protein A resin used for large-scale purification of monoclonal antibody

    Institute of Scientific and Technical Information of China (English)

    陈晓虹; 吴建国; 曹传平; 唐亮; 曾贤; 范佳君; 李玉彬; 赵丽丽; 刘增田

    2014-01-01

    目的 比较不同型号Protein A填料的性能,筛选合适的Protein A填料,用于大规模单克隆抗体(简称单抗)纯化.方法 采用表达H02单抗的CHO细胞培养上清,测试不同供应商Protein A填料的蛋白结合载量,选择合适的填料,用于大规模单抗纯化;检测宿主细胞蛋白(host cell protein,HCP)、外源DNA及Protein A残留量,以评价其纯化效果;检测Protein A柱在位清洗(cleaning in place,CIP)效果,并验证其循环使用次数.结果 Amsphere Protein A JWT203填料为合适的ProteinA填料,H02单抗大规模纯化后,可有效去除HCP、DNA及脱落Protein A;采用0.1 mol/LNaOH进行CIP,经过300个循环,对H02单抗仍保持80%以上动态结合载量,Protein A脱落无明显增加,对HCP、外源DNA、聚合体的去除能力无明显改变,层析图谱与初始基本一致,Amsphere Protein A JWT203填料的理化性质较稳定.结论 AmsphereProtein A JWT203是一种可用于大规模单抗纯化的新型Protein A填料.

  10. The lowest-energy chlorophyll of photosystem II is adjacent to the peripheral antenna: Emitting states of CP47 assigned via circularly polarized luminescence.

    Science.gov (United States)

    Hall, Jeremy; Renger, Thomas; Müh, Frank; Picorel, Rafael; Krausz, Elmars

    2016-09-01

    The identification of low-energy chlorophyll pigments in photosystem II (PSII) is critical to our understanding of the kinetics and mechanism of this important enzyme. We report parallel circular dichroism (CD) and circularly polarized luminescence (CPL) measurements at liquid helium temperatures of the proximal antenna protein CP47. This assembly hosts the lowest-energy chlorophylls in PSII, responsible for the well-known "F695" fluorescence band of thylakoids and PSII core complexes. Our new spectra enable a clear identification of the lowest-energy exciton state of CP47. This state exhibits a small but measurable excitonic delocalization, as predicated by its CD and CPL. Using structure-based simulations incorporating the new spectra, we propose a revised set of site energies for the 16 chlorophylls of CP47. The significant difference from previous analyses is that the lowest-energy pigment is assigned as Chl 612 (alternately numbered Chl 11). The new assignment is readily reconciled with the large number of experimental observations in the literature, while the most common previous assignment for the lowest energy pigment, Chl 627(29), is shown to be inconsistent with CD and CPL results. Chl 612(11) is near the peripheral light-harvesting system in higher plants, in a lumen-exposed region of the thylakoid membrane. The low-energy pigment is also near a recently proposed binding site of the PsbS protein. This result consequently has significant implications for our understanding of the kinetics and regulation of energy transfer in PSII. PMID:27342201

  11. NMR assignments of the macro domain from Middle East respiratory syndrome coronavirus (MERS-CoV).

    Science.gov (United States)

    Huang, Yi-Ping; Cho, Chao-Cheng; Chang, Chi-Fon; Hsu, Chun-Hua

    2016-10-01

    The newly emerging human pathogen, Middle East respiratory syndrome coronavirus (MERS-CoV), contains a macro domain in the highly conserved N-terminal region of non-structural protein 3. Intense research has shown that macro domains bind ADP-ribose and other derivatives, but it still remains intangible about their exact function. In this study we report the preliminary structural analysis through solution NMR spectroscopy of the MERS-CoV macro domain. The near complete NMR assignments of MERS-CoV macro domain provide the basis for subsequent structural and biochemical investigation in the context of protein function.

  12. 45 CFR 302.50 - Assignment of rights to support.

    Science.gov (United States)

    2010-10-01

    ... process; or (2) Except for obligations assigned under 42 CFR 433.146, other legal process as established... competent jurisdiction or administrative process which covers the assigned support rights. (2) If there is no court or administrative order, an amount determined in writing by the IV-D agency as part of...

  13. 76 FR 34658 - The Internet Assigned Numbers Authority (IANA) Functions

    Science.gov (United States)

    2011-06-14

    ... of Inquiry, Request for Comments on the Internet Assigned Numbers Authority (IANA) Functions, 76 FR 10569 (Feb. 25, 2011), available at http://www.ntia.doc.gov/frnotices/2011/fr_ianafunctionsnoi_02252011... National Telecommunications and Information Administration The Internet Assigned Numbers Authority...

  14. On the Use of Writing Assignments in Intermediate Microeconomic Theory

    Science.gov (United States)

    O'Neill, Patrick B.

    2009-01-01

    A typical writing assignment in upper level required courses is a term paper. However many economics majors, particularly those in business schools, need to develop skill at writing shorter pieces. In this paper I describe numerous examples of shorter writing assignments that I have incorporated into an Intermediate Microeconomic Theory course.…

  15. 14 CFR 1245.109 - Assignment of title to NASA.

    Science.gov (United States)

    2010-01-01

    ... 14 Aeronautics and Space 5 2010-01-01 2010-01-01 false Assignment of title to NASA. 1245.109... INTELLECTUAL PROPERTY RIGHTS Patent Waiver Regulations § 1245.109 Assignment of title to NASA. (a) The instrument of waiver set forth in § 1245.115(c) shall be voided by NASA with respect to the domestic title...

  16. 47 CFR 52.111 - Toll free number assignment.

    Science.gov (United States)

    2010-10-01

    ... 47 Telecommunication 3 2010-10-01 2010-10-01 false Toll free number assignment. 52.111 Section 52...) NUMBERING Toll Free Numbers § 52.111 Toll free number assignment. Toll free numbers shall be made available on a first-come, first-served basis unless otherwise directed by the Commission....

  17. Genetics of traffic assignment models for strategic transport planning

    NARCIS (Netherlands)

    Bliemer, M.C.J.; Raadsen, M.; Brederode, L.; Bell, M.; Wismans, L.J.J.; Smith, M.

    2016-01-01

    This paper presents a review and classification of traffic assignment models for strategic transport planning purposes by using concepts analogous to genetics in biology. Traffic assignment models share the same theoretical framework (DNA), but differ in capability (genes). We argue that all traffic

  18. Dynamic Equilibrium Assignment Convergence by En-route Flow Smoothing

    NARCIS (Netherlands)

    Pel, A.J.; Bliemer, M.C.J.; Hoogendoorn, S.P.

    2010-01-01

    An essential feature in many dynamic traffic assignment (DTA) models used for planning purposes is to compute the (dynamic) equilibrium assignment, where travellers follow user-optimal routes, leading to minimal experienced route travel times. To compute these time-varying route flows in the equilib

  19. 24 CFR 203.350 - Assignment of mortgage.

    Science.gov (United States)

    2010-04-01

    ... 24 Housing and Urban Development 2 2010-04-01 2010-04-01 false Assignment of mortgage. 203.350... URBAN DEVELOPMENT MORTGAGE AND LOAN INSURANCE PROGRAMS UNDER NATIONAL HOUSING ACT AND OTHER AUTHORITIES SINGLE FAMILY MORTGAGE INSURANCE Contract Rights and Obligations Assignment of Mortgage §...

  20. 25 CFR 212.34 - Individual tribal assignments excluded.

    Science.gov (United States)

    2010-04-01

    ... 25 Indians 1 2010-04-01 2010-04-01 false Individual tribal assignments excluded. 212.34 Section... ALLOTTED LANDS FOR MINERAL DEVELOPMENT How To Acquire Leases § 212.34 Individual tribal assignments... made pursuant to tribal constitutions or ordinances for the use of individual Indians and assignees...

  1. Assignment Situations with Multiple Ownership and their Games

    NARCIS (Netherlands)

    Miquel, S.; van Velzen, S.; Hamers, H.J.M.; Norde, H.W.

    2005-01-01

    An assignment situation can be considered as a two-sided market consisting of two disjoint sets of objects.A non-negative reward matrix describes the profit if an object of one group is assigned to an object of the other group. Assuming that each object is owned by a different agent, Shapley and Shu

  2. Semi-Infinite Assignment Problems and Related Games

    NARCIS (Netherlands)

    Llorca, N.; Tijs, S.H.; Timmer, J.B.

    1999-01-01

    In 1972 Shapley and Shubik introduced assignment games associated to finite assignment problems in which two types of agents were involved and they proved that these games have a non-empty core. In this paper we look at the situation where the set of one type is infinite and investigatewhen the core

  3. Negotiating Languages and Cultures: Enacting Translingualism through a Translation Assignment

    Science.gov (United States)

    Kiernan, Julia; Meier, Joyce; Wang, Xiqiao

    2016-01-01

    This collaborative project explores the affordances of a translation assignment in the context of a learner-centered pedagogy that places composition students' movement among languages and cultures as both a site for inquiry and subject of analysis. The translation assignment asks students to translate scholarly articles or culture stories from…

  4. The Eco-Sculpture Assignment: Using Art to Scaffold Metacognition

    Science.gov (United States)

    Polegato, Rosemary

    2014-01-01

    The Eco-Sculpture Assignment demonstrates that art may be used as a conduit to scaffold metacognition in marketing courses. Theoretical underpinnings are drawn from the literature on pedagogy used in general, marketing, and art education contexts. The assignment is described in detail, followed by examples of learner response that illustrate…

  5. The Pedagogy of Assignments in Social Justice Teacher Education

    Science.gov (United States)

    McDonald, Morva A.

    2008-01-01

    This article examines the pedagogy of assignments in social justice teacher education programs. Employing a programmatic view, this study aims to understand the collective representation of social justice provided by assignments across multiple courses. Findings come from a qualitative case study of two social justice programs. Drawing on concepts…

  6. Graduate Writing Assignments across Faculties in a Canadian University

    Science.gov (United States)

    Shi, Ling; Dong, Yanning

    2015-01-01

    This study examines 143 graduate assignments across 12 faculties or schools in a Canadian university in order to identify types of writing tasks. Based on the descriptions provided by the instructors, we identified nine types of assignments, with scholarly essay being the most common, followed by summary and response, literature review, project,…

  7. Automating Formative and Summative Feedback for Individualised Assignments

    Science.gov (United States)

    Hamilton, Ian Robert

    2009-01-01

    Purpose: The purpose of this paper is to report on the rationale behind the use of a unique paper-based individualised accounting assignment, which automated the provision to students of immediate formative and timely summative feedback. Design/methodology/approach: As students worked towards completing their assignment, the package provided…

  8. Case Assignment in the Inalienable Possession Construction in Korean.

    Science.gov (United States)

    Maling, Joan; Kim, Soowon

    1992-01-01

    Investigates principles for assigning case to the Noun Phrases (NP) in the Part-Whole Construction in Korean. It is shown that the case marking on the part-NP is a function of the case-assigning properties of the matrix verb, even when this is lexically governed. (41 references) (Author/LB)

  9. 7 CFR 1530.115 - Paperwork Reduction Act assigned number.

    Science.gov (United States)

    2010-01-01

    ... 7 Agriculture 10 2010-01-01 2010-01-01 false Paperwork Reduction Act assigned number. 1530.115... SERVICE, DEPARTMENT OF AGRICULTURE THE REFINED SUGAR RE-EXPORT PROGRAM, THE SUGAR CONTAINING PRODUCTS RE-EXPORT PROGRAM, AND THE POLYHYDRIC ALCOHOL PROGRAM § 1530.115 Paperwork Reduction Act assigned...

  10. Individual Assignments and Academic Dishonesty--Exploring the Learning Conundrum

    Science.gov (United States)

    Leonard, Valorie; LeBrasseur, Rolland

    2008-01-01

    A survey of university business professors focused on their use of individual assignments in courses and their views on cheating and its impact on student learning. Based on responses from 456 professors (37% response rate) from Ontario, Canada, it was concluded that most faculty believe that individual assignments are effective learning tools and…

  11. 42 CFR 435.610 - Assignment of rights to benefits.

    Science.gov (United States)

    2010-10-01

    ... 42 Public Health 4 2010-10-01 2010-10-01 false Assignment of rights to benefits. 435.610 Section....610 Assignment of rights to benefits. (a) As a condition of eligibility, the agency must require... paternity and in obtaining medical support and payments, unless the individual establishes good cause...

  12. 42 CFR 436.610 - Assignment of rights to benefits.

    Science.gov (United States)

    2010-10-01

    ... 42 Public Health 4 2010-10-01 2010-10-01 false Assignment of rights to benefits. 436.610 Section... General Financial Eligibility Requirements and Options § 436.610 Assignment of rights to benefits. (a) As... payments, unless the individual establishes good cause for not cooperating, and except for...

  13. 48 CFR 1442.202 - Assignment of contract administration.

    Science.gov (United States)

    2010-10-01

    ... Assignment of contract administration. (a) The decision to withhold normal individual contract administration... 48 Federal Acquisition Regulations System 5 2010-10-01 2010-10-01 false Assignment of contract administration. 1442.202 Section 1442.202 Federal Acquisition Regulations System DEPARTMENT OF THE...

  14. Accounting for experimental noise reveals that mRNA levels, amplified by post-transcriptional processes, largely determine steady-state protein levels in yeast.

    Directory of Open Access Journals (Sweden)

    Gábor Csárdi

    2015-05-01

    Full Text Available Cells respond to their environment by modulating protein levels through mRNA transcription and post-transcriptional control. Modest observed correlations between global steady-state mRNA and protein measurements have been interpreted as evidence that mRNA levels determine roughly 40% of the variation in protein levels, indicating dominant post-transcriptional effects. However, the techniques underlying these conclusions, such as correlation and regression, yield biased results when data are noisy, missing systematically, and collinear---properties of mRNA and protein measurements---which motivated us to revisit this subject. Noise-robust analyses of 24 studies of budding yeast reveal that mRNA levels explain more than 85% of the variation in steady-state protein levels. Protein levels are not proportional to mRNA levels, but rise much more rapidly. Regulation of translation suffices to explain this nonlinear effect, revealing post-transcriptional amplification of, rather than competition with, transcriptional signals. These results substantially revise widely credited models of protein-level regulation, and introduce multiple noise-aware approaches essential for proper analysis of many biological phenomena.

  15. Multiobjective controller synthesis via eigenstructure assignment with state feedback

    Science.gov (United States)

    Li, Zhao; Lam, James

    2016-10-01

    A general parameter scheme for multiobjective controller synthesis via eigenstructure assignment with state feedback is proposed. The scheme provides total pole configurability, that is, pole assignment constraints, partial pole assignment constraints, generalised regional pole assignment constraints can be dealt with simultaneously without introducing essential conservatism. The scheme is derived from the pole assignment approach using Sylvester equations, and the parameter space is the Cartesian product of some subspaces characterising the free parameters. Under the scheme, the controller design problems are formulated as nonlinear optimisation problems with both objectives and constraints being differentiable and can be solved by derivative-based nonlinear programming technique. Numerical examples are given to illustrate the efficiency of the proposed method.

  16. Evaluation of a UMLS Auditing Process of Semantic Type Assignments

    Science.gov (United States)

    Gu, Huanying; Hripcsak, George; Chen, Yan; Morrey, C. Paul; Elhanan, Gai; Cimino, James J.; Geller, James; Perl, Yehoshua

    2007-01-01

    The UMLS is a terminological system that integrates many source terminologies. Each concept in the UMLS is assigned one or more semantic types from the Semantic Network, an upper level ontology for biomedicine. Due to the complexity of the UMLS, errors exist in the semantic type assignments. Finding assignment errors may unearth modeling errors. Even with sophisticated tools, discovering assignment errors requires manual review. In this paper we describe the evaluation of an auditing project of UMLS semantic type assignments. We studied the performance of the auditors who reviewed potential errors. We found that four auditors, interacting according to a multi-step protocol, identified a high rate of errors (one or more errors in 81% of concepts studied) and that results were sufficiently reliable (0.67 to 0.70) for the two most common types of errors. However, reliability was low for each individual auditor, suggesting that review of potential errors is resource-intensive. PMID:18693845

  17. Fast assignment reduction in inconsistent incomplete decision systems

    Institute of Scientific and Technical Information of China (English)

    Min Li; Shaobo Deng; Shengzhong Feng; Jianping Fan

    2014-01-01

    This paper focuses on fast algorithm for computing the assignment reduct in inconsistent incomplete decision systems. It is quite inconvenient to judge the assignment reduct directly ac-cording to its definition. We propose the judgment theorem for the assignment reduct in the inconsistent incomplete decision system, which greatly simplifies judging this type reduct. On such basis, we derive a novel attribute significance measure and construct the fast assignment reduction algorithm (F-ARA), intended for com-puting the assignment reduct in inconsistent incomplete decision systems. Final y, we make a comparison between F-ARA and the discernibility matrix-based method by experiments on 13 Univer-sity of California at Irvine (UCI) datasets, and the experimental results prove that F-ARA is efficient and feasible.

  18. Sequential extraction results in improved proteome profiling of medicinal plant Pinellia ternata tubers, which contain large amounts of high-abundance proteins.

    Directory of Open Access Journals (Sweden)

    Xiaolin Wu

    Full Text Available Pinellia ternata tuber is one of the well-known Chinese traditional medicines. In order to understand the pharmacological properties of tuber proteins, it is necessary to perform proteome analysis of P. ternata tubers. However, a few high-abundance proteins (HAPs, mainly mannose-binding lectin (agglutinin, exist in aggregates of various sizes in the tubers and seriously interfere with proteome profiling by two-dimensional electrophoresis (2-DE. Therefore, selective depletion of these HAPs is a prerequisite for enhanced proteome analysis of P. ternata tubers. Based on differential protein solubility, we developed a novel protocol involving two sequential extractions for depletion of some HAPs and prefractionation of tuber proteins prior to 2-DE. The first extraction using 10% acetic acid selectively extracted acid-soluble HAPs and the second extraction using the SDS-containing buffer extracted remaining acid-insoluble proteins. After application of the protocol, 2-DE profiles of P. ternata tuber proteins were greatly improved and more protein spots were detected, especially low-abundance proteins. Moreover, the subunit composition of P. ternata lectin was analyzed by electrophoresis. Native lectin consists of two hydrogen-bonded subunits (11 kDa and 25 kDa and the 11 kDa subunit was a glycoprotein. Subsequently, major HAPs in the tubers were analyzed by mass spectrometry, with nine protein spots being identified as lectin isoforms. The methodology was easy to perform and required no specialized apparatus. It would be useful for proteome analysis of other tuber plants of Araceae.

  19. Methods of using the quadratic assignment problem solution

    Directory of Open Access Journals (Sweden)

    Izabela Kudelska

    2012-09-01

    Full Text Available Background: Quadratic assignment problem (QAP is one of the most interesting of combinatorial optimization. Was presented by Koopman and Beckamanna in 1957, as a mathematical model of the location of indivisible tasks. This problem belongs to the class NP-hard issues. This forces the application to the solution already approximate methods for tasks with a small size (over 30. Even though it is much harder than other combinatorial optimization problems, it enjoys wide interest because it models the important class of decision problems. Material and methods: The discussion was an artificial intelligence tool that allowed to solve the problem QAP, among others are: genetic algorithms, Tabu Search, Branch and Bound. Results and conclusions: QAP did not arise directly as a model for certain actions, but he found its application in many areas. Examples of applications of the problem is: arrangement of buildings on the campus of the university, layout design of electronic components in systems with large scale integration (VLSI, design a hospital, arrangement of keys on the keyboard.

  20. Cerebellar Synaptic Plasticity and the Credit Assignment Problem.

    Science.gov (United States)

    Jörntell, Henrik

    2016-04-01

    The mechanism by which a learnt synaptic weight change can contribute to learning or adaptation of brain function is a type of credit assignment problem, which is a key issue for many parts of the brain. In the cerebellum, detailed knowledge not only of the local circuitry connectivity but also of the topography of different sources of afferent/external information makes this problem particularly tractable. In addition, multiple forms of synaptic plasticity and their general rules of induction have been identified. In this review, we will discuss the possible roles of synaptic and cellular plasticity at specific locations in contributing to behavioral changes. Focus will be on the parts of the cerebellum that are devoted to limb control, which constitute a large proportion of the cortex and where the knowledge of the external connectivity is particularly well known. From this perspective, a number of sites of synaptic plasticity appear to primarily have the function of balancing the overall level of activity in the cerebellar circuitry, whereas the locations at which synaptic plasticity leads to functional changes in terms of limb control are more limited. Specifically, the postsynaptic forms of long-term potentiation (LTP) and long-term depression (LTD) at the parallel fiber synapses made on interneurons and Purkinje cells, respectively, are the types of plasticity that mediate the widest associative capacity and the tightest link between the synaptic change and the external functions that are to be controlled. PMID:25417189

  1. Yeast Interacting Proteins Database: YJL199C, YJL199C [Yeast Interacting Proteins Database

    Lifescience Database Archive (English)

    Full Text Available d in closely related Saccharomyces species; protein detected in large-scale protein-protein interaction studies...cies; protein detected in large-scale protein-protein interaction studies Rows with this prey as prey (4) Ro...n; not conserved in closely related Saccharomyces species; protein detected in large-scale protein-protein interaction studies... species; protein detected in large-scale protein-protein interaction studies Rows with this prey as prey Ro

  2. Assignment of congested NMR spectra: Carbonyl backbone enrichment via the Entner Doudoroff pathway

    Science.gov (United States)

    Goldbourt, Amir; Day, Loren A.; McDermott, Ann E.

    2007-12-01

    In NMR spectra of complex proteins, sparse isotope enrichment can be important, in that the removal of many 13C- 13C homonuclear J-couplings can narrow the lines and thereby facilitate the process of spectral assignment and structure elucidation. We present a simple scheme for selective yet extensive isotopic enrichment applicable for production of proteins in organisms utilizing the Entner-Doudoroff (ED) metabolic pathway. An enrichment scheme so derived is demonstrated in the context of a magic-angle spinning solid-state NMR (MAS SSNMR) study of Pf1 bacteriophage, the host of which is Pseudomonas aeruginosa, strain K (PAK), an organism that uses the ED pathway for glucose catabolism. The intact and infectious Pf1 phage in this study was produced by infected PAK cells grown on a minimal medium containing 1- 13C D-glucose ( 13C in position 1) as the sole carbon source, as well as 15NH 4Cl as the only nitrogen source. The 37 MDa Pf1 phage consists of about 93% major coat protein, 1% minor coat proteins, and 6% single-stranded, circular DNA. As a consequence of this composition and the enrichment scheme, the resonances in the MAS SSNMR spectra of the Pf1 sample were almost exclusively due to carbonyl carbons in the major coat protein. Moreover, 3D heteronuclear NCOCX correlation experiments also show that the amino acids leucine, serine, glycine, and tyrosine were not isotopically enriched in their carbonyl positions (although most other amino acids were), which is as expected based upon considerations of the ED metabolic pathway. 3D NCOCX NMR data and 2D 15N- 15N data provided strong verification of many previous assignments of 15N amide and 13C carbonyl shifts in this highly congested spectrum; both the semi-selective enrichment patterns and the narrowed linewidths allowed for greater certainty in the assignments as compared with use of uniformly enriched samples alone.

  3. An economical method for production of (2H, (13CH3-threonine for solution NMR studies of large protein complexes: application to the 670 kDa proteasome.

    Directory of Open Access Journals (Sweden)

    Algirdas Velyvis

    Full Text Available NMR studies of very high molecular weight protein complexes have been greatly facilitated through the development of labeling strategies whereby (13CH(3 methyl groups are introduced into highly deuterated proteins. Robust and cost-effective labeling methods are well established for all methyl containing amino acids with the exception of Thr. Here we describe an inexpensive biosynthetic strategy for the production of L-[α-(2H; β-(2H;γ-(13C]-Thr that can then be directly added during protein expression to produce highly deuterated proteins with Thr methyl group probes of structure and dynamics. These reporters are particularly valuable, because unlike other methyl containing amino acids, Thr residues are localized predominantly to the surfaces of proteins, have unique hydrogen bonding capabilities, have a higher propensity to be found at protein nucleic acid interfaces and can play important roles in signaling pathways through phosphorylation. The utility of the labeling methodology is demonstrated with an application to the 670 kDa proteasome core particle, where high quality Thr (13C,(1H correlation spectra are obtained that could not be generated from samples prepared with commercially available U-[(13C,(1H]-Thr.

  4. Heuristic for Task-Worker Assignment with Varying Learning Slopes

    Directory of Open Access Journals (Sweden)

    Wipawee Tharmmaphornphilas

    2010-04-01

    Full Text Available Fashion industry has variety products, so the multi-skilled workers are required to improve flexibility in production and assignment. Generally the supervisor will assign task to the workers based on skill and skill levels of worker. Since in fashion industry new product styles are launched more frequently and the order size tends to be smaller, the workers always learn when the raw material and the production process changes. Consequently they require less time to produce the succeeding units of a task based on their learning ability. Since the workers have both experience and inexperience workers, so each worker has different skill level and learning ability. Consequently, the assignment which assumed constant skill level is not proper to use. This paper proposes a task-worker assignment considering worker skill levels and learning abilities. Processing time of each worker changes along production period due to a worker learning ability. We focus on a task-worker assignment in a fashion industry where tasks are ordered in series; the number of tasks is greater than the number of workers. Therefore, workers can perform multiple assignments followed the precedence restriction as an assembly line balancing problem. The problem is formulated in an integer linear programming model with objective to minimize makespan. A heuristic is proposed to determine the lower bound (LB and the upper bound (UB of the problem and the best assignment is determined. The performance of the heuristic method is tested by comparing quality of solution and computational time to optimal solutions.

  5. A New Approach to Pointer Analysis for Assignments

    Institute of Scientific and Technical Information of China (English)

    HUANG Bo; ZANG Binyu; LI Jing; ZHU Chuanqi

    2001-01-01

    Pointer analysis is a technique to identify at compile-time the po tential values of the pointer expressions in a program, which promises significant benefits for optimizing and parallelizing compilers. In this paper, a new approach to pointer analysis for assignments is presented. In this approach, assignments are clas sified into three categories: pointer assignments, structure (union) assignments and normal assignments which don't affect the point-to information. Pointer analyses for these three kinds of assignments respectively make up the integrated algorithm. When analyzing a pointer assignment, a new method called expression expansion is used to calculate both the left targets and the right targets. The integration of recursive data structure analysis into pointer analysis is a significant originality of this paper, which uniforms the pointer analysis for heap variables and the pointer analysis for stack variables. This algorithm is implemented in Agassiz, an analyzing tool for C programs developed by Institute of Parallel Processing, Fudan University. Its accuracy and effectiveness are illustrated by experimental data.

  6. Clonify: unseeded antibody lineage assignment from next-generation sequencing data.

    Science.gov (United States)

    Briney, Bryan; Le, Khoa; Zhu, Jiang; Burton, Dennis R

    2016-01-01

    Defining the dynamics and maturation processes of antibody clonal lineages is crucial to understanding the humoral response to infection and immunization. Although individual antibody lineages have been previously analyzed in isolation, these studies provide only a narrow view of the total antibody response. Comprehensive study of antibody lineages has been limited by the lack of an accurate clonal lineage assignment algorithm capable of operating on next-generation sequencing datasets. To address this shortcoming, we developed Clonify, which is able to perform unseeded lineage assignment on very large sets of antibody sequences. Application of Clonify to IgG+ memory repertoires from healthy individuals revealed a surprising lack of influence of large extended lineages on the overall repertoire composition, indicating that this composition is driven less by the order and frequency of pathogen encounters than previously thought. Clonify is freely available at www.github.com/briney/clonify-python. PMID:27102563

  7. Clonify: unseeded antibody lineage assignment from next-generation sequencing data.

    Science.gov (United States)

    Briney, Bryan; Le, Khoa; Zhu, Jiang; Burton, Dennis R

    2016-04-22

    Defining the dynamics and maturation processes of antibody clonal lineages is crucial to understanding the humoral response to infection and immunization. Although individual antibody lineages have been previously analyzed in isolation, these studies provide only a narrow view of the total antibody response. Comprehensive study of antibody lineages has been limited by the lack of an accurate clonal lineage assignment algorithm capable of operating on next-generation sequencing datasets. To address this shortcoming, we developed Clonify, which is able to perform unseeded lineage assignment on very large sets of antibody sequences. Application of Clonify to IgG+ memory repertoires from healthy individuals revealed a surprising lack of influence of large extended lineages on the overall repertoire composition, indicating that this composition is driven less by the order and frequency of pathogen encounters than previously thought. Clonify is freely available at www.github.com/briney/clonify-python.

  8. Linear assignment maps for correlated system-environment states

    Energy Technology Data Exchange (ETDEWEB)

    Rodriguez-Rosario, Cesar A.; Modi, Kavan; Aspuru-Guzik, Alan

    2010-01-20

    Assignment maps are mathematical operators that describe initial system-environment states for open quantum systems. We re-examine the notion of assignments that account for correlations between the system and the environment and show that these maps can be made linear at the expense of giving up positivity or consistency of the map. We study the role of positivity and consistency of the map and show the effects of relaxing these. Finally, we establish a connection between the violation of the positivity of linear assignments and the no-broadcasting theorem.

  9. Optimal assignment of incoming flights to baggage carousels at airports

    DEFF Research Database (Denmark)

    Barth, Torben C.

    The problem considered in this report is an assignment problem occurring at airports. This problem concerns the assignment of baggage carousels in baggage claim halls to arriving aircraft (baggage carousel assignment problem). This is a highly dynamic problem since disruptions frequently occur du...... and in general is a substantial support in decision making....... during operations. We introduce a basic static model that can be adapted to the layout of different airports. Afterwards we show how a decision support system based on a MIP-model can be designed in a dynamic real world environment. The system supports the decisions of the dispatcher during daily...

  10. Localized Algorithm for Channel Assignment in Cognitive Radio Networks

    Directory of Open Access Journals (Sweden)

    Harpreet Kaur

    2015-12-01

    Full Text Available Cognitive Radio has been emerged as a revolutionary solution to migrate the current shortage of spectrum allocation in wireless networks. In this paper, an improved localized channel allocation algorithm based on channel weight is proposed. A factor of channel stability is introduced based on link environment, which efficiently assigns the best channels to the links. Based on the framework, a conflict resolution strategy is used to make the scheme adaptable to different network conditions. Calculations indicate that this algorithm can reduce the conflicts, increase the delivery rate and link assignment rate compared with the basic channel assignment algorithm.

  11. Application of k-person and k-task maximal efficiency assignment algorithm to water piping repair

    Institute of Scientific and Technical Information of China (English)

    Su-juan ZHENG; Xiu-ming YU; Li-qing CAO

    2009-01-01

    Solving the absent assignment problern of the shortest time limit in a weighted bipartite graph with the minimal weighted k-matching algorithm is unsuitable for situations in which large numbers of problems need to be addressed by large numbers of parties. This paper simplifies the algorithm of searching for the even alternating path that contains a maximal element using the minimal weighted k-matching theorem and intercept graph. A program for solving the maximal efficiency assignment problem was compiled. As a case study, the program was used to solve the assignment problem of water piping repair in the case of a large number of companies and broken pipes, and the validity of the program was verified.

  12. Robust Pole Assignment of Digital Control System with Output Multirate Sampled

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Multirate digital control system is a periodically time-variant (PTV)system in its essence. It has many “super capability”, such as obtaining arbitrarily-large gain-margin,simultaneous stabilization, strong stabilization, decentralized control, etc. Utilizing freedom aroused from the multirate sampling of system output, this paper assigns poles of the closedloop system robustly, and so improves the resistance of the system to perturbation.

  13. Site-specific protein O-glycosylation modulates proprotein processing - Deciphering specific functions of the large polypeptide GalNAc-transferase gene family

    DEFF Research Database (Denmark)

    Schjoldager, Katrine Ter-Borch Gram; Clausen, Henrik

    2012-01-01

    Posttranslational modifications (PTMs) greatly expand the function and regulation of proteins, and glycosylation is the most abundant and diverse PTM. Of the many different types of protein glycosylation, one is quite unique; GalNAc-type (or mucin-type) O-glycosylation, where biosynthesis...... is initiated in the Golgi by up to twenty distinct UDP-N-acetyl-a-d-galactosamine:polypeptide N-acetylgalactosaminyltransferases (GalNAc-Ts). These GalNAc-Ts are differentially expressed in cells and have different (although partly overlapping) substrate specificities, which provide for both unique functions...... and considerable redundancy. Recently we have begun to uncover human diseases associated with deficiencies in GalNAc-T genes (GALNTs). Thus deficiencies in individual GALNTs produce cell and protein specific effects and subtle distinct phenotypes such as hyperphosphatemia with hyperostosis (GALNT3...

  14. PseAAC-General: Fast Building Various Modes of General Form of Chou’s Pseudo-Amino Acid Composition for Large-Scale Protein Datasets

    Directory of Open Access Journals (Sweden)

    Pufeng Du

    2014-02-01

    Full Text Available The general form pseudo-amino acid composition (PseAAC has been widely used to represent protein sequences in predicting protein structural and functional attributes. We developed the program PseAAC-General to generate various different modes of Chou’s general PseAAC, such as the gene ontology mode, the functional domain mode, and the sequential evolution mode. This program allows the users to define their own desired modes. In every mode, 544 physicochemical properties of the amino acids are available for choosing. The computing efficiency is at least 100 times that of existing programs, which makes it able to facilitate the extensive studies on proteins and peptides. The PseAAC-General is freely available via SourceForge. It runs on both Linux and Windows.

  15. Using Computer-Assisted Personalized Assignment System in a Large-Enrollment General Physics

    Science.gov (United States)

    Gok, Tolga

    2010-01-01

    The on-line tutoring system, LON-CAPA, was implemented in Introductory Calculus-Based Physics-II course at Colorado School of Mines in fall 2008 and spring 2009. In this paper, the features and the case study of the LON-CAPA implementation were described. The performance data obtained from the scores of students enrolled in the course represented…

  16. Effect of long-term feeding with a plant protein mixture based diet on growth and body/fillet quality traits of large rainbow trout (Oncorhynchus mykiss)

    OpenAIRE

    De Francesco, Matilde; Parisi, Giuliana; Medale, Francoise; Lupi, Paola; Kaushik, Sadasivam; Poli, Bianca

    2004-01-01

    Two diets based oil fish meal (FM) or oil a mixture of plant protein sources supplemented with free amino acids (PP) were fed to rainbow trout having a mean initial body weight of 162.5 g reared at 17 +/- 1 degreesC for 24 weeks to evaluate the effect of plant protein sources oil fish growth, morphological and body quality traits. At the end of the growth study, samples were withdrawn for chemical analyses as well as measurements of different parameters of quality. Compared to the FM gro...

  17. Sequential backbone assignment based on dipolar amide-to-amide correlation experiments

    Energy Technology Data Exchange (ETDEWEB)

    Xiang, ShengQi; Grohe, Kristof; Rovó, Petra; Vasa, Suresh Kumar; Giller, Karin; Becker, Stefan; Linser, Rasmus, E-mail: rali@nmr.mpibpc.mpg.de [Max Planck Institute for Biophysical Chemistry, Department for NMR-Based Structural Biology (Germany)

    2015-07-15

    Proton detection in solid-state NMR has seen a tremendous increase in popularity in the last years. New experimental techniques allow to exploit protons as an additional source of information on structure, dynamics, and protein interactions with their surroundings. In addition, sensitivity is mostly improved and ambiguity in assignment experiments reduced. We show here that, in the solid state, sequential amide-to-amide correlations turn out to be an excellent, complementary way to exploit amide shifts for unambiguous backbone assignment. For a general assessment, we compare amide-to-amide experiments with the more common {sup 13}C-shift-based methods. Exploiting efficient CP magnetization transfers rather than less efficient INEPT periods, our results suggest that the approach is very feasible for solid-state NMR.

  18. Optimization of Fleet Assignment: A Case Study in Turkey

    Directory of Open Access Journals (Sweden)

    Yavuz OZDEMIR

    2012-01-01

    Full Text Available Since poor fleet assignment can cause a great increase in costs for airline companies, asolution of the type ‘right fleet for the right flight’ would be very useful. In this paper, a fleetassignment model is set up using the data of the largest Airline Company in Turkey, TurkishAirlines. The aim of this model is to assign the most appropriate fleet type to flights whileminimizing the cost and determining the optimal number of aircraft grounded overnight at eachairport. We set up a model with constraints with thinking all airline operations and solve ourproblem using integer linear programming. Finally, we get an optimum solution which minimizesthe total cost while assigning the fleet type to the flight leg. Using optimization software (Lindo6.1, the solution to this problem generates a minimum daily cost of fleet assignment.

  19. 28 CFR 544.74 - Work assignment limitations.

    Science.gov (United States)

    2010-07-01

    ... appointment and promotion apply to all inmates, including those exempted from required participation in the... of the assignment. Local Federal Prison Industry (FPI) management may elect to retain the...

  20. 34 CFR 682.508 - Assignment of a loan.

    Science.gov (United States)

    2010-07-01

    ... FISL Program loan is to result in a change in the identity of the party to whom the borrower must send..., provide separate notices to the borrower of— (i) The assignment; (ii) The identity of the assignee;...